50 results on '"Dehbi M"'
Search Results
2. Multiplexed analysis of inflammatory, metabolic and stress markers in obese subjects before and after a defined exercise program: P042
- Author
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Baturcam, E., Abubakr, J., Abu-Farha, M., Al-Arouj, M., Al-Ghimlas, F., Al-Khairi, I., Al-Mass, A., Al-Mudhaf, D., Bennakhi, A., Cherian, P., Hammad, M., John, J., Kavalakatt, S., Khadir, A., Tiss, A., Warsame, S., Dermime, S., and Dehbi, M.
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- 2012
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3. Genome annotation and intraviral interactome for the streptococcus pneumoniae virulent phage Dp-1
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Sabri, M., Häuser, R., Ouellette, M., Liu, J., Dehbi, M., Moeck, G., García, Ernesto, Titz, B., Uetz, P., Moineau, S., Sabri, M., Häuser, R., Ouellette, M., Liu, J., Dehbi, M., Moeck, G., García, Ernesto, Titz, B., Uetz, P., and Moineau, S.
- Abstract
Streptococcus pneumoniae causes several diseases, including pneumonia, septicemia, and meningitis. Phage Dp-1 is one of the very few isolated virulent S. pneumoniae bacteriophages, but only a partial characterization is currently available. Here, we confirmed that Dp-1 belongs to the family Siphoviridae. Then, we determined its complete genomic sequence of 56,506 bp. It encodes 72 open reading frames, of which 44 have been assigned a function. We have identified putative promoters, Rho-independent terminators, and several genomic clusters. We provide evidence that Dp-1 may be using a novel DNA replication system as well as redirecting host protein synthesis through queuosine-containing tRNAs. Liquid chromatography-mass spectrometry analysis of purified phage Dp-1 particles identified at least eight structural proteins. Finally, using comprehensive yeast two-hybrid screens, we identified 156 phage protein interactions, and this intraviral interactome was used to propose a structural model of Dp-1.
- Published
- 2011
4. Designing a gestural interface for Mentor robot control
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Dehbi, M., primary and Ahmed Foitih, Z., additional
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- 2013
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5. P042 Multiplexed analysis of inflammatory, metabolic and stress markers in obese subjects before and after a defined exercise program
- Author
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Baturcam, E., primary, Abubakr, J., additional, Abu-Farha, M., additional, Al-Arouj, M., additional, Al-Ghimlas, F., additional, Al-Khairi, I., additional, Al-Mass, A., additional, Al-Mudhaf, D., additional, Bennakhi, A., additional, Cherian, P., additional, Hammad, M., additional, John, J., additional, Kavalakatt, S., additional, Khadir, A., additional, Tiss, A., additional, Warsame, S., additional, Dermime, S., additional, and Dehbi, M., additional
- Published
- 2012
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6. Overlapping DNA recognition motifs between Sp1 and a novel trans-acting factor within the wt1 tumour suppressor gene promoter
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Teresa Discenza, M., primary, Dehbi, M., additional, and Pelletier, J., additional
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- 1997
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7. PAX8-mediated activation of the wt1 tumor suppressor gene.
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Dehbi, M., primary and Pelletier, J., additional
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- 1996
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8. Biosynthesis and polarized distribution of neutral endopeptidase in primary cultures of kidney proximal tubule cells
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Jalal, F, primary, Dehbi, M, additional, Berteloot, A, additional, and Crine, P, additional
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- 1994
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9. Transcriptional activation of the CEF-4/9E3 cytokine gene by pp60v-src
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Dehbi, M, primary, Mbiguino, A, additional, Beauchemin, M, additional, Chatelain, G, additional, and Bédard, P A, additional
- Published
- 1992
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10. Tissue factor/factor VIIa pathway mediates coagulation activation in induced-heat stroke in the baboon.
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Bouchama A, Al-Mohanna F, Assad L, Baturcam E, Eldali A, Owaidah T, and Dehbi M
- Published
- 2012
11. Prognostic factors in heat wave-related deaths: a meta-analysis.
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Bouchama A, Dehbi M, Mohamed G, Matthies F, Shoukri M, and Menne B
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- 2007
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12. Association of obesity with down-regulation of heat shock protein 40 expression and evidence that exercise retrieves its normal expression
- Author
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Abu-Farha Mohamed, Abubakr Jehad, Kavalakatt Sina, Khadir Abdelkrim, Al-Arouj Monira, Al-Ghimlas Fahad, Al-Khairi Irina, Al-Mudhaf Dalal, Baturcam Engin, Bennakhi Abdullah, Cherian Preethi, Hammad Maha, John Jeena, Tiss Ali, Warsame Samia, Dermime Said, and Dehbi Mohammed
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Medicine ,Science - Published
- 2012
- Full Text
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13. Genetic Deletion of DNAJB3 Using CRISPR-Cas9, Produced Discordant Phenotypes.
- Author
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Nejat S, Menikdiwela KR, Efotte A, Scoggin S, Vandanmagsar B, Thornalley PJ, Dehbi M, and Moustaid-Moussa N
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- Animals, Female, Male, Mice, Body Weight genetics, CRISPR-Cas Systems genetics, Diet, High-Fat adverse effects, Glucose metabolism, HSP40 Heat-Shock Proteins genetics, HSP40 Heat-Shock Proteins metabolism, Insulin genetics, Insulin metabolism, Mice, Knockout, Obesity genetics, Obesity metabolism, Phenotype, RNA, Messenger, Diabetes Mellitus, Type 2 genetics, Diabetes Mellitus, Type 2 metabolism, Insulin Resistance genetics
- Abstract
Several pathways and/or genes have been shown to be dysregulated in obesity-induced insulin resistance (IR) and type 2 diabetes (T2D). We previously showed, for the first time, impaired expression of DNAJB3 mRNA and protein in subjects with obesity, which was concomitant with increased metabolic stress. Restoring the normal expression of DNAJB3 attenuated metabolic stress and improved insulin signaling both in vivo and in vitro, suggesting a protective role of DNAJB3 against obesity and T2D. The precise underlying mechanisms remained, however, unclear. This study was designed to confirm the human studies in a mouse model of dietary obesity-induced insulin resistance, and, if validated, to understand the underlying mechanisms. We hypothesized that mice lacking DNAJB3 would be more prone to high-fat (HF)-diet-induced increase in body weight and body fat, inflammation, glucose intolerance and insulin resistance as compared with wild-type (WT) littermates. Three DNAJB3 knockout (KO) lines were generated (KO 30, 44 and 47), using CRISPR-Cas9. Male and female KO and WT mice were fed a HF diet (45% kcal fat) for 16 weeks. Body weight was measured biweekly, and a glucose tolerance test (GTT) and insulin tolerance test (ITT) were conducted at week 13 and 14, respectively. Body composition was determined monthly by nuclear magnetic resonance (NMR). Following euthanasia, white adipose tissue (WAT) and skeletal muscle were harvested for further analyses. Compared with WT mice, male and female KO 47 mice demonstrated higher body weight and fat mass. Similarly, KO 47 mice also showed a slower rate of glucose clearance in GTT that was consistent with decreased mRNA expression of the GLUT4 gene in WAT but not in the muscle. Both male and female KO 47 mice exhibited higher mRNA levels of the pro-inflammatory marker TNF-a in WAT only, whereas increased mRNA levels of MCP1 chemokine and the ER stress marker BiP/Grp78 were observed in male but not in female KO 47 mice. However, we did not observe the same changes in the other KO lines. Taken together, the phenotype of the DNAJB3 KO 47 mice was consistent with the metabolic changes and low levels of DNAJB3 reported in human subjects. These findings suggest that DNAJB3 may play an important role in metabolic functions and glucose homeostasis, which warrants further phenotyping and intervention studies in other KO 47 and other KO mice, as well as investigating this protein as a potential therapeutic target for obesity and T2D.
- Published
- 2023
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14. DNAJB3 attenuates ER stress through direct interaction with AKT.
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Islam Z, Diane A, Khattab N, Dehbi M, Thornalley P, and Kolatkar PR
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- Humans, I-kappa B Kinase, Proto-Oncogene Proteins c-akt, Protein Serine-Threonine Kinases, Biological Transport, Heat-Shock Proteins, HSP40 Heat-Shock Proteins, Diabetes Mellitus, Type 2, Insulin Resistance
- Abstract
Metabolic stress involved in several dysregulation disorders such as type 2 diabetes mellitus (T2DM) results in down regulation of several heat shock proteins (HSPs) including DNAJB3. This down regulation of HSPs is associated with insulin resistance (IR) and interventions which induce the heat shock response (HSR) help to increase the insulin sensitivity. Metabolic stress leads to changes in signaling pathways through increased activation of both c-jun N-terminal kinase-1 (JNK1) and the inhibitor of κB inflammatory kinase (IKKβ) which in turn leads to inactivation of insulin receptor substrates 1 and 2 (IRS-1 and IRS-2). DNAJB3 interacts with both JNK1 and IKKβ kinases to mitigate metabolic stress. In addition DNAJB3 also activates the PI3K-PKB/AKT pathway through increased phosphorylation of AKT1 and its substrate AS160, a Rab GTPase-activating protein, which results in mobilization of GLUT4 transporter protein and improved glucose uptake. We show through pull down that AK T1 is an interacting partner of DNAJB3, further confirmed by isothermal titration calorimetry (ITC) which quantified the avidity of AKT1 for DNAJB3. The binding interface was identified by combining protein modelling with docking of the AKT1-DNAJB3 complex. DNAJB3 is localized in the cytoplasm and ER, where it interacts directly with AKT1 and mobilizes AS160 for glucose transport. Inhibition of AKT1 resulted in loss of GLUT4 translocation activity mediated by DNAJB3 and also abolished the protective effect of DNAJB3 on tunicamycin-induced ER stress. Taken together, our findings provide evidence for a direct protein-protein interaction between DNAJB3 and AKT1 upon which DNAJB3 alleviates ER stress and promotes GLUT4 translocation., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Islam et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)
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- 2023
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15. Role of the DNAJ/HSP40 family in the pathogenesis of insulin resistance and type 2 diabetes.
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Diane A, Abunada H, Khattab N, Moin ASM, Butler AE, and Dehbi M
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- Animals, Glucose, Humans, Insulin, Obesity, Diabetes Mellitus, Type 2, Insulin Resistance
- Abstract
Insulin resistance (IR) underpins a wide range of metabolic disorders including type 2 diabetes (T2D), metabolic syndrome and cardiovascular diseases. IR is characterized by a marked reduction in the magnitude and/or delayed onset of insulin to stimulate glucose disposal. This condition is due to defects in one or several intracellular intermediates of the insulin signaling cascade, ranging from insulin receptor substrate (IRS) inactivation to reduced glucose phosphorylation and oxidation. Genetic predisposition, as well as other precipitating factors such as aging, obesity, and sedentary lifestyles are among the risk factors underlying the pathogenesis of IR and its subsequent progression to T2D. One of the cardinal hallmarks of T2D is the impairment of the heat shock response (HSR). Human and animal studies provided compelling evidence of reduced expression of several components of the HSR (i.e. Heat shock proteins or HSPs) in diabetic samples in a manner that correlates with the degree of IR. Interventions that induce the HSR, irrespective of the means to achieve it, proved their effectiveness in enhancing insulin sensitivity and improving glycemic index. However, most of these studies have been focused on HSP70 family. In this review, we will focus on the novel role of DNAJ/HSP40 cochaperone family in metabolic diseases associated with IR., (Copyright © 2021 Elsevier B.V. All rights reserved.)
