Biology subjects

Saleem, A.

Publications and source records attributed to Saleem, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Plasma-derived extracellular vesicles as potential biomarkers and mediators of functional alterations in MELAS

Mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS) syndrome is a genetic disorder characterized by progressive neuromuscular and multisystem symptoms. MELAS typically manifests during childhood, can be difficult to diagnose, and has no cure. Extracellular vesicles (EVs) are lipid-enclosed nanoparticles secreted from cells that contain biological cargo and have demonstrated potential as biomarkers. We investigated the potential of plasma-derived EVs as diagnostic biomarkers of MELAS and examined their functional effects on mitochondrial respiration in treated skeletal muscle myotubes. Plasma-derived EVs were isolated from MELAS patients and age- and sex-matched control individuals, and biophysical characteristics and cargo of EVs analyzed. A Mito Stress Test was performed to assess oxygen consumption rate (OCR) in healthy myotubes treated with Control- or MELAS-EVs to determine the functional effects of circulatory EVs. Nine MELAS patients from two families were studied, and the results were categorized by age, sex and mtDNA heteroplasmy level. EV size and zeta potential remained unchanged. However, total EV concentration was higher in MELAS patients, particularly for small-EVs (<200 nm) and in younger patients (<25 years old). Relative protein yield per EV was lower in the MELAS group, especially among female and younger individuals. EV double-stranded DNA (dsDNA) concentration did not differ between MELAS- and Control-EVs overall, but was higher in male MELAS patients. Protein markers typically enriched in small-EVs showed altered expression in MELAS EVs: TSG101 and CD63 were lower, while flotillin-1 was higher compared to Control-EVs. A decrease in basal OCR was shown in cells treated with MELAS-EVs, with a similar response noted in the group treated with EVs from female MELAS patients. Post-treatment analysis showed no differences in oxidative phosphorylation (OXPHOS) subunit levels between cells treated with MELAS- and Control-EVs. In conclusion, plasma-derived EVs show promise as potential biomarkers for MELAS, and circulating EVs in this patient population may contribute to systemic metabolic dysfunction.

Cell Biology↗

bioRxiv2026

Local mechanical gradients underlie coordinated cascades of epithelial cell-cycle advance

Cell division mechanically perturbs the local environment in epithelial tissues, yet whether and how these perturbations propagate to coordinate cell-cycle progression across neighboring cells remains unclear. Here, we combine live cell-cycle tracking with mechanical analysis to examine how division events organize in space and time. We find that coordinated changes and spatial gradients in forces, morphology, and dynamics precede synchronized G1[->]S transitions in nearby cells. These transitions emerge within localized high-tension zones and give rise to spatiotemporal clusters of cell divisions, indicating that division events are mechanically coupled and propagate across neighboring cells. Supporting this biophysical picture, similar mechanical patterns arising from cell extrusion are sufficient to induce cell-cycle re-entry in neighboring cells. Together, these findings suggest a mechanically mediated framework for coordinated proliferation, possibly driven by positive mechanical feedback.

biophysics↗

bioRxiv2026

Human breast milk extracellular vesicles from mothers with asthma differentially modulate the release of inflammatory cytokines by primary human airway smooth muscle cells in a recipient-cell specific manner

