Kanwaljit Chopra

Beneficial modulation of the microbiota-gut-brain axis (MGBA) using dietary fibres and psychobiotics has emerged as a novel strategy for the management of various neurological conditions including epilepsy. Herein, we investigated the effects of supplementation of modified kodo millet bran (mKMB) along with GABA-producing Levilactobacillus brevis LAB6 MTCC 25662 in mitigating epileptic seizures, neuroinflammation and colonic aberrations in pentylenetetrazol (PTZ)-induced epilepsy in SD rats. Chronic PTZ kindling induced gut–brain dysregulation, whereas supplementation with mKMB and LAB6 reduced seizure severity, preserved neuronal spines, and restored hippocampal GABAergic function. It also effectively attenuated neuro- and systemic inflammation (~ 1.77-fold overall cytokine reduction), associated with enhanced FFAR2/3 expression and increased gut GABA and SCFAs. Microbiome analysis showed that supplementation mitigated kindling-induced dysbiosis by reducing Aerococcus, Clostridium, Enterococcus and enriching beneficial microbes; notably, Turicibacter abundance increased to 21.39% in Syn + PTZ compared to 13.6% in PTZ (log₂FC = + 0.65, p = 0.056) and 17.21% in mKMB + PTZ (log₂FC = + 0.56, p = 0.032), alongside an increase in Lactobacillus, particularly Levilactobacillus brevis. These findings highlight the protective potential of dietary fibres and rationale psychobiotics supplementation in modulating MGBA and developing an effective management strategy for epilepsy.

Reduced levels of circulating gonadal hormones in post-menopausal women can negatively affect various physiological functions, including brain and gut deficits. There is an urgent need to find novel strategies to mitigate estrogen–gut–microbiome–brain axis (EGMBA) dysfunction. This study aimed to investigate the effect of fructooligosaccharide (FOS), a non-digestible prebiotic fiber, on estrogen deficiency-induced alterations in the EGMBA using an ovariectomized (OVX) rat model. Adult female SD rats were bilaterally OVX to induce estrogen deficiency and associated EGMBA dysfunction. Rats were administered FOS (50 mg kg⁻¹ p.o.⁻¹) for 28 consecutive days. To assess EGMBA dysfunction, after 28 days, we performed behavioral tests, biochemical estimations (oxidative stress), molecular estimations (inflammatory markers via ELISA), gene expression analysis (HPA axis, monoamine neurotransmission, apoptosis, gut microbiota alterations, & gut barrier integrity via RT-PCR/qPCR), and histopathological analysis. Administration of FOS significantly improved behavioral outcomes (reducing anxiety and depression, and improving memory). FOS also attenuates oxidative stress and inflammatory markers. FOS regulates apoptosis (upregulation of BCL-2 and downregulation of Bax), HPA axis functioning (corticosterone, GR, MR, & CRH), and monoamine neurotransmission (MAO-A & COMT) in the hippocampus of OVX rats. FOS also promoted healthy cell growth and prevented apoptosis. Additionally, FOS restored gut microbial eubiosis, improved mucus secretion (MUC-2), preserved tight junction protein expression (Lipocalin-2, Claudin, & TLR-4), and maintained the colon microstructure. FOS exerts multifaceted protective effects on the EGMBA by modulating gut and brain functions. These findings support its potential as a non-hormonal therapeutic approach for managing postmenopausal complications.

Background Diabetic foot ulcers (DFUs), a debilitating manifestation of diabetes mellitus, involve persistent inflammation, oxidative stress, and impaired wound healing, largely driven by NF-κB overactivation. Amlexanox (ALX), a synthetic anti-inflammatory and antioxidant agent, was evaluated for its therapeutic potential in DFUs. Methods In-silico molecular docking and pharmacokinetic studies were conducted to predict ALX’s interactions with major DFU targets and to assess its topical suitability. STZ-induced diabetic rats with full-thickness foot ulcers (5 mm) received topical ALX (2.5% and 5%) or silver sulfadiazine ointments for 14 days. Wound closure, antioxidant enzymes, oxidative stress markers, connective tissue markers, pro-inflammatory markers and NF-κB expression were assessed in the wound tissue. Results ALX demonstrated strong binding to the pathological targets (NF-κB, MMP-9, MPO, and COX-2) and displayed a favourable PK profile. Moreover, ALX topical treatment did not normalise diabetic metabolic alterations, but it dose-dependently promoted wound healing. Indeed, ALX 5% significantly accelerated wound closure (p < 0.0001) and led to marked suppression of NF-κB and MMP-9 expression (p < 0.0001). It also enhanced the levels of connective tissue markers, including hexosamine (p < 0.001), hydroxyproline, and hexuronic acid (p < 0.0001). Robust anti-inflammatory effects were also observed, as ALX reduced IL-1β, IL-6, TNF-α levels and COX-2 activity (p < 0.0001). Furthermore, ALX 5% diminished oxidative stress by lowering LPO, PCO, and MPO levels (p < 0.0001), and restoring GSH levels and SOD and CAT (p < 0.001) activities indicating its anti-oxidant properties. Conclusion Taken together, the present work highlights the mechanisms associated with ALX mediated wound healing in diabetic rats thereby demonstrating its potential as a promising strategy for DFU management.

