Biology subjects

Saini, C.

Publications and source records attributed to Saini, C..

10 recordsLinked to original sources

bioRxiv2026

Gut-Associated Metabolites (GAMs) revitalize dysfunctional CD8⁺ T lymphocytes in Osteosarcoma

Osteosarcoma (OS) is one of the top ranking and deadliest primary malignant bone tumor of youngsters and adolescents. OS has poor prognostic features due to its immunosuppressive "cold" tumor microenvironment, obstructing the anti-tumor effector functions of immune cells (including cytotoxic CD8 T lymphocytes). Recent developments have focused on the role of Gut Microbiota in modulating effector immune responses in various cancers. However, the immunomodulatory role of gut microbiota and its derived gut-associated metabolites (GAMs) in OS still remains unclear. Here we report that peripheral CD8 T lymphocytes in osteosarcoma patients are less frequent in circulation, hypo-producers of effector cytokines (IFN-{gamma} and TNF-), and have compromised metabolic fitness. We characterized and investigated the effect of a panel of microbiota-derived GAMs on CD8 T lymphocytes, confirming their immunomodulatory role with respect to CD8 T lymphocytes activation, cellular metabolism and effector functions. Indole-3-lactic acid (ILA; product of tryptophan metabolism), was observed to be the strongest immunostimulant among all the GAMs studied. ILA enhanced the anti-tumor effector functions of CD8 T lymphocytes through increased glucose uptake, a higher mitochondrial bio-mass and increased production of IFN-{gamma} and TNF-. Moreover, ILA-primed cytotoxic T lymphocytes exhibited considerably higher cytotoxicity and increased apoptosis of osteosarcoma cells (U2OS). Altogether, our data demonstrates that ILA acts as a microbial-derived immune modulator to metabolically reprogram and restore dysfunctional CD8 T lymphocytes against osteosarcoma. This study for the first time demonstrates the therapeutic potential of exploiting the nexus between "Gut-Immune-Bone Tumor" ternary as a safe and cost-effective combinatorial immunotherapy against osteosarcoma. Graphical AbstractIndole-3-Lactic Acid (ILA) Reinvigorates CD8+ T-Cell Immunity in Osteosarcoma Circulating CD8+ T cells from osteosarcoma patients exhibit impaired metabolic fitness, reduced effector cytokine production, and diminished tumoricidal activity. Screening of gut-associated metabolites identified the microbial tryptophan metabolite indole-3-lactic acid (ILA) as the most potent immunomodulator. ILA restores glucose uptake and mitochondrial biomass, enhances IFN-{gamma} and TNF- production, promotes polyfunctional CD8+ T-cell responses, and significantly improves CTL-mediated killing of osteosarcoma cells, highlighting its potential as a microbiota-derived immunometabolic therapeutic for osteosarcoma. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/741694v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@19809aforg.highwire.dtl.DTLVardef@a3eb09org.highwire.dtl.DTLVardef@19bada3org.highwire.dtl.DTLVardef@1a4c4da_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2026

Vitamin D mitigates Inflammatory Bone Loss in Postmenopausal Osteoporosis via modulating the Gut-Immune-Bone axis

