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Recent advances in gene expression analysis techniques and increased access to technologies such as microarrays, qPCR arrays, and next-generation sequencing, in the last decade, have led to increased awareness of the complexity of the inflammatory responses that lead to pathology. This finding is also the case for rheumatic diseases, importantly and specifically, rheumatoid arthritis (RA). The coincidence in major genetic and epigenetic regulatory events leading to RA's inflammatory state is now well-recognized. Research groups have characterized the gene expression profile of early RA patients and identified a group of miRNAs that is particularly abundant in the early stages of the disease and miRNAs associated with treatment responses. In this perspective, we summarize the current state of RNA-based biomarker discovery and the context of technology adoption/implementation due to the COVID-19 pandemic. These advances have great potential for clinical application and could provide preclinical disease detection, follow-up, treatment targets, and biomarkers for treatment response monitoring. [ABSTRACT FROM AUTHOR]

Viral sepsis has been proposed as an accurate term to describe all multisystemic dysregulations and clinical findings in severe and critically ill COVID-19 patients. The adoption of this term may help the implementation of more accurate strategies of early diagnosis, prognosis, and in-hospital treatment. We accurately quantified 110 metabolites using targeted metabolomics, and 13 cytokines/chemokines in plasma samples of 121 COVID-19 patients with different levels of severity, and 37 non-COVID-19 individuals. Analyses revealed an integrated host-dependent dysregulation of inflammatory cytokines, neutrophil activation chemokines, glycolysis, mitochondrial metabolism, amino acid metabolism, polyamine synthesis, and lipid metabolism typical of sepsis processes distinctive of a mild disease. Dysregulated metabolites and cytokines/chemokines showed differential correlation patterns in mild and critically ill patients, indicating a crosstalk between metabolism and hyperinflammation. Using multivariate analysis, powerful models for diagnosis and prognosis of COVID-19 induced sepsis were generated, as well as for mortality prediction among septic patients. A metabolite panel made of kynurenine/tryptophan ratio, IL-6, LysoPC a C18:2, and phenylalanine discriminated non-COVID-19 from sepsis patients with an area under the curve (AUC (95%CI)) of 0.991 (0.986–0.995), with sensitivity of 0.978 (0.963–0.992) and specificity of 0.920 (0.890–0.949). The panel that included C10:2, IL-6, NLR, and C5 discriminated mild patients from sepsis patients with an AUC (95%CI) of 0.965 (0.952–0.977), with sensitivity of 0.993(0.984–1.000) and specificity of 0.851 (0.815–0.887). The panel with citric acid, LysoPC a C28:1, neutrophil-lymphocyte ratio (NLR) and kynurenine/tryptophan ratio discriminated severe patients from sepsis patients with an AUC (95%CI) of 0.829 (0.800–0.858), with sensitivity of 0.738 (0.695–0.781) and specificity of 0.781 (0.735–0.827). Septic patients who survived were different from those that did not survive with a model consisting of hippuric acid, along with the presence of Type II diabetes, with an AUC (95%CI) of 0.831 (0.788–0.874), with sensitivity of 0.765 (0.697–0.832) and specificity of 0.817 (0.770–0.865). [ABSTRACT FROM AUTHOR]

Skin insult and damage start a complex healing process that involves a myriad of coordinated reactions at both the cellular and molecular level occurring simultaneously. These processes can be divided into that of cell migration and tissue remodeling of the wound. In addition, it is well known that deep wounds that derive from surgical procedures need a multidisciplinary approach to have a successful wound healing process. Recently, there has been a renowned interest in the identification of active compounds derived from ornamental, edible, and wild plants being used in the cosmetic and skin product industry. Recent reports suggest that active components of several plants such as Propolis and Aloe vera could be used to induce the process of wound healing and tissue regeneration and reducing therefore the time to complete wound closure. Other plant species such as Achillea millefolium or Salvia officinalis have anti-inflammatory properties and promote cellular proliferation contributing to faster tissue regeneration. It has been described that Malva sylvestris influences the formation of fibrosis-free granulation tissue in the skin. Recent observations suggest that Casearia sylvestris induces the angiogenic process. These effects have been evaluated in cell lines, different animal models, and some in randomized clinical trials. In this review we summarize the evidence of plant extracts and their active components (when known) in the acceleration of the wound closure process and tissue repair. [ABSTRACT FROM AUTHOR]

Aging is a major health issue due to the increased susceptibility of elderly people to infectious, autoimmune, and cardiovascular diseases. Innate immunity is an important mechanism to avoid primary infections; therefore, decreasing of its activity may lead to development of infections. Antimicrobial peptides (AMPs) are effector molecules of innate immunity that can eliminate microbial invaders. The role that cytokines play in the regulation of these innate immune mechanisms needs to be explored. Serum determinations of Th1, Th2, and Th17 cytokines were performed in order to evaluate their association with AMPs human beta-defensin (HBD)-2 and LL-37 in young adults, elder adults, and elder adults with recurrent infections. Our results showed differences in interleukin (IL)-10 and IL-6 among the different groups. Inverse correlations in serum cytokine levels and HBD-2 production were identified for IL-10, IL-2, IL-4, tumor necrosis factor-α, and IL-6. Also inverse correlations were identified for IL-10, IL-4, and cathelicidin (LL-37). Such results could impact the development of immunomodulators that promote AMP production to prevent and/or contain infectious diseases in this population. [ABSTRACT FROM PUBLISHER]