- Published
- 2021
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16. The Role of Heat Shock Proteins in Type 1 Diabetes.
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Moin ASM, Nandakumar M, Diane A, Dehbi M, and Butler AE
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- Animals, Autoantigens immunology, Autoimmune Diseases immunology, Humans, T-Lymphocytes immunology, Diabetes Mellitus, Type 1 immunology, Heat-Shock Proteins immunology
- Abstract
Type 1 diabetes (T1D) is a T-cell mediated autoimmune disease characterized by recognition of pancreatic β-cell proteins as self-antigens, called autoantigens (AAgs), followed by loss of pancreatic β-cells. (Pre-)proinsulin ([P]PI), glutamic acid decarboxylase (GAD), tyrosine phosphatase IA-2, and the zinc transporter ZnT8 are key molecules in T1D pathogenesis and are recognized by autoantibodies detected in routine clinical laboratory assays. However, generation of new autoantigens (neoantigens) from β-cells has also been reported, against which the autoreactive T cells show activity. Heat shock proteins (HSPs) were originally described as "cellular stress responders" for their role as chaperones that regulate the conformation and function of a large number of cellular proteins to protect the body from stress. HSPs participate in key cellular functions under both physiological and stressful conditions, including suppression of protein aggregation, assisting folding and stability of nascent and damaged proteins, translocation of proteins into cellular compartments and targeting irreversibly damaged proteins for degradation. Low HSP expression impacts many pathological conditions associated with diabetes and could play a role in diabetic complications. HSPs have beneficial effects in preventing insulin resistance and hyperglycemia in type 2 diabetes (T2D). HSPs are, however, additionally involved in antigen presentation, presenting immunogenic peptides to class I and class II major histocompatibility molecules; thus, an opportunity exists for HSPs to be employed as modulators of immunologic responses in T1D and other autoimmune disorders. In this review, we discuss the multifaceted roles of HSPs in the pathogenesis of T1D and in autoantigen-specific immune protection against T1D development., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Moin, Nandakumar, Diane, Dehbi and Butler.)
- Published
- 2021
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17. Alpha lipoic acid attenuates ER stress and improves glucose uptake through DNAJB3 cochaperone.
- Author
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Diane A, Mahmoud N, Bensmail I, Khattab N, Abunada HA, and Dehbi M
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- Animals, Biomarkers metabolism, Endoplasmic Reticulum Chaperone BiP, Gene Silencing drug effects, Hep G2 Cells, Humans, Mice, Mitochondria drug effects, Mitochondria metabolism, Molecular Chaperones metabolism, Oxidative Stress drug effects, RNA, Small Interfering metabolism, Tunicamycin pharmacology, Endoplasmic Reticulum Stress drug effects, Glucose metabolism, HSP40 Heat-Shock Proteins metabolism, Thioctic Acid pharmacology
- Abstract
Persistent ER stress, mitochondrial dysfunction and failure of the heat shock response (HSR) are fundamental hallmarks of insulin resistance (IR); one of the early core metabolic aberrations that leads to type 2 diabetes (T2D). The antioxidant α-lipoic acid (ALA) has been shown to attenuate metabolic stress and improve insulin sensitivity in part through activation of the heat shock response (HSR). However, these studies have been focused on a subset of heat shock proteins (HSPs). In the current investigation, we assessed whether ALA has an effect on modulating the expression of DNAJB3/HSP40 cochaperone; a potential therapeutic target with a novel role in mitigating metabolic stress and promoting insulin signaling. Treatment of C2C12 cells with 0.3 mM of ALA triggers a significant increase in the expression of DNAJB3 mRNA and protein. A similar increase in DNAJB3 mRNA was also observed in HepG2 cells. We next investigated the significance of such activation on endoplasmic reticulum (ER) stress and glucose uptake. ALA pre-treatment significantly reduced the expression of ER stress markers namely, GRP78, XBP1, sXBP1 and ATF4 in response to tunicamycin. In functional assays, ALA treatment abrogated significantly the tunicamycin-mediated transcriptional activation of ATF6 while it enhanced the insulin-stimulated glucose uptake and Glut4 translocation. Silencing the expression of DNAJB3 but not HSP72 abolished the protective effect of ALA on tunicamycin-induced ER stress, suggesting thus that DNAJB3 is a key mediator of ALA-alleviated tunicamycin-induced ER stress. Furthermore, the effect of ALA on insulin-stimulated glucose uptake is significantly reduced in C2C12 and HepG2 cells transfected with DNAJB3 siRNA. In summary, our results are supportive of an essential role of DNAJB3 as a molecular target through which ALA alleviates ER stress and improves glucose uptake.
- Published
- 2020
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18. The GLP-1 analog exendin-4 modulates HSP72 expression and ERK1/2 activity in BTC6 mouse pancreatic cells.
- Author
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Madhu D, Khadir A, Hammad M, Kavalakatt S, Dehbi M, Al-Mulla F, Abubaker J, and Tiss A
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- Animals, Apoptosis drug effects, Cell Line, Cell Proliferation drug effects, Cell Survival drug effects, Diabetes Mellitus, Type 2 metabolism, Endoplasmic Reticulum drug effects, Endoplasmic Reticulum metabolism, Exenatide pharmacology, HSP40 Heat-Shock Proteins, Membrane Glycoproteins, Mice, Molecular Chaperones, Phosphorylation, Protective Agents pharmacology, Protein Interaction Maps, Up-Regulation, Exenatide metabolism, Glucagon-Like Peptide 1 analogs & derivatives, HSP72 Heat-Shock Proteins metabolism, Insulin-Secreting Cells metabolism, MAP Kinase Signaling System physiology
- Abstract
Lipotoxicity, an important factor in the pathogenesis of diabetes, leads to defective β-cell proliferation and increased apoptosis. Glucagon-like peptide-1 (GLP-1) analogs, which are used to treat type 2 diabetes, reduce endoplasmic reticulum stress and inflammation in pancreatic β-cells and improve their survival. However, their effects on the heat shock response (HSR) have not been elucidated yet. We investigated whether the GLP-1 analog exendin-4 exerts its protective effect by modulating the HSR and mitogen-activated protein kinases (MAPKs) in BTC-6 mouse pancreatic cells under palmitic acid (PA) stress. Expression patterns were analyzed using mass spectrometry, Western blotting, and qRT-PCR in the presence of 250 or 400 μM PA and 100 nM exendin-4. Additionally, we measured MAPK expression and phosphorylation using qRT-PCR and Western blotting, respectively. Upregulation of heat shock protein (HSP), notably HSP72, in the presence of PA, was attenuated by exendin-4. Despite the absence of global effects on the HSR system, exendin-4 attenuated the expression of other non-classical HSPs (GRP94, DNAJA1, and DNAJB6) in the presence of PA. Regarding MAPKs, only extracellular signal-regulated kinase (ERK) phosphorylation was highly increased by exendin-4 in both the presence and absence of PA. Furthermore, exendin-4 significantly alleviated PA-induced cell death, which was further confirmed with proteomics analysis where key cellular functions, including cellular growth, assembly, and organization, were improved by exendin-4 treatment. Thus, our results expand the protective role of GLP-1 analogs to include other cellular mechanisms involved in restoring normal β-cell homeostasis., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)
- Published
- 2020
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19. DNAJB3 attenuates metabolic stress and promotes glucose uptake by eliciting Glut4 translocation.
- Author
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Arredouani A, Diane A, Khattab N, Bensmail I, Aoude I, Chikri M, Mohammad R, Abou-Samra AB, and Dehbi M
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- 3T3 Cells, Animals, Cell Line, Tumor, HEK293 Cells, HSP40 Heat-Shock Proteins genetics, HSP70 Heat-Shock Proteins genetics, Hep G2 Cells, Humans, I-kappa B Kinase metabolism, Mice, Mitogen-Activated Protein Kinase 8 metabolism, RNA Interference, RNA, Small Interfering genetics, Stress, Physiological physiology, Transcription Factor RelA metabolism, Tumor Necrosis Factor-alpha metabolism, Active Transport, Cell Nucleus physiology, Diabetes Mellitus, Type 2 pathology, Glucose metabolism, Glucose Transporter Type 4 metabolism, HSP40 Heat-Shock Proteins metabolism
- Abstract
Failure of the heat shock response is a key event that leads to insulin resistance and type 2 diabetes. We recently showed that DNAJB3 co-chaperone is downregulated in obese and diabetic patients and that physical exercise restores its normal expression with a significant improvement of the clinical outcomes. In 3T3-L1 adipocytes, DNAJB3 has a role in improving the sensitivity to insulin and glucose uptake. In co-immunoprecipitation assays, DNAJB3 interacts with both JNK1 and IKKβ kinases. However, the functional impact of such interaction on their activities has not been investigated. Here, we assessed the effect of DNAJB3 on the respective activity of JNK1 and IKKβ in cell-based assays. Using JNK1- and IKKβ-dependent luciferase reporters, we show a marked decrease in luciferase activity by DNAJB3 in response to PMA and TNF-α that was consistent with a decrease in the translocation of p65/NF-κB to the nucleus in response to LPS. Furthermore, TNF-α-mediated IL-6 promoter activation and endogenous mRNA expression are significantly abrogated by DNAJB3 both in 3T3-L1 and C2C12 cells. The ability of DNAJB3 to mitigate ER stress and oxidative stress was also investigated and our data show a significant improvement of both forms of stress. Finally, we examined the effect of overexpressing and knocking down the expression of DNAJB3 on glucose uptake in C2C12 as well as the molecular determinants. Accordingly, we provide evidence for a role of DNAJB3 in promoting both basal and insulin-stimulated glucose uptake. Our finding reveals also a novel role of DNAJB3 in eliciting Glut4 translocation to the plasma membrane. These results suggest a physiological role of DNAJB3 in mitigating metabolic stress and improving glucose homeostasis and could therefore represent a novel therapeutic target for type 2 diabetes.
- Published
- 2019
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20. DUSP1 Is a Potential Marker of Chronic Inflammation in Arabs with Cardiovascular Diseases.
