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Brucella, a zoonotic facultative intracellular pathogenic bacterium, poses a significant threat both to human health and to the development of the livestock industry. Alanine racemase (Alr), the enzyme responsible for alanine racemization, plays a pivotal role in regulating virulence in this bacterium. Moreover, Brucella mutants with alr gene deletions (Δalr) exhibit potential as vaccine candidates. However, the mechanisms that underlie the detrimental effects of alr knockouts on Brucella pathogenicity remain elusive. Here, initially, we conducted a bioinformatics analysis of Alr, which demonstrated a high degree of conservation of the protein within Brucella spp. Subsequent metabolomics studies unveiled alterations in amino acid pathways following deletion of the alr gene. Furthermore, alr deletion in Brucella suis S2 induced decreased resistance to stress, antibiotics, and other factors. Transmission electron microscopy of simulated macrophage intracellular infection revealed damage to the cell wall in the Δalr strain, whereas propidium iodide staining and alkaline phosphatase and lactate dehydrogenase assays demonstrated alterations in cell membrane permeability. Changes in cell wall properties were revealed by measurements of cell surface hydrophobicity and zeta potential. Finally, the diminished adhesion capacity of the Δalr strain was shown by immunofluorescence and bacterial enumeration assays. In summary, our findings indicate that the alr gene that regulates amino acid metabolism in Brucella influences the properties of the cell wall, which modulates bacterial adherence capability. This study is the first demonstration that Alr impacts virulence by modulating bacterial metabolism, thereby providing novel insights into the pathogenic mechanisms of Brucella spp. [ABSTRACT FROM AUTHOR]

Ezzat Nabil Abbas Ibrahim,1,* Ehab Tharwat,2,* Mahmoud Mohammed Ahmed Ali Khalil,1 Ahmed Rabie Mohammed,1 Mostafa F Mohammed,1 Ahmed Mohammed Madinah Alkady,1 Ezzeldin Ramadan Ezzeldin,2 Riad Elzaher Hassan Ahmed,2 Yasser H Al-Faky,3 Mohamed Hassanein,1 Ahmed N Elsayed,1 Mohammed Eid Abd El-Salam1,* 1Department of Ophthalmology, Faculty of Medicine, Al-Azhar University, Cairo, Egypt; 2Department of Ophthalmology, Faculty of Medicine, Al-Azhar University, Damietta, Egypt; 3Department of Ophthalmology, King Faisal Specialist Hospital and Research Center, Riyadh, Saudia Arabia*These authors contributed equally to this workCorrespondence: Ehab Tharwat, Department of Ophthalmology, Faculty of Medicine, Al-Azhar University, Alkawthar Building at Alshenawy St, from Algeish St, Mansoura, Dakahlia, Egypt, Tel +20 10 06952497, Fax +20 50 2507370, Email ehabtharwat71@domazhermedicine.edu.egPurpose: This study aims to assess the combination of anterior lamellar recession (ALR) with blepharoplasty, suprasternal fixation, and internal eyelash bulb extirpation of aberrant lashes posteriorly located in patients with any grade of upper eyelid trachomatous cicatricial entropion.Patients and Methods: We reviewed the clinical data of eighty-six patients (143 eyelids) including age, gender, systemic medical illnesses, and comprehensive ophthalmological assessment. Eyelid evaluation was recorded, including laterality, previous surgical technique used, possible trichiasis etiology, abnormality of the lid margin, tarsal plate consistency (shrinkage or loosening), skin fold overhanging, laxity of the pretarsal skin, margin reflex distance 1 (MRD1), lagophthalmos, and lid retraction. The success rate was assessed at 3, 6, 9, and 12 months postoperative.Results: The success rate was 97.2% in the third month, which decreased significantly to 92.3% in the 6th month and 90.2% in the 9th month (P = 0.01, and 0.001 respectively). In the 9th month, we had fourteen failed cases. All of them were submitted for a second intervention. Three underwent electrolysis, four cases underwent re-internal bulb extirpation, four cases underwent the same procedure, and three cases underwent epilation. The success of the failed cases after the second intervention was significantly associated with the type of reintervention (P = 0.03), in which all of them succeed except two cases that underwent epilation. Kaplan-Meier analysis showed that the mean recurrence time in our study was 6.8 months (95% CI = 5.8– 7.7 months).Conclusion: This study showed the combination of ALR with blepharoplasty, suprasternal fixation, and cauterization or internal bulb extirpation of posteriorly located lashes procedure resulted in a high success rate in patients with any form of UCE with no increase in incidence or degree of lagophthalmos associated with UCE.Keywords: anterior lamellar recession, ALR, blepharoplasty, cicatricial entropion, suprasternal fixation

Background: Acute T lymphoblastic leukemia (T‐ALL) occurs in 25% of adults diagnosed with Acute lymphocytic leukemia (ALL), and drug resistance is still a clinical obstacle. Augmenter of liver regeneration (ALR) is important to ALL drug resistance and is involved in the regulation of mitochondrial function; we speculated that the high expression of ALR in T‐ALL promotes drug resistance through the alteration of mitochondrial function and the inhibition of the mitochondrial apoptosis pathway. Method: We silenced and overexpressed ALR in the T‐ALL cell lines that were untreated or treated with dexamethasone (DXM) or methotrexate (MTX). Apoptosis, proliferation, reactive oxygen species and ATP productions, mitochondrial membrane potential, and mitochondrial respiratory chain complex expression in cells were examined. The data were collated to comprehensively evaluate the effects of ALR expression change on mitochondrial function and drug resistance in T‐ALL cells. Results: ALR knockdown led to the inhibition of proliferation, an increase in apoptosis, and the promotion of the cells' sensitivity to drugs. It also showed mitochondrial dysfunction. ALR knockdown actived the mitochondrial apoptosis pathway. The treatment of ALR knockdown T‐ALL cells with MTX or DXM further altered the mitochondrial function of T‐ALL cells and actived the mitochondrial apoptosis pathway. Overexpression of ALR promoted cell proliferation and drug resistance, reduced apoptosis, protected mitochondrial function, and inhibited the mitochondrial apoptosis pathway. Conclusion: T‐ALL resistance caused by ALR through the alteration of mitochondrial function is associated with the inhibition of the mitochondrial apoptosis pathway. [ABSTRACT FROM AUTHOR]