Structure–activity relationship (SAR) studies allow the evaluation of the relationship between structural chemical changes and biological activity. Fluoroquinolones have chemical characteristics that allow their structure to be modified and new analogs with different therapeutic properties to be generated. The objective of this research is to identify and select the C-7 heterocycle fluoroquinolone analog (FQH 1–5) with antibacterial activity similar to the reference fluoroquinolone through in vitro, in silico, and in vivo evaluations. First, SAR analysis was conducted on the FQH 1–5, using an in vitro antimicrobial sensibility model in order to select the best compound. Then, an in silico model mechanism of action analysis was carried out by molecular docking. The non-bacterial cell cytotoxicity was evaluated, and finally, the antimicrobial potential was determined by an in vivo model of topical infection in mice. The results showed antimicrobial differences between the FQH 1–5 and Gram-positive and Gram-negative bacteria, identifying the 7-benzimidazol-1-yl-fluoroquinolone (FQH-2) as the most active against S. aureus. Suggesting the same mechanism of action as the other fluoroquinolones; no cytotoxic effects on non-bacterial cells were found. FQH-2 was demonstrated to decrease the amount of bacteria in infected wound tissue. [ABSTRACT FROM AUTHOR]

Tuberculosis is an ongoing threat to global health, especially with the emergence of multi drug-resistant (MDR) and extremely drug-resistant strains that are motivating the search for new treatment strategies. One potential strategy is immunotherapy using Innate Defence Regulator (IDR) peptides that selectively modulate innate immunity, enhancing chemokine induction and cell recruitment while suppressing potentially harmful inflammatory responses. IDR peptides possess only modest antimicrobial activity but have profound immunomodulatory functions that appear to be influential in resolving animal model infections. The IDR peptides HH2, 1018 and 1002 were tested for their activity against two M. tuberculosis strains, one drug-sensitive and the other MDR in both in vitro and in vivo models. All peptides showed no cytotoxic activity and only modest direct antimicrobial activity versus M. tuberculosis (MIC of 15–30 µg/ml). Nevertheless peptides HH2 and 1018 reduced bacillary loads in animal models with both the virulent drug susceptible H37Rv strain and an MDR isolate and, especially 1018 led to a considerable reduction in lung inflammation as revealed by decreased pneumonia. These results indicate that IDR peptides have potential as a novel immunotherapy against TB. [ABSTRACT FROM AUTHOR]

Abstract: Tuberculosis (TB) is one of the most important infectious diseases, causing 1.8 million deaths annually worldwide. This problem has increased because of the association with human immmunodeficiency virus and diabetes mellitus type 2, mainly in developing countries. In the past few years it has been highlighted the significance of antimicrobial peptides in the immunopathogenesis of TB ex vivo and in experimental models studies. In this study we analyzed the expression of CAMP, DEFA1, DEFB4, and DEFB103A in patients with latent TB and progressive TB with and without comorbidity with diabetes mellitus type 2. Antimicrobial peptide gene expression increased during progressive TB, which could be used as a biomarker for reactivation. By contrast, patients with diabetes mellitus type 2 have lower antimicrobial peptides gene expression, suggesting that the lack of its proper production in these patients contribute to enhance the risk for TB reactivation. [Copyright &y& Elsevier]

Autoantibodies have a central role in the physiopathology of Rheumatoid Arthritis (RA). However, the responsible factors that trigger and perpetuate the autoantibodies production are unknown. Toll-like receptors (TLRs) have been considered as promotors of autoantibodies production to break down the immunotolerance in RA. Evaluate the expression levels of TLR7 and TLR9 as well as their correlation with autoantibodies in first-degree relatives (FDR) of RA patients (seropositive and seronegative to ACPA), respect to early RA (eRA) and chronic RA (cRA) patients. We selected 32 RA patients (16 as eRA and 16 as cRA) and 32 FDR of RA patients (16 seropositive and 16 seronegative to ACPA). Expression levels of TLR7 and TLR9 in whole blood samples from each group were measured by real-time PCR using total RNA extracted from each subject. Also, correlation analysis between TLRs expression and autoantibodies was performed. The expression of TLR7 and TLR9 was diminished in RA patients (p <0.01) but elevated in ACPA– FDR (p <0.0001) and ACPA+ FDR (p <0.05) with a positive correlation between them (r = 0.749, p <0.000). Moreover, the expression levels of TLR7 correlate positively with ACPA levels in both seropositive ACPA+ FDR subjects (r = 0.582, p = 0.018) and eRA patients (r = 0.593, p = 0.020). Our results showed overexpression of TLR7 and TLR9 may occur in preclinical RA subjects. TLR7 overexpression correlated with ACPA levels' production, suggesting TLR7 may play a role in ACPA development. [Display omitted] [ABSTRACT FROM AUTHOR]

Type 2 diabetes (DM2) is an increasingly prevalent disease that challenges tuberculosis (TB) control strategies worldwide. It is significant that DM2 patients with poor glycemic control (PDM2) are prone to developing tuberculosis. Furthermore, elucidating the molecular mechanisms that govern this susceptibility is imperative to address this problem. Therefore, a pilot transcriptomic study was performed. Human blood samples from healthy controls (CTRL, HbA1c < 6.5%), tuberculosis (TB), comorbidity TB-DM2, DM2 (HbA1c 6.5–8.9%), and PDM2 (HbA1c > 10%) groups (n = 4 each) were analyzed by differential expression using microarrays. We use a network strategy to identify potential molecular patterns linking the differentially expressed genes (DEGs) specific for TB-DM2 and PDM2 (p-value < 0.05, fold change > 2). We define OSM, PRKCD, and SOCS3 as key regulatory genes (KRGs) that modulate the immune system and related pathways. RT-qPCR assays confirmed upregulation of OSM, PRKCD, and SOCS3 genes (p < 0.05) in TB-DM2 patients (n = 18) compared to CTRL, DM2, PDM2, or TB groups (n = 17, 19, 15, and 9, respectively). Furthermore, OSM, PRKCD, and SOCS3 were associated with PDM2 susceptibility pathways toward TB-DM2 and formed a putative protein–protein interaction confirmed in STRING. Our results reveal potential molecular patterns where OSM, PRKCD, and SOCS3 are KRGs underlying the compromised immune response and susceptibility of patients with PDM2 to develop tuberculosis. Therefore, this work paved the way for fundamental research of new molecular targets in TB-DM2. Addressing their cellular implications, and the impact on the diagnosis, treatment, and clinical management of TB-DM2 could help improve the strategy to end tuberculosis for this vulnerable population. [ABSTRACT FROM AUTHOR]