Breastfeeding provides health benefits in childhood, reducing the frequency of gastrointestinal and respiratory infections. Breastmilk (BM) is a rich source of bioactive molecules including extracellular vesicles (EVs), which exert immunomodulatory signalling in recipient cells, with cargo that is affected by maternal characteristics. Here we investigated the biophysical characteristics of BM-EVs from mothers with (asthmatic BM-EVs) or without asthma (control BM-EVs) and their effect on the release of cytokines from primary human hTERT-immortalized airway smooth muscle cells (hASMs) from asthmatic or non-asthmatic (control) donors. BM-EVs were isolated using size exclusion chromatography (N=5/group), characterized biophysically and by EV-specific protein markers. In addition, BM-EV were co-cultured (48h) with primary hASM cells from both non-asthmatic (control) and asthmatic donors to determine the effect on cytokine release. All participants were Caucasian and the BM was collected 12-15 weeks postpartum. BM-EVs showed the presence of intact and small-EVs ([~]100 nm). Asthmatic BM-EVs appeared to have a smaller average EV size (135.6 nm) vs. controls (148.3 nm, p=0.0613), but [~]5-fold higher concentration of both total (p=0.0014) and small EVs (p=0.0016). The expression of EV subtype protein expression was reduced in asthmatic BM-EVs vs. control BM-EVs: CD63 by 86% (p=0.0224), flotillin-1 by 40% (p=0.0196), CD9 by 24% (p=0.0646) and HSP70 by 69% (p=0.0873). Asthmatic BM-EVs co-cultured with hASMs from control donors decreased pro-inflammatory cytokine release: MCP-1 by 55% (p=0.0286), IL-6 by 45% (p=0.0801) and IL-2 by 32% (p=0.0970) vs. control-BM-EVs. Conversely, asthmatic BM-EVs co-cultured with hASMs from asthmatic donors increased secretion of anti-inflammatory cytokine IL-10 by 32% (p=0.0660), and IL-1Ra by 75% (p=0.0875), and pro-inflammatory IL-2 by 57% (p=0.0688) vs. control-BM-EVs. Internalization of control and asthmatic BM-EVs was confirmed by labelled EV uptake experiments. No detrimental effects on cell viability with BM-EV treatment were observed. In summary, asthmatic BM-EVs are smaller and enriched in BM, and exert differential effects on cytokine release in a BM-donor and recipient-cell specific manner. Given that BM can enter infant airways, the immunomodulatory effects of BM-EVs on hASMs warrants further investigation to delineate the under underlying mechanisms.

cell biology↗

bioRxiv2025

3D-printed toolkit for regulating epithelial unjamming

During wound healing and cancer invasion, epithelial monolayers transition from a solid-like, jammed state to a fluid-like unjammed state. However, affordable and accessible tools for modeling and precise tuning of this response remain limited. Here, we present a simple, lithography-free 3D-printed toolkit of customizable inserts and scratchers for fabricating micro-gaps with defined geometries and widths down to [~]50 {micro}m, without requiring cleanroom facilities or specialized expertise. Using epithelial monolayers, we demonstrate that gap width fundamentally regulates unjamming dynamics. Narrow gaps induced rapid, coordinated migration with high initial velocities, whereas wide gaps suppressed early unjamming and produced slower, spatially heterogeneous closure. Cell shape dynamics confirmed conserved elongation, cell rounding, and closure behaviors across conditions. The presented toolkit provides new opportunities for investigating the mechanical and molecular mechanisms that regulate epithelial plasticity, enabling fine control of unjamming transitions and allowing the modeling of both healthy and pathological wound-response regimes.

biophysics↗

bioRxiv2025

Competence of DNA Tetrahedron and Exosomes as Nanocarriers for Epirubicin drug delivery in Breast Cancer Cells

Epirubicin, a chemotherapeutic agent, is used in treatment of metastatic breast cancer treatment. However, poor intracellular delivery and systemic toxicity are its major limitations. Our study evaluates the use of DNA tetrahedron (TD) structures and exosomes for targeted delivery of epirubicin to enhance its therapeutic efficacy. Increased intracellular concentration of epirubicin was observed in Flow cytometer when delivered via the TD: Epi system compared to free epirubicin. Cell viability assays using Annexin V-FITC and propidium iodide staining revealed higher apoptosis rates in the TD: Epi system. Exosomes, primarily 100 {+/-} 20 nm in size (99% of particles), exhibit a - 28.5 mV zeta potential, ensuring colloidal stability. SEM and TEM confirm their spherical shape, uniform size, and double membrane structure. The TD: Epi system showed the highest efficacy in RT-qPCR analysis, followed by Epirubicin + Exosomes systems and epirubicin alone, further corroborated with western blot results. While exosomes also provided improved delivery and therapeutic outcomes compared to free epirubicin, they were less effective than the TD: Epi system. These results suggest that the TD carrier significantly enhances the delivery and apoptotic efficacy of epirubicin when compared to exosome-based delivery or epirubicin alone offering a promising strategy for improving treatment outcomes. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=178 HEIGHT=200 SRC="FIGDIR/small/637654v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@10e0393org.highwire.dtl.DTLVardef@10ce3eeorg.highwire.dtl.DTLVardef@148552aorg.highwire.dtl.DTLVardef@1cf4664_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2024

