Polyglutamine disorders: Pathogenesis and potential drug interventions.

Polyglutamine/poly(Q) diseases are a group nine hereditary neurodegenerative disorders caused due to abnormally expanded stretches of CAG trinucleotide in functionally distinct genes. All human poly(Q) diseases are characterized by the formation of microscopically discernable poly(Q) positive aggregates, the inclusion bodies. These toxic inclusion bodies are responsible for the impairment of several cellular pathways such as autophagy, transcription, cell death, etc., that culminate in disease manifestation. Although, these diseases remain largely without treatment, extensive research has generated mounting evidences that various events of poly(Q) pathogenesis can be developed as potential drug targets. The present review article briefly discusses the key events of disease pathogenesis, model system-based investigations that support the development of effective therapeutic interventions against pathogenesis of human poly(Q) disorders, and a comprehensive list of pharmacological and bioactive compounds that have been experimentally shown to alleviate poly(Q)-mediated neurotoxicity. Interestingly, due to the common cause of pathogenesis, all poly(Q) diseases share etiology, thus, findings from one disease can be potentially extrapolated to other poly(Q) diseases as well.

Glutamine stimulates the S6K/4E-BP branch of insulin signalling pathway to mitigate human poly(Q) disorders in Drosophila disease models.

OBJECTIVE AND METHODS: Since, the S6K/4E-BP sub-pathway can be stimulated by various amino acids; we extended our investigation to examine if oral feeding of amino acids delivers rescue against human poly(Q) toxicity in Drosophila. We utilised Drosophila models of two different poly(Q) disorders to test our hypothesis. Glutamine was fed to the test flies orally mixed in the food. Control and treated flies were then tested for different parameters, such as formation of poly(Q) aggregates and neurodegeneration, to evaluate glutamine's proficiency in mitigating poly(Q) neurotoxicity. RESULTS: Our study, for the first time, reports that glutamine feeding stimulates the growth promoting S6K/4E-BP branch of insulin signalling pathway and restricts pathogenesis of poly(Q) disorders in Drosophila disease models. We noted that glutamine treatment restricts the formation of neurotoxic poly(Q) aggregates and minimises neuronal deaths. Further, glutamine treatment re-establishes the chromatin architecture by improving the histone acetylation which is otherwise compromised in poly(Q) expressing neuronal cells. DISCUSSION: Since, the insulin signalling pathway as well as mechanism of action of glutamine are fairly conserved between human and Drosophila, our finding strongly suggests that glutamine holds immense potential to be developed as an intervention therapy against the incurable human poly(Q) disorders.

Glipizide ameliorates human poly(Q) mediated neurotoxicity by upregulating insulin signalling in Drosophila disease models.

Increasing reports suggest insulin signalling pathway as a putative drug target against polyglutamine [poly(Q)] disorders, such as Huntington's disease (HD), Spinocerebellar ataxias (SCA) 1, 2, 3 etc. However, studies on drug-based stimulation of insulin signalling cascade to mitigate poly(Q) pathogenesis are lacking. In our study, we adopted an evidence-based approach to examine if some established insulin stimulating drug can be utilized to restrict poly(Q) aetiology in Drosophila disease models. For the first time, we report that glipizide, an FDA approved anti-diabetic drug upregulates insulin signalling in poly(Q) expressing tissues and restricts formation of inclusion bodies and neurodegeneration. Moreover, it reinstates the chromatin architecture by improving histone acetylation, which is otherwise abrogated due to poly(Q) toxicity. In view of the functional conservation of insulin signalling pathway in Drosophila and humans, our finding strongly suggests that glipizide can be repurposed as an effective treatment strategy against the neurodegenerative poly(Q) disorders. Also, with appropriate validation studies in mammalian disease models, glipizide could be subsequently considered for the clinical trials in human patients.

Differential Functional Responses of Neutrophil Subsets in Severe COVID-19 Patients.