The first degree relatives of rheumatoid arthritis (RA) patients have a higher risk of developing RA, which is related to the expression of autoantibodies against citrullinated proteins (ACPA). Remarkably, prior to the onset of RA, cartilage damage is already initiated, whereas ACPA autoantibodies are already expressed. Here we show that both TNF-α and IL-6 are also increased prior to the onset of RA. Furthermore, when the levels of DKK1 and Sclerostin were evaluated in first degree relatives of RA patients, we found that the serum levels of TNF- α correlate with the expression levels of both DKK1 and Sclerostin. Interestingly, when the disease is already established, the correlation of TNF- α with DKK1 is lost in RA patients, whereas the correlation of Sclerostin with both TNF- α and IL-6 is further increased. Our data suggest a subclinical inflammation in patients at high risk of developing RA, which might lead to an increase in the levels of both DKK1 and Sclerostin, contributing to joint damage in the preclinical phase of the disease linked to the expression of ACPA autoantibodies. [ABSTRACT FROM AUTHOR]

Genomics has significantly revolutionized pathogen surveillance, particularly in epidemiological studies, the detection of drug-resistant strains, and disease control. Despite its potential, the representation of Latin American countries in the genomic catalogues of Mycobacterium tuberculosis (Mtb), the bacteria responsible for Tuberculosis (TB), remains limited. In this study, we present a whole genome sequencing (WGS)-based analysis of 85 Mtb clinical strains from 17 Mexican states, providing insights into local adaptations and drug resistance signatures in the region. Our results reveal that the Euro-American lineage (L4) accounts for 94% of our dataset, showing 4.1.2.1 (Haarlem, n = 32), and 4.1.1.3 (X-type, n = 34) sublineages as the most prevalent. We report the presence of the 4.1.1.3 sublineage, which is endemic to Mexico, in six additional locations beyond previous reports. Phenotypic drug resistance tests showed that 34 out of 85 Mtb samples were resistant, exhibiting a variety of resistance profiles to the first-line antibiotics tested. We observed high levels of discrepancy between phenotype and genotype associated with drug resistance in our dataset, including pyrazinamide-monoresistant Mtb strains lacking canonical variants of drug resistance. Expanding the Latin American Mtb genome databases will enhance our understanding of TB epidemiology and potentially provide new avenues for controlling the disease in the region. [ABSTRACT FROM AUTHOR]

The early diagnosis of diabetic nephropathy (DN) is essential for improving the prognosis and effectively manage patients affected with this disease. The standard biomarkers, including albuminuria and glomerular filtration rate, are not very precise. New molecular biomarkers are needed to more accurately identify DN and better predict disease progression. Characteristic DN biomarkers can be identified using transcriptomic analysis. To evaluate the transcriptomic profile of controls (CTRLs, n = 15), patients with prediabetes (PREDM, n = 15), patients with type-2 diabetes mellitus (DM2, n = 15), and patients with DN (n = 15) by microarray analysis to find new biomarkers. RT-PCR was then used to confirm gene biomarkers specific for DN. Blood samples were used to isolate RNA for microarray expression analysis. 26,803 unique gene sequences and 30,606 LncRNA sequences were evaluated-Selected gene biomarkers for DN were validated using qPCR assays. Sensitivity, specificity, and area under the curve (AUC) were calculated as measures of diagnostic accuracy. The DN transcriptome was composed of 300 induced genes, compared to CTRLs, PREDM, and DM-2 groups. RT-qPCR assays validated that METLL22, PFKL, CCNB1 and CASP2 genes were induced in the DN group compared to CTRLs, PREDM, and DM-2 groups. The ROC analysis for these four genes showed 0.9719, 0.8853, 0.8533 and 0.7748 AUC values, respectively. Among induced genes in the DN group, we found that CASP2, PFKL and CCNB1 may potentially be used as biomarkers to diagnose DN. Of these, METLL22 had the highest AUC score, at 0.9719. [ABSTRACT FROM AUTHOR]

Background and Aims Type 2 diabetes mellitus (DM2) confers a higher risk for active tuberculosis (TB). However, information on associated risk factors for latent tuberculosis infection (LTBI) inpatients with DM2 is limited. We conducted a cross-sectional study to elucidate the prevalence of LTBI and its associated factors on Mexican adults with DM2 receiving medical care at the Mexican Social Security Institute (IMSS). Methods Six hundred patients with DM2 without a prior history of TB from outpatient diabetes clinics were enrolled in the study. The tuberculin-skin-test (TST) was performed. The presence of LTBI was defined by a TST value of ≥5 mm. A standardized interview and physical examination were conducted to obtain clinical, demographic, and LTBI risk factor information; all subjects were laboratory tested to determine the presence of exclusion criteria. Microscopic examination of sputum samples and chest x-rays was performed to identify potential active TB. Subjects with any finding suggesting active TB or malignancy were excluded. A logistic regression model was used to identify variables associated with LTBI. Results LTBI prevalence among patients with DM2 was 51.3%. Risk factors for LTBI were living with a relative with TB, having been in prison, having hemoglobin values >14 g/dL, and glycosylated hemoglobin (HbA1c) values of >7%. Blood pressure, economic income, or anthropometric measurements were not associated risk factors. Conclusions Over one half of patients with DM harbor LTBI. Exposure to certain environmental conditions and poorly controlled DM2 (HbA1c >7.0%) were risk factors for having LTBI in persons with DM2. [ABSTRACT FROM AUTHOR]