- Author
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Khadir A, Kavalakatt S, Dehbi M, Alarouj M, Bennakhi A, Tiss A, and Elkum N
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- Adult, Aged, Arabs, C-Reactive Protein metabolism, Cardiovascular Diseases blood, Case-Control Studies, Female, Humans, Kuwait, Lipoproteins, LDL blood, Logistic Models, Male, Middle Aged, Biomarkers blood, Cardiovascular Diseases metabolism, Dual Specificity Phosphatase 1 blood
- Abstract
Background: Cardiovascular disease (CVD) risks persist in patients despite the use of conventional treatments. This might be due to chronic inflammation as reflected in epidemiological studies associating circulating low-grade inflammatory markers with CVD recurrent events. Here, we explored this potential link by assessing plasma dual-specificity phosphatase 1 (DUSP1) levels and comparing them to high-sensitivity CRP (hsCRP) and oxidized low-density lipoprotein (oxLDL) levels and their associations to conventional CVD risk factors in confirmed CVD patients., Methods: Human adults with reported CVD ( n = 207) and controls ( n = 70) living in Kuwait were used in this study. Anthropometric and classical biochemical parameters were determined. Plasma levels of DUSP1, oxLDL, and hsCRP were measured using human enzyme-linked immunosorbent assay kits., Results: DUSP1 and hsCRP plasma levels and their least square means were higher in CVD cases, while oxLDL plasma levels were lower ( p < 0.05). Multivariate logistic regression analysis showed that DUSP1 and hsCRP are independently associated with CVD in the studied population, as reflected by 2-fold and 1.5-fold increased risks with increased levels of DUSP1 and hsCRP, respectively. In our study, DUSP1 levels were found to be associated with CVD despite statin treatment and diabetes status ( p < 0.05), whereas hsCRP mainly correlated with obesity markers., Conclusions: Circulating DUSP1 might be a predictor of chronic subclinical inflammation and residual risk in CVD patients, whereas our data suggest that the association between hsCRP and CVD is largely accounted for adiposity risk factors.
- Published
- 2018
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21. Physical Exercise Enhanced Heat Shock Protein 60 Expression and Attenuated Inflammation in the Adipose Tissue of Human Diabetic Obese.
- Author
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Khadir A, Kavalakatt S, Cherian P, Warsame S, Abubaker JA, Dehbi M, and Tiss A
- Abstract
Heat shock protein 60 (HSP60) is a key protein in the crosstalk between cellular stress and inflammation. However, the status of HSP60 in diabetes and obesity is unclear. In the present study, we investigated the hypothesis that HSP60 expression levels in the adipose tissue of human obese adults with and without diabetes are different and physical exercise might affect these levels. Subcutaneous adipose tissue (SAT) and blood samples were collected from obese adults with and without diabetes ( n = 138 and n = 92, respectively, at baseline; n = 43 for both groups after 3 months of physical exercise). Conventional RT-PCR, immunohistochemistry, immunofluorescence, and ELISA were used to assess the expression and secretion of HSP60. Compared with obese adults without diabetes, HSP60 mRNA and protein levels were decreased in SAT in diabetic obese together with increased inflammatory marker expression and glycemic levels but lower VO
2 Max . More interestingly, a 3-month physical exercise differentially affected HSP60 expression and the heat shock response but attenuated inflammation in both groups, as reflected by decreased endogenous levels of IL-6 and TNF-α. Indeed, HSP60 expression levels in SAT were significantly increased by exercise in the diabetes group, whereas they were decreased in the non-diabetes group. These results were further confirmed using immunofluorescence microscopy and anti-HSP60 antibody in SAT. Exercise had only marginal effects on HSP60 secretion and HSP60 autoantibody levels in plasma in both obese with and without diabetes. Physical exercise differentially alleviates cellular stress in obese adults with and without diabetes despite concomitant attenuation of the inflammatory response.- Published
- 2018
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22. Physical exercise alleviates ER stress in obese humans through reduction in the expression and release of GRP78 chaperone.
- Author
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Khadir A, Kavalakatt S, Abubaker J, Cherian P, Madhu D, Al-Khairi I, Abu-Farha M, Warsame S, Elkum N, Dehbi M, and Tiss A
- Subjects
- 3T3 Cells, Adult, Anaerobic Threshold, Animals, Body Mass Index, C-Reactive Protein metabolism, Endoplasmic Reticulum Chaperone BiP, Female, Gene Expression Profiling, Heat-Shock Proteins biosynthesis, Heat-Shock Proteins genetics, Humans, Male, Mice, Middle Aged, Obesity genetics, Signal Transduction, Subcutaneous Fat metabolism, Unfolded Protein Response genetics, Waist Circumference, Endoplasmic Reticulum Stress genetics, Exercise, Heat-Shock Proteins metabolism, Obesity metabolism, Obesity therapy
- Abstract
Background and Objectives: Perturbation of the endoplasmic reticulum (ER) homeostasis has emerged as one of the prominent features of obesity and diabetes. This occurs when the adaptive unfolded protein response (UPR) fails to restore ER function in key metabolic tissues. We previously reported increased inflammation and impaired heat shock response (HSR) in obese human subjects that were restored by physical exercise. Here, we investigated the status of ER stress chaperone; glucose-regulated protein 78 (GRP78) and its downstream UPR pathways in human obese, and their modulation by a supervised 3-month physical exercise., Methods: Subcutaneous adipose tissue (SAT) and blood samples were collected from non-diabetic adult human lean (n=40) and obese (n=40, at baseline and after 3months of physical exercise). Transcriptomic profiling was used as a primary screen to identify differentially expressed genes and it was carried out on SAT samples using the UPR RT(2) Profiler PCR Array. Conventional RT-PCR, immunohistochemistry, immunofluorescence, Western blot and ELISA were used to validate the transcriptomic data. Correlation analyses with the physical, clinical and biochemical outcomes were performed using Pearson's rank correlation coefficient., Results: Levels of GRP78 and its three downstream UPR arms; activating transcription factor-6 (ATF6), inositol-requiring enzyme-1α (IRE1α) and protein kinase RNA-like endoplasmic reticulum kinase (PERK) were increased in obese subjects. More interestingly, higher levels of circulating GRP78 protein were found in obese compared to lean subjects which correlated negatively with maximum oxygen uptake (VO2 Max) but positively with high-sensitivity C-reactive protein (hsCRP) and obesity indicators such as BMI, percentage body fat (PBF) and waist circumference. GRP78 increased secretion in obese was further confirmed in vitro using 3T3-L1 preadipocyte cells under ER stress. Finally, we showed that physical exercise significantly attenuated the expression and release of GRP78 with a concomitant reduction in the phosphorylation of IRE1α and eukaryotic initiation factor-2α (eIF2α)., Conclusion: Our results suggest that physical exercise alleviates ER stress in human obese through attenuation of GRP78 signaling network., (Copyright © 2016 Elsevier Inc. All rights reserved.)
- Published
- 2016
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23. DNAJB3/HSP-40 cochaperone improves insulin signaling and enhances glucose uptake in vitro through JNK repression.
- Author
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Abu-Farha M, Cherian P, Al-Khairi I, Tiss A, Khadir A, Kavalakatt S, Warsame S, Dehbi M, Behbehani K, and Abubaker J
- Subjects
- 3T3-L1 Cells, Adipose Tissue metabolism, Animals, Cell Line, Diabetes Mellitus, Type 2 genetics, Diabetes Mellitus, Type 2 metabolism, Gene Expression Regulation, HEK293 Cells, HSP40 Heat-Shock Proteins genetics, Humans, Leukocytes, Mononuclear metabolism, Mice, Models, Biological, Obesity genetics, Obesity metabolism, Phosphorylation, Protein Binding, Glucose metabolism, HSP40 Heat-Shock Proteins metabolism, Insulin metabolism, JNK Mitogen-Activated Protein Kinases metabolism, Signal Transduction
- Abstract
Heat shock response (HSR) is an essential host-defense mechanism that is dysregulated in obesity-induced insulin resistance and type 2 diabetes (T2D). Our recent data demonstrated that DNAJB3 was downregulated in obese human subjects and showed negative correlation with inflammatory markers. Nevertheless, DNAJB3 expression pattern in diabetic subjects and its mode of action are not yet known. In this study, we showed reduction in DNAJB3 transcript and protein levels in PBMC and subcutaneous adipose tissue of obese T2D compared to obese non-diabetic subjects. Overexpression of DNAJB3 in HEK293 and 3T3-L1 cells reduced JNK, IRS-1 Ser-307 phosphorylation and enhanced Tyr-612 phosphorylation suggesting an improvement in IRS-1 signaling. Furthermore, DNAJB3 mediated the PI3K/AKT pathway activation through increasing AKT and AS160 phosphorylation. AS160 mediates the mobilization of GLUT4 transporter to the cell membrane and thereby improves glucose uptake. Using pre-adipocytes cells we showed that DNAJB3 overexpression caused a significant increase in the glucose uptake, possibly through its phosphorylation of AS160. In summary, our results shed the light on the possible role of DNAJB3 in improving insulin sensitivity and glucose uptake through JNK repression and suggest that DNAJB3 could be a potential target for therapeutic treatment of obesity-induced insulin resistance.
- Published
- 2015
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24. MAP kinase phosphatase DUSP1 is overexpressed in obese humans and modulated by physical exercise.
- Author
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Khadir A, Tiss A, Abubaker J, Abu-Farha M, Al-Khairi I, Cherian P, John J, Kavalakatt S, Warsame S, Al-Madhoun A, Al-Ghimlas F, Elkum N, Behbehani K, Dermime S, and Dehbi M
- Subjects
- Adiposity genetics, Adult, Cohort Studies, Dual Specificity Phosphatase 1 metabolism, Female, Gene Expression Regulation, Enzymologic, Humans, Male, Middle Aged, Obesity metabolism, Thinness genetics, Thinness metabolism, Up-Regulation genetics, Dual Specificity Phosphatase 1 genetics, Exercise physiology, Obesity genetics
- Abstract
Chronic low-grade inflammation and dysregulation of the stress defense system are cardinal features of obesity, a major risk factor for the development of insulin resistance and diabetes. Dual-specificity protein phosphatase 1 (DUSP1), known also as MAP kinase phosphatase 1 (MKP1), is implicated in metabolism and energy expenditure. Mice lacking DUSP1 are resistant to high-fat diet-induced obesity. However, the expression of DUSP1 has not been investigated in human obesity. In the current study, we compared the expression pattern of DUSP1 between lean and obese nondiabetic human subjects using subcutaneous adipose tissue (SAT) and peripheral blood mononuclear cells (PBMCs). The levels of DUSP1 mRNA and protein were significantly increased in obese subjects with concomitant decrease in the phosphorylation of p38 MAPK (p-p38 MAPK) and PGC-1α and an increase in the levels of phospho-JNK (p-JNK) and phospho-ERK (p-ERK). Moreover, obese subjects had higher levels of circulating DUSP1 protein that correlated positively with various obesity indicators, triglycerides, glucagon, insulin, leptin, and PAI-1 (P < 0.05) but negatively with V̇O(2max) and high-density lipoprotein (P < 0.05). The observation that DUSP1 was overexpressed in obese subjects prompted us to investigate whether physical exercise could reduce its expression. In this study, we report for the first time that physical exercise significantly attenuated the expression of DUSP1 in both the SAT and PBMCs, with a parallel increase in the expression of PGC-1α and a reduction in the levels of p-JNK and p-ERK along with attenuated inflammatory response. Collectively, our data suggest that DUSP1 upregulation is strongly linked to adiposity and that physical exercise modulates its expression. This gives further evidence that exercise might be useful as a strategy for managing obesity and preventing its associated complications., (Copyright © 2015 the American Physiological Society.)