Characteristics of early career health researchers and experiences of burnout during the COVID-19 pandemic in Canada

IntroductionThe COVID-19 pandemic disrupted research globally. How it impacted Canadian early-career health researchers (ECHRs) remains unclear. We administered a survey to understand the composition of ECHRs in Canada, their job experiences, and experiences of burnout during the COVID-19 pandemic. MethodsA cross-sectional survey was conducted in May 2023 of Canadian ECHRs defined as within 7 years of their first independent research position. Quantitative analyses included a description of respondents by research pillar, socio-demographic and workplace characteristics, and the prevalence of burnout, disengagement or exhaustion. Sample characteristics were compared to national data on ECHRs from a Canadian funding agency. Thematic analysis of free-text responses was also conducted. ResultsA total of 225 respondents met the eligibility criteria. Most respondents were assistant professors and characteristics of our sample were like the national data. The COVID-19 pandemic posed many challenges to student recruitment, and emotional support of students, with over half of the respondents reporting a moderate to significant decline in mental health compared to pre-pandemic. A significant proportion of respondents were experiencing high burnout (62%, 95%CI:56-67%), exhaustion (64%, 95%CI: 57-70%) or disengagement (91%, 95%CI: 87-95%). Thematic analysis identified three themes: ongoing benefits/problems preceding the pandemic, unintended outcomes of strategies to manage/prevent/contain COVID-19, and reasons to stay in their current position. ConclusionsOur survey revealed that Canadian ECHRs reported many diverse challenges during the COVID-19 pandemic and high burnout, putting the sustainability of this workforce at risk. Improved systems are needed to understand the long-term impacts and support the future of the Canadian health research ecosystem.

scientific communication and education↗

bioRxiv2024

Exercise-induced extracellular vesicles mediate apoptosis in human colon cancer cells in an exercise intensity-dependent manner

Regular exercise is known to reduce incidence rates and improve the prognosis of all cancers, but the underlying mechanisms remain elusive. Ample evidence suggests that exercise exerts therapeutic effects through extracellular vesicles (EVs), essential for cellular communication. Here, we hypothesized that exercise-induced EVs from serum of healthy young male participants will exert anti-tumorigenic effects on human colon cancer HT-29 cells, in an exercise intensity-dependent manner. 10 healthy young active males (25.4{+/-}6.2yrs, with maximal oxygen consumption (VO2max) = 45{+/-}3.7 ml.min-1.kg-1 participated in a randomized crossover trial. Participants underwent two different workload-matched, acute bouts of exercise: (1) moderate-intensity continuous exercise (MICE) at 50-55% V02max, and (2) high-intensity interval exercise (HIIE) at 90% V02max on a cycle ergometer. A control session of rest (PRE) was included. EVs were isolated from serum samples collected during PRE and immediately after each exercise session. EVs were co-incubated with colon cancer HT-29 cells (100 {micro}g EVs/ml, 48-72h), and effect on cell viability, migration, and apoptosis measured. EV treatment reduced cell viability in all groups (PRE, MICE, HIIE) by 35%, 43% and 47% respectively, vs. PBS. EVs from HIIE group showed a significantly greater reduction in cell viability vs. PRE, therefore further analysis used these groups only. PRE-EVs reduced migration by 27%, and HIIE-EVs by 39%. EV from HIIE group increased expression of pro-apoptotic markers: Bax/Bcl-2 ratio by 56% and Caspase-3 by 30% vs. PBS, with no change observed in the PRE group. Further, 16% of cells in PRE, and 28% of cells in HIIE were TUNEL-positive, indicating DNA fragmentation, a hallmark feature of apoptosis. Our data show that exercise-induced EVs reduced cell viability, in an exercise intensity-dependent manner. HIIE-derived EVs exerted the most anti-tumorigenic effects: decreased cell viability, reduced cell migration, increase in pro-apoptotic protein expression, and elevated DNA fragmentation. It is likely these changes were mediated by altered EV Cargo induced by exercise, as the amount of EVs was the same in each treatment group. To our knowledge, this is the first human study that illustrates the therapeutic potential of exercise-induced EVs in cancer treatment.