Peptidyl arginine deiminases (PAD) are proteins that modify arginine residues to citrulline. In humans there are five isoforms PAD1-4 and PAD6. PAD4 is involved in several human diseases such as Alzheimer, breast cancer, rheumatoid arthritis and therefore has been recognized as a potential pharmacological target. In this work, 3175 molecules retrieved from ZINC12 Food and Drug Administration (FDA) catalog database were assayed through a structure-based virtual screening to identify potential PAD4 binders. Three were confirmed through enzyme inhibition studies. Flow cytometry analysis were used to check for drug cytotoxic effects on blood cells from three healthy donors. Molecular docking in a wild-type PAD4 structure showed the top 20 FDA molecules with the best binding energies. Those molecules were distributed in clusters of mixed binding energies but with similar chemical structures. These patterns along with the FDA information were used to aid in the rational selection of methotrexate, testosterone and leucovorin compounds for in vitro evaluations. Time course fluorescent enzyme kinetics confirmed that methotrexate (0.01–1 mmol) and testosterone enanthate (49.9–12.5 mmol) inhibit PAD4 at similar levels to the known inhibitor BB-Cl-amidine (8.8 mmol), except leucovorin. The chosen concentrations lack cytotoxic effects on blood cells. These findings encourage repositioning campaigns in the quest of PAD4 inhibitors. [ABSTRACT FROM AUTHOR]

The increasing number of people with type 2 diabetes (DM2) is alarming and if it is taken into account that the relative odds of developing tuberculosis in diabetic patients ranges from 2.44 to 8.33 compared with non-diabetic patients, thus in developing countries where these two diseases are encountering face to face, there is a need for prophylaxis strategies. The role of vitamin D has been widely implicated in growth control of Mycobacterium tuberculosis (Mtb) during primary infection mainly through the induction of certain antimicrobial peptides (AMPs). In this study we evaluated the vitamin D serum levels, CYP27B1-hydroxylase enzyme, vitamin D receptor (VDR) and AMPs gene expression in Healthy donors, DM2 and TB patients. Results showed that DM2 group has lower VDR and AMPs expression levels. When Monocytes Derived Macrophages (MDM) from DM2 patients with low VDR expression were supplemented with vitamin D, MDMs eliminate efficiently M. tuberculosis. This preliminary study suggests the use of vitamin D as prophylaxis for tuberculosis in high DM2 endemic countries. [ABSTRACT FROM AUTHOR]

Antibiotics resistance is a health problem worldwide, which is characterized by low or no effective treatments against bacterial infections that increase morbidity and mortality in septicemia patients. In Mexico, part of the problem has been attributed to self-medication and uncontrolled antibiotics sale. Today is necessary to emphasize regulation of pharmaceuticals classified as generic or similar drugs because they lack of quality control, which may cause low effectiveness of the drug and thus promote drug resistance in the Mexican population. In this paper we analyzed the drug resistance prevalence in bacteria identified in hemocultures obtained of patients with septicemia from Mexico City. We found that drugs such as linezolid, imipenem and gatifloxacin are the most effective drugs against bacterial strains with the highest rate of drug resistance in this population. However, penicillins were less effective drugs because of drug resistant bacteria. We also found that the bacteria with more drug resistance index in our studies were Staphylococcus hominis, Acinetobacter baumannii and Pseudomonas aeruginosa. [ABSTRACT FROM AUTHOR]

Abstract: Five variants of human β-defensins (HBDs) were expressed in Escherichia coli using two vector systems (pET28a(+) and pQE30) with inducible expression by IPTG. The last vector has not been previously reported as an expression system for HBDs. The recombinant peptides were different in their lengths and overall charge. The HBDs were expressed as soluble or insoluble proteins depending on the expression system used, and the final protein yields ranged from 0.5 to 1.6mg of peptide/g of wet weight cells, with purities higher than 90%. The recombinant HBDs demonstrated a direct correlation between antimicrobial activity and the number of basic charged residues; that is, their antimicrobial activity was as follows: HBD3-M-HBD2>HBD3=HBD3-M=HB2-KLK>HBD2 when assayed against E. coli, Staphylococcus aureus and Pseudomonas aeruginosa. Interestingly, HBD2 had the best antimicrobial activity against the Mycobacterium tuberculosis strain H37Rv (1.5μM) and the heterologous tandem peptide, HBD3-M-HBD2, had the best minimal inhibitory concentration (MIC) value (2.7μM) against a multidrug resistance strain (MDR) of M. tuberculosis, demonstrating the feasibility of the use of HBDs against pathogenic M. tuberculosis reported to be resistant to commercial antibiotics. [Copyright &y& Elsevier]