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- 2015
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25. Gender-specific association of oxidative stress and inflammation with cardiovascular risk factors in Arab population.
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Khadir A, Tiss A, Kavalakatt S, Behbehani K, Dehbi M, and Elkum N
- Subjects
- Adult, Anthropometry, Arabs, Body Mass Index, Cross-Sectional Studies, Female, Humans, Male, Middle Aged, Obesity, Abdominal metabolism, Reactive Oxygen Species metabolism, Risk Factors, Waist Circumference, Cardiovascular Diseases ethnology, Cardiovascular Diseases metabolism, Inflammation pathology, Oxidative Stress, Sex Factors
- Abstract
Background: The impact of gender difference on the association between metabolic stress and cardiovascular disease (CVD) remains unclear. We have investigated, for the first time, the gender effect on the oxidative and inflammatory stress responses and assessed their correlation with classical cardiometabolites in Arab population., Methods: A total of 378 adult Arab participants (193 females) were enrolled in this cross-sectional study. Plasma levels of CRP, IL-6, IL-8, TNF-α, ROS, TBARs, and PON1 were measured and correlated with anthropometric and cardiometabolite parameters of the study population., Results: Compared to females, males had significantly higher FBG, HbA1c, TG, and blood pressure but lower BMI, TC, and HDL (P < 0.05). After adjustment for BMI and WC, females had higher levels of ROS, TBARS, and CRP (P < 0.001) whereas males had increased levels of IL-8, IL-6, and TNF-α (P < 0.05). Moreover, after adjustment for age, BMI, and gender, the levels of TNF-α, IL-6, and ROS were associated with central obesity but not general obesity., Conclusion: Inflammation and oxidative stress contribution to CVD risk in Arab population linked to gender and this risk is better reflected by central obesity. Arab females might be at risk of CVD complications due to increased oxidative stress.
- Published
- 2015
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26. Immunohistochemical profiling of the heat shock response in obese non-diabetic subjects revealed impaired expression of heat shock proteins in the adipose tissue.
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Tiss A, Khadir A, Abubaker J, Abu-Farha M, Al-Khairi I, Cherian P, John J, Kavalakatt S, Warsame S, Al-Ghimlas F, Elkum N, Behbehani K, Dermime S, and Dehbi M
- Subjects
- HSP72 Heat-Shock Proteins metabolism, Humans, Interleukin-6 blood, Obesity blood, Tumor Necrosis Factor-alpha blood, Adipose Tissue metabolism, Heat-Shock Proteins metabolism, Heat-Shock Response physiology, Immunohistochemistry methods, Obesity metabolism
- Abstract
Background: Obesity is characterized by a chronic low-grade inflammation and altered stress responses in key metabolic tissues. Impairment of heat shock response (HSR) has been already linked to diabetes and insulin resistance as reflected by decrease in heat shock proteins (HSPs) expression. However, the status of HSR in non-diabetic human obese has not yet been elucidated. The aim of the current study was to investigate whether obesity triggers a change in the HSR pattern and the impact of physical exercise on this pattern at protein and mRNA levels., Methods: Two groups of adult non-diabetic human subjects consisting of lean and obese (n = 47 for each group) were enrolled in this study. The expression pattern of HSP-27, DNAJB3/HSP-40, HSP-60, HSC-70, HSP72, HSP-90 and GRP-94 in the adipose tissue was primarily investigated by immunohistochemistry and then complemented by western blot and qRT-PCR in Peripheral blood mononuclear cells (PBMCs). HSPs expression levels were correlated with various physical, clinical and biochemical parameters. We have also explored the effect of a 3-month moderate physical exercise on the HSPs expression pattern in obese subjects., Results: Obese subjects displayed increased expression of HSP-60, HSC-70, HSP-72, HSP-90 and GRP-94 and lower expression of DNAJB3/HSP-40 (P < 0.05). No differential expression was observed for HSP-27 between the two groups. Higher levels of HSP-72 and GRP-94 proteins correlated positively with the indices of obesity (body mass index and percent body fat) and circulating levels of IFN-gamma-inducible protein 10 (IP-10) and RANTES chemokines. This expression pattern was concomitant with increased inflammatory response in the adipose tissue as monitored by increased levels of Interleukin-6 (IL-6), Tumor necrosis factor-α (TNF-α), and RANTES (P < 0.05). Physical exercise reduced the expression of various HSPs in obese to normal levels observed in lean subjects with a parallel decrease in the endogenous levels of IL-6, TNF-α, and RANTES., Conclusion: Taken together, these data indicate that obesity triggers differential regulation of various components of the HSR in non-diabetic subjects and a 3-month physical moderate exercise was sufficient to restore the normal expression of HSPs in the adipose tissue with concomitant attenuation in the inflammatory response.
- Published
- 2014
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27. IL-33 is negatively associated with the BMI and confers a protective lipid/metabolic profile in non-diabetic but not diabetic subjects.
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Hasan A, Al-Ghimlas F, Warsame S, Al-Hubail A, Ahmad R, Bennakhi A, Al-Arouj M, Behbehani K, Dehbi M, and Dermime S
- Subjects
- Adult, Biomarkers, Body Weight, Diabetes Mellitus blood, Diabetes Mellitus metabolism, Glycated Hemoglobin metabolism, Humans, Interleukin-33, Lipids blood, Middle Aged, Obesity blood, Obesity metabolism, Young Adult, Body Mass Index, Interleukins blood, Lipid Metabolism, Metabolome
- Abstract
Objective: Recent studies have demonstrated a protective role for IL-33 against obesity-associated inflammation, atherosclerosis and metabolic abnormalities. IL-33 promotes the production of T helper type 2 (Th2) cytokines, polarizes macrophages towards a protective alternatively activated phenotype, reduces lipid storage and decreases the expression of genes associated with lipid metabolism and adipogenesis. Our objective was to determine the level of serum IL-33 in non-diabetic and diabetic subjects, and to correlate these levels with clinical (BMI and body weight) and metabolic (serum lipids and HbA1c) parameters., Methods: The level of IL-33 was measured in the serum of lean, overweight and obese non-diabetic and diabetic subjects, and then correlated with clinical and metabolic parameters., Results: Non-lean subjects had significantly (P = 0.01) lower levels of IL-33 compared to lean controls. IL-33 was negatively correlated with the BMI and body weight in lean and overweight, but not obese (non-diabetic and diabetic), subjects. IL-33 is associated with protective lipid profiles, and is negatively correlated with HbA1c, in non-diabetic (lean, overweight and obese) but not diabetic subjects., Conclusions: Our data support previous findings showing a protective role for IL-33 against adiposity and atherosclerosis, and further suggest that reduced levels of IL-33 may put certain individuals at increased risk of developing atherosclerosis and insulin resistance. Therefore, IL-33 may serve as a novel marker to predict those who may be at increased risk of developing atherosclerosis.
- Published
- 2014
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28. Physical exercise reduces the expression of RANTES and its CCR5 receptor in the adipose tissue of obese humans.
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Baturcam E, Abubaker J, Tiss A, Abu-Farha M, Khadir A, Al-Ghimlas F, Al-Khairi I, Cherian P, Elkum N, Hammad M, John J, Kavalakatt S, Lehe C, Warsame S, Behbehani K, Dermime S, and Dehbi M
- Subjects
- Adult, Anthropometry, Body Mass Index, Body Weight, Chemokine CXCL10 blood, Female, Humans, Inflammation blood, Interleukin-6 blood, MAP Kinase Signaling System, Male, Middle Aged, RNA, Messenger metabolism, Signal Transduction, Thiobarbituric Acid Reactive Substances, Tumor Necrosis Factor-alpha blood, Adipose Tissue metabolism, Chemokine CCL5 blood, Exercise, Gene Expression Regulation, Obesity metabolism, Receptors, CCR5 blood
- Abstract
RANTES and its CCR5 receptor trigger inflammation and its progression to insulin resistance in obese. In the present study, we investigated for the first time the effect of physical exercise on the expression of RANTES and CCR5 in obese humans. Fifty-seven adult nondiabetic subjects (17 lean and 40 obese) were enrolled in a 3-month supervised physical exercise. RANTES and CCR5 expressions were measured in PBMCs and subcutaneous adipose tissue before and after exercise. Circulating plasma levels of RANTES were also investigated. There was a significant increase in RANTES and CCR5 expression in the subcutaneous adipose tissue of obese compared to lean. In PBMCs, however, while the levels of RANTES mRNA and protein were comparable between both groups, CCR5 mRNA was downregulated in obese subjects (P < 0.05). Physical exercise significantly reduced the expression of both RANTES and CCR5 (P < 0.05) in the adipose tissue of obese individuals with a concomitant decrease in the levels of the inflammatory markers TNF- α , IL-6, and P-JNK. Circulating RANTES correlated negatively with anti-inflammatory IL-1 ra (P = 0.001) and positively with proinflammatory IP-10 and TBARS levels (P < 0.05). Therefore, physical exercise may provide an effective approach for combating the deleterious effects associated with obesity through RANTES signaling in the adipose tissue.
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- 2014
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29. Gender differences in ghrelin association with cardiometabolic risk factors in arab population.
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Abu-Farha M, Dehbi M, Noronha F, Tiss A, Alarouj M, Behbehani K, Bennakhi A, and Elkum N
- Abstract
Ghrelin is a stomach produced hormone that has been shown to have protective role against development of CVD which is a leading cause of death in the Arab world. The objective of this study is to examine the gender difference in association between traditional CVD risk factors and plasma ghrelin among Arabs. 359 Arab residents in Kuwait participated in a cross-sectional survey (≥20 years old): 191 were females and 168 were males. Plasma level of ghrelin was assessed using Luminex-based assay. Ghrelin levels were significantly higher in females (935 ± 78 pg/mL) than males (763 ± 65 pg/mL) (P = 0.0007). Females showed inverse association with WC (r = -0.23, P = 0.001) and HbA1C (r = -0.19, P = 0.0102) as well as SBP (r = -0.15, P = 0.0383) and DBP (r = -0.16, P = 0.0230), respectively. Higher levels of ghrelin were shown to associate with increased insulin resistance, as measured by HOMAIR, in male Arab subjects (P-trend = 0.0202) but not in females. In this study we show that higher ghrelin level was negatively associated with measures of obesity, HbA1C, and blood pressure in females and positively associated with increased insulin resistance in Arab males.
- Published
- 2014
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30. Proteomics analysis of human obesity reveals the epigenetic factor HDAC4 as a potential target for obesity.
