Biology subjects

Mendiratta, M.

Publications and source records attributed to Mendiratta, M..

7 recordsLinked to original sources

bioRxiv2025

Dual Preconditioned Mesenchymal Stem Cells-Derived Culture Conditioned Media Augment Immunomodulation and Drive Metabolic Reprogramming in Acute Graft-versus-Host Disease

BackgroundThe toxicity associated with conventional conditioning regimens limits treatment outcomes in autoimmune disorders such as acute graft-versus-host disease (aGVHD). Developing effective adjunctive interventions to restore immune balance is therefore essential. This study investigated the immunomodulatory potential of mesenchymal stem cells (MSCs)-derived culture-conditioned media (CCM) from naive and preconditioned MSCs-hypoxic (MSCsHYP), apoptotic (MSCsAPO), and dual preconditioned (MSCsHYP+APO) in aGVHD. MethodsHuman MSCs isolated from bone marrow and Whartons Jelly were preconditioned under hypoxia (1% O2), apoptosis (1 {micro}M staurosporine, 24 h), or both. The immunoregulatory and antioxidant properties of their CCM were evaluated through T-cell proliferation assays, Treg induction, macrophage polarization, mitochondrial function, and T-cell bioenergetics. Comparative proteomic profiling of CCM from WJ-MSCs and WJ-MSCsHYP+APO co-cultured with aGVHD patient-derived activated PBMNCs was performed using LC-MS/MS, alongside in vivo validation in a chemotherapy-induced aGVHD murine model. ResultsWJ-MSCsHYP+APO-CCM exhibited superior immunomodulatory efficacy, suppressing T-cell proliferation, promoting Treg and Th2/Th9 differentiation, and driving M2 macrophage polarization. It reduced mitochondrial ROS, enhanced mitochondrial polarization, and shifted T-cell metabolism from glycolysis toward oxidative phosphorylation. Proteomic analysis revealed modulation of IL-12, IL-17, and JAK-STAT pathways, along with regulation of complement, coagulation, and metabolic cascades. Interaction with immune cells further enhanced its antioxidant and tissue-reparative properties via extracellular matrix remodeling. ConclusionDual preconditioning under hypoxia and apoptosis amplifies the immunomodulatory, antioxidant, and reparative efficacy of WJ-MSC-CCM, offering a potent non-cellular therapeutic strategy for aGVHD management. GRAPHICAL ABSTRACTImmunomodulatory and immune metabolic reprogramming potential of naive and preconditioned MSCs (MSCs, MSCsHYP, MSCsAPO, MSCsHYP+APO) derived CCM in Acute GVHD (created using Biorender.com) O_FIG O_LINKSMALLFIG WIDTH=95 HEIGHT=200 SRC="FIGDIR/small/686182v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@f7ed73org.highwire.dtl.DTLVardef@1063015org.highwire.dtl.DTLVardef@13ed827org.highwire.dtl.DTLVardef@e103c0_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Predictive Modeling of acute Graft-versus-Host-Disease using Machine Learning on Immune Cell and Cytokine Profiles at Engraftment