cancer biology↗

bioRxiv2024

Identification of novel myokines and putative protein targets that mediate functional adaptations in response to chronic contractile activity induced skeletal muscle-extracellular vesicle treatment

We have previously shown that skeletal muscle-derived extracellular vesicles (EVs) released post-chronic contractile activity (CCA) increased mitochondrial biogenesis in murine myoblasts, and decreased cell viability and induced apoptosis and senescence in non-small cell lung cancer cells. While the underlying mechanisms are unknown, the effects perpetuated were dependent on membrane-bound proteins. Here, we performed an extensive LC-MS/MS proteomic analysis on EVs from control and CCA myotubes. A total of 2900 proteins were identified in CON-EVs and CCA-EVs, including EV-associated proteins such as TSG101, tetraspanins (CD9, CD81, and CD63), flotillin-1, and annexins. Of these, 856 proteins are novel and not listed in EV databases (ExoCarta and Vesiclepedia), indicating that myotube-EVs harbor proteins not yet identified in EVs of different origin. Additionally, we identified 2062 unique proteins that have not yet been previously reported in myotube-EVs to date. Remarkably, of the 2900 total proteins identified, we observed 46 upregulated, and 25 downregulated differentially expressed proteins (DEPs) in CCA-EVs vs. control-EVs. Most of upregulated DEPs include EV-associated proteins. Comparing the 71 DEPs with proteins expressed in skeletal muscle indicated 61 of these as potential myokines. We identified actin cytoskeleton signaling, integrin signaling and muscle contraction as the most enriched pathways among the DEPs using different databases/software including FunRich, KEGG, STRING and Ingenuity Pathway Analysis. Using a relevance score that prioritized membrane-bound proteins with known function in mitochondrial biogenesis and inhibition of cancer growth, we identified top-scoring highly enriched DEPs of interest: IGF1R, ATP7A, PFN1, GJA1, PRKCA and ITGA6. We confirmed upregulation of these targets in EVs using immunoblotting. Among these top-scoring DEPs, PFN1, and ITGA6 are associated with EVs, with expression upregulated following acute exercise. In summary, we report the first comprehensive analysis of skeletal muscle-EV proteome following CCA, with identification of putative protein targets and signaling pathways that may execute the pro-metabolic and anti-tumorigenic effects of CCA-EVs.

cell biology↗

bioRxiv2024

The pro-apoptotic effect of chronic contractile activity-induced extracellular vesicles on Lewis Lung Carcinoma cells

Regular exercise reduces tumor growth in vivo and in vitro, but the exact mechanisms have yet to be fully elucidated. We have previously shown that chronic contractile activity (CCA) increases the concentration of skeletal muscle-derived EVs, and these in turn increased mitochondrial biogenesis in myoblasts. Here, we hypothesized that skeletal muscle-EVs derived post-CCA will mediate the anti-tumorigenic effects associated with chronic exercise. C2C12 myoblasts were differentiated into myotubes, electrically paced, and EVs isolated from conditioned media from control and CCA myotubes using differential ultracentrifugation. Lewis lung carcinoma (LLC) cells were treated with the total number of control-EVs or CCA-EVs isolated after each day of contractile activity for 4 days. Permeabilized CCA-EVs with or without proteinase K before co-culture with LLC cells were used as controls. Effect of EV treatment on cell count, viability, apoptosis, senescence, migration, and mitochondrial content was measured. CCA-EV treatment reduced cell count by 18% and cell viability by 6% vs. control-EVs. CCA-EVs increased the incidence of apoptotic hallmarks: DNA fragmentation by 13%, Annexin V+/PI+ cells by 21%, and the expression of pro-apoptotic Bax (by 25%) and Bax/Bcl-2 ratio (by 60%) vs. control-EVs. CCA-EVs increased number of senescent cells by 29%, and senescence markers, HMGB1 (by 49%) and p16 (by 92%) vs. control-EVs. When CCA-EVs were pretreated with Triton X-100 with or without proteinase-K, the increase in apoptosis and senescence was abrogated, confirming the effect is due to intact EVs and likely through EV membrane-proteins. CCA-EVs did not have any effect on cell migration and mitochondrial content vs. control-EVs. This study illustrates for the first time the potential of CCA-induced skeletal muscle-EVs in mediating anti-tumorigenic effects traditionally linked with chronic exercise.