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Abu-Farha M, Tiss A, Abubaker J, Khadir A, Al-Ghimlas F, Al-Khairi I, Baturcam E, Cherian P, Elkum N, Hammad M, John J, Kavalakatt S, Warsame S, Behbehani K, Dermime S, and Dehbi M
- Subjects
- Adipose Tissue metabolism, Blotting, Western, Body Composition, Body Mass Index, Chemokine CCL5 metabolism, Epigenesis, Genetic genetics, Gene Expression Profiling, Humans, Immunohistochemistry, Leukocytes, Mononuclear metabolism, Obesity genetics, Oxygen Consumption physiology, Real-Time Polymerase Chain Reaction, Exercise physiology, Gene Expression Regulation physiology, Histone Deacetylases metabolism, Obesity metabolism, Proteomics methods, Repressor Proteins metabolism, Thrombospondin 1 metabolism
- Abstract
Sedentary lifestyle and excessive energy intake are prominent contributors to obesity; a major risk factors for the development of insulin resistance, type 2 diabetes and cardiovascular diseases. Elucidating the molecular mechanisms underlying these chronic conditions is of relevant importance as it might lead to the identification of novel anti-obesity targets. The purpose of the current study is to investigate differentially expressed proteins between lean and obese subjects through a shot-gun quantitative proteomics approach using peripheral blood mononuclear cells (PBMCs) extracts as well as potential modulation of those proteins by physical exercise. Using this approach, a total of 47 proteins showed at least 1.5 fold change between lean and obese subjects. In obese, the proteomic profiling before and after 3 months of physical exercise showed differential expression of 38 proteins. Thrombospondin 1 (TSP1) was among the proteins that were upregulated in obese subjects and then decreased by physical exercise. Conversely, the histone deacetylase 4 (HDAC4) was downregulated in obese subjects and then induced by physical exercise. The proteomic data was further validated by qRT-PCR, Western blot and immunohistochemistry in both PBMCs and adipose tissue. We also showed that HDAC4 levels correlated positively with maximum oxygen consumption (VO2 Max) but negatively with body mass index, percent body fat, and the inflammatory chemokine RANTES. In functional assays, our data indicated that ectopic expression of HDAC4 significantly impaired TNF-α-dependent activation of NF-κB, establishing thus a link between HDAC4 and regulation of the immune system. Together, the expression pattern of HDAC4 in obese subjects before and after physical exercise, its correlation with various physical, clinical and metabolic parameters along with its inhibitory effect on NF-κB are suggestive of a protective role of HDAC4 against obesity. HDAC4 could therefore represent a potential therapeutic target for the control and management of obesity and presumably insulin resistance.
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- 2013
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31. DNAJB3/HSP-40 cochaperone is downregulated in obese humans and is restored by physical exercise.
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Abubaker J, Tiss A, Abu-Farha M, Al-Ghimlas F, Al-Khairi I, Baturcam E, Cherian P, Elkum N, Hammad M, John J, Kavalakatt S, Khadir A, Warsame S, Dermime S, Behbehani K, and Dehbi M
- Subjects
- Adipose Tissue metabolism, Adult, Female, HSP40 Heat-Shock Proteins genetics, Humans, Immunoprecipitation, In Vitro Techniques, Male, Middle Aged, Obesity genetics, Oxygen Consumption physiology, Palmitates pharmacology, Tunicamycin pharmacology, Exercise physiology, HSP40 Heat-Shock Proteins metabolism, Obesity metabolism
- Abstract
Obesity is a major risk factor for a myriad of disorders such as insulin resistance and diabetes. The mechanisms underlying these chronic conditions are complex but low grade inflammation and alteration of the endogenous stress defense system are well established. Previous studies indicated that impairment of HSP-25 and HSP-72 was linked to obesity, insulin resistance and diabetes in humans and animals while their induction was associated with improved clinical outcomes. In an attempt to identify additional components of the heat shock response that may be dysregulated by obesity, we used the RT(2)-Profiler PCR heat shock array, complemented with RT-PCR and validated by Western blot and immunohistochemistry. Using adipose tissue biopsies and PBMC of non-diabetic lean and obese subjects, we report the downregulation of DNAJB3 cochaperone mRNA and protein in obese that negatively correlated with percent body fat (P = 0.0001), triglycerides (P = 0.035) and the inflammatory chemokines IP-10 and RANTES (P = 0.036 and P = 0.02, respectively). DNAJB positively correlated with maximum oxygen consumption (P = 0.031). Based on the beneficial effect of physical exercise, we investigated its possible impact on DNAJB3 expression and indeed, we found that exercise restored the expression of DNAJB3 in obese subjects with a concomitant decrease of phosphorylated JNK. Using cell lines, DNAJB3 protein was reduced following treatment with palmitate and tunicamycin which is suggestive of the link between the expression of DNAJB3 and the activation of the endoplasmic reticulum stress. DNAJB3 was also shown to coimmunoprecipiate with JNK and IKKβ stress kinases along with HSP-72 and thus, suggesting its potential role in modulating their activities. Taken together, these data suggest that DNAJB3 can potentially play a protective role against obesity.
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- 2013
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32. Toll-like receptor 4 and high-mobility group box 1 are critical mediators of tissue injury and survival in a mouse model for heatstroke.
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Dehbi M, Uzzaman T, Baturcam E, Eldali A, Ventura W, and Bouchama A
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- Animals, Biomarkers metabolism, Body Temperature, Disease Models, Animal, Female, Gene Expression, HMGB1 Protein administration & dosage, HMGB1 Protein genetics, Heat Stroke etiology, Heat Stroke genetics, Heat Stroke prevention & control, Hot Temperature adverse effects, Interleukin-1beta genetics, Interleukin-1beta metabolism, Interleukin-6 genetics, Interleukin-6 metabolism, Male, Mice, Mice, Inbred C3H, Mice, Transgenic, Protein Structure, Tertiary, Recombinant Proteins administration & dosage, Recombinant Proteins genetics, Recombinant Proteins metabolism, Signal Transduction, Toll-Like Receptor 4 genetics, HMGB1 Protein metabolism, Heat Stroke metabolism, Toll-Like Receptor 4 metabolism
- Abstract
The molecular mechanisms that initiate the inflammatory response in heatstroke and their relation with tissue injury and lethality are not fully elucidated. We examined whether endogenous ligands released by damaged/stressed cells such as high-mobility group box 1 (HMGB1) signaling through Toll-like receptor 4 (TLR4) may play a pathogenic role in heatstroke. Mutant TLR4-defective (C3H/HeJ) and wild type (C3H/HeOuJ) mice were subjected to heat stress in an environmental chamber pre-warmed at 43.5 °C until their core temperature reached 42.7°C, which was taken as the onset of heatstroke. The animals were then allowed to recover passively at ambient temperature. A sham-heated group served as a control. Mutant mice displayed more histological liver damage and higher mortality compared with wild type mice (73% vs. 27%, respectively, P<0.001). Compared to wild type mice, mutant mice exhibited earlier plasma release of markers of systemic inflammation such as HMGB1 (206 ± 105 vs. 63 ± 21 ng/ml; P = 0.0018 and 209 ± 100 vs. 46 ± 32 ng/ml; P<0.0001), IL-6 (144 ± 40 vs. 46 ± 20 pg/ml; P<0.001 and 184 ± 21 vs. 84 ± 54 pg/ml; P = 0.04), and IL-1β (27 ± 4 vs. 1.7 ± 2.3 pg/ml; P<0.0001 at 1 hour). Both strains of mice displayed early release of HMGB1 into the circulation upstream of IL-1β and IL-6 responses which remained elevated up to 24 h. Specific inhibition of HMGB1 activity with DNA-binding A Box (600 µg/mouse) protected the mutant mice against the lethal effect of heat stress (60% A Box vs. 18% GST protein, P = 0.04). These findings suggest a protective role for the TLR4 in the host response to severe heat stress. They also suggest that HMGB1 is an early mediator of inflammation, tissue injury and lethality in heatstroke in the presence of defective TLR4 signaling.
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- 2012
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33. sRAGE in diabetic and non-diabetic critically ill patients: effects of intensive insulin therapy.
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Arabi YM, Dehbi M, Rishu AH, Baturcam E, Kahoul SH, Brits RJ, Naidu B, and Bouchama A
- Subjects
- Aged, Blood Glucose analysis, Female, HMGB1 Protein metabolism, Hospital Mortality, Humans, Hyperglycemia mortality, Hypoglycemic Agents administration & dosage, Insulin administration & dosage, Male, Middle Aged, Prospective Studies, Receptor for Advanced Glycation End Products, Regression Analysis, Critical Illness mortality, Diabetes Mellitus, Type 2 drug therapy, Hyperglycemia drug therapy, Hypoglycemic Agents therapeutic use, Insulin therapeutic use, Receptors, Immunologic metabolism
- Abstract
Introduction: Hyperglycemia represents an independent prognostic factor in critically ill non-diabetic patients but not in those with diabetes. In this context, there is an ongoing debate on the benefit of an intensive insulin therapy, particularly in diabetic patients. We tested the hypothesis that expression of the receptor for advanced glycation end-products (RAGE), an important signal transduction receptor that elicits long-lasting nuclear factor kappa B (NF-κB) activation, may underlie this difference. RAGE expression is regulated by multiple ligands, including high mobility group box-1 (HMGB-1), and is reflected by its released soluble form (sRAGE)., Methods: A predesigned analysis was conducted of prospectively collected samples from 76 hyperglycemic critically ill patients (33 type-2 diabetes, 43 non-diabetes) aged ≥ 18 years with blood glucose of > 6.1 mmol/L enrolled in a randomized controlled trial comparing intensive insulin therapy with conventional insulin therapy. sRAGE and its ligand HMGB-1 together with IL-6, and soluble thrombomodulin (as markers of inflammation and endothelial cell injury, respectively) were evaluated in ICU, at Days 1, 3, 5 and 7. Plasma samples from 18 healthy subjects were used as controls., Results: Both diabetic and non-diabetic hyperglycemic patients showed increased plasma sRAGE, HMGB-1 and soluble thrombomodulin levels at the time of admission to ICU. Plasma IL-6 concentration was only increased in non-diabetic patients. Plasma levels of sRAGE were higher in diabetic compared with non-diabetic patients. Intensive insulin therapy resulted in a significant decrease of sRAGE and thrombomodulin at Day 7, in diabetic but not in non-diabetic patients. Circulating sRAGE levels correlated positively with IL-6 and soluble thrombomodulin levels and inversely with HMGB-1. Multivariate regression analysis demonstrated that sRAGE remains independently correlated with HMGB-1 only in diabetic patients. Neither sRAGE nor any inflammatory markers are associated with mortality., Conclusions: These findings support the hypothesis that sRAGE release, time-course and response to intensive insulin therapy differ between hyperglycemic diabetic and non-diabetic critically ill patients. Whether this difference underlies the dissimilarity in clinical outcome of hyperglycemia in these two conditions warrants further studies.
- Published
- 2011
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34. Genome annotation and intraviral interactome for the Streptococcus pneumoniae virulent phage Dp-1.