Brucella suis, the causative agent of brucellosis, poses a significant public health and animal husbandry threat. However, the role of the alanine racemase (alr) gene, which encodes alanine racemase in Brucella, remains unclear. Here, we analyzed an alr deletion mutant and a complemented strain of Brucella suis S2. The knockout strain displayed an unaltered, smooth phenotype in acriflavine agglutination tests but lacked the core polysaccharide portion of lipopolysaccharide (LPS). Genes involved in the LPS synthesis were significantly upregulated in the deletion mutant. The alr deletion strain exhibited reduced intracellular viability in the macrophages, increased macrophage-mediated killing, and upregulation of the apoptosis markers. Bcl2, an anti-apoptotic protein, was downregulated, while the pro-apoptotic proteins, Bax, Caspase-9, and Caspase-3, were upregulated in the macrophages infected with the deletion strain. The infected macrophages showed increased mitochondrial membrane permeability, Cytochrome C release, and reactive oxygen species, activating the mitochondrial apoptosis pathway. These findings revealed that alanine racemase was dispensable in B. suis S2 but influenced the strain's rough features and triggered the mitochondrial apoptosis pathway during macrophage invasion. The deletion of the alr gene reduced the intracellular survival and virulence. This study enhances our understanding of the molecular mechanism underlying Brucella's survival and virulence and, specifically, how alr gene affects host immune evasion by regulating bacterial LPS biosynthesis. [ABSTRACT FROM AUTHOR]

SPG15 is a hereditary spastic paraplegia subtype caused by mutations in Spastizin, a protein encoded by the ZFYVE26 gene. Spastizin is involved in autophagosome maturation and autophagic lysosome reformation and SPG15-related mutations lead to autophagic lysosome reformation defects with lysosome enlargement, free lysosome depletion and autophagosome accumulation. Symptomatic and rehabilitative treatments are the only therapy currently available for patients. Here, we targeted autophagy and lysosomes in SPG15 patient-derived cells by using a library of autophagy-modulating compounds. We identified a rose of compounds affecting intracellular calcium levels, the calcium-calpain pathway or lysosomal functions, which reduced autophagosome accumulation. The six most effective compounds were tested in vivo in a new SPG15 loss of function Drosophila model that mimicked the reported SPG15 phenotype, with autophagosome accumulation, enlarged lysosomes, reduced free lysosomes, autophagic lysosome reformation defects and locomotor deficit. These compounds, namely verapamil, Bay K8644, 2′,5′-dideoxyadenosine, trehalose, Small-Molecule Enhancer of Rapamycin 28 and trifluoperazine, improved lysosome biogenesis and function in vivo , demonstrating that lysosomes are a key pharmacological target to rescue SPG15 phenotype. Among the others, the Small-Molecule Enhancer of Rapamycin 28 was the most effective, rescuing both autophagic lysosome reformation defects and locomotor deficit, and could be considered as a potential therapeutic compound for this hereditary spastic paraplegia subtype. [ABSTRACT FROM AUTHOR]

Brucella, a zoonotic facultative intracellular pathogenic bacterium, poses a significant threat both to human health and to the development of the livestock industry. Alanine racemase (Alr), the enzyme responsible for alanine racemization, plays a pivotal role in regulating virulence in this bacterium. Moreover, Brucella mutants with alr gene deletions (Δalr) exhibit potential as vaccine candidates. However, the mechanisms that underlie the detrimental effects of alr knockouts on Brucella pathogenicity remain elusive. Here, initially, we conducted a bioinformatics analysis of Alr, which demonstrated a high degree of conservation of the protein within Brucella spp. Subsequent metabolomics studies unveiled alterations in amino acid pathways following deletion of the alr gene. Furthermore, alr deletion in Brucella suis S2 induced decreased resistance to stress, antibiotics, and other factors. Transmission electron microscopy of simulated macrophage intracellular infection revealed damage to the cell wall in the Δalr strain, whereas propidium iodide staining and alkaline phosphatase and lactate dehydrogenase assays demonstrated alterations in cell membrane permeability. Changes in cell wall properties were revealed by measurements of cell surface hydrophobicity and zeta potential. Finally, the diminished adhesion capacity of the Δalr strain was shown by immunofluorescence and bacterial enumeration assays. In summary, our findings indicate that the alr gene that regulates amino acid metabolism in Brucella influences the properties of the cell wall, which modulates bacterial adherence capability. This study is the first demonstration that Alr impacts virulence by modulating bacterial metabolism, thereby providing novel insights into the pathogenic mechanisms of Brucella spp.

• In this study, we continue our exploration of the alr gene, investigating its role in the interaction between Brucella and the host through murine and cellular experiments. • Our findings indicate that the extracellular substances produced by Δ alr effectively activate the TLR4-NF-κB-NLRP3 pathway in macrophages, inducing a more robust inflammatory response and subsequently reducing bacterial survival within cells. • Furthermore, Δ alr exhibits lower toxicity and enhanced immunogenicity in mice. • Importantly, we further elucidate the pivotal role played by alr in immune evasion during the interaction between Brucella and the host, providing insights for both complementing alr function and advancing Brucella vaccine development. Brucella is an intracellular parasitic bacterium lacking typical virulence factors, and its pathogenicity primarily relies on replication within host cells. In this study, we observed a significant increase in spleen weight in mice immunized with a Brucella strain deleted of the gene for alanine racemase (Alr), the enzyme responsible for alanine racemization (Δ alr). However, the bacterial load in the spleen markedly decreased in the mutant strain. Concurrently, the ratio of white pulp to red pulp in the spleen was increased, serum IgG levels were elevated, but no significant damage to other organs was observed. In addition, the inflammatory response was potentiated and the NF-κB-NLRP3 signaling pathway was activated in macrophages (RAW264.7 Cells and Bone Marrow-Derived Cells) infect ed with the Δ alr mutant. Further investigation revealed that the Δ alr mutant released substantial amounts of protein in a simulated intracellular environment which resulted in heightened inflammation and activation of the TLR4-NF-κB-NLRP3 pathway in macrophages. The consequent cytoplasmic exocytosis reduced intracellular Brucella survival. In summary, cytoplasmic exocytosis products resulting from infection with a Brucella strain deleted of the alr gene effectively activated the TLR4-NFκB-NLRP3 pathway, triggered a robust inflammatory response, and reduced bacterial survival within host cells. Moreover, the Δ alr strain exhibits lower toxicity and stronger immunogenicity in mice. [ABSTRACT FROM AUTHOR]