cancer biology↗

bioRxiv2024

Chronic contractile activity induced skeletal muscle-derived extracellular vesicles increase mitochondrial biogenesis in recipient myocytes via transmembrane or peripheral membrane proteins

The effect of chronic contractile activity (CCA) on the biophysical properties and functional activity of skeletal muscle extracellular vesicles (Skm-EVs) is poorly understood due to challenges in distinguishing Skm-EVs originating from exercising muscle in vivo. To address this, myoblasts were differentiated into myotubes, and electrically paced (3 h/day, 4 days @ 14 V). CCA evoked an increase in mitochondrial biogenesis in stimulated vs. non-stimulated (CON) myotubes as expected. EVs were isolated from conditioned media from control and stimulated myotubes using differential ultracentrifugation and characterized biophysically using tunable resistive pulse sensing (TRPS, Exoid), TEM and western blotting. TEM images confirmed isolated round-shaped vesicles of about 30 - 150 nm with an intact lipid bilayer. The mean size of EVs ranged from 98 -138 nm, and was not altered by CCA. Zeta potential and total EV protein yield remained unchanged between groups, and total EV secretion increased after 4 days of CCA. Concomitant analysis of EVs after each day of CCA also demonstrated a progressive increase in CCA-EV concentration, while size and zeta potential remained unaltered, and EV protein yield increased in both CON-EVs and CCA groups. CCA-EVs were enriched with small-EVs vs. CON-EVs, concomitant with higher expression of small-EV markers CD81, Tsg101 and HSP70. In whole cell lysates, CD63 and ApoA1 were reduced with CCA in myotubes, whereas CD81, Tsg101, Flotillin-1 and HSP70 levels remained unchanged. To evaluate the functional effect of EVs secreted post-CCA, we treated C2C12 myoblasts with all EVs isolated from CON or CCA myotubes after each day of stimulation, and measured cell count, cell viability, protein yield and mitochondrial biogenesis in recipient cells. There was no effect on cell count, viability and protein yield. Myoblasts treated with CCA-EVs exhibited increased mitochondrial biogenesis as indicated by enhanced MitoTracker Red staining, cytochrome c oxidase activity, and protein expression of electron transport chain subunit, CIV-MTCO1. Further, CCA-EV treatment enhanced maximal oxygen consumption rates (OCR), and ATP production in treated myoblasts. This increase in maximal OCR was abrogated when CCA-EVs pre-treated with proteinase K were co-cultured with myoblasts, indicating the pro-metabolic effect was likely mediated by transmembrane or peripheral membrane proteins in CCA-EVs. Our data highlight the novel effect of Skm-EVs isolated post-CCA in mediating pro-metabolic effects in recipient cells and thereby transmitting the effects associated with traditional exercise. Further investigation to interrogate the underlying mechanisms involved in downstream cellular metabolic adaptations is warranted.

cell biology↗

bioRxiv2024

PI3K-dependant reprogramming of hexokinase isoforms controls glucose metabolism and functional responses of B lymphocytes

B lymphocyte metabolic reprogramming is essential for B cell differentiation and mounting a healthy immune response. The PI3K signaling pathway regulates B cell metabolism, but the mechanisms involved are not well understood. Here we report that signaling via PI3K8 can impact B cell glucose metabolism and immune functions via selective upregulation of hexokinase 2 (HK2). Three HK enzymes can catalyze the critical first step for glucose utilization and may selectively direct glucose into specific catabolic and anabolic pathways. While HK1 is constitutively expressed in B cells, HK2 is strikingly upregulated during B cell activation in a PI3K8-dependent manner. HK2 shows a unique distribution between mitochondrial and cytoplasmic pools that is also regulated by PI3K. Genetic deletion of HK2 significantly impairs extracellular acidification rate and glycolytic ATP production despite strong expression of HK1. B cell-specific deletion of HK2 in mice caused mild perturbations in B cell development but did not prevent generation of mature B cell subsets. HK2-deficient B cells show altered functional responses in vitro and evidence of metabolic adaptation to become less dependent on glucose and more dependent on glutamine. HK2-deficient B cells exhibit impaired glycolysis, altered metabolite profiles and altered flux of labeled glucose carbons into the pentose phosphate pathway. Upon immunization, HK2-deficient mice exhibit impaired generation of germinal centre B cells, plasmablasts and antibody responses. We further found that HK2 expression in primary human chronic lymphocytic leukemia (CLL) cells was associated with recent proliferation and could be reduced by PI3K inhibition. Our study identifies hexokinase 2 upregulation as a functionally important component of B cell metabolic reprogramming dependent on the PI3K pathway.