Abstract: Tuberculosis (TB) is a major worldwide health problem in part due to the lack of development of new treatments and the emergence of new strains such as multidrug-resistant (MDR) and extensively drug-resistant strains that are threatening and impairing the control of this disease. In this study, the efficacy of natural and synthetic cationic antimicrobial (host defence) peptides that have been shown often to possess broad-spectrum antimicrobial activity was tested. The natural antimicrobial peptides human LL-37 and mouse CRAMP as well as synthetic peptides E2, E6 and CP26 were tested for their activity against Mycobacterium tuberculosis both in in vitro and in vivo models. The peptides had moderate antimicrobial activities, with minimum inhibitory concentrations ranging from 2μg/mL to 10μg/mL. In a virulent model of M. tuberculosis lung infection, intratracheal therapeutic application of these peptides three times a week at doses of ca. 1mg/kg led to significant 3–10-fold reductions in lung bacilli after 28–30 days of treatment. The treatments worked both against the drug-sensitive H37Rv strain and a MDR strain. These results indicate that antimicrobial peptides might constitute a novel therapy against TB. [Copyright &y& Elsevier]

Type 2 diabetes mellitus (T2D) is a risk factor for the development of tuberculosis (TB) through mechanisms poorly understood. Monocytes and macrophages are key effector cells to control TB, but they are also subverted by Mycobacterium tuberculosis (Mtb). Specifically, Mtb can induce a bystander effect that skews monocyte differentiation towards macrophages with a permissive phenotype to infection. Here, we evaluated whether T2D impacts this TB aspect. Our approach was to differentiate monocytes from healthy control (HC) subjects and T2D patients into macrophages (MDM), and then assess their response to Mtb infection, including their secretome content and bystander effect capacity. Through flow cytometric analyses, we found a lower level of activation markers in MDM from T2D patients than from HC in response to mock (HLA‐DR, CD86 and CD163) or Mtb challenge (CD14 and CD80). In spite of high TGF‐β1 levels in mock‐infected MDM from T2D patients, cytometric bead arrays indicated that there were no major differences in the secretome cytokine content in these cells relative to HC‐MDM, even in response to Mtb. Mimicking a bystander effect, the secretome of Mtb‐infected HC‐MDM drove HC monocytes towards MDM with a permissive phenotype for Mtb intracellular growth. However, the secretome from Mtb‐infected T2D‐MDM did not exacerbate the Mtb load compared to secretome from Mtb‐infected HC‐MDM, possibly due to the high IL‐1β production relative to Mtb‐infected HC‐MDM. Collectively, despite T2D affecting the basal MDM activation, our approach revealed that it has no major consequence on their response to Mtb or capacity to generate a bystander effect influencing monocyte differentiation. [ABSTRACT FROM AUTHOR]

Introduction: The formation of neutrophil extracellular traps (NETs) is a process in which several kinds of enzymes participate generating posttranslational modifications of proteins. NETs have been associated with infectious, autoimmune, and inflammatory diseases. Inhibition of several proteases reduces the formation of NETs. In the present work, we analyzed the role of several broad-acting and specific inhibitors of proteases in the formation of NETs. Methods: Neutrophils were isolated from peripheral blood of healthy individuals by density gradient. The neutrophils were quantified and seeded into cell culture plates. Phorbol myristate acetate and A23187 were used as NETs inducers, and several specific inhibitors of proteases were used. The cells were stained for cytoskeleton or DNA. The cell-free supernatants were used to assess DNA release. Statistical analysis was carried out by a Kruskal-Wallis or ANOVA test. Results: We observed marked changes in actin organization after the induction of NETs, suggesting that the cytoskeleton is being actively regulated. When we used protease inhibitors, the release of DNA was reduced, suggesting the participation of actin remodeling in the process. Further characterization of the specific proteases revealed that calpain modulates the reorganization of actin cytoskeleton and DNA release. Preservation of part of the actin cytoskeleton suggests that DNA release is not only a mechanic process associated to the chromatin decondensation; rather the process is highly regulated by active proteases that promote cytoskeleton reorganization and chromatin decondensation that culminates in DNA release. Conclusion: Calpain mediates the DNA release in the NET formation process by the modification of cortical actin cytoskeleton in a calcium-dependent manner. [ABSTRACT FROM AUTHOR]

The MERS-CoV, SARS-CoV, and SARS-CoV-2 are highly pathogenic viruses that can cause severe pneumonic diseases in humans. Unfortunately, there is a non-available effective treatment to combat these viruses. Domain-motif interactions (DMIs) are an essential means by which viruses mimic and hijack the biological processes of host cells. To disentangle how viruses achieve this process can help to develop new rational therapies. Data mining was performed to obtain DMIs stored as regular expressions (regexp) in 3DID and ELM databases. The mined regexp information was mapped on the coronaviruses' proteomes. Most motifs on viral protein that could interact with human proteins are shared across the coronavirus species, indicating that molecular mimicry is a common strategy for coronavirus infection. Enrichment ontology analysis for protein domains showed a shared biological process and molecular function terms related to carbon source utilization and potassium channel regulation. Some of the mapped motifs were nested on B, and T cell epitopes, suggesting that it could be as an alternative way for reverse vaccinology. The information obtained in this study could be used for further theoretic and experimental explorations on coronavirus infection mechanism and development of medicines for treatment. [ABSTRACT FROM AUTHOR]

Type 2 diabetes mellitus (DM2) is strongly associated with other comorbidities such as obesity, atherosclerosis, and hypertension. Obesity is associated with sustained low-grade inflammatory response due to the production of proinflammatory cytokines. This inflammatory process promotes the differentiation of some myeloid cells, including myeloid-derived suppressor cells (MDSCs). In this study, two groups of individuals were included: DM2 patients and non-DM2 individuals with similar characteristics. Immunolabeling of CD15+ CD14- and CD33+ HLA-DR-/low was performed from whole peripheral blood, and samples were analyzed by flow cytometry, and frequencies of MDSCs and the relationship of these with clinical variables, cytokine profile (measured by cytometric bead array), and anthropometric variables were analyzed. The frequency of CD33+ HLA-DR-/low MDSCs (that produce IL-10 and TGF-β, according to an intracellular detection) is higher in patients with DM2 (P<0.05), and there is a positive correlation between the frequency of CD15+ CD14- and CD33+ HLA-DR-/low MDSC phenotypes. DM2 patients have an increased concentration of serum IL-5 (P<0.05). Also, a negative correlation between the frequency of CD15+ CD14- MDSCs and LDL cholesterol was found. Our group of DM2 patients have an increased frequency of mononuclear MDSC CD33+ HLA-DR-/low that produce TGF-β and IL-10. These cytokines have been associated with immune modulation and reduced T cell responses. DM2 and non-DM2 subjects show a similar cytokine profile, but the DM2 patients have an increased concentration of IL-5. [ABSTRACT FROM AUTHOR]