Estrogen deficiency in postmenopausal women disrupts reproductive, metabolic, brain, and gut health, partly by promoting inflammation, oxidative stress, and gut dysbiosis. Together, responsible for the development of gut-brain axis (GBA) dysfunction. Daily life stressors in women, particularly chronic stress, may further exacerbate this dysfunction; however, their synergistic effects with estrogen deficiency remain poorly understood. The current study aimed to develop an animal model of GBA dysfunction that mimics postmenopausal conditions. To induce GBA dysfunction, female Sprague Dawley rats were bilaterally ovariectomized (OVX) and exposed to chronic unpredictable mild stress (CUMS) for 28 days. To confirm GBA dysfunction, neurobehavioral, biochemical, molecular, and histopathological parameters were performed. We observed significant changes in physiological, & neurobehavioral parameters in OVX, CUMS, and OVX + CUMS group rats. We also observed marked enhancement in oxidative stress, neuroinflammation, and reduced acetylcholinesterase activity in the brain, and increased corticosterone levels in serum of OVX, CUMS, and OVX + CUMS group rats. Furthermore, we also observed a marked increase in pro-inflammatory cytokines, oxidative stress, reduction in MUC-2 and tight junction gene expression in the proximal colon, and changes in gut bacterial abundances in the feces of experimental groups. Histopathological examination revealed pronounced morphological damage in the proximal colon and brain of OVX, CUMS, and OVX + CUMS group rats. Thus, estrogen deficiency and chronic stress for one month synergistically induce GBA dysfunction. This developed animal model provides a robust platform for exploring novel therapeutic strategies to counteract GBA dysfunction arising from estrogen deficiency and chronic stress. Graphical Abstract

The present study aimed to evaluate the wound healing potential of phycocyanin (PC) in diabetic rats. STZ (55 mg/kg) was injected intraperitoneally in Wistar rats for the induction of diabetes. A sterile biopsy punch created a foot ulcer (5 mm). Diabetic rats were treated with topical PC (1.5% and 2.5%) for 2 weeks. Body weight and metabolic parameters were measured. Various biochemical estimations, including oxidative stress markers (LPO, PCO), antioxidant indices (GSH, SOD), collagen content, and anti-inflammatory markers (IL-1β, TNF-α, NF-κβ, IL-6), along with MMP-9, VEGF, Nrf2, and HO-1 levels were assessed in the wound tissue. MTT and in-vitro scratch assays were also performed. The results revealed that PC significantly increased wound closure and collagen content but failed to alter the STZ-induced increase in metabolic parameters. Furthermore, PC suppressed oxidative stress pro-inflammatory markers and MMP-9 levels and enhanced anti-oxidant defences, VEGF, Nrf2, and HO-1 levels in the wound tissue. In addition, in vitro studies demonstrated a significant increase in cell migratory activity in the wound-healing scratch assay. Collectively, this study suggests that PC accelerated wound healing by reducing oxidative stress and inflammation, possibly via upregulation of the Nrf2 signaling pathway, making it an effective approach for treating diabetic foot ulcers.

Gut dysbiosis, an imbalance in the gut microbiota (GM), has emerged as a key factor contributing to metabolic dysfunction following estrogen deficiency in menopause. Disruptions in gut microbial composition, triggered by hormonal imbalances and chronic stress, can exacerbate metabolic disturbances, such as obesity, dyslipidemia, and diabetes etc. Thus strategies, which modulates gut dysbiosis may proves important for the treatment of metabolic dysfunctions in postmenopausal state. However, the physiological, biochemical and molecular changes occurring due to estrogen deficiency and stress are yet to be elucidated. Thus, aim of the present study was to develop a pre-clinical model to investigate the effect of estrogen deficiency and chronic unpredictable mild stress (CUMS) alone and in combination on GM alterations and associated metabolic dysfunction. Ovariectomized (OVX) rats were subjected to CUMS for 28 days to induce metabolic alterations and gut dysbiosis. Estrogen deficiency and chronic stress altered physiological changes, adiposity, and anthropometric markers. Furthermore, OVX and CUMS elevated oxidative stress in liver and colon and increased inflammatory markers as detected in serum, Liver, and colon. This, in turn, led to damage of the mucosal layer and disruption of tight junction proteins. OVX and CUMS also caused gut dysbiosis, as indicated by an increased ratio of pathogenic to beneficial gut bacterial populations. Histopathological examination showed colon and liver tissues damage in OVX, CUMS, and OVX + CUMS group rats. OVX and CUMS, both individually and synergistically, contributed to the development of gut and metabolic dysfunction, collectively referred to as gut-metabolic axis dysfunction in OVX-stressed rats.