The immune system interacts with many nominal 'danger' signals, endogenous danger‐associated (DAMP), exogenous pathogen (PAMP) and allergen (AAMP)‐associated molecular patterns. The immune context under which these are received can promote or prevent immune activating or inflammatory mechanisms and may orchestrate diverse immune responses in allergy and cancer. Each can act either by favouring a respective pathology or by supporting the immune response to confer protective effects, depending on acuity or chronicity. In this Position Paper under the collective term danger signals or DAMPs, PAMPs and AAMPs, we consider their diverse roles in allergy and cancer and the connection between these in AllergoOncology. We focus on their interactions with different immune cells of the innate and adaptive immune system and how these promote immune responses with juxtaposing clinical outcomes in allergy and cancer. While danger signals present potential targets to overcome inflammatory responses in allergy, these may be reconsidered in relation to a history of allergy, chronic inflammation and autoimmunity linked to the risk of developing cancer, and with regard to clinical responses to anti‐cancer immune and targeted therapies. Cross‐disciplinary insights in AllergoOncology derived from dissecting clinical phenotypes of common danger signal pathways may improve allergy and cancer clinical outcomes. [ABSTRACT FROM AUTHOR]

Synergistic interactions between chemical inhibitors, whilst informative, can be difficult to interpret, as chemical inhibitors can often have multiple targets, many of which can be unknown. Here, using multiplexed transcriptional repression, we have validated that the simultaneous repression of glutamate racemase and alanine racemase has a synergistic interaction in Mycobacterium tuberculosis . This confirms prior observations from chemical interaction studies and highlights the potential of targeting multiple enzymes involved in mycobacterial cell wall synthesis.

Brucella suis, the causative agent of brucellosis, poses a significant public health and animal husbandry threat. However, the role of the alanine racemase (alr) gene, which encodes alanine racemase in Brucella, remains unclear. Here, we analyzed an alr deletion mutant and a complemented strain of Brucella suis S2. The knockout strain displayed an unaltered, smooth phenotype in acriflavine agglutination tests but lacked the core polysaccharide portion of lipopolysaccharide (LPS). Genes involved in the LPS synthesis were significantly upregulated in the deletion mutant. The alr deletion strain exhibited reduced intracellular viability in the macrophages, increased macrophage-mediated killing, and upregulation of the apoptosis markers. Bcl2, an anti-apoptotic protein, was downregulated, while the pro-apoptotic proteins, Bax, Caspase-9, and Caspase-3, were upregulated in the macrophages infected with the deletion strain. The infected macrophages showed increased mitochondrial membrane permeability, Cytochrome C release, and reactive oxygen species, activating the mitochondrial apoptosis pathway. These findings revealed that alanine racemase was dispensable in B. suis S2 but influenced the strain’s rough features and triggered the mitochondrial apoptosis pathway during macrophage invasion. The deletion of the alr gene reduced the intracellular survival and virulence. This study enhances our understanding of the molecular mechanism underlying Brucella’s survival and virulence and, specifically, how alr gene affects host immune evasion by regulating bacterial LPS biosynthesis.

The mitochondrial intermembrane space hosts a machinery for oxidative protein folding, the mitochondrial disulfide relay. This machinery imports a large number of soluble proteins into the compartment, where they are retained through oxidative folding. Additionally, the disulfide relay enhances the stability of many proteins by forming disulfide bonds. In this review, we describe the mitochondrial disulfide relay in human cells, its components, and their coordinated collaboration in mechanistic detail. We also discuss the human pathologies associated with defects in this machinery and its protein substrates, providing a comprehensive overview of its biological importance and implications for health., (© 2024 The Authors. FEBS Open Bio published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

To report a novel modified technique for severe recurrent cicatricial entropion correction based on anterior lamellar recession (ALR) and grafting. Six eyelids of five patients (9–48 years; three females) with severe cicatricial entropion (three upper and three lower eyelids) had surgical correction using ALR and labial mucosal grafting for spacing the ciliary margin away from the lid margin and reconstruction of the lid margin and posterior lamella. The modified technique included using mucous membrane as a single unit for anterior lamella, lid margin, and posterior lamella reconstruction employing a different suturing technique. The indications for surgery included Stevens–Johnson syndrome (3), chemical injury (2), and post-surgical scarring in congenital distichiasis with lymphedema (1). Entropion resolved in all patients with restoration of eyelid margin and reduction in ocular discomfort. Trichiasis was present in all six eyelids (100%) preoperatively and resolved completely in all but one case (83% success rate) with three residual focal trichiatic lashes in the temporal area, which was successfully managed with radiofrequency ablation. There were no recurrences of trichiatic or distichiatic lashes at a median follow-up duration of 16 months (range, 12–18 months). Successful resolution of the recurrent cicatricial entropion can be achieved with the use of labial mucosa as one solution for spacing the anterior lamella and reconstruction of the lid margin and posterior lamella with minimal recurrence rate. [ABSTRACT FROM AUTHOR]