Chronic exposure to arsenic (As), whether directly through the consumption of contaminated drinking water or indirectly through the daily intake of As-contaminated food, is a health threat for more than 150 million people worldwide. Epidemiological studies found an association between chronic consumption of As and several pathologies, the most common being cancer-related disorders. However, As consumption has also been associated with metabolic disorders that could lead to diverse pathologies, such as type 2 diabetes mellitus, nonalcoholic fatty liver disease, and obesity. Here, we used ultra-performance liquid chromatography (UPLC) coupled to electrospray ionization/quadrupole time-of-flight mass spectrometry (ESI-QToF) to assess the effect of chronic intergenerational As exposure on the lipid metabolism profiles of serum from 4-month-old Wistar rats exposed to As prenatally and also during early life in drinking water (3 ppm). Significant differences in the levels of certain identified lysophospholipids, phosphatidylcholines, and triglycerides were found between the exposed rats and the control groups, as well as between the sexes. Significantly increased lipid oxidation determined by the malondialdehyde (MDA) method was found in exposed rats compared with controls. Chronic intergenerational As exposure alters the rat lipidome, increases lipid oxidation, and dysregulates metabolic pathways, the factors associated with the chronic inflammation present in different diseases associated with chronic exposure to As (i.e., keratosis, Bowen's disease, and kidney, liver, bladder, and lung cancer). [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA) is a systemic autoimmune disorder characterized by chronic and symmetrical inflammation of synovial tissue with subsequent joint destruction. SUMO1 is an important regulator of apoptosis through non‐canonical mechanism in synovial fibroblasts, and POU2AF1 is a known B‐cell transcriptional co‐activator. The specific objective of this study was to measure the expression of SUMO1 and POU2AF1 on first‐degree relatives of patients with RA and also in the preclinical and clinical stages of RA and describe their possible role in RA physiopathology. Blood samples were collected from ACPA+, ACPA‐, early and established RA subjects recruited. ACPAs and CarP autoantibodies were determined by ELISA Eurodiagnostica CCplus kit according to previously described protocols. RNA was isolated from blood samples; the purity as integrity was determined. Gene expression analysis was made by RT‐qPCR using specific primers for SUMO1 and POU2AF1 mRNAs; relative expression was determined according to the 2‐ΔΔct method procedure. Significant differences in the expression of both, SUMO1 and POU2AF1 were identified when comparing arthritis versus healthy or ACPA+ individuals, suggesting that the down regulation of such genes starts after the onset of symptoms in RA patients. Also, a significant correlation was identified for POU2AF1 and disease progression whit a downward trend for those with established RA. The implications of such gene down regulation are discussed in the context of RA physiopathology. [ABSTRACT FROM AUTHOR]

Type-2 Diabetes (T2D) is a predisposing cause for developing tuberculosis (TB) in low- and middle-income countries. TB-T2D comorbidity worsens clinical control and prognosis of the affected individuals. The underlying metabolic alterations for this infectious-metabolic disease are still largely unknown. Possible mediators of the increased susceptibility to TB in diabetic patients are lipids levels, which are altered in individuals with T2D. To evaluate the modulation of glycerophospholipids in patients with TB-T2D, an untargeted lipidomic approach was developed by means of ultra-performance liquid chromatography (UPLC) coupled to electrospray ionization/quadrupole time-of-flight mass spectrometry (ESI-QToF). In addition, tandem mass spectrometry was performed to determine the identity of the differentially expressed metabolites. We found that TB infected individuals with or without T2D share a common glycerophospholipid profile characterized by a decrease in phosphatidylcholines. A total of 14 glycerophospholipids were differentially deregulated in TB and TB-T2D patients and could potentially be considered biomarkers. It is necessary to further validate these identified lipids as biomarkers, focusing on the anticipate diagnosis for TB development in T2D predisposed individuals. [ABSTRACT FROM AUTHOR]

Abstract The physiopathology of rheumatoid arthritis (RA) is mediated by proinflammatory cytokines, some of which are regulated by the JAK/STAT pathway. Tofacitinib is a JAK inhibitor, but its role in the regulation of microRNAs (miRNAs) is unknown. There is also no information regarding the role of miRNAs in the clinical relapse/remission of RA. The present project aims to identify a signature profile of miRNA expression in a subgroup of RA patients who had to discontinue tofacitinib treatment (because of the ending of a 5-year open-label clinical trial) and to describe the expression of miRNAs during RA remission or flare-up. The relative expression of 61 miRNAs was determined in serum samples with the Firefly™ BioWorks assay. Statistical analysis was performed by means of Student's t -test and heatmap analysis was performed with Firefly™ Analysis Workbench software and in the software GraphPad® Prism v5.0. Target prediction and Gene Ontology analysis were carried out using bioinformatic tools. We found a distinctive signature of miRNA expression associated with relapse, featuring upregulated expression of hsa‑miR‑432‑5p ( p < 0.05). We also found upregulation of hsa‑miR‑194‑5p ( p < 0.05) in samples of patients with RA flare-up. Gene Ontology analysis of the target genes for hsa‑miR‑432‑5p was performed to identify relevant pathways associated with relapse; the implications of these pathways in the physiopathology of RA are discussed. Tofacitinib treatment does not have a direct effect on the expression of measured miRNAs. The changes in hsa‑miR‑432‑5p and hsa‑miR‑194‑5p are associated with the regulation of proinflammatory pathways and RA flare-up. Highlights • hsa‑miR‑432‑5p is downregulated in patients achieving clinical remission. • Treatment with Tofacitinib does not affect miRNA expression directly. • Patients with relapse have a specific miRNA signature as measured by Firefly Bioworks. • hsa‑miR‑194-5p is upregulated during RA flare-up patients suggesting that this miRNA could be used as a biomarker for relapse. [ABSTRACT FROM AUTHOR]