immunology↗

bioRxiv2024

Omega-3 polyunsaturated fatty acids modify glucose metabolism in THP-1 monocytes

Chronic inflammation is a driving factor in diseases like obesity and type 2 diabetes. Enhanced glucose metabolism, including via oxidative phosphorylation, may contribute to heightened immune activation. A recent clinical trial showed that supplementation with the n-3 fatty acid -linolenic acid (ALA) reduced oxidative phosphorylation rates in human monocytes. However, the mechanism remains unknown. Therefore, our objective was to explore the direct effects of ALA and docosahexaenoic acid (DHA) on glucose metabolism in a cell culture model and to explore the molecular mechanism. THP-1 monocytes were treated for 48h with 10-40 M of ALA or DHA and compared with vehicle and oleic acid controls. The Seahorse XFe24 system was used to approximate catabolic rates in the presence of glucose, including glycolysis and oxidative phosphorylation. The latter was validated by respirometry using an Oroboros O2k Oxygraph. Both ALA and DHA treatments reduced oxidative phosphorylation and increased glycolytic rates relative to control conditions. We identified pyruvate dehydrogenase kinase 4 (PDK4), an enzyme that inhibits the conversion of pyruvate to acetyl-CoA, as a possible mechanistic candidate. This gene was significantly upregulated by ALA and, to a greater extent, by DHA. Using fluorescent indicators, we also found that DHA increased reactive oxygen species while ALA had no effect. Our data suggest that ALA and DHA trigger a re-wiring of bioenergetic pathways in monocytes, possibly via the upregulation of PKD4. Given the close relationship between cell metabolism and immune cell activation, this may represent a novel mechanism by which n-3 fatty acids modulate immune function and inflammation.

cell biology↗

bioRxiv2023

A cell-type-specific nitric oxide-cGMP pathway regulates proprioceptor morphology

Neurons show remarkable morphological diversity facilitated by transcriptional regulation during development and fitting with their connectivity and function. Second messenger pathways mediated by cyclic 3,5 adenosine or guanosine monophosphate (cAMP or cGMP) may serve to integrate extrinsic signals and regulate branching and growth of neurites. We investigated the regulation of cGMP signaling in neuronal morphogenesis in Drosophila somatosensory neurons and found cell type-specificity in expression of the enzyme soluble guanylate cyclase (sGC) in different sensory neuron classes. sGC is responsible for the production of cGMP in response to the diffusible free radical messenger nitric oxide (NO) such that different levels of sGC impart cell type-specific sensitivity to NO. Supporting a role for the NO-cGMP pathway in neuronal morphogenesis, we find that knockout of sGC or nitric oxide synthase (NOS) impacts the morphology of multidendritic proprioceptors. The homeodomain transcription factor Cut shows different levels of expression in different somatosensory neuron types and we find that Cut regulates the level of cGMP signaling via repression of sGC expression. Thus, transcription factor levels contribute to cell type diversification by regulating levels of a signaling pathway to mediate somatosensory neuron morphogenesis.

neuroscience↗

bioRxiv2023

The mitophagy receptor Nix coordinates nuclear calcium signaling to modulate the muscle phenotype.