Neutrophils play a significant role in determining disease severity following SARS-CoV-2 infection. Gene and protein expression defines several neutrophil clusters in COVID-19, including the emergence of low density neutrophils (LDN) that are associated with severe disease. The functional capabilities of these neutrophil clusters and correlation with gene and protein expression are unknown. To define host defense and immunosuppressive functions of normal density neutrophils (NDN) and LDN from COVID-19 patients, we recruited 64 patients with severe COVID-19 and 26 healthy donors (HD). Phagocytosis, respiratory burst activity, degranulation, neutrophil extracellular trap (NET) formation, and T-cell suppression in those neutrophil subsets were measured. NDN from severe/critical COVID-19 patients showed evidence of priming with enhanced phagocytosis, respiratory burst activity, and degranulation of secretory vesicles and gelatinase and specific granules, while NET formation was similar to HD NDN. COVID LDN response was impaired except for enhanced NET formation. A subset of COVID LDN with intermediate CD16 expression (CD16Int LDN) promoted T cell proliferation to a level similar to HD NDN, while COVID NDN and the CD16Hi LDN failed to stimulate T-cell activation. All 3 COVID-19 neutrophil populations suppressed stimulation of IFN-γ production, compared to HD NDN. We conclude that NDN and LDN from COVID-19 patients possess complementary functional capabilities that may act cooperatively to determine disease severity. We predict that global neutrophil responses that induce COVID-19 ARDS will vary depending on the proportion of neutrophil subsets.

Serum and Urine Interferon Gamma-Induced Protein 10 (IP-10) Levels in Lupus Nephritis.

Background: Lupus nephritis (LN) is a prevalent and severe complication of systemic lupus erythematosus (SLE). Non-invasive diagnostics are limited, and current therapies have inadequate response rates. Expression of the chemokine Interferon-γ-induced protein 10 (IP-10) is regulated by Interferon-γ signaling and NF-κB, and its molecular activity and enhanced urine concentrations are implicated in LN, but its utility as a diagnostic marker and association with demographic, clinical, or pathologic features is not defined. Methods: 38 LN patients and 11 patients with non-LN glomerular diseases (GD) with active disease were included. Eighteen of the LN patients had achieved remission at one follow-up during the study time. Serum and urine were obtained from these samples, and the IP-10 levels were measured. Results: Serum and urine IP-10 levels are significantly enhanced in LN patients with active disease as compared with normal individuals (serum average 179.7 pg/mL vs. 7.2 pg/mL, p < 0.0001; urine average 28.7 pg/mg vs. 1.6 pg/mg, p = 0.0019) and patients with other forms of glomerular disease (serum average 179.7 pg/mL vs. 84.9 pg/mL, p = 0.0176; urine average 28.7 pg/mg vs. 0.18 pg/mg, p = 0.0011). Urine IP-10 levels are significantly higher in patients with proliferative LN (PLN) than those with membranous LN (MLN) (average 32.8 pg/mg vs. 7.6 pg/mg, p = 0.0155). Urine IP-10 levels are also higher in MLN versus primary membranous nephropathy (MN) (average 7.6 pg/mg vs. 0.2 pg/mg, p = 0.0193). Importantly, serum IP-10 levels remain elevated during active LN and LN remission, but urine IP-10 levels are decreased from active LN to remission in 72% of our patients. Lastly, serum, but not urine IP-10 levels are significantly higher in African American than White American LN patients in active LN (average 227.8 pg/mL vs. 103.4 pg/mL, p = 0.0309) and during LN remission (average 254.6 pg/mL vs. 89.2 pg/mL, p = 0.0399). Conclusions: Our findings suggest that serum and urine IP-10 measurements provide promising tests for monitoring LN activity, differentiation between classifications of LN, and differentiation between LN and other forms of glomerular disease. We also conclude that further assessment of elevated IP-10 levels in the serum and urine of high-risk populations (i.e., African American) could be beneficial in determining why many of these patients have worse outcomes and are non-responsive to standard therapeutics.

The S6k/4E-BP mediated growth promoting sub-pathway of insulin signalling cascade is essential to restrict pathogenesis of poly(Q) disorders in Drosophila.