Osteoporosis is a prevalent skeletal disorder characterized by deterioration of bone microarchitecture and loss of bone mineral density, leading to increased fracture risk and substantial health and economic burdens, particularly among older adults. Bone remodeling is orchestrated by a complex interplay of systemic and local regulators, among which vitamin D plays a central role in maintaining skeletal homeostasis. Although numerous studies have examined the effects of vitamin D on bone metabolism, outcomes have been inconsistent across populations, dosing regimens, and experimental models. To clarify the net skeletal impact of vitamin D, we investigated its effects in postmenopausal osteoporosis (PMO). Vitamin D (1,25-dihydroxyvitamin D3- active form of vitamin D) supplementation effectively prevented bone loss in ovariectomized mice, at both lower and higher concentrations. Mechanistically, vitamin D promoted osteoclast differentiation in vitro, consistent with its RANKL-dependent pro-osteoclastogenic activity, yet paradoxically conferred bone protection in vivo. This discrepancy was explained by vitamin Ds profound immunomodulatory effects, which reshaped both innate and adaptive immune responses to suppress osteoclast formation and function. Concurrently, vitamin D improved intestinal barrier integrity and restored gut microbial composition, thereby stabilizing the gut-immune-bone axis and reducing pro-resorptive inflammatory signaling. Together, these findings demonstrate that vitamin D prevents bone loss through the coordinated regulation of immune and gut homeostasis, reconciling its apparent pro-resorptive effects in vitro with its overall anti-resorptive outcomes in vivo. This integrated mechanism highlights immune-gut microbial modulation as a key mediator of vitamin D-induced bone preservation and supports the development of vitamin D as an immunotherapeutic adjunct for the prevention and management of PMO. Altogether, our findings for the first time dissect the paradox surrounding the osteoprotective property of vitamin D supplementation. O_FIG O_LINKSMALLFIG WIDTH=186 HEIGHT=200 SRC="FIGDIR/small/735433v1_ufig1.gif" ALT="Figure 1"> View larger version (73K): org.highwire.dtl.DTLVardef@1550497org.highwire.dtl.DTLVardef@a06ab4org.highwire.dtl.DTLVardef@1bd55e5org.highwire.dtl.DTLVardef@1526acd_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical Abstract:C_FLOATNO Vitamin D strengthens intestinal barrier integrity and restores gut microbial composition, which stabilizes the gut-bone axis and suppresses pro-resorptive inflammatory signaling. Vitamin D enhances osteoclastogenesis both directly and through ILC3- and Th17-mediated pathways, while inhibiting osteoclastogenesis via Treg- and Breg-dependent mechanisms. In circulation, vitamin D maintains the homeostasis between IL-10 and IL-17 cytokines. Collectively, these results indicate that vitamin D helps prevent bone loss by orchestrating the immune system and maintaining gut homeostasis. C_FIG

immunology↗

bioRxiv2026

Enhanced strand transfer and mismatch extension by HIV-1C reverse transcriptase promote sequence motif duplication

Genetic diversification of HIV-1 is driven largely by the error-prone activity of reverse transcriptase (RT) and frequent template switching during reverse transcription. A rare outcome of nonhomologous recombination is sequence motif duplication, which can alter viral gene regulation and protein function. Previous studies have shown that such duplications occur at significantly higher frequencies in HIV-1 subtype C (HIV-1C), particularly within the long terminal repeat (LTR) and p6-Gag regions, where they can confer replication advantages. However, the mechanistic basis for this subtype-specific bias remains unclear. We therefore investigated whether intrinsic biochemical properties of HIV-1C RT contribute to its elevated duplication frequency. Bioinformatic analysis of 6,877 full-length HIV-1 genomes identified four duplication hotspots, with the highest frequencies in HIV-1C. Comparative sequence analysis of RT revealed several subtype-specific residues, including a highly conserved threonine at position 359 (T359) in the connection domain of HIV-1C RT. Structural modeling suggested that T359 can form an additional hydrogen bond with the nascent cDNA, potentially stabilizing the RT-template complex. Biochemical characterization of recombinant RT variants demonstrated that residue 359 modulates polymerase activity and maintains subtype-specific optimal catalytic function. Functional assays further revealed that HIV-1C RT exhibits enhanced template strand transfer compared with HIV-1B RT. In addition, next-generation sequencing-based primer extension assays showed that HIV-1C RT extends mismatched 3' termini more efficiently across multiple mismatch types. Together, these findings indicate that subtype-specific biochemical properties of HIV-1C RT, particularly enhanced strand transfer and mismatch extension mediated in part by T359, promote nonhomologous recombination events that generate sequence motif duplications. This work provides a mechanistic explanation for the elevated duplication frequency characteristic of HIV-1C and highlights how subtle RT polymorphisms can shape viral evolutionary trajectories.

microbiology↗

bioRxiv2026

A highly stringent high-throughput screening assay for identifying transcription-modulating agents of HIV-1 latency