Mitochondrial quality control is critical in muscle to ensure both contractile and metabolic function. Nix is a BCL-2 family member, mitophagy receptor, and has been implicated in muscle atrophy. Human GWAS suggests altered Nix expression could predispose to manifestations of mitochondrial disease. To interrogate Nix function, we generated a muscle-specific knockout model. Nix knockout mice displayed a ragged-red fibre phenotype, along with accumulation of mitochondria and endo/sarcoplasmic reticulum with altered morphology. Intriguingly, Nix knockout mice were more insulin sensitive with a corresponding increase in glycogen-rich muscle fibres. Kinome- and gene expression analyses revealed that Nix knockout impairs NFAT and canonical myostatin signaling, with alterations in muscle fibre-type composition and evidence of regeneration. Mechanistic experiments demonstrated that Nix modulates mitophagy, along with ER-phagy leading to altered nuclear calcium signaling. Collectively, these observations identify novel roles for Nix coordinating selective autophagy, oxidative gene expression, and signaling pathways that maintain the muscle phenotype.

cell biology↗

bioRxiv2023

Functional Connectivity of the Cerebellar Vermis in Bipolar Disorder and Associations with Mood

PurposeStudies of the neural underpinnings of bipolar type I disorder have focused on the emotional control network. However, there is also growing evidence for cerebellar involvement, including abnormal structure, function, and metabolism. Here, we sought to assess functional connectivity of the cerebellum with the cerebrum in bipolar disorder and to assess whether any effects might depend on mood. MethodsThis cross-sectional study enrolled 128 participants with bipolar type I disorder and 83 control comparison participants who completed a 3T MRI scan, which included anatomical imaging as well as resting state BOLD imaging. Functional connectivity of the cerebellar vermis to all other brain regions was assessed. Based on quality control metrics of the fMRI data, 109 participants with bipolar disorder and 79 controls were used to in the statistical analysis comparing connectivity of the vermis as well as associations with mood. Potential impacts of medications were also explored. ResultsFunctional connectivity of the cerebellar vermis in bipolar disorder was found to differ significantly between brain regions known to be involved in the control of emotion, motor function, and language. While connections with emotion and motor control areas were significantly stronger in bipolar disorder, connection to a region associated language production was significantly weaker. In the participants with bipolar disorder, ratings of depression and mania were inversely associated with vermis functional connectivity. No effect of medications on these connections were observed. ConclusionTogether the findings suggest cerebellum may play a compensatory role in bipolar disorder and when it can no longer fulfill this role, depression and mania develop. The proximity of the cerebellar vermis to the skull may make this region a potential target for treatment with transcranial magnetic stimulation.

neuroscience↗

bioRxiv2022

In-silico comparative modeling and interaction studies of PRSV proteins - Accelerated towards dissection of structure based evolutionary divergence and functional interaction of virus within the host

Function based structure analysis of viral proteins reinforce their distinctive and unanticipated role within the host. The interaction dynamics of virus protein and host which is prerequisite for complete infectivity as well as systemic spread of invading virus demands to explore viral proteins in framework of their interacting partners. Papaya ringspot virus strain from Pakistan (PRSV-PK) spreading as atypical variant and causes drastic reduction in papaya production. The in-depth knowledge of the virus variant and effective management is obligatory. The desired objective is achievable once the evolutionary dynamics, molecular characterization and physicochemical structural properties conceived from the 3D protein structures are comprehended. Although the diversity studies on PRSV-PK strain been established but still there is a niche regarding structural based evolutionary dynamics of virus proteins and their probable interaction mode inside the host. The present investigations provided insights into the in-silico analysis of the functionally significant genes Coat protein (CP), Helper component proteinase (HC-Pro) and Nuclear Inclusion b protein (NIb) of PRSV-PK. The protein structure has been modeled using Phyre2, Swiss-Model and i-TASSER. Phyre2 built model showed 100% confidence for 67%, 63% and 20% sequence identity residues for PRSV CP, HC-Pro and NIb proteins respectively. The Swiss model showed identity values of 63.40%, 62.42% and 16.49% for CP, HC-Pro and NIb protein. whereas, i-TASSER server exhibited identity values of 67%, 63% and 19% for CP, HC-Pro and NIb proteins respectively. These structures provided a base line for functional analysis of experimentally derived crystal structures. The predicted models were validated using protein structure checking tools PROCHECK. Further the PRSV-PK-CP structures were compared with the PRSV-CP structures of representative isolates from different geographical regions. Nevertheless, the comparative modeling provides the insight into the evolutionary characteristics and proposed genetic diversity of PRSV based on protein structures. In addition, the conserved functional motifs have been mapped on aligned protein sequences of CP, HC-Pro and NIb, and their critical function within the host has been highlighted. Eventually, the interaction of papaya protein with the invading PRSV-CP has been predicted through in-silico protein-protein docking to elucidate their possible role in virus inhibition. The established structural-functional relation provided a basis to propose probable host-virus interactions in terms of virus infectivity, resultant host adaptability, and host defense response activation to counteract the virus invasion.