Human neurodegenerative polyglutamine [poly(Q)] disorders, such as Huntington's disease (HD) and spinocerebellar ataxias (SCA), are characterised by an abnormal expansion of CAG repeats in the affected gene. The mutated proteins misfold and aggregate to form inclusion bodies that sequester important factors involved in cellular transcription, growth, stress and autophagic response and other essential functions. The insulin signalling pathway has been demonstrated as a major modifier and a potential drug target to ameliorate the poly(Q) mediated neurotoxicity in various model systems. Insulin signalling cascade harbours several downstream sub-pathways, which are synergistically involved in discharging indispensable biological functions such as growth and proliferation, metabolism, autophagy, regulation of cell death pathways etc. Hence, it is difficult to conclude whether the mitigation of poly(Q) neurotoxicity is an accumulative outcome of the insulin cascade, or the result of a specific sub-pathway. For the first time, we report that the ligand binding domain of insulin receptor mediated downstream growth promoting sub-pathway plays the pivotal role in operating the rescue event. We show that the growth promoting activity of insulin cascade is essential to minimize the abundance of inclusion bodies, to restrict neurodegeneration, and to restore the cellular transcriptional balance. Subsequently, we noted the involvement of the mTOR/S6k/4E-BP candidates in mitigating poly(Q) mediated neurotoxicity. Due to the conserved cellular functioning of the insulin cascade across species, and availability of several growth promoting molecules, our results in Drosophila poly(Q) models indicate towards a possibility of designing novel therapeutic strategies to restrict the pathogenesis of devastating human poly(Q) disorders.

Neutrophil exocytosis induces podocyte cytoskeletal reorganization and proteinuria in experimental glomerulonephritis.

Acute glomerulonephritis is characterized by rapid glomerular neutrophil recruitment, proteinuria, and glomerular hypercellularity. The current study tested the hypothesis that the release of neutrophil granule contents plays a role in both the loss of filtration barrier leading to proteinuria and the increase in glomerular cells. Inhibition of neutrophil exocytosis with a peptide inhibitor prevented proteinuria and attenuated podocyte and endothelial cell injury but had no effect on glomerular hypercellularity in an experimental acute glomerulonephritis model in mice. Cultivation of podocytes with neutrophil granule contents disrupted cytoskeletal organization, an in vitro model for podocyte effacement and loss of filtration barrier. Activated, cultured podocytes released cytokines that stimulated neutrophil chemotaxis, primed respiratory burst activity, and stimulated neutrophil exocytosis. We conclude that crosstalk between podocytes and neutrophils contributes to disruption of the glomerular filtration barrier in acute glomerulonephritis. Neutrophil granule products induce podocyte injury but do not participate in the proliferative response of intrinsic glomerular cells.

Endocytosis is required for exocytosis and priming of respiratory burst activity in human neutrophils.

OBJECTIVE AND DESIGN: Neutrophil generation of reactive oxygen species (ROS) is enhanced by exposure to pro-inflammatory agents in a process termed priming. Priming is depending on exocytosis of neutrophil granules and p47phox phosphorylation-dependent translocation of cytosolic NADPH oxidase components. Clathrin-mediated endocytosis was recently reported to be necessary for priming, but the mechanism linking endocytosis to priming was not identified. The present study examined the hypothesis that endocytosis regulates neutrophil priming by controlling granule exocytosis. MATERIALS AND METHODS: Clathrin-mediated endocytosis by isolated human neutrophils was inhibited by chlorpromazine, monodansylcadaverine, and sucrose. Exocytosis of granule subsets was measured as release of granule components by ELISA or chemiluminescence. ROS generation was measured as extracellular release of superoxide as reduction of ferrocytochrome c. p38 MAPK activation and p47phox phosphorylation were measured by immunoblot analysis. Statistical analysis was performed using a one-way ANOVA with the Tukey-Kramer multiple-comparison test. RESULTS: Inhibition of endocytosis prevented priming of superoxide release by TNFα and inhibited TNFα stimulation and priming of exocytosis of all four granule subsets. Inhibition of endocytosis did not reduce TNFα-stimulated p38 MAPK activation or p47phox phosphorylation. Inhibition of NADPH oxidase activity blocked TNFα stimulation of secretory vesicle and gelatinase granule exocytosis. CONCLUSIONS: Endocytosis is linked to priming of respiratory burst activity through ROS-mediated control of granule exocytosis.

Frontline Science: Tumor necrosis factor-α stimulation and priming of human neutrophil granule exocytosis.