Background: Several epidemiological studies in diabetic patients have demonstrated a protective effect of metformin to the development of several types of cancer. The underlying mechanisms of such phenomenon is related to the effect of metformin on cell proliferation among which, mTOR, AMPK and other targets have been identified. However, little is known about the role that metformin treatment have on other cell types such as keratinocytes and whether exposure to metformin of these cells might have serious repercussions in wound healing delay and in the development of complications in diabetic patients with foot ulcers or in their exacerbation. Material and Methods: HaCaT Cells were exposed to various concentrations of metformin and cell viability was evaluated by a Resazurin assay; Proliferation was also evaluated with a colony formation assay and with CFSE dilution assay by flow cytometry. Cell cycle was also evaluated by flow cytometry by PI staining. An animal model of wound healing was used to evaluate the effect of metformin in wound closure. Also, an analysis of patients receiving metformin treatment was performed to determine the effect of metformin treatment on the outcome and wound area. Statistical analysis was performed on SPSS v. 18 and GraphPad software v.5. Results: Metformin treatment significantly reduces cell proliferation; colony formation and alterations of the cell cycle are observed also in the metformin treated cells, particularly in the S phase. There is a significant increase in the area of the wound of the metformin treated animals at different time points (P<0.05). There is also a significant increase in the size and wound area of the patients with diabetic foot ulcers at the time of hospitalization. A protective effect of metformin was observed for amputation, probably associated with the anti inflammatory effects reported of metformin. Conclusions: Metformin treatment reduces cell proliferation and reduces wound healing in an animal model and affects clinical outcomes in diabetic foot ulcer patients. Chronic use of this drug should be further investigated to provide evidence of their security in association with DFU. [ABSTRACT FROM AUTHOR]

Abstract: The World Health Organization (WHO) has estimated that there are about 8 million new cases annually of active Tuberculosis (TB). Despite its irregular effectiveness (0–89%), the Bacillus Calmette-Guérin) BCG is the only vaccine available worldwide for prevention of TB; thus, the design is important of novel and more efficient vaccination strategies. Considering that β-defensin-2 is an antimicrobial peptide that induces dendritic cell maturation through the TLR-4 receptor and that both ESAT-6 and Ag85B are immunodominant mycobacterial antigens and efficient activators of the protective immune response, we constructed two DNA vaccines by the fusion of the gene encoding β-defensin-2 and antigens ESAT6 (pDE) and 85B (pDA). After confirming efficient local antigen expression that induced high and stable Interferon gamma (IFN-γ) production in intramuscular (i.m.) vaccinated Balb/c mice, groups of mice were vaccinated with DNA vaccines in a prime-boost regimen with BCG and with BCG alone, and 2 months later were challenged with the mild virulence reference strain H37Rv and the highly virulent clinical isolate LAM 5186. The level of protection was evaluated by survival, lung bacilli burdens, and extension of tissue damage (pneumonia). Vaccination with both DNA vaccines showed similar protection to that of BCG. After the challenge with the highly virulent Mycobacterium tuberculosis strain, animals that were prime-boosted with BCG and then boosted with both DNA vaccines showed significant higher survival and less tissue damage than mice vaccinated only with BCG. These results suggest that improvement of BCG vaccination, such as the prime-boost DNA vaccine, represents a more efficient vaccination scheme against TB. [Copyright &y& Elsevier]

Abstract: Background: Foot ulcers are one of the main diabetes complications due to its high frequency and difficulty of complete healing. There are several factors that participate in diabetic ulcers development and limited information exists about the role of antimicrobial peptides (AMP) in its pathogenesis. Objective: The aim of this study was to analyze the expression pattern of the main AMPs: Human Neutrophil Peptide (HNP)-1, Human β-defensin (HBD)-1, HBD-2, HBD-3, HBD-4 and cathelicidin LL-37 in biopsies from diabetic foot ulcers (DFU). Methods: 20 biopsies from DFU grade 3 according to Wagner''s classification and 20 biopsies from healthy donors were obtained. Real time PCR, immunohistochemistry and primary cell cultures were performed. Results: β-Defensins were overexpressed in DFU, whereas LL-37 has low or none expression in comparison with healthy skin. When primary cell culture from these biopsies were performed and infected with Staphylococcus aureus, epidermal cell from diabetic ulcers showed lower LL-37 expression compared with cell cultures from healthy donors skin. Conclusion: These results suggest that though most AMPs are expressed in DFU, this production is not appropriate to promote wound healing and contain secondary infections. [Copyright &y& Elsevier]