In this study, l‐isoleucine production in Corynebacterium glutamicum WM001 was improved by deleting three genes in the genome, replacing the native promoter of ilvA in the genome, and overexpression of five genes in an alr‐based auxotrophic complementation expression system. The three genes deleted in the genome are alaT, brnQ, and alr. Deletion of alaT improved l‐isoleucine production by increasing the supply of pyruvate, whereas deletion of brnQ improved l‐isoleucine production by blocking the uptake of extracellular l‐isoleucine. Exchange of the native promoter of ilvA with promoter tac or tacM could contribute to l‐isoleucine production by increasing 2‐ketobutyric acid; tac is better than tacM for improving l‐isoleucine yield. Different combinations of the genes ilvBN, ppnK, lrp, and brnFE were overexpressed in an alr‐based auxotrophic complementation expression system to further improve l‐isoleucine production, and the best yield after 72‐H flask fermentation was obtained from the strain WM005/pYCW‐1‐ilvBN2‐ppnK1. Without addition of any antibiotics, WM005/pYCW‐1‐ilvBN2‐ppnK1 could produce 32.1 g/L l‐isoleucine after 72‐H fed‐batch fermentation, which is 34.3% increase compared with the original strain WM001. [ABSTRACT FROM AUTHOR]

Hereditary spastic paraplegia is a spastic gait disorder that arises from degeneration of corticospinal axons. The subtype SPG48 is associated with mutations in the zeta subunit of the adaptor protein complex five (AP5). AP5 function and the pathophysiology of SPG48 are only poorly understood. Here, we report an AP5 zeta knockout mouse, which shows an age-dependent degeneration of corticospinal axons. Our analysis of knockout fibroblasts supports a trafficking defect from late endosomes to the transGolgi network and reveals a structural defect of the Golgi. We further show that both autophagic flux and the recycling of lysosomes from autolysosomes were impaired in knockout cells. In vivo, we observe an increase of autophagosomes and autolysosomes and, at later stages, the accumulation of intracellular waste in neurons. Taken together, we propose that loss of AP5 function blocks autophagy and thus leads to the aberrant accumulation of autophagic cargo, which finally results in axon degeneration.

Abstract Background Childhood morbidities are a major cause of mortality of children in the developing countries particularly in Ethiopia. Regardless of the noticeable improvement in the reduction of under-five death in Ethiopia, childhood diarrhea and fever are still the leading cause of death. In Ethiopia, the burden of child mortality is alarming and calls for determined efforts in combating such health problems. Therefore, this study aimed to investigate the risk factors for childhood morbidity specifically for diarrhea and fever. Methods To gain insight into children’s health issues, the 2016 Ethiopian Demographic and Health Survey data were used. Among the marginal models, alternating logistic regression that is an extension of the generalized estimating equation model was used to investigate the risk factors of childhood morbidity explicitly for diarrhea and fever. Results The results show that the child’s sex, child’s age, anemia level, husband education level, mother’s work status, mother’s marital status, breastfeeding status and region are all chosen significant risk factors related with childhood diarrhea disease and fever disease. Conclusion The study indicated that male children, 0–11 months aged children, 12–23 months aged children, anemic children, husband with a lower education, mothers paid employment, non-breastfed children, regions of Amhara, Afar, Dire Dawa, Benishangul, Gambela, Oromia, SNNPR, Somali and Tigray were significantly associated with higher odds of morbidity in Ethiopia. Therefore, there is a need for children morbidity interventions intended to improve child health outcomes in the country.

Abstract Background Non-typhoidal Salmonella (NTS) infections caused primarily by S. Enteritidis and S. Typhimurium particularly in immunocompromised hosts have accounted for a large percentage of fatalities in developed nations. Antibiotics have revolutionized the cure of enteric infections but have also led to the rapid emergence of pathogen resistance. New powerful therapeutics involving metabolic enzymes are expected to be potential targets for combating microbial infections and ensuring effective health management. Therefore, the need for new antimicrobials to fight such health emergencies is paramount. Enteric bacteria successfully evade the gut and colonize their hosts through specialized virulence strategies. An important player, alanine racemase is a key enzyme facilitating bacterial survival. Results This study aims at understanding the contribution of alanine racemase genes alr, dadX and SEN3897 to Salmonella survival in vitro and in vivo. We have shown SEN3897 to function as a unique alanine racemase in S. Enteritidis which displayed essential alanine racemase activity. Interestingly, the sole presence of this gene in alr dadX double mutant showed a strict dependence on d-alanine supplementation both in vitro and in vivo. However, Alr complementation in d-alanine auxotrophic strain restored the alanine racemase deficiency. The Km and Vmax of SEN3897 was 89.15 ± 10.2 mM, 400 ± 25.6 µmol/(min mg) for l-alanine and 35 ± 6 mM, 132.5 ± 11.3 µmol/(min mg) for d-alanine, respectively. In vitro assays for invasion and survival as well as in vivo virulence assays involving SEN3897 mutant showed attenuated phenotypes. Further, this study also showed attenuation of d-alanine auxotrophic strain in vivo for the development of potential targets against Salmonella that can be investigated further. Conclusion This study identified a third alanine racemase gene unique in S. Enteritidis which had a potential effect on survival and pathogenesis in vitro and in vivo. Our results also confirmed that SEN3897 by itself wasn’t able to rescue d-alanine auxotrophy in S. Enteritidis which further contributed to its virulence properties.