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Sabri M, Häuser R, Ouellette M, Liu J, Dehbi M, Moeck G, García E, Titz B, Uetz P, and Moineau S
- Subjects
- Chromatography, Liquid, DNA Replication, DNA, Viral chemistry, DNA, Viral genetics, Gene Order, Genes, Viral, Mass Spectrometry, Molecular Sequence Data, Multigene Family, Open Reading Frames, Promoter Regions, Genetic, Protein Biosynthesis, Sequence Analysis, DNA, Siphoviridae classification, Siphoviridae ultrastructure, Streptococcus Phages classification, Streptococcus Phages ultrastructure, Terminator Regions, Genetic, Viral Structural Proteins analysis, Genome, Viral, Streptococcus Phages genetics, Streptococcus pneumoniae virology
- Abstract
Streptococcus pneumoniae causes several diseases, including pneumonia, septicemia, and meningitis. Phage Dp-1 is one of the very few isolated virulent S. pneumoniae bacteriophages, but only a partial characterization is currently available. Here, we confirmed that Dp-1 belongs to the family Siphoviridae. Then, we determined its complete genomic sequence of 56,506 bp. It encodes 72 open reading frames, of which 44 have been assigned a function. We have identified putative promoters, Rho-independent terminators, and several genomic clusters. We provide evidence that Dp-1 may be using a novel DNA replication system as well as redirecting host protein synthesis through queuosine-containing tRNAs. Liquid chromatography-mass spectrometry analysis of purified phage Dp-1 particles identified at least eight structural proteins. Finally, using comprehensive yeast two-hybrid screens, we identified 156 phage protein interactions, and this intraviral interactome was used to propose a structural model of Dp-1.
- Published
- 2011
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35. Hsp-72, a candidate prognostic indicator of heatstroke.
- Author
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Dehbi M, Baturcam E, Eldali A, Ahmed M, Kwaasi A, Chishti MA, and Bouchama A
- Subjects
- Animals, Blotting, Western, Enzyme-Linked Immunosorbent Assay, Fever metabolism, Papio, HSP72 Heat-Shock Proteins metabolism, Heat Stroke metabolism
- Abstract
Exposure of rats to environmental heat enhances the expression of heat shock protein-72 (Hsp-72) in most of their organs proportionally to heat stress severity. Pre-induction or over-expression of Hsp-72 prevents organ damage and lethality, suggesting that heat shock proteins (Hsps) may have a pathogenic role in this condition. We investigated the expression profile of Hsps in baboons subjected to environmental heat stress until the core temperature attained 42.5 degrees C (moderate heatstroke) or occurrence of hypotension associated with core temperature > or = 43.5 degrees C (severe heatstroke). Western blot analysis demonstrated a differential induction of Hsp-72 among organs of heat-stressed animals with the highest induction in the liver and the lowest in lung. Hsp-60 and Hsc-70 expression was similar between control and heat-stressed animals. ELISA studies indicated a marked release of Hsp-72 into the circulation of baboons with severe heatstroke with a peak at 24 h post-heatstroke onset and remained sustained up to 72 h. Hsp-72 release was not associated with core temperature or systolic blood pressure, but correlated with markers of liver, myocardium, and skeletal muscle tissue necrosis. Non-survivors displayed significantly higher Hsp-72 levels than survivors. No Hsp-60 was detected in the circulation. These findings add further evidence that increased expression of Hsp-72 may be an important component of the host response to severe heatstroke. They also suggest that extracellular Hsp-72 is a marker of multiple organs tissue damage. Whether extracellular Hsp-72 plays a role in the host immune response to heat stress merits further studies.
- Published
- 2010
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36. Inhibition of transcription in Staphylococcus aureus by a primary sigma factor-binding polypeptide from phage G1.
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Dehbi M, Moeck G, Arhin FF, Bauda P, Bergeron D, Kwan T, Liu J, McCarty J, Dubow M, and Pelletier J
- Subjects
- Bacterial Proteins chemistry, Bacterial Proteins genetics, Bacterial Proteins metabolism, DNA-Directed RNA Polymerases genetics, DNA-Directed RNA Polymerases metabolism, Gene Expression Regulation, Bacterial, Peptides genetics, Protein Binding, Protein Structure, Tertiary, Sigma Factor chemistry, Sigma Factor genetics, Staphylococcus Phages genetics, Staphylococcus aureus metabolism, Staphylococcus aureus virology, Viral Proteins genetics, Down-Regulation, Peptides metabolism, Sigma Factor metabolism, Staphylococcus Phages metabolism, Staphylococcus aureus genetics, Transcription, Genetic, Viral Proteins metabolism
- Abstract
The primary sigma factor of Staphylococcus aureus, sigma(SA), regulates the transcription of many genes, including several essential genes, in this bacterium via specific recognition of exponential growth phase promoters. In this study, we report the existence of a novel staphylococcal phage G1-derived growth inhibitory polypeptide, referred to as G1ORF67, that interacts with sigma(SA) both in vivo and in vitro and regulates its activity. Delineation of the minimal domain of sigma(SA) that is required for its interaction with G1ORF67 as amino acids 294 to 360 near the carboxy terminus suggests that the G1 phage-encoded anti-sigma factor may occlude the -35 element recognition domain of sigma(SA). As would be predicted by this hypothesis, the G1ORF67 polypeptide abolished both RNA polymerase core-dependent binding of sigma(SA) to DNA and sigma(SA)-dependent transcription in vitro. While G1ORF67 profoundly inhibits transcription when expressed in S. aureus cells in mode of action studies, our finding that G1ORF67 was unable to inhibit transcription when expressed in Escherichia coli concurs with its inability to inhibit transcription by the E. coli holoenzyme in vitro. These features demonstrate the selectivity of G1ORF67 for S. aureus RNA polymerase. We predict that G1ORF67 is one of the central polypeptides in the phage G1 strategy to appropriate host RNA polymerase and redirect it to phage reproduction.
- Published
- 2009
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37. Recombinant activated protein C attenuates endothelial injury and inhibits procoagulant microparticles release in baboon heatstroke.
- Author
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Bouchama A, Kunzelmann C, Dehbi M, Kwaasi A, Eldali A, Zobairi F, Freyssinet JM, and de Prost D
- Subjects
- Animals, Antithrombin III, Cytoprotection, Disease Models, Animal, Endothelium, Vascular metabolism, Endothelium, Vascular pathology, Fibrin Fibrinogen Degradation Products metabolism, Fibrinolysis drug effects, Fibrinolytic Agents administration & dosage, Heat Stroke blood, Heat Stroke complications, Heat Stroke metabolism, Heat Stroke pathology, Humans, Infusions, Intravenous, Interleukin-6 blood, Multiple Organ Failure etiology, Multiple Organ Failure metabolism, Multiple Organ Failure prevention & control, Papio hamadryas, Peptide Hydrolases blood, Protein C administration & dosage, Recombinant Proteins pharmacology, Severity of Illness Index, Thrombomodulin blood, Time Factors, Tissue Plasminogen Activator blood, Transport Vesicles drug effects, Blood Coagulation drug effects, Endothelium, Vascular drug effects, Fibrinolytic Agents pharmacology, Heat Stroke prevention & control, Protein C pharmacology, Transport Vesicles metabolism
- Abstract
Objective: We tested the hypothesis that the antithrombotic and cytoprotective effects of recombinant human activated protein C (rhAPC) protect baboons against the lethal effects of heatstroke., Methods and Results: Fourteen anesthetized baboons assigned randomly to rhAPC (n=7) or control group (n=7) were heat-stressed in a prewarmed incubator at 44 to 47 degrees C until systolic blood pressure fell below 90 mm Hg, which signaled severe heatstroke. rhAPC was administered intravenously (24 microg/kg/h) for 12 hours at onset of heatstroke. Heat stress induced coagulation and fibrinolysis activation as evidenced by a significant increase from baseline levels in plasma levels of thrombin-antithrombin (TAT) complexes, tissue plasminogen activator, and D-dimer. Heat stress elicited cell activation/injury as assessed by the release of interleukin (IL)-6, soluble thrombomodulin, and procoagulant microparticles (MPs). rhAPC did not significantly reduce heatstroke-induced thrombin generation, and D-dimer and had no effect on fibrinolytic activity. In contrast, rhAPC infusion attenuated significantly the plasma rise of IL-6 and inhibited the release of soluble thrombomodulin and MPs as compared with control group. No difference in survival was observed between rhAPC-treated and control group., Conclusions: rhAPC given to heatstroke baboons provided cytoprotection, but had no effect on heatstroke-induced coagulation activation and fibrin formation. Inhibition of MPs by rhAPC suggested a novel mechanism of action for this protein.
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- 2008
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38. Glucocorticoids do not protect against the lethal effects of experimental heatstroke in baboons.
- Author
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Bouchama A, Kwaasi A, Dehbi M, Al Mohanna F, Eldali A, El-Sayed R, Tbakhi A, Alzahrani AS, and Roberts AG
- Subjects
- Alanine Transaminase blood, Analysis of Variance, Animals, Bilirubin blood, Blood Pressure drug effects, Complement C3 metabolism, Complement C4 metabolism, Creatine Kinase blood, Dexamethasone pharmacology, Dexamethasone therapeutic use, Enzyme-Linked Immunosorbent Assay, Glucocorticoids therapeutic use, Heart Rate drug effects, Heat Stroke blood, Heat Stroke physiopathology, Interleukin-6 blood, Interleukin-6 metabolism, L-Lactate Dehydrogenase blood, Lactic Acid blood, Papio, Random Allocation, Temperature, Time Factors, Glucocorticoids pharmacology, Heat Stroke drug therapy
- Abstract
The mortality and neurological morbidity in heatstroke have been attributed to the host's inflammatory responses to heat stress, suggesting that anti-inflammatory therapy may improve outcome. We tested the hypothesis that a high dose of dexamethasone protects baboons against the lethal effects of heatstroke. Ten anesthetized baboons (Papio hamadryas) were assigned randomly to dexamethasone (n = 5) or control group (n = 5). Dexamethasone (2 mg/kg i.v.) was administered in four divided doses every 6 h starting immediately before heat stress and continuing during cooling. All animals were heat-stressed in a prewarmed neonatal incubator at 44 degrees C to 47 degrees C until systolic blood pressure fell less than 90 mmHg and then cooled passively at the ambient temperature. Mortality and neurological morbidity were noted, and biochemical markers of tissue injury/organ dysfunction were determined. Circulating interleukin (IL) 6 and complement components (C3 and C4) were measured sequentially. All heat-stressed animals had systemic inflammation indicated by increased plasma IL-6 and decreased C3 and C4 levels. Dexamethasone attenuated the complement system activation and maintained a higher plasma concentration of IL-6, with a significant augmentation of arterial blood pressure. Dexamethasone did not prevent the occurrence of severe heatstroke but unexpectedly aggravated significantly the tissue injury and multiorgan system dysfunction. Two animals (40%) in the control group and one in the steroid group survived (P > 0.05). Dexamethasone failed to protect the baboons from the lethal effects of heatstroke. These results do not support clinical testing of corticosteroids as beneficial in preventive or therapeutic strategies for the treatment of heatstroke in humans.