Neutrophil granule exocytosis plays an important role in innate and adaptive immune responses. The present study examined TNF-α stimulation or priming of exocytosis of the 4 neutrophil granule subsets. TNF-α stimulated exocytosis of secretory vesicles and gelatinase granules and primed specific and azurophilic granule exocytosis to fMLF stimulation. Both stimulation and priming of exocytosis by TNF-α were dependent on p38 MAPK activity. Bioinformatic analysis of 1115 neutrophil proteins identified by mass spectrometry as being phosphorylated by TNF-α exposure found that actin cytoskeleton regulation was a major biologic function. A role for p38 MAPK regulation of the actin cytoskeleton was confirmed experimentally. Thirteen phosphoproteins regulated secretory vesicle quantity, formation, or release, 4 of which-Raf1, myristoylated alanine-rich protein kinase C (PKC) substrate (MARCKS), Abelson murine leukemia interactor 1 (ABI1), and myosin VI-were targets of the p38 MAPK pathway. Pharmacologic inhibition of Raf1 reduced stimulated exocytosis of gelatinase granules and priming of specific granule exocytosis. We conclude that differential regulation of exocytosis by TNF-α involves the actin cytoskeleton and is a necessary component for priming of the 2 major neutrophil antimicrobial defense mechanisms: oxygen radical generation and release of toxic granule contents.

Exocytosis of neutrophil granule subsets and activation of prolyl isomerase 1 are required for respiratory burst priming.

This study tested the hypothesis that priming the neutrophil respiratory burst requires both granule exocytosis and activation of the prolyl isomerase Pin1. Fusion proteins containing the TAT cell permeability sequence and either the SNARE domain of syntaxin-4 or the N-terminal SNARE domain of SNAP-23 were used to examine the role of granule subsets in TNF-mediated respiratory burst priming using human neutrophils. Concentration-inhibition curves for exocytosis of individual granule subsets and for priming of fMLF-stimulated superoxide release and phagocytosis-stimulated H2O2 production were generated. Maximal inhibition of priming ranged from 72 to 88%. Linear regression lines for inhibition of priming versus inhibition of exocytosis did not differ from the line of identity for secretory vesicles and gelatinase granules, while the slopes or the y-intercepts were different from the line of identity for specific and azurophilic granules. Inhibition of Pin1 reduced priming by 56%, while exocytosis of secretory vesicles and specific granules was not affected. These findings indicate that exocytosis of secretory vesicles and gelatinase granules and activation of Pin1 are independent events required for TNF-mediated priming of neutrophil respiratory burst.

Comparative study of psychiatric morbidity among the displaced and non-displaced populations in the Andaman and Nicobar Islands following the tsunami.

OBJECTIVE: The objective of this study was to compare the psychiatric morbidity between the displaced and non-displaced populations of the Andaman and Nicobar Islands during the first three months following the 2004 earthquake and tsunami. METHODS: The study was conducted at the 74 relief camps in the Andaman and Nicobar Islands. Port Blair had 12 camps, which provided shelter to 4,684 displaced survivors. There were 62 camps on Car-Nicobar Island, which provided shelter to approximately 8,100 survivors who continued to stay in their habitat (non-displaced population). The study sample included all of the survivors who sought mental health assistance inside the camp. A psychiatrist diagnosed the patients using the ICD-10 criteria. RESULTS: Psychiatric morbidity was 5.2% in the displaced population and 2.8% in the non-displaced population. The overall psychiatric morbidity was 3.7%. The displaced survivors had significantly higher psychiatric morbidity than did the non-displaced population. The disorders included panic disorder, anxiety disorders not otherwise specified, and somatic complaints. The existence of an adjustment disorder was significantly higher in the non-displaced survivors. Depression and post-traumatic stress disorder (PTSD) were distributed equally in both groups. CONCLUSIONS: Psychiatric morbidity was found to be highest in the displaced population. However, the incidence of depression and PTSD were distributed equally in both groups. Involvement of community leaders and survivors in shared decision-making processes and culturally acceptable interventions improved the community participation. Cohesive community, family systems, social support, altruistic behavior of the community leaders, and religious faith and spirituality were factors that helped survivors cope during the early phase of the disaster.

Psychological impact of the tsunami on children and adolescents from the andaman and nicobar islands.

OBJECTIVE: The aim of this article is to present the assessment of the presentation of symptoms and psychiatric morbidity of children and adolescents from the Andaman and Nicobar islands during the first 3 months following the December 2004 earthquake and tsunami. METHOD: According to predefined criteria, a primary survivor is one who was exposed directly to the earthquake and tsunami, a secondary survivor is one with close family and personal ties to primary survivors, and tertiary survivors are individuals from the communities beyond the impact area, a majority of which were exposed to the earthquake only. This study included 37 primary and secondary survivors (aged