Mia40/CHCHD4 and Erv1/ALR are essential for oxidative protein folding in the mitochondrial intermembrane space of yeast and mammals. In contrast, many protists, including important apicomplexan and kinetoplastid parasites, lack Mia40. Furthermore, the Erv homolog of the model parasite Leishmania tarentolae (LtErv) was shown to be incompatible with Saccharomyces cerevisiae Mia40 (ScMia40). Here we addressed structure-function relationships of ScErv1 and LtErv as well as their compatibility with the oxidative protein folding system in yeast using chimeric, truncated, and mutant Erv constructs. Chimeras between the N-terminal arm of ScErv1 and a variety of truncated LtErv constructs were able to rescue yeast cells that lack ScErv1. Yeast cells were also viable when only a single cysteine residue was replaced in LtErvC17S. Thus, the presence and position of the C-terminal arm and the kinetoplastida-specific second (KISS) domain of LtErv did not interfere with its functionality in the yeast system, whereas a relatively conserved cysteine residue before the flavodomain rendered LtErv incompatible with ScMia40. The question whether parasite Erv homologs might also exert the function of Mia40 was addressed in another set of complementation assays. However, neither the KISS domain nor other truncated or mutant LtErv constructs were able to rescue yeast cells that lack ScMia40. The general relevance of Erv and its candidate substrate small Tim1 was analyzed for the related parasite L. infantum. Repeated unsuccessful knockout attempts suggest that both genes are essential in this human pathogen and underline the potential of mitochondrial protein import pathways for future intervention strategies.

Background: Identifying objective changes following an auditory training program is central to the assessment of the program's efficacy. Purpose: This study aimed (1) to objectively determine the efficacy of a 12-week auditory processing training (APT) program in individuals with autism spectrumdisorder using auditory evoked potentials (AEPs) and (2) to identify the top central AEP predictors of the overall score on the Test of AuditoryProcessing Skills-3 (TAPS-3), the primary behavioral outcome measure of the APT program published in our earlier article. Research Design: A one-group pretraining, posttraining design was used. Study Sample: The sample included 15 children and young adults diagnosed with autism spectrum disorder. Participants underwent the APT program consisting of computerized dichotic training, one-on-one therapist-directed auditory training, and the use of remote microphone technology at home and in the classroom. Data Collection and Analysis: All participants underwent pre- and posttraining auditory brain stem responses (ABRs), complex auditory brain stem responses (cABRs), and auditory late responses (ALRs). Test results from ABRs and ALRs were grouped based on scores obtained in their dominant and nondominant ears. Paired t-tests were used to assess the efficacy of the training program, and least absolute shrinkage and selection operator regression was used to assess the relationship between ALRs and the TAPS-3 overall summed raw score reported in our earlier article. Results and Conclusions: When compared with pretraining results, posttraining results showed shorter ABR latencies and larger amplitudes. The cABRs showed decreased latencies of the frequency following waves, a reduction in pitch error, and enhancement of pitch strength and phase shift. ALR results indicated shorter latencies and larger amplitudes. Our earlier article showed that the TAPS-3 overall score was significantly higher after training. This study showed that the top three ALR predictors of TAPS-3 outcomes were P1 amplitude in the dominant ear, and N1 amplitude in the dominant and nondominant ears. [ABSTRACT FROM AUTHOR]

• Silencing TMPRSS11D reduced the immunosuppression effect of MDSCs on T cells. • TMPRSS11D induced endoplasmic reticulum stress in MDSCs. • TMPRSS11D induced endoplasmic reticulum stress in MDSCs through negative regulation of ALR. • TMPRSS11D-mediated ER stress promoted the immunosuppressive effect of MDSCs on T cells. • Silencing TMPRSS11D hindered tumor growth in tumor-bearing mice. Myeloid-derived suppressor cells (MDSCs) contribute to tumor immune evasion, and have been identified as immunosuppressive cells in cervical cancer. The effect of TMPRSS11D (transmembrane serine protease 11D) in some cancers has been reported, but its role in immune escape of cervical cancer is still unclear. This study aims to elucidate the regulatory mechanism of TMPRSS11D on the immunosuppressive function of MDSCs in cervical cancer. Our data showed that the proportion of polymorphonucleoid MDSCs (PMN-MDSCs), the contents of immunosuppressive factors (including INOS, IDO, and ARG-1) secreted by MDSCs, and TMPRSS11D mRNA level in peripheral blood mononuclear cells (PBMCs) of malignant cervical cancer patients was significantly higher than that of benign tumor patients. Next, CD3+ T cells from PBMCs of cervical cancer patients were stimulated with anti-CD3 and anti-CD28, and then co-cultured with PMN-MDSCs from the same donors at a ratio of 1:2 for 3 days. PMN-MDSCs from malignant tumors produced more ROS, while TMPRSS11D knockdown blocked ROS production. PMN-MDSCs inhibited T cell proliferation and IFN-γ production, while silencing TMPRSS11D in PMN-MDSCs hindered the immunosuppressive effect of PMN-MDSCs. Mechanistically, TMPRSS11D bound to ALR (Augmenter of liver regeneration) and negatively regulated ALR expression, inducing ER stress in PMN-MDSCs, thereby enhancing the immunosuppressive effect of PMN-MDSCs on T cells. Additionally, mouse xenograft tumor assay was conducted to assess the role of TMPRSS11D in tumor growth and MDSC accumulation in vivo. Silencing TMPRSS11D impeded the growth of cervical cancer xenografts and reduced the accumulation of MDSCs in tumor tissues. In conclusion, TMPRSS11D induced ER stress in MDSCs through negative regulation of ALR, thus enhancing the immunosuppressive effect of MDSCs on T cells, so as to promote the growth of cervical cancer tumors. [ABSTRACT FROM AUTHOR]

Hereditary spastic paraplegia is a spastic gait disorder that arises from degeneration of corticospinal axons. The subtype SPG48 is associated with mutations in the zeta subunit of the adaptor protein complex five (AP5). AP5 function and the pathophysiology of SPG48 are only poorly understood. Here, we report an AP5 zeta knockout mouse, which shows an age-dependent degeneration of corticospinal axons. Our analysis of knockout fibroblasts supports a trafficking defect from late endosomes to the trans Golgi network and reveals a structural defect of the Golgi. We further show that both autophagic flux and the recycling of lysosomes from autolysosomes were impaired in knockout cells. In vivo , we observe an increase of autophagosomes and autolysosomes and, at later stages, the accumulation of intracellular waste in neurons. Taken together, we propose that loss of AP5 function blocks autophagy and thus leads to the aberrant accumulation of autophagic cargo, which finally results in axon degeneration. • First in vivo disease model for AP5-deficency. • Redistribution of Golgi related proteins to lysosomes upon knockout of AP5 zeta. • Structural alterations of the trans Golgi network. • Block of autophagic flux and impaired recycling of lysosomes from autolyosomes under challenged conditions. [ABSTRACT FROM AUTHOR]