- Published
- 2007
- Full Text
- View/download PDF
39. Cooling and hemodynamic management in heatstroke: practical recommendations.
- Author
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Bouchama A, Dehbi M, and Chaves-Carballo E
- Subjects
- Adult, Aged, Aged, 80 and over, Dantrolene therapeutic use, Heat Stroke physiopathology, Hemodynamics, Humans, Hydrotherapy instrumentation, Hydrotherapy methods, Hypothermia, Induced instrumentation, Middle Aged, Muscle Relaxants, Central therapeutic use, Treatment Outcome, Heat Stroke therapy, Hypothermia, Induced methods
- Abstract
Introduction: Although rapid cooling and management of circulatory failure are crucial to the prevention of irreversible tissue damage and death in heatstroke, the evidence supporting the optimal cooling method and hemodynamic management has yet to be established., Methods: A systematic review of all clinical studies published in Medline (1966 to 2006), CINAHL (Cumulative Index to Nursing & Allied Health Literature) (1982 to 2006), and Cochrane Database was performed using the OVID interface without language restriction. Search terms included heatstroke, sunstroke, and heat stress disorders., Results: Fourteen articles reported populations subjected to cooling treatment for classic or exertional heatstroke and included data on cooling time, neurologic morbidity, or mortality. Five additional articles described invasive monitoring with central venous or pulmonary artery catheters. The four clinical trials and 15 observational studies covered a total of 556 patients. A careful analysis of the results obtained indicated that the cooling method based on conduction, namely immersion in iced water, was effective among young people, military personnel, and athletes with exertional heatstroke. There was no evidence to support the superiority of any one cooling technique in classic heatstroke. The effects of non-invasive, evaporative, or conductive-based cooling techniques, singly or combined, appeared to be comparable. No evidence of a specific endpoint temperature for safe cessation of cooling was found. The circulatory alterations in heatstroke were due mostly to a form of distributive shock associated with relative or absolute hypovolemia. Myocardial failure was found to be rare., Conclusion: A systematic review of the literature failed to identify reliable clinical data on the optimum treatment of heatstroke. Nonetheless, the findings of this study could serve as a framework for preliminary recommendations in cooling and hemodynamic management of heatstroke until more evidence-based data are generated.
- Published
- 2007
- Full Text
- View/download PDF
40. Competition of bacteriophage polypeptides with native replicase proteins for binding to the DNA sliding clamp reveals a novel mechanism for DNA replication arrest in Staphylococcus aureus.
- Author
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Belley A, Callejo M, Arhin F, Dehbi M, Fadhil I, Liu J, McKay G, Srikumar R, Bauda P, Bergeron D, Ha N, Dubow M, Gros P, Pelletier J, and Moeck G
- Subjects
- Amino Acid Sequence, Bacterial Proteins metabolism, Binding, Competitive, DNA Replication physiology, DNA, Bacterial metabolism, DNA-Directed DNA Polymerase metabolism, Fluorescence Resonance Energy Transfer, Molecular Sequence Data, Staphylococcus aureus genetics, Two-Hybrid System Techniques, DNA Polymerase III metabolism, DNA, Bacterial biosynthesis, Staphylococcus aureus virology, Streptococcus Phages physiology, Viral Proteins metabolism
- Abstract
Bacteriophages have evolved specific mechanisms that redirect bacterial metabolic pathways to the bacteriophage reproduction cycle. In this study, we characterized the bactericidal mechanism of two polypeptides from bacteriophages Twort and G1 that target the DNA sliding clamp of Staphylococcus aureus. The DNA sliding clamp, which tethers DNA polymerase to its template and thereby confers processivity upon the enzyme, was found to be essential for the viability of S. aureus. Expression of polypeptides TwortORF168 and G1ORF240 in S. aureus selectively inhibited DNA replication which in turn resulted in cell death. Both polypeptides specifically inhibited the S. aureus DNA replicase that was reconstituted in vitro but not the corresponding replicase of Streptococcus pyogenes. We demonstrated that inhibition of DNA synthesis is multifaceted and occurs via binding the DNA sliding clamp: TwortORF168 and G1ORF240 bound tightly to the DNA sliding clamp and prevented both its loading onto DNA and its interaction with DNA polymerase C. These results elucidate the impact of bacteriophage polypeptide expression upon DNA replication in the growing cell.
- Published
- 2006
- Full Text
- View/download PDF
41. A new class of small molecule RNA polymerase inhibitors with activity against rifampicin-resistant Staphylococcus aureus.
- Author
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Arhin F, Bélanger O, Ciblat S, Dehbi M, Delorme D, Dietrich E, Dixit D, Lafontaine Y, Lehoux D, Liu J, McKay GA, Moeck G, Reddy R, Rose Y, Srikumar R, Tanaka KS, Williams DM, Gros P, Pelletier J, Parr TR Jr, and Far AR
- Subjects
- Anti-Bacterial Agents chemistry, Anti-Bacterial Agents pharmacology, DNA-Directed RNA Polymerases metabolism, Molecular Structure, Molecular Weight, Structure-Activity Relationship, Anti-Bacterial Agents classification, Carboxylic Acids chemistry, Carboxylic Acids pharmacology, DNA-Directed RNA Polymerases antagonists & inhibitors, Drug Resistance, Bacterial, Rifampin pharmacology, Staphylococcus aureus drug effects, Thiophenes chemistry, Thiophenes pharmacology
- Abstract
The RNA polymerase holoenzyme is a proven target for antibacterial agents. A high-throughput screening program based on this enzyme from Staphylococcus aureus had previously identified a 2-ureidothiophene-3-carboxylate as a low micromolar inhibitor. An investigation of the relationships between the structures of this class of compounds and their inhibitory- and antibacterial activities is described here, leading to a set of potent RNA polymerase inhibitors with antibacterial activity. Characterization of this bioactivity, including studies of the mechanism of action, is provided, highlighting the power of the reverse chemical genetics approach in providing tools to inhibit the bacterial RNA polymerase.
- Published
- 2006
- Full Text
- View/download PDF
42. Antimicrobial drug discovery through bacteriophage genomics.
- Author
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Liu J, Dehbi M, Moeck G, Arhin F, Bauda P, Bergeron D, Callejo M, Ferretti V, Ha N, Kwan T, McCarty J, Srikumar R, Williams D, Wu JJ, Gros P, Pelletier J, and DuBow M
- Subjects
- Anti-Bacterial Agents therapeutic use, Bacterial Infections drug therapy, Bacterial Infections metabolism, Bacterial Infections virology, Bacteriophages metabolism, Drug Design, Gene Expression Profiling methods, Genome, Viral, Humans, Staphylococcal Infections drug therapy, Staphylococcal Infections metabolism, Staphylococcal Infections virology, Bacterial Proteins metabolism, Drug Delivery Systems methods, Proteome metabolism, Staphylococcus Phages metabolism, Staphylococcus aureus metabolism, Staphylococcus aureus virology, Viral Proteins metabolism
- Abstract
Over evolutionary time bacteriophages have developed unique proteins that arrest critical cellular processes to commit bacterial host metabolism to phage reproduction. Here, we apply this concept of phage-mediated bacterial growth inhibition to antibiotic discovery. We sequenced 26 Staphylococcus aureus phages and identified 31 novel polypeptide families that inhibited growth upon expression in S. aureus. The cellular targets for some of these polypeptides were identified and several were shown to be essential components of the host DNA replication and transcription machineries. The interaction between a prototypic pair, ORF104 of phage 77 and DnaI, the putative helicase loader of S. aureus, was then used to screen for small molecule inhibitors. Several compounds were subsequently found to inhibit both bacterial growth and DNA synthesis. Our results suggest that mimicking the growth-inhibitory effect of phage polypeptides by a chemical compound, coupled with the plethora of phages on earth, will yield new antibiotics to combat infectious diseases.
- Published
- 2004
- Full Text
- View/download PDF
43. Activation of the wt1 Wilms' tumor suppressor gene by NF-kappaB.
- Author
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Dehbi M, Hiscott J, and Pelletier J
- Subjects
- 3T3 Cells, Animals, Binding Sites, Cell Line, Transformed, Gene Expression Regulation, Mice, NF-kappa B p50 Subunit, Recombinant Fusion Proteins metabolism, Transcription Factor RelA, Tumor Cells, Cultured, WT1 Proteins, DNA-Binding Proteins genetics, Genes, Wilms Tumor, NF-kappa B metabolism, Promoter Regions, Genetic, Transcription Factors genetics, Transcriptional Activation
- Abstract
The Wilm's tumor suppressor gene, wt1, is expressed in a very defined spatial-temporal fashion and plays a key role in development of the urogenital system. Transacting factors governing wt1 expression are poorly defined. The presence of putative kappa-B binding sites within the wt1 gene prompted us to investigate whether members of the NF-kappaB/Rel family of transcription factors are involved in regulating wt1 expression. In transient transfection assays, ectopic expression of p50 and p65 subunits of NF-kappaB stimulated wt1 promoter activity 10-30-fold. Deletion mutagenesis revealed that NF-kappa-B responsiveness is mediated by a short DNA fragment located within promoter proximal sequences of the major transcription start site. Two kappaB-binding sites are present in this region and form specific complexes with purified NF-kappaB proteins, as revealed by electrophoretic mobility gel shift assays. Ectopic expression of p50 and p65 resulted in increased transcription of the endogenous wt1 gene, as revealed by nuclear run-on experiments. Taken together, these results indicate that members of the NF-kappaB/Rel family are important for activating expression of wt1 and reside upstream of the regulatory cascade leading to wt1 activation.
- Published
- 1998
- Full Text
- View/download PDF
44. Overlapping DNA recognition motifs between Sp1 and a novel trans-acting factor within the wt1 tumour suppressor gene promoter.
- Author
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Discenza MT, Dehbi M, and Pelletier J
- Subjects
- Animals, Base Sequence, Binding Sites, Cell Extracts, Cell Line, Cell Nucleus, Conserved Sequence, DNA metabolism, DNA-Binding Proteins chemistry, HeLa Cells, Humans, Mice, Molecular Weight, Organ Specificity, Point Mutation, Species Specificity, Transcriptional Activation physiology, DNA-Binding Proteins metabolism, Genes, Wilms Tumor genetics, Promoter Regions, Genetic genetics, Sp1 Transcription Factor metabolism
- Abstract
The Wilms' tumor suppressor gene, wt1 , encodes a zinc finger transcription factor which has been shown to regulate the expression of several genes involved in cellular proliferation and differentiation. Expression of wt1 is developmentally regulated and restricted to a small set of tissues which include the fetal urogenital system, mesothelium and spleen. A highly conserved motif within the wt1 promoter, located between nucleotides -34 and -71 relative to the first transcription start site in the murine promoter, harbors consensus binding sites for Sp1 and members of the paired-box transcription factor family. Pax-2 and Pax-8 are known to enhance expression of wt1 through this conserved regulatory element. In this report, we demonstrate that Sp1 is able to bind to two sites within the 38 bp conserved region (CR). By electrophoretic mobility shift assays (EMSAs), we have identified a novel binding activity, referred to as complex D, which recognizes sequences overlapping one of the Sp1 sites in the CR. EMSA competition experiments indicate that binding of complex D and Sp1 to the CR is mutually exclusive and Sp1 is able to displace complex D binding. In situ UV crosslinking and molecular mass determinations indicate that complex D is a complex of approximately 130 kDa, consisting of at least two proteins of approximately 62 and approximately 70 kDa. Transient transfections suggest that complex D may function as an activator.