HIV-1 latency remains a central obstacle to curing infection, and current latency-modulating agents (LMAs) suffer from poor specificity and inconsistent efficacy. To enable discovery of small molecules (SMs) that directly target the viral master transcriptional regulatory circuit (MTRC), we developed a highly stringent, dual-reporter high-throughput screening (HTS) assay based on a natural HIV-1 subtype C long terminal repeat (LTR) variant, LRhR-HC, which exhibits markedly reduced transcriptional noise and a high activation threshold. We engineered Jurkat cells to stably harbour two independent reporter cassettes driven by the tough-to-activate LRhR-HC-LTR and the canonical LR-HHC-LTR, enabling simultaneous detection of latency-promoting and latency-reversing activities through both fluorescent and secreted enzymatic reporters. This dual-reporter line responded robustly and predictably to conventional latency-reversal agents (LRAs) and latency-promoting agents (LPAs), validating assay responsiveness. Z'-factor measurements demonstrated excellent assay performance, with values ranging from approximately 0.7 across different formats, confirming a strong dynamic range and reproducibility. Screening of the 1,520-compound Prestwick chemical library (PCL), which includes FDA-approved drugs, identified 27 candidate LPAs, including known agents such as Spironolactone and Aminacrine, thereby validating the assay specificity while revealing several novel inhibitory molecules. A mechanistically focused panel of 10 additional compounds yielded two putative LPAs and one LRA, with secondary analyses confirming their latency-modulating activities and cytotoxicity profiles. Collectively, this HIV-1C-derived dual-reporter platform provides a stringent and flexible HTS system for identifying LMAs with potential applications in both block-and-lock and shock-and-kill cure strategies.

molecular biology↗

bioRxiv2025

Deoxycholic acid (DCA) alleviates LPS-induced Inflammatory Bone Loss via modulating the Gut-Bone homeostasis

Osteoporosis and other forms of inflammatory bone loss are marked by disrupted bone remodelling resulting from an imbalance between osteoclast-mediated bone resorption and osteoblast-driven bone formation. This imbalance is often exacerbated by chronic inflammation and gut microbiota dysbiosis. Recently, attention has turned to gut-associated metabolites (GAMs) such as secondary bile acids, particularly deoxycholic acid (DCA), which act as immunomodulators influencing both systemic inflammation and bone metabolism. In this study, we investigated the role of DCA in inflammatory bone loss using an in vivo model, with a focus on osteoblast and osteoclast function, gut barrier integrity, and gut-microbiota diversity. DCA administration significantly suppressed osteoclastogenesis along with enhancing osteoblastogenesis, indicating its dual regulatory role in bone remodelling. Furthermore, DCA treatment enhanced gut integrity, reversed dysbiosis and reduced systemic inflammation by downregulating osteoclastogenic cytokines (TNF-, IL-6, IL-17, RANKL, etc.). These findings suggest that DCA mitigates LPS-induced inflammatory bone loss through a multifaceted mechanism involving direct effects on bone cells and restoration of gut integrity and homeostasis. Our results highlight the therapeutic potential of targeting the gut microbiota-derived bile acid pathway, particularly DCA, as a novel strategy for managing osteoporosis and other inflammatory bone disorders. O_FIG O_LINKSMALLFIG WIDTH=131 HEIGHT=200 SRC="FIGDIR/small/678724v1_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@1702fddorg.highwire.dtl.DTLVardef@568b38org.highwire.dtl.DTLVardef@1ce463aorg.highwire.dtl.DTLVardef@164a04d_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 9.C_FLOATNO Graphical Abstract. Secondary bile acids act via FXR and TGR5 receptors on osteoclast and osteoblast cells and thereby regulate bone remodelling. C_FIG

cell biology↗

bioRxiv2025

ILC3 inhibits Osteoclastogenesis and ameliorates Inflammatory Bone Loss in Post-menopausal Osteoporosis

Although the significance of T lymphocytes in maintaining bone homeostasis is well established, the role of innate lymphoid cells (ILCs), the innate counterparts of T cells, in maintaining bone homeostasis is uncertain. In this study, we examined, for the first time, the anti-osteoclastogenic property of ILC3 in vitro. We observed that ILC3 inhibits RANKL-mediated osteoclastogenesis in a cell ratio-dependent manner. We further employed an ovariectomized (ovx) mouse model, which mimics postmenopausal osteoporosis (PMO), to investigate the role of ILC3 in inflammatory bone loss. Notably, our in vivo data unequivocally validate that the ovx mice have a markedly lower frequency of ILC3 in the bone marrow (BM). Furthermore, the temporal-kinetic analysis revealed that alterations in ILC3 cell dynamics and function in the BM are associated with the development and progression of inflammatory bone loss in PMO. Additionally, our in vivo results demonstrate that dysbiosis in PMO drives significant alterations in the frequency of ILC3 subsets by promoting the expansion of IL-17-producing Nkp46- ILC3 and inhibiting the development of IL-22-producing Nkp46+ ILC3. Moreover, T-bet+ ILC3s are significantly increased in ovariectomized (ovx) mice. Altogether, the present study for the first time, reports the critical role of dysregulated ILC3 in the progression of PMO and offers a novel immunotherapeutic approach targeting ILC3 for treating and managing PMO. Graphical abstractPMO is associated with leaky gut and dysbiosis, which leads to a deficiency of SCFAs in the ovx mice. Dysbiosis in ovx mice disturbs the homeostatic balance of the ILC3 with an increase in IL-17-producing ILC3 and a decrease in IL-22-producing ILC3. Further Tbet expression is enhanced in the ILC3 of the ovx mice, which is associated with disease pathogenesis. Additionally, IL-2 expression is decreased in the ILC3 of the ovx mice. Decreased expression of ILC3 migratory molecules reduced the migration of ILC3 in bone marrow. ILC3 inhibits osteoclast differentiation. However, in the case of ovx mice, ILC3 are compromised and promote osteoclastogenesis. O_FIG O_LINKSMALLFIG WIDTH=154 HEIGHT=200 SRC="FIGDIR/small/677486v1_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@10543ccorg.highwire.dtl.DTLVardef@a6452dorg.highwire.dtl.DTLVardef@798f8org.highwire.dtl.DTLVardef@cc026c_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Promoter evolution in HIV-1C establishes latent reservoirs highly resistant to reversal