Graphical abstract With HFP-gas sparger in the downcomer zone of ALR liquid mixing is achieved with lower number of liquid circulations than in regular ALR. Highlights • Mixing in the airlift bioreactor with helical flow promoters was studied. • Air input through a helical flow promoter prolonged the circulation time. • Axial mixing was intensified in the airlift bioreactor with a helical flow promoter. • Mixing was more related to dispersion than to liquid circulation speed. Abstract Cultivation of aerobic microorganisms typically requires rapid, but gentle and simultaneously energy efficient mixing in bioreactors to prevent high gradients between local concentrations. The mixing efficiency in a pilot scale airlift bioreactor with helical flow promoters was investigated with varying air inputs. Helical flow promoters were helical tubes attached to riser or downcomer, or both and besides acting as flow promoter they were used as additional gas spargers. Mixing time, circulation time, dimensionless mixing time and Bodenstein number were used as assessment parameters to analyze the mixing efficiency of different airlift setups with helical flow promoters. Introducing air through the helical flow promoter in the downcomer zone prolonged the circulation time, but shortened the mixing time and decreased the Bodenstein number. Low Bodenstein number indicated an improved axial dispersion. Mixing was more related to dispersion in the airlift reactor with helical flow promoter in the downcomer zone whereas in a regular airlift reactor mixing was related to liquid circulation speed. [ABSTRACT FROM AUTHOR]

Inflammasomes are protein scaffolds required for the activation of caspase-1 and the subsequent release of interleukin (IL)-1β, IL-18, and danger signals, as well as the induction of pyroptotic cell death to restore homeostasis following infection and sterile tissue damage. However, excessive inflammasome activation also causes detrimental inflammatory disease. Therefore, extensive control mechanisms are necessary to prevent improper inflammasome responses and inflammatory disease. Inflammasomes are assembled by sequential nucleated polymerization of Pyrin domain (PYD) and caspase recruitment domain (CARD)-containing inflammasome components. Once polymerization is nucleated, this process proceeds in a self-perpetuating manner and represents a point of no return. Therefore, regulation of this key step is crucial for a controlled inflammasome response. Here, we provide an update on two single domain protein families containing either a PYD or a CARD, the PYD-only proteins (POPs) and CARD-only proteins (COPs), respectively. Their structure allows them to occupy and block access to key protein–protein interaction domains necessary for inflammasome assembly, thereby regulating the threshold of these nucleated polymerization events, and consequently, the inflammatory host response.

Retroviruses are major causes of disease in animals and human. Better understanding of the initial host immune response to these viruses could provide insight into how to limit infection. Mouse retroviruses that are endemic in their hosts provide an important genetic tool to dissect the different arms of the innate immune system that recognize retroviruses as foreign. Here, we review what is known about the major branches of the innate immune system that respond to mouse retrovirus infection, Toll-like receptors and nucleic acid sensors, and discuss the importance of these responses in activating adaptive immunity and controlling infection.

Text se zabývá otázkou, zda datum u závěti by mělo být jeho obligatorní, nebo fakultativní náležitostí. Tuto otázku ve vztahu k OZ 2012 zkoumá z pohledu historického, kdy srovnává starší platnou právní úpravu na našem území, zároveň však ale poukazuje na její inspirační zdroje, zejména právo německé, rakouské, polské a uherské. Rovněž se zabývá některými otázkami, jež plynou z toho, jak je požadavek na dataci závěti v právní právě kodifikován, zejména jako podobu může ona datace mít.

Liver is constantly exposed to pathogens, viruses, chemicals, and toxins, and several of them cause injury, leading to the loss of liver mass and sometimes resulting in cirrhosis and cancer. Under physiological conditions, liver can regenerate if the loss of cells is less than the proliferation of hepatocytes. If the loss is more than the proliferation, the radical treatment available is liver transplantation. Due to this reason, the search for an alternative therapeutic agent has been the focus of liver research. Liver regeneration is regulated by several growth factors; one of the key factors is augmenter of liver regeneration (ALR). Involvement of ALR has been reported in crucial processes such as oxidative phosphorylation, maintenance of mitochondria and mitochondrial biogenesis, and regulation of autophagy and cell proliferation. Augmenter of liver regeneration has been observed to be involved in liver regeneration by not only overcoming cell cycle inhibition but by maintaining the stem cell pool as well. These observations have created curiosity regarding the possible role of ALR in maintenance of liver health. Thus, this review brings a concise presentation of the work done in areas exploring the role of ALR in normal liver physiology and in liver health maintenance by fighting liver diseases, such as liver failure, non‐alcoholic fatty liver disease/non‐alcoholic steatohepatitis, viral infections, cirrhosis, and hepatocellular carcinoma. [ABSTRACT FROM AUTHOR]

ABSTRACT Cytosolic DNAs derived from retrotransposons serve as pathogen-associated molecular patterns for pattern recognition receptors (PRRs) that stimulate the induction of interferons (IFNs) and other cytokines, leading to autoimmune disease. Cyclic GMP-AMP synthase is one PRR that senses retrotransposon DNA, activating type I IFN responses through the stimulator of IFN genes (STING). Absent in melanoma 2 (AIM2)-like receptors (ALRs) have also been implicated in these pathways. Here we show that the mouse ALR IFI205 senses cytosolic retrotransposon DNA independently of cyclic GMP-AMP production. AIM2 antagonizes IFI205-mediated IFN induction activity by sequestering it from STING. We also found that the complement of genes located in the ALR locus in C57BL/6 and AIM2 knockout mice are different and unique, which has implications for interpretation of the sensing of pathogens in different mouse strains. Our data suggest that members of the ALR family are critical to the host IFN response to endogenous DNA. IMPORTANCE Autoimmune diseases like Aicardi-Goutières syndrome and lupus erythematosus arise when cells of the immune system become activated and attack host cells and tissues. We found that DNA generated by endogenous retroviruses and retroelements in inbred mice and mouse cells is recognized by several host proteins found in macrophages that are members of the ALR family and that these proteins both suppress and activate the pathways leading to the generation of cytokines and IFNs. We also show that there is great genetic diversity between different inbred mouse strains in the ALR genes, which might contribute to differential susceptibility to autoimmunity. Understanding how immune cells become activated is important to the control of disease.