evolutionary biology↗

bioRxiv2022

Unique Pakistani gut microbiota highlights population-specific microbiota signatures of type 2 diabetes mellitus

Biogeographic variations in the gut microbiota reflect host and environmental factors delineating human populations, and are pivotal to understand global patterns of host-microbiota interactions in health and prevalent lifestyle-related diseases, such as type 2 diabetes mellitus (T2D). Pakistani adults, having an exceptionally high prevalence of T2D, are one of the most understudied populations in microbiota research to date. The aim of the present study is to examine the gut microbiota across individuals from Pakistan and other populations of non-industrialized and industrialized lifestyles with a focus on T2D. The fecal samples from 94 urban-dwelling Pakistani adults with and without T2D were profiled by 16S ribosomal RNA gene amplicon sequencing and qPCR, and plasma samples quantified for circulating levels of lipopolysaccharide binding protein (LBP) and the activation ability of Toll-like receptor (TLR)-signaling. Publicly available datasets generated with comparable molecular methods were retrieved for comparative analysis. Overall, urbanized Pakistanis gut microbiota was similar to that of transitional or non-industrialized populations, depleted in Akkermansiaceae and enriched in Prevotellaceae (dominated by the non-Westernized clades of Prevotella copri). The relatively high proportion of Atopobiaceae appeared to be a unique characteristic of the Pakistani gut microbiota. The Pakistanis with T2D had elevated levels of LBP and TLR-signaling in circulation as well as gut microbial signatures atypical of other populations e.g., increased relative abundance of Libanicoccus/Parolsenella, limiting the inter-population extrapolation of gut microbiota-based classifiers for T2D. Taken together, our findings call for more global representation of understudied populations to extend the applicability of microbiota-based diagnostics and therapeutics.

microbiology↗

bioRxiv2022

Skeletal muscle-derived extracellular vesicles are altered with chronic contractile activity

Extracellular vesicles (EVs) are small lipid bilayer-delimited particles that are secreted by all cells, playing a central role in cellular communication. EVs are released from skeletal muscle during exercise, but the effects of contractile activity on skeletal muscle-derived EVs (Skm-EVs) are poorly understood due to the challenges in distinguishing Skm-EVs derived from exercising muscle in vivo. Using tunable resistive pulse sensing (TRPS), we previously demonstrated that chronic contractile activity (CCA) increased the secretion of Skm-EVs from C2C12 myotubes, while their size and zeta potential remained unchanged. Here, we aimed to determine whether similar results could be obtained using an alternative method of EV characterization, dynamic light scattering (DLS). C2C12 myoblasts were differentiated into myotubes, and electrically paced (3h/day x 4days @14V, C-PACE EM, IonOptix) to mimic chronic exercise in vitro. EVs were isolated from conditioned media of control and stimulated myotubes using differential ultracentrifugation, and characterized based on size and zeta potential. The mean size of EVs from chronically stimulated myotubes (CCA-EVs, 132 nm) was 26% smaller than control (CON-EVs, 178 nm). Size distribution analysis revealed that CCA-EVs were enriched in small EVs (100-150 nm), while CON-EVs were largely composed of 200-250 nm sized vesicles. Additionally, zeta potential was 27% lower in CCA-EVs compared to CON-EVs. Our data indicate that the effect of CCA on facilitating the release of smaller, more stable EVs, is a robust finding, reproducible by multiple methods of EV characterization. Future studies investigating the mechanisms by which CCA influences Skm-EV biogenesis and secretion are warranted.

cell biology↗