BackgroundAcute Graft-versus-Host-Disease (aGvHD) is a major immune complication following allogenic hematopoietic stem cell transplantation (Allo-HSCT), initiated by conditioning regimen-associated tissue damage. It involves the complex interplay of immune cells and cytokines. Our study aims to leverage machine learning (ML) algorithms on the immune and cytokine profile of Allo-HSCT recipients to develop biomarker-based classification models to predict the onset of aGvHD at the time of engraftment. Materials and MethodsSeventy patients diagnosed with hematological disorders who had undergone Ist Allo-HSCT were recruited from All India Institute of Medical Sciences, New Delhi, India. Peripheral blood (PB) was collected from the patients at the time of engraftment, and the immune cell subtypes and cytokine profiles were analyzed using flow cytometry and ELISA respectively. The individual cell counts were then processed using basic ML models, including support vector classifier with RBF kernel, Decision Tree, and Random Forest, chosen for their mathematical simplicity and feature importance advantage of Decision Trees and Random Forests. Various data settings were utilized in the study: combined immune and cytokine counts, immune cell counts only, cytokine counts only, T-cell counts only, NK cell counts only, dendritic cell counts only, and B-cell counts only. These configurations were selected to investigate how different data sets impact the prediction of aGvHD before its onset. ResultsAt the engraftment flow cytometric analysis of reconstituted lymphocytes in patients who developed aGvHD revealed that there was a remarkable decrease in the ratio of CD4+/CD8+ T-cell and Tregs, with an increase in the cytotoxic regulatory NK-cell, dendritic cells, and B-cell. The levels of pro-inflammatory cytokines (IFN-{gamma}, IL-1{beta}, IP-10, TNF-, IL-17, IL-12p70, MIP-1, MIP-1{beta}, RANTES), and Th17-and Th1-cells were elevated with consequent decline of the levels of anti-inflammatory cytokine IL-10, IL-2, IL-4 and Th2-, Th9-cells. Machine learning based on 48 parameters [all immune cell subsets n=34 and all cytokines (n=14)]. The correlation heat map shows a higher correlation of aGvHD with the cytokine profile with or without immune cells (accuracy: 1), T-cell alone (accuracy: 0.96); NK-cell alone (accuracy: 0.93); dendritic cells alone (accuracy: 0.90), B-cell alone (accuracy: 0.86). ConclusionThe current models classify perfectly, indicating the potential for a ML algorithm in predicting the onset of aGvHD. However, a study with a larger sample size is required to validate these classification models and mitigate the risk of overfitting observed due to the consistently high performance. The study also highlights the potential of cytokine profiles as a viable alternative to T-cell counts, as evidenced by the correlation heat map and classifier models. These findings provide valuable insights into dataset requirements and future directions for integrating ML models into aGvHD prediction.

immunology↗

bioRxiv2025

Dose-Response Immunomodulatory Effects of Mesenchymal Stem Cells-Derived Culture-Conditioned Media in Acute Graft-versus-Host-Disease

BackgroundMesenchymal stem cell-based therapy faces challenges that have driven interest in MSCs-derived culture-conditioned media (CCM) as a cell-free alternative. Our study aims to optimize the dose, and collection timing of CCM to enhance its therapeutic efficacy in aGVHD, while also standardizing co-culture conditions for CD3+ T-cell interaction with CCM. Material and MethodsHuman MSCs were isolated from BM and WJ and subsequently preconditioned under hypoxic conditions (1% O2) for 24 hours in a tri-gas incubator. Culture-conditioned media (CCM) was collected from both naive and hypoxia-preconditioned MSCs at 24, 48, and 72 hours and filtered using a 0.2 m membrane filter. CD3+ T-cell were isolated from PBMNCs derived from aGVHD patients. These T-cell were co-cultured at varying densities (2*106, 5*106, and 10*106 cells/ml) with different concentrations of CCM (25%, 50%, and 100%), and cell proliferation was assessed using the MTS assay. Furthermore, CD3+ T-cell proliferation and activation status were evaluated in a 2D co-culture model of CD3+ T-cell and CCM using flow cytometry. ResultsOur findings revealed that CCM collected at 48 hours, at a 50% concentration, exerted the most pronounced inhibitory effect on CD3+ T-cell proliferation, particularly at a density of 5*106 cells/ml, irrespective of the MSCs source. Hypoxia preconditioning significantly enhanced the immunomodulatory effects, with WJ-MSCsHYP-CCM demonstrating superior efficacy in suppressing T-cell proliferation, increasing the CD4+/CD8+ T-cell ratio, and reducing CD4+ T-cell activation compared to BM-MSCsHYP-CCM. ConclusionThese results emphasize the critical role of optimizing CCM collection timing and concentration to maximize therapeutic potential. Our study paves the way for the development of standardized, scalable, and effective cell-free therapies for aGVHD. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=198 HEIGHT=200 SRC="FIGDIR/small/640019v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@16ecc48org.highwire.dtl.DTLVardef@12591dborg.highwire.dtl.DTLVardef@18af4beorg.highwire.dtl.DTLVardef@1c9e8c9_HPS_FORMAT_FIGEXP M_FIG C_FIG Optimum conditions for CCM dose, collection time point, and CD3+ T-cell dose for effective immunomodulatory effect of MSCs-derived CCM in aGVHD. (created using Biorender.com)

immunology↗

bioRxiv2025

Hypoxia-Mediated Molecular Interactions of Tissue-Specific Mesenchymal Stem Cells Drive Metabolic Reprogramming and Immunomodulation in Acute Graft-versus-Host Disease