Pathogenic mtDNA mutations associated with alterations of respiratory complex I, mitochondrial proliferation (oncocytic-like phenotype) and increase in antioxidant response were previously reported in type I endometrial carcinoma (EC). To evaluate whether in the presence of pathogenic mtDNA mutations other mitochondrial adaptive processes are triggered by cancer cells, the expression level of proteins involved in mitochondrial dynamics, mitophagy, proteolysis and apoptosis were evaluated in type I ECs harboring pathogenic mtDNA mutations and complex I deficiency. An increase in the fission protein Drp1, in the mitophagy protein BNIP3, in the mitochondrial protease CLPP, in the antioxidant and anti-apoptotic protein ALR and in Bcl-2 as well as a decrease in the fusion protein Mfn2 were found in cancer compared to matched non malignant tissue. Moreover, the level of these proteins was measured in type I EC, in hyperplastic (the premalignant form) and in non malignant tissues to verify whether the altered expression of these proteins is a common feature of endometrial cancer and of hyperplastic tissue. This analysis confirmed in type I EC samples, but not in hyperplasia, an alteration of the expression level of these proteins. These results suggest that in this cancer mitochondrial fission, antioxidant and anti-apoptotic response may be activated, as well as the discharge of damaged mitochondrial proteins as adaptation processes to mitochondrial dysfunction. [ABSTRACT FROM AUTHOR]

Clustered binary responses are often found in ecological studies. Data analysis may include modeling the marginal probability response. However, when the association is the main scientific focus, modeling the correlation structure between pairs of responses is the key part of the analysis. Second-order generalized estimating equations (GEE) are established in the literature. Some of them are more efficient in computational terms, especially facing large clusters. Alternating logistic regression (ALR) and orthogonalized residual (ORTH) GEE methods are presented and compared in this paper. Simulation results show a slightly superiority of ALR over ORTH. Marginal probabilities and odds ratios are also estimated and compared in a real ecological study involving a three-level hierarchical clustering. ALR and ORTH models are useful for modeling complex association structure with large cluster sizes. [ABSTRACT FROM AUTHOR]

Eukaryotic cells harbor membrane-enclosed compartments to spatially separate different biochemical processes. As a result, proteins that become synthesized in the cytosol but fulfill their function in another compartment require translocation machineries. In the intermembrane space (IMS) of mitochondria, the mitochondrial disulfide relay is responsible for the import of many soluble proteins in an oxidation-dependent manner. These IMS proteins carry out important tasks and therefore their import, folding and maintenance are crucial for the remainder of the cell. In this review, we first describe the machinery for oxidative protein folding in the IMS and then focus on recent developments, which especially concern the mammalian machinery, its substrates and its physiological role. [ABSTRACT FROM AUTHOR]

A novel pilot scale helix airlift reactor (helix-ALR) consisting of a regular draft tube airlift reactor (A d /A r = 1.17) and helical tubes attached to riser or downcomer, or both, was studied in different configurations with varying air inputs. The helical tubes acted as helical flow promoters (HFP) and additional gas spargers. The aim was to investigate the performance of these HFP-gas spargers and study their effect on oxygen transfer in ALR. HFP-gas spargers improved the volumetric oxygen transfer coefficients of the ALR radically even without additional energy, reaching up to 3 times higher k L a-values (over 0.1 l/s) when compared to regular ALR. Small bubbles from the HFP-gas spargers played an important role in the oxygen transfer, and downcomer helix had greater effect in general on k L a and hydrodynamics than riser helix. Empirical correlations regarding mass transfer and gas holdups were formed. When compared to stirred tank reactors (STR) the required specific power input of the helix-ALR for a level of k L a 0.1 l/s was around 0.34 kW/m 3 while for STR 2.0 kW/m 3 , suggesting that the helix-ALR is very energy efficient. HFP-gas spargers can significantly improve the performance of ALRs regarding oxygen transfer, lower their operating costs and improve the total energy economy. [ABSTRACT FROM AUTHOR]

The Brazilian deep space mission ASTER, as temporarily named, plans to send a small spacecraft to encounter and investigate the triple asteroid 2001-SN263. The launch is scheduled (initially) to occur in 2015, arriving in 2018. The main motivation of the mission is the development of technology and expertise to leverage the national space sector. Within the scientific goals, the investigation of the still unknown asteroid 2001-SN263. The main project guideline is to aggregate the widest possible Brazilian involvement in the platform, the development and operation of subsystems, integration, payload, as well as in the tracking, navigation, guidance and control of the probe. To meet this guideline, among others, the decision for the development of a laser altimeter in Brazil to fly in the mission was taken. This effort is currently coordinated by a group of researchers from the aerospace engineering personnel of UFABC. This article presents the preliminary development plan for the design of this instrument, which was called ALR (ASTER Laser Rangefinder).