Differences in clinical manifestations, immune response, metabolic alterations, and outcomes (including disease severity and mortality) between men and women with COVID-19 have been reported since the pandemic outbreak, making it necessary to implement sex-specific biomarkers for disease diagnosis and treatment. This study aimed to identify sex-associated differences in COVID-19 patients by means of a genetic algorithm (GALGO) and machine learning, employing support vector machine (SVM) and logistic regression (LR) for the data analysis. Both algorithms identified kynurenine and hemoglobin as the most important variables to distinguish between men and women with COVID-19. LR and SVM identified C10:1, cough, and lysoPC a 14:0 to discriminate between men with COVID-19 from men without, with LR being the best model. In the case of women with COVID-19 vs. women without, SVM had a higher performance, and both models identified a higher number of variables, including 10:2, lysoPC a C26:0, lysoPC a C28:0, alpha-ketoglutaric acid, lactic acid, cough, fever, anosmia, and dysgeusia. Our results demonstrate that differences in sexes have implications in the diagnosis and outcome of the disease. Further, genetic and machine learning algorithms are useful tools to predict sex-associated differences in COVID-19. [ABSTRACT FROM AUTHOR]

Ultraviolet (UV) exposure has been linked to skin damage and carcinogenesis, but recently UVB has been proposed as a therapeutic approach for cancer. Herein, we investigated the cellular and molecular effects of UVB in immortal and tumorigenic HPV positive and negative cells. Cells were irradiated with 220.5 to 1102.5 J/m2 of UVB and cell proliferation was evaluated by crystal violet, while cell cycle arrest and apoptosis analysis were performed through flow cytometry. UVB effect on cells was recorded at 661.5 J/m2 and it was exacerbated at 1102.5 J/m2. All cell lines were affected by proliferation inhibition, cell cycle ablation and apoptosis induction, with different degrees depending on tumorigenesis level or HPV type. Analysis of the well-known UV-responsive p53, E2F1 and microtubules system proteins was performed in SiHa cells in response to UVB through Western-blotting assays. E2F1 and the Microtubule-associated protein 2 (MAP2) expression decrease correlated with cellular processes alteration while p53 and Microtubule-associated Protein 1S (MAP1S) expression switch was observed since 882 J/m2, suggesting they were required under more severe cellular damage. However, expression transition of α-Tubulin3C and β-Tubulin was abruptly noticed until 1102.5 J/m2 and particularly, γ-Tubulin protein expression remained without alteration. This study provides insights into the effect of UVB in cervical cancer cell lines. [ABSTRACT FROM AUTHOR]

The micro RNA (miR)-34 family is composed of 5p and 3p strands of miR-34a, miR-34b, and miR-34c. The 5p strand's expression and function is studied in cervical cancer. The 3p strand's function and regulation remain to be elucidated. To study the function of the passenger strands of miR-34 family members, we overexpressed 5p and 3p strands using a synthetic miRNA in cervical cell lines. Cell proliferation was evaluated using crystal violet. Migration and invasion were tested using transwell assays, Western blot, and zymography. Possible specific targets and cell signaling were investigated for each strand. We found that miR-34a-5p inhibited proliferation, migration, and cell invasion accompanied by matrix metalloproteinase 9 (MMP9) activity and microtubule-associated protein 2 (MAP2) protein reduction. We also found that miR-34b-5p and miR-34c-5p inhibit proliferation and migration, but not invasion. In contrast, miR-34c-5p inhibits MMP9 activity and MAP2 protein, while miR-34b-5p has no effect on these genes. Furthermore, miR-34a-3p and miR-34b-3p inhibit proliferation and migration, but not invasion, despite the later reducing MMP2 activity, while miR-34c-3p inhibit proliferation, migration, and cell invasion accompanied by MMP9 activity and MAP2 protein inhibition. The difference in cellular processes, MMP2 and MMP9 activity, and MAP2 protein inhibition by miR-34 family members suggests the participation of other regulated genes. This study provides insights into the roles of passenger strands (strand*) of the miR-34 family in cervical cancer. [ABSTRACT FROM AUTHOR]

Background Pulmonary tuberculosis (PTB) is a public health problem with 10.4 million new cases reported in 2017 (1). According to the World Health Organization (WHO), accurate diagnostic tests based in serum biomarkers to detect new cases of tuberculosis are necessary. Aim of the study To evaluate antibodies against Mycobacterium. tuberculosis (Mtb) peptides (Ab-Mtb) and three soluble host biomarkers by ELISA serial multiple test in sera from non-infected controls (NIC, n = 31), latent tuberculosis (LTB, n = 37) and PTB (n = 28) patients in a diagnosis tuberculosis assay. Materials and methods Levels of four Ab-Mtb peptides derived from Mtb and three host response molecules in serum from NIC, LTB and PTB were evaluated by ELISA as tuberculosis biomarkers. Multiple comparisons tests, determination of diagnostic values and ROC curves were performed. Serial and parallel multiple tests were performed with the biomarkers with the highest discriminatory capacity to improve diagnostic values of the test. Results We found significant differences between biomarkers levels in PTB comparing LTB and NIC to all candidate biomarkers; peptides P12033, P12037, and serum biomarkers such as sCD14 and chemokine CXCL9 showed the best sensitivity and specificity, the highest discriminatory power, and the best area under the curve (AUC) individually. In serial multiple tests, P12037 and sCD14 together have 92% of sensitivity and 91% of specificity, with positive and negative likelihood ratios greater than 10. Conclusions Ab-Mtb peptide P12037 and sCD14 could be applied in a diagnostic test for suspected PTB to improve accuracy and time to diagnosis and could be implemented in a POCT device which can be affordable. [ABSTRACT FROM AUTHOR]

An abstract of the article "nduction of Defensins by L-Isoleucine as Novel Immunotherapy in Experimental Murine Tuberculosis" by Bruno Rivas-Santiago, Diana Aguilar-Leon, and colleagues is presented.