BackgroundMesenchymal stem cells (MSCs) mediate immunomodulation through various mechanisms, including apoptosis, efferocytosis, and mitochondrial transfer. Our study investigates the impact of hypoxia preconditioning on the immune metabolic reprogramming and immunomodulatory potential of MSCs in acute graft-versus-host disease (aGVHD). Additionally, we explored the differential immunomodulatory effects of tissue-specific MSCs, specifically bone marrow (BM) and Whartons Jelly (WJ), and elucidated the mechanisms underlying variability in their therapeutic efficacy. MethodsMSCs were isolated from BM and WJ and subjected to hypoxia preconditioning. Their immunometabolic programming potential was assessed by evaluating T-cell proliferation, regulatory T-cell (Treg) induction, effector T-cell differentiation toward Th2, Th9 phenotypes, and macrophage polarization, T-cell bioenergetics in the direct co-culture systems. ResultsWJ-MSCsHYP exhibited superior immunomodulatory properties compared to BM-MSCsHYP, by inhibiting T-cell proliferation, enhancing Treg induction, and promoting anti-inflammatory macrophage polarization. WJ-MSCsHYP demonstrated enhanced mitochondrial transfer to T-cell, improving mitochondrial health, reducing ROS, and promoting oxidative phosphorylation, leading to immune homeostasis. Unlike BM-MSCs, WJ-MSCs exhibited higher rates of apoptosis, which facilitated immune modulation through mechanisms independent of efferocytosis. ConclusionOur findings highlight that WJ-MSCsHYP is a superior candidate for aGVHD by utilizing apoptosis, mitochondrial transfer, and metabolic reprogramming to achieve immune regulation.

immunology↗

bioRxiv2024

A Comparative assessment of T-Cell response of Healthy donors and Acute Graft-versus-Host-Disease Patients: Customizing immune monitoring platform

BackgroundT-cell activation and proliferation are critical for understanding immune responses in both healthy and pathological conditions such as acute graft-versus-host disease (aGVHD). Phytohemagglutinin (PHA) and interleukin-2 (IL-2) are commonly used in in vitro assays to study T-cell responses. In view of the discrete response of T cells from aGvHD patients cohorts, our study optimized PHA / IL-2 based T-cell response among healthy individuals versus aGVHD patients. MethodsPeripheral blood was collected from age- and sex-matched healthy individuals (n=10) and aGVHD patients (n=10). CD3+ T-cell were isolated and stimulated with varying concentrations of PHA (1-10g/ml) and IL-2 (50-500 IU/ml). Cell proliferation was assessed using MTS and CFSE assays, while their apoptosis was evaluated with Annexin V/7-AAD staining. ResultsWe observed enhanced proliferation of healthy individuals at higher PHA concentrations (5-10g/ml), whereas aGVHD patients exhibited heightened proliferation even at lower PHA concentrations (1-2.5g/ml) at 48 hours. Prolonged exposure of T cells from GvHD patients to PHA led to decreased proliferation while it increased in the T cells from healthy donors.IL-2 supplementation (50 IU/ml) of T-cells from healthy donors significantly enhanced their proliferation and survival, with the optimal concentration supporting robust proliferation over extended culture periods. ConclusionOur study optimized PHA and IL-2 concentrations required for T-cell proliferation studies among healthy individuals and aGVHD patients. and underscored experimental conditions required for studying T-cell behavior/dysregulation in aGVHD condition. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/611044v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@ab3c47org.highwire.dtl.DTLVardef@2786forg.highwire.dtl.DTLVardef@30c698org.highwire.dtl.DTLVardef@3e4821_HPS_FORMAT_FIGEXP M_FIG C_FIG Highlights of the studyO_LILower doses of PHA (1.0g/ml) and IL-2 (50IU/ml) are optimum conditions for aGVHD patients derived CD3+ T-cell proliferation under in vitro conditions. C_LIO_LIThe maximum T-cell proliferation in healthy individuals occurs with 7.5g/ml PHA and 50IU/ml IL-2. C_LIO_LIHigher doses of PHA induce cytotoxicity in both cohorts. C_LIO_LIIL-2 significantly enhances T-cell survival, with 50IU/ml maintaining robust proliferation over extended periods. C_LI

immunology↗

bioRxiv2024

Uncovering the bequeathing potential of Apoptotic Mesenchymal Stem Cells via small Extracellular Vesicles for its enhanced immunomodulatory ability