KMT2D, which encodes a histone H3K4methyltransferase, has been implicated in human congenital heart disease in the context of Kabuki syndrome. However, its role in heart development is not understood. Here, we demonstrate a requirement for KMT2D in cardiac precursors and cardiomyocytes during cardiogenesis in mice. Gene expression analysis revealed downregulation of ion transport and cell cycle genes, leading to altered calcium handling and cell cycle defects.We further determined that myocardial Kmt2d deletion led to decreased H3K4me1 and H3K4me2 at enhancers and promoters. Finally, we identified KMT2D-bound regions in cardiomyocytes, of which a subset was associated with decreased gene expression and decreased H3K4me2 in mutant hearts. This subset included genes related to ion transport, hypoxia-reoxygenation and cell cycle regulation, suggesting that KMT2D is important for these processes. Our findings indicate that KMT2D is essential for regulating cardiac gene expression during heart development primarily via H3K4 di-methylation. [ABSTRACT FROM AUTHOR]

Cytosolic pattern recognition receptors (PRRs) sense a wide range of endogenous danger-associated molecular patterns (DAMPs) as well as exogenous pathogen-associated molecular patterns (PAMPs). In particular, Nod-like receptors (NLRs) containing a pyrin domain (PYD), called NLRPs, and AIM2-like receptors (ALRs) have been shown to play a critical role in host defense by facilitating clearance of pathogens and maintaining a healthy gut microflora. NLRPs and ALRs both encode a PYD, which is crucial for relaying signals that result in an efficient innate immune response through activation of several key innate immune signaling pathways. However, mutations in these PRRs have been linked to the development of auto-inflammatory and autoimmune diseases. In addition, they have been implicated in metabolic diseases. In this review, we summarize the function of PYD-containing NLRPs and ALRs and address their contribution to innate immunity, host defense and immune-linked diseases.

Among the scientific instruments that will fly on the upcoming mission ASTER, the first Brazilian deep space mission, the laser altimeter will play an important role in the surveys regarding shapes, topographies and masses of the asteroids of the triple system. The development of the instrument in partnership among universities (UNICAMP, UFABC) and companies of the Brazilian aerospace sector is in progress. This paper describes and presents the set of studies that were conducted aiming at the creation of software to embark on the control and signal processing unit of the instrument, with respect to the modeling and simulation of its operation and regarding the emission of laser pulses along with the detection and processing of return pulses. As the main result, a package of computer programs was created to simulate the operation of a pulsed laser altimeter with principle of operation based on the measurement of the pulse time of flight. The simulator software was named ALR_Sim. The results obtained with the ALR_Sim helped to better understand some key features and parameters of the operation of the future instrument, such as the sampling time of the detected signal and the definition of the type of detector suitable for the expected intensity of the return signal. The simulator software was successfully tested with respect to the most common expected situations. [ABSTRACT FROM AUTHOR]

Receptors of the innate immune system detect conserved determinants of microbial and viral origin. Activation of these receptors initiates signaling events that culminate in an effective immune response. Recently, the view that innate immune signaling events rely on and operate within a complex cellular infrastructure has become an important framework for understanding the regulation of innate immunity. Compartmentalization within this infrastructure provides the cell with the ability to assign spatial information to microbial detection and regulate immune responses. Several cell biological processes play a role in the regulation of innate signaling responses; at the same time, innate signaling can engage cellular processes as a form of defense or to promote immunological memory. In this review, we highlight these aspects of cell biology in pattern-recognition receptor signaling by focusing on signals that originate from the cell surface, from endosomal compartments, and from within the cytosol. [ABSTRACT FROM AUTHOR]

Mitochondria biogenesis is a fundamental process for the organization and normal function of all cells. Since the majority of mitochondrial proteins are synthesized in the cytosol, protein import is the major mechanism for mitochondria biogenesis. We describe the different pathways that ensure correct targeting and intra mitochondrial sorting of mitochondrial proteins. The import process of several proteins of the mitochondrial intermembrane space relies on the Mitochondrial Import and Assembly 40 and Essential for respiration and vegetative growth 1 (Erv1) proteins that together constitute the oxidative folding machinery of the mitochondrial intermembrane space. Recent work has implicated the FAD-oxidase protein Erv1 (ad its human homolog Augmenter of Liver Regeneration) as an anti-apoptotic factor in mammalian cells (including neuronal cells) that undergo Reactive Oxygen Species-triggered apoptosis. The different roles of this protein as a key factor in mitochondria biogenesis, iron-sulfur cluster biogenesis and in neuronal protection against apoptosis are discussed. [ABSTRACT FROM AUTHOR]

The clinical use of a bioartificial liver (BAL) device strongly depends on the development of human liver cell lines. The aim of this study was to establish and assess the potential use of the stable HepG2 cell line expressing human augmenter of liver regeneration (hALR). The cDNA encoding hALR protein was inserted into pcDNA3.1 to generate pcDNA3.1/hALR, following which pcDNA3.1/hALR was transfected to HepG2 to establish a cell line that stably expressed hALR (HepG2 hALR). A total of 800 million HepG2 hALR cells were loaded into laboratory-scale BAL bioreactors and cultured for 4 days, during which time the parameters of hepatocyte-specific function and general metabolism were determined. The cell line that stably expressed human ALR was successfully established. The expression of recombinant hALR was higher in the HepG2 hALR cell line than in the HepG2 cell line based on immunofluorescence and immunoblot assays. In samples removed from the BAL bioreactor on day 4, compared to HepG2 cells, HepG2 hALR cells produced significantly more alpha-fetoprotein (127.3 %; P < 0.05) and urea (128.8 %; P < 0.05) and eliminated more glucose (135.7 %; P < 0.05); the level of human albumin was also higher (117 %) in HepG2 hALR cells, but the difference was not significant ( P > 0.05). After 24 h of culture, the mean lidocaine removal rate was 65.1 and 57.3 % in culture supernatants of HepG2 hALR and HepG2 cell lines, respectively ( P < 0.01). Based on these results, we conclude that HepG2 hALR cells showed liver-specific functionality when cultured inside the bioreactor and would therefore be a potential cell source for BAL. [ABSTRACT FROM AUTHOR]