Latent viral reservoirs remain a major barrier to curing HIV-1, with the long-terminal repeat (LTR) and Tat playing crucial roles in regulating viral transcription. Subtype-specific transcription factor binding site (TFBS) variations within the LTR significantly influence latency and reservoir stability. In earlier work, we identified HIV-1C LTR variants with duplicated TFBS motifs, including NF-{kappa}B, AP1, RBEIII, and TCF-1/LEF-1. Using five cell models, including Jurkat and primary CD4 T cells, we compared canonical R-LTR and variant R2-LTR strains. Across sub-genomic reporters, single-round infections, and full-length viral vectors, we found that the balance between RBEIII and NF-{kappa}B motifs governs stability of latency. The two-viruses-one-cell system that normalized confounding environmental factors further revealed that latency is primarily controlled by intrinsic transcriptional circuits rather than external stimuli. In longitudinal studies of HIV-1 individuals from acute and chronic infection phases, we observed dominant R strains during early infection and the spontaneous emergence of R2 strains in nearly half of chronic-phase subjects, a process accelerated by ART. Upon CD4 T cell activation, R strains preferentially rebounded, while R2 strains showed strong resistance to reversal, even in subjects harbouring a co-infection. Together, these findings establish the clinical significance of LTR variation in latency regulation and identify the R2 phenotype as a critical determinant of reservoir persistence. These results underscore the importance of addressing reservoir heterogeneity in cure strategies, particularly in HIV-1C-prevalent regions.

microbiology↗

bioRxiv2023

Mitigating inflammatory bone loss in post-menopausal osteoporosis via targeting the IL-9 producing osteoclastogenic Th9 cells

Recent discoveries have established the pivotal role of IL-9-secreting Th9 cells in a wide spectrum of inflammatory and autoimmune diseases. However, little is known about how Th9 cells contribute to the etiology of inflammatory bone loss in post-menopausal osteoporosis (PMO). We observed that IL-9 has a pathological impact on inflammatory bone loss in ovariectomized (Ovx) mice. Our in vivo temporal kinetics analysis further revealed that estrogen deprivation increased the release of IL-9 from Th which in turn enhances the IL-17-producing Th17 cells. Both ex vivo and in vivo studies corroborated these findings in Ovx mice, as estrogen diminishes IL-9s effect on the differentiation of Th17 cells as well as the potential of Th9 cells to produce IL-9. Mechanistically, Th9 cells in an IL-9-dependent manner enhance osteoclastogenesis and thereby establish themselves as a novel and independent osteoclastogenic Th subset. Blocking IL-9 improves bone health in Ovx mice by inhibiting the differentiation and function of both osteoclasts and Th9/Th17 cells. Our clinical findings further attested to the osteoporotic role of Th9 cells in post-menopausal osteoporotic human subjects. Collectively, our study establishes IL-9-secreting Th cells as the critical regulator of bone loss observed in PMO and highlights the fundamental implications of IL-9/Th9 targeted immunotherapies as an innovative approach for the treatment of inflammatory bone loss observed in osteoporosis.

immunology↗

bioRxiv2023

Lactobacillus rhamnosus (LR) ameliorates acute respiratory distress syndrome (ARDS) via modulating the lung Innate Lymphoid Cells (ILCs)-Mononuclear Phagocytic System (MPS)