Small Extracellular Vesicles (sEVs) derived from Mesenchymal Stem Cells (MSCs) have emerged as a promising avenue for cell-free therapeutics in regenerative medicine. These vesicles, endowed with regenerative cargo inherited from their parent cells, have attracted attention for their role in immunomodulation and ROS alleviation. Notably, the deliberate induction of apoptosis in MSCs prior to Extracellular Vesicles (EVs) isolation has been identified as a strategy to augment the regenerative capabilities of MSCs-EVs, as certain reports have suggested that MSCs undergo apoptosis to exert their therapeutic effect post-transplantation. Moreover, selecting an optimal tissue source for deriving MSC-sEVs is equally crucial to ensure consistent and improved clinical outcomes. Multiple attributes of MSCs like their antioxidant, Immunomodulatory & regenerative properties make them particularly appealing for clinical studies, wherein mechanisms such as paracrine secretions and efferocytosis play pivotal roles. This investigation meticulously explores the comparative immunomodulatory & antioxidant capabilities of Apoptotic sEVs (Apo-sEVs) with Viable sEVs (V-sEVs) obtained from both Bone Marrow (BM) and Whartons Jelly (WJ)-derived MSCs, using an in vitro liver injury model. The findings from the present study contribute valuable insights into the comparative efficacy of Apo-sEVs and V-sEVs. This will aid in addressing a critical gap in understanding the role of apoptosis in enhancing the reparative capability of MSCs-sEVs. It also aims to shed light on the optimal source of MSCs for generating Apo-sEVs in translational applications. GRAPHICAL ABSTARCT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/590581v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@1392183org.highwire.dtl.DTLVardef@1c72d6borg.highwire.dtl.DTLVardef@176ee90org.highwire.dtl.DTLVardef@128559e_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2024

Synergistic Hypoxia and Apoptosis Conditioning Unleashes Superior Mesenchymal Stem Cells Efficacy in Acute Graft-versus-Host-Disease

Mesenchymal stem cells (MSCs) have emerged as promising candidates for immune modulation in various diseases that are associated with dysregulated immune responses like Graft-versus-Host-Disease (GVHD). MSCs are pleiotropic and the fate of MSCs following administration is a major determinant of their therapeutic efficacy. In this context, we here demonstrate that hypoxia preconditioned apoptotic MSCs [bone marrow (BM), Whartons Jelly (WJ)] bear more immune programming ability in a cellular model of acute Graft-versus-Host-Disease (aGVHD). To this purpose, we programmed MSCs by exposing them to hypoxia and inducing apoptosis both sequentially as well as simultaneously. Our findings demonstrated that WJ MSCs that were conditioned with indicated approaches simultaneously induced the differentiation of CD4+T-cell towards Tregs, enhanced Th2 effector, and concomitantly mitigated Th1 and Th17, with polarization of M1 effector macrophages towards their M2 phenotype, and more interestingly enhanced efferocytosis by macrophages indicated Th2 programming ability of MSCs programmed by conjunctional approaches Overall, our study highlights the potential of WJ-MSCs conditioned with hypoxia and apoptosis concurrently, as a promising therapeutic strategy for aGVHD and underscores the importance of considering MSC apoptosis in optimizing MSCs-based cellular therapy protocols for enhanced therapeutic efficacy in aGvHD. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=123 HEIGHT=200 SRC="FIGDIR/small/588248v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@92c102org.highwire.dtl.DTLVardef@bd895forg.highwire.dtl.DTLVardef@185e041org.highwire.dtl.DTLVardef@45ef0f_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