To ascertain safety, the assessment of bacterial strains to be used as probiotics needs rigorous and well-designed in vitro and in vivo studies. Here, the safety of a GABA-producing Levilactobacillus brevis LAB6 MTCC 25662 was assessed using in (silico, vitro and vivo) approaches. Firstly, the genomic analysis suggested that LAB6 is non-pathogenic to humans, as it does not harbour the genes for virulence, pathogenesis-related and horizontally transferable antimicrobial resistance. LAB6 neither produces biogenic amines nor degrades mucin, has no haemolytic activity and does not exert cytotoxicity on HEK-293 cells. In vivo safety of LAB6 was assessed in acute, subacute and sub-chronic oral feeding experiments following revised OECD guidelines 425, 407 and 408, respectively. Histopathological, blood biochemical, haematological parameters, gut permeability and oxidative stress levels were assessed. In vivo studies indicated that LAB6 did not negatively impact haematological markers or cause deleterious histological alterations in the vital organs. The anti-inflammatory potential of LAB6 in alleviating lipopolysaccharide (LPS)-induced inflammation in murine macrophages was assessed in the presence of GABA_A and GABA_B receptor antagonists. LAB6, owing to its GABA-producing ability, prevented LPS-induced inflammation by reducing TNF-α, IL-6 and IL-1β levels by 62.3%, 27.2% and 74.8%, respectively. Antagonism of the GABA_A receptor with bicuculline methiodide (BMI) partially blunted the protective effects of LAB6, while GABA_B antagonism has no significant impact in curtailing its protective effects. Overall results indicated that oral consumption of anti-inflammatory and GABA-producing LAB6 is safe to test in human studies further.

Understanding the communication between the gut and brain has emerged as a new research avenue in neurological disorders like epilepsy, with increasing evidence pointing towards the influence of intestinal microbes on the central nervous system. Here, the protective effects of a psychobiotic strain Levilactobacillus brevis LAB6 MTCC 25662 supplementation against pentylenetetrazole (PTZ)‐induced kindling in male Sprague Dawley rats was studied. Two GABA‐producing strains Levilactobacillus brevis LAB6 and Limosilactobacillus fermentum LAB19 were screened for their protective effects against chronic PTZ‐induced kindling. Chronic subconvulsive PTZ administration increased the convulsive activity, ultimately leading to generalized tonic–clonic seizures as indicated by a progressive rise in the seizure score, changes in endogenous antioxidants, and changes in inflammatory cytokines. Intervention with LAB6, but not LAB19, significantly attenuated these alterations and beneficially modulated gut health by increasing the production of short‐chain fatty acids and promoting healthy gut microbes, suggesting strain‐specific effects of LAB6. Further, colonization study with indocyanine green (ICG) labeled LAB6 was done, followed by dose‐dependent protective efficacy of LAB6 in PTZ‐induced kindling. A single dose administration (10¹⁰ CFU) showed that LAB6 translocated and adhered throughout the gastrointestinal (GI) tract within 8 h of administration and reached the distal colon and cecum. The supplementation of LAB6 at a medium dose of 2 × 10¹⁰ CFU was most efficient in restoring altered gut barrier functions, behavioral deficits, and modulation of peripheral GABA levels. Hence, it is a promising psychobiotic candidate for positively modulating gut‐brain functions in epilepsy. image

Purpose The regulatory landscape in rare diseases and oncology has evolved to address unmet medical needs by implementing expedited approval pathways. The US FDA’s Accelerated Approval and the EMA’s Conditional Marketing Authorization facilitate earlier patient access to therapies through reliance on surrogate endpoints derived from early-phase clinical trials. The review aims to provide a comprehensive review of the role and utilization of surrogate endpoints in accelerated drug approvals, highlighting their strengths, limitations, and the varying perspectives of stakeholders on their validity and utility. Methods This article reviews existing literature and regulatory guidelines to assess the effectiveness and challenges associated with surrogate endpoints in expedited approval pathways. It also examines the post-approval commitment adherence required by regulatory bodies, exploring discrepancies among stakeholder perspectives. Results Findings indicate that while surrogate endpoints enable faster market access, uncertainties remain regarding post-approval commitments and their consistency. Differences in stakeholder opinions also persist, reflecting varying levels of confidence in the validity and applicability of surrogate endpoints. Conclusion Surrogate endpoints play a crucial role in accelerating drug approvals in areas with high unmet needs, yet challenges around post-approval commitments and stakeholder acceptance suggest the need for enhanced regulatory clarity and ongoing assessment of surrogate endpoint validity.