Acute-respiratory-distress-syndrome (ARDS), the ultimate manifestation of acute-lung-injury (ALI) is a life-threatening respiratory failure with a significantly higher incidence and mortality worldwide. Recent discoveries have emphasized the existence of a potential nexus between gut and lung-health wherein physiology of the gut is directly linked with the outcomes of the lung pathologies. These discoveries fuel novel approaches including probiotics for the treatment of several respiratory disorders including ALI/ARDS. Lactobacillus rhamnosus (LR) is a preferred probiotic of choice as it has been reported to exhibit potent anti-inflammatory activities in various inflammatory diseases. In the present study, we investigated the prophylactic-potential of LR in lipopolysaccharide (LPS)-induced ALI/ARDS mice model, which mimics the pathophysiology of several respiratory disorders including respiratory tract infections, COVID-19, influenza, pneumonia, asthma, tuberculosis, cystic fibrosis, chronic obstructive pulmonary disease (COPD) etc. Our in vivo findings revealed that pretreatment with LR significantly attenuated lung inflammation and improved the pathophysiology of lung-tissues in ALI/ARDS mice. We observed that LR-administration suppressed the LPS-induced inflammatory cell infiltration in the lungs via ameliorating vascular-permeability (edema) of the lungs. Acute and chronic lung-disorders, including ARDS, are largely governed by innate-immune response. Interestingly, we observed that LR via modulating different ILCs (first responder to infections) subsets viz. ILC1, ILC2 and ILC3 prevent lung-fibrosis and maintain vascular permeability in LPS induced ALI/ARDS mice model. Of note, we observed a significant-enhancement in the percentage of inflammatory IL-17 producing CD3-Ror{gamma}t+NKp46- ILC3-fraction along with a significant reduction in IL-22 (responsible for vascular integrity) producing CD3-Ror{gamma}t+NKp46+ ILC3 in both the BALF and lung-tissues. These ILCs would further augment the activation and recruitment of mononuclear phagocytic system (MPS-monocytes, macrophages and DCs) along with neutrophils and eosinophils in the lungs and BALF in ALI/ARDS mice model. Furthermore, gene expression and protein-analysis demonstrated that LR treatment significantly reduces the expression of inflammatory-cytokines in lung tissue and serum, thereby suggesting its potent immunomodulatory activity in attenuating ALI/ARDS. Summarily, our research convincingly establishes the prophylactic-role of LR in the prevention and management of respiratory-distress syndromes driven by the diverse inflammatory insults via modulating the lungs "ILCs-MPS" axis with significant clinical-implications for the management of COVID-19, Influenza, COPD etc.

immunology↗

bioRxiv2022

Bifidobacterium longum attenuates ovariectomy-induced bone loss via modulating the Immunoporotic Breg-Treg-Th17 cell axis

Discoveries in the last few years have emphasized the existence of an enormous breadth of communication between osteo-immune system. These discoveries fuel novel approaches for the treatment of several bone-pathologies including osteoporosis, an inflammatory bone anomaly affecting more than 500 million people globally. Bifidobacterium longum (BL) is preferred probiotic of choice due to its varied immunomodulatory potential in alleviating various inflammatory diseases. Here, we evaluate the effect of BL in ovariectomy (ovx)-induced post-menopausal osteoporotic mice model. Our in vitro findings reveal that BL suppresses the differentiation and functional activity of RANKL-induced osteoclastogenesis in both mouse bone marrow cells and human PBMCs. Our in vivo data clearly establish that BL exhibits osteoprotective potential via modulating the "immunoporotic" Breg-Treg-Th17 cell-axis. Furthermore, {micro}CT and bone mechanical strength data support that BL supplementation significantly enhanced bone mass and strength, and improved microarchitecture in ovx mice. Remarkably, alteration in frequencies of CD19+CD1dhiCD5+ Bregs, CD4+Foxp3+IL-10+ Tregs, and CD4+Ror{gamma}t+IL-17+ Th17 immune cells in distinct lymphoid organs along with serum-cytokine data (enhanced anti-osteoclastogenic cytokines IFN-{gamma} and IL-10 and reduced osteoclastogenic-cytokines IL-6, IL-17, and TNF-) strongly support the immunomodulatory potential of BL. Altogether our findings establish a novel osteo-protective and immunoporotic potential of BL in augmenting bone health under osteoporotic conditions.

immunology↗