- Published
- 1997
- Full Text
- View/download PDF
45. Multiple signaling pathways control the activation of the CEF-4/9E3 cytokine gene by pp60v-src.
- Author
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Bojović B, Rodrigues N, Dehbi M, and Bédard PA
- Subjects
- Animals, Avian Sarcoma Viruses, Blotting, Northern, Cell Transformation, Viral, Chick Embryo, DNA-Binding Proteins metabolism, Kinetics, Naphthalenes metabolism, Nuclear Proteins metabolism, Promoter Regions, Genetic, Protein Kinase C metabolism, Protein Serine-Threonine Kinases metabolism, Proto-Oncogene Proteins metabolism, Proto-Oncogene Proteins c-raf, Proto-Oncogene Proteins p21(ras) metabolism, RNA, Messenger metabolism, Regulatory Factor X Transcription Factors, Tetradecanoylphorbol Acetate pharmacology, Transcription Factor AP-1 metabolism, Avian Proteins, Cytokines genetics, Gene Expression Regulation, Oncogene Protein pp60(v-src) metabolism, Signal Transduction, Transcription Factors
- Abstract
The CEF-4/9E3 cytokine gene is expressed aberrantly in chicken embryo fibroblasts (CEF) transformed by the Rous sarcoma virus. The expression of CEF-4 is dependent on both transcriptional and post-transcriptional mechanisms of regulation. The characterization of the promoter region indicated that three distinct regulatory elements corresponding to an AP-1 binding site (or TRE), a PRDII/kappaB domain, and a CAAT box are involved in the activation by pp60(v-)src. In this report we investigate the signaling pathways controlling the expression of the TRE and PRDII domain. The expression of a dominant negative mutant of p21(ras) reduced the activity of both elements. In contrast a similar mutant of c-Raf-1 affected modestly the activation dependent on the TRE but not PRDII. The stress-activated protein kinase (SAPK)/Jun N-terminal kinase (JNK) pathway was important for the activity of PRDII and the TRE but was not markedly stimulated by pp60(v-)src. The addition of calphostin C and the inhibition of protein kinase C (PKC) diminished the accumulation of the CEF-4 mRNA and reduced the activity of a TRE-controlled promoter. Likewise, the depletion of PKC by chronic treatment with phorbol esters inhibited the activation of the TRE. Rous sarcoma virus-transformed CEF treated with calphostin C were also flatter, did not display a high degree of criss-crossing, and appeared morphologically normal. Hence PKC was important for the activation of AP-1 and the morphological transformation of CEF. The constitutive expression of CEF-4 was correlated with transformation only when dependent on the TRE. This was not true for PRDII, which was the only element required for the constitutive activation to the CEF-4 promoter in nontransformed cells treated chronically with phorbol esters.
- Published
- 1996
- Full Text
- View/download PDF
46. The paired-box transcription factor, PAX2, positively modulates expression of the Wilms' tumor suppressor gene (WT1).
- Author
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Dehbi M, Ghahremani M, Lechner M, Dressler G, and Pelletier J
- Subjects
- 3T3 Cells physiology, Animals, Base Sequence, DNA-Binding Proteins genetics, DNA-Binding Proteins pharmacology, Gene Expression Regulation, Neoplastic drug effects, Gene Expression Regulation, Neoplastic physiology, Humans, Mice, Molecular Sequence Data, PAX2 Transcription Factor, Promoter Regions, Genetic, Transcription Factors genetics, Transcription Factors pharmacology, Transcriptional Activation, Transfection, WT1 Proteins, DNA-Binding Proteins biosynthesis, DNA-Binding Proteins physiology, Gene Expression Regulation, Developmental physiology, Genes, Wilms Tumor, Transcription Factors biosynthesis, Transcription Factors physiology
- Abstract
The Wilms' tumor suppressor gene, wt1, encodes a zinc finger protein which functions as a transcriptional regulator. Expression of the wt1 gene is developmentally regulated and restricted to a small set of tissues which include the fetal urogenital system, mesothelium, and spleen. In the developing kidney, induction of neprohogenesis by the ureter is accompanied by an increase in expression levels of the Pax-2 gene, a developmentally and spatially regulated paired-box member. This is followed by an increase in wt1 expression as mesenchymal cells condense and differentiate. In this report, we demonstrate that PAX2 isoforms are capable of transactivating the wt1 promoter. Deletion mutagenesis of the wt1 promoter identified an element responsible for mediating PAX2 responsiveness, located between nucleotides -33 and -71 relative to the first wt1 transcription start site. Consistent with its identity as a PAX responsive element, multimerization of this mofit upstream of a heterologous minimal promoter enhanced reporter activity when co-transfected with a Pax-2 expression vector. Finally, we demonstrate that PAX2 can stimulate expression of the endogenous wt1 gene. These results suggest that a role for PAX2 during mesenchyme-to-epithelium transition in renal development is to induce wt1 expression.
- Published
- 1996
47. Repression of the retinoic acid receptor-alpha gene by the Wilms' tumor suppressor gene product, wt1.
- Author
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Goodyer P, Dehbi M, Torban E, Bruening W, and Pelletier J
- Subjects
- Base Sequence, Cell Line, DNA, Humans, Molecular Sequence Data, Retinoic Acid Receptor alpha, WT1 Proteins, Wilms Tumor genetics, DNA-Binding Proteins genetics, Gene Expression Regulation genetics, Receptors, Retinoic Acid genetics, Transcription Factors genetics, Zinc Fingers genetics
- Abstract
The Wilms' tumor (WT) suppressor gene, WT1, encodes a zinc finger DNA binding protein (wt1) which functions as a transcriptional regulator. Germline WT1 mutations predispose to WTs and in many cases are associated with urogenital anomalies. Identification of wt1 downstream targets is essential to understanding regulatory processes involved in development of this system. In this study, we demonstrate that wt1 can repress transcription of the retinoic acid receptor-alpha 1 (RAR-alpha 1) promoter. Transient transfection, deletion mutagenesis, and mobility shift assays suggest that wt1 mediates repression of the human RAR-alpha 1 promoter through a GC-rich DNA binding motif (5'-GCGGGGGCG-3'), at positions -111 to -120 bp (relative to the transcription initiation site). In contrast, the murine RAR-alpha 1 promoter contains a cryptic binding motif and is not responsive to wt1. These results indicate that some wt1-regulatory pathways are not conserved across species, suggesting a molecular basis for differences in phenotypes between humans and mice harboring WT1 lesions.
- Published
- 1995
48. Activation of human immunodeficiency virus 1 gene expression by the src oncoprotein.
- Author
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Dehbi M, Beaulieu S, Cohen EA, and Bédard PA
- Subjects
- Animals, Base Sequence, Chick Embryo, HIV Long Terminal Repeat, Molecular Sequence Data, NF-kappa B physiology, Promoter Regions, Genetic, TATA Box, Gene Expression Regulation, Viral, HIV-1 genetics, Oncogene Protein pp60(v-src) physiology
- Abstract
Several genes are induced constitutively in cells transformed by the v-src oncoprotein. This induction is generally dependent on the activation of transcription factors binding to src-responsive elements of the promoter. In previous studies, we showed that the induction of the CEF-4/9E3 cytokine gene by pp60v-src is dependent on the PRDII/kappa B domain of the promoter (Dehbi et al., 1992). In this investigation, we describe the activation of the HIV-1 LTR by pp60v-src and show that a region of 30 bp containing the two NF-kappa B binding sites is critical for activation of the promoter. The induction was dependent on transformation since non-transforming forms of pp60v-src had little or no effect on the promoter. The expression of proviral DNA and the release of p24 antigen were also increased by v-src indicating that viral replication was stimulated in src-transformed cells. The effect of v-src on HIV-1 gene expression occurred in the presence or in the absence of the tat viral trans-activator, in fibroblasts and in Jurkat T lymphocytes. These results indicate that several promoters controlled by PRDII/kappa B may be activated constitutively in v-src transformed cells and suggest that oncogenic tyrosine kinases may play a role in the induction of viruses with a PRDII/kappa B-controlled promoter.
- Published
- 1994
49. Regulation of gene expression in oncogenically transformed cells.
- Author
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Dehbi M and Bédard PA
- Subjects
- Animals, Base Sequence, Cell Transformation, Neoplastic metabolism, Humans, Molecular Sequence Data, Neoplasm Proteins biosynthesis, Neoplasm Proteins genetics, Oncogenes, Signal Transduction genetics, Transcription, Genetic physiology, Transcriptional Activation, Cell Transformation, Neoplastic genetics, Gene Expression Regulation, Neoplastic
- Abstract
Several genes expressed in response to growth factors are also regulated aberrantly in oncogenically transformed cells. The constitutive expression of genes encoding extracellular proteases, transcription factors, and cytokines is often correlated with cell transformation. In several instances, the uncontrolled expression of these genes is the result of transcriptional activation. Therefore, much attention has been devoted to the study of promoter function in transformed cells. We now review the results of recent investigations on transformation-dependent gene expression. The activation of several transcription factors in oncogenically-transformed cells is described. Results regarding the regulation of promoters through PRD II/kappa B are presented for cells transformed by a variety of oncogenes. Finally, we discuss the significance of transcription factor activation in the process of cell transformation.
- Published
- 1992
- Full Text
- View/download PDF
50. The stringent response blocks DNA replication outside the ori region in Bacillus subtilis and at the origin in Escherichia coli.
- Author
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Levine A, Vannier F, Dehbi M, Henckes G, and Séror SJ
- Subjects
- Arginine analogs & derivatives, Arginine pharmacology, Bacillus subtilis metabolism, Bacterial Proteins biosynthesis, DNA, Bacterial biosynthesis, Escherichia coli metabolism, Kinetics, Mutation, Nucleic Acid Hybridization, Temperature, Bacillus subtilis genetics, Chromosomes, Bacterial metabolism, DNA Replication, Escherichia coli genetics, Gene Expression Regulation, Bacterial
- Abstract
When the Bacillus subtilis dnaB37 mutant, defective in initiation, is returned to permissive temperature after growth at 45 degrees C, DNA replication is synchronized. Under these conditions, we have shown previously that DNA replication is inhibited when the Stringent Response is induced by the amino acid analogue, arginine hydroxamate. We have now shown, using DNA-DNA hybridization analysis, that substantial replication of the oriC region nevertheless occurs during the Stringent Response, and that replication inhibition is therefore implemented downstream from the origin. On the left arm, replication continues for at least 190 x 10(3) base-pairs to the gnt gene and for a similar distance on the right arm to the gerD gene. When the Stringent Response is lifted, DNA replication resumed downstream from oriC on both arms, confirming that DNA replication is regulated at a post-initiation level during the Stringent Response in B. subtilis. Resumption of DNA synthesis following the lifting of the Stringent Response did not require protein or RNA synthesis or the initiation protein DnaB. We suggest, therefore, that a specific control region, involving Stringent Control sites, facilitate reversible inhibition of fork movement downstream from the origin via modifications of a replisome component during the Stringent Response. In contrast, in Escherichia coli, induction of the Stringent Response appears to block initiation of DNA replication at oriC itself. No DNA synthesis was detected in the oriC region and, upon lifting the Stringent Response, replication occurred from oriC. Post-initiation control in B. subtilis therefore results in duplication of many key genes involved in growth and sporulation. We discuss the possibility that such a control might be linked to differentiation in this organism.
- Published
- 1991
- Full Text
- View/download PDF
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