Biology subjects

Patil, R.

Publications and source records attributed to Patil, R..

12 recordsLinked to original sources

bioRxiv2025

Chronic delivery of buprenorphine during abstinence decreases incubation of heroin seeking and neuronal activation in medial prefrontal cortex and striatum in male and female rats

Rationale and ObjectiveBuprenorphine is an FDA-approved medication for opioid addiction, but the brain regions underlying its therapeutic effects remain unknown. We previously found that chronic buprenorphine treatment decreases several relapse-related behaviors in rats. Here, we tested whether chronic buprenorphine decreases the time-dependent increase (incubation) in heroin seeking during abstinence. We also used the activity marker Fos to test whether buprenorphines inhibition of incubation of heroin seeking is associated with decreased activation of cortical and striatal regions. MethodsWe trained Oprm1-Cre rats and their wild-type littermates to self-administer heroin for 12 days (6 h/day). On abstinence Day 1, we tested for heroin seeking under extinction conditions. On Day 14, we implanted osmotic minipumps containing saline or buprenorphine (6 mg/kg/day). On abstinence Days 21-22, we tested the rats (or not tested, baseline condition) for incubation of heroin seeking, after which brains were collected for Fos immunohistochemistry. ResultsOprm1-Cre rats did not differ from wild-type littermates in heroin self-administration or non-incubated heroin seeking on abstinence Day 1. In both genotypes, chronic buprenorphine decreased incubated heroin seeking on Days 21-22. Buprenorphine also decreased incubation-related neuronal activation in several cortical areas (anterior cingulate and dorsal peduncular, but not prelimbic, infralimbic, or orbitofrontal cortex) and striatal areas (dorsolateral and dorsomedial striosomes and nucleus accumbens core, but not shell). ConclusionsChronic buprenorphine decreased incubation of heroin seeking, supporting the predictive validity of the rat model. This effect was associated with decreased neuronal activity in specific subregions of the medial prefrontal cortex and striatum.

neuroscience↗

bioRxiv2024

High fat diet induces differential age- and gender-dependent changes in neuronal function linked to redox stress.

The prevalence of neurodegenerative diseases, such as Alzheimers disease and Parkinsons disease, is steadily increasing, posing significant challenges to global healthcare systems. Emerging evidence suggests that dietary habits, particularly consumption of high-fat diets specify which fats, may play a pivotal role in the development and progression of neurodegenerative disorders. Moreover, several studies have shed light on the intricate communication between the gut and the brain, known as the gut-brain axis and its involvement in neurodegenerative processes. This study aims to assess the effects of a high-fat dietary intake on various aspects of neuronal function during aging and following gender separation to help understand the potential contributions of diet to neuronal function. To investigate the effects of a high-fat diet, Drosophila melanogaster was used and exposed to standard normal food diet (NF) and high-fat diet (HF). Adults were grouped at 10 and 45 days of age in male and female flies reared under the same conditions. Multiple assays were conducted, showing differential gender- and HF diet-induced oxidative stress levels as determined by malondialdehyde (MDA) measurements, enhanced caspase-3 expression and reduced climbing activity. Adult lifespan under both dietary conditions was unchanged but odour-associated learning ability was reduced in larvae reared in a HF diet. This is the first study to characterise effects of diet on neuronal phenotypes in an age- and gender-specific manner in a Drosophila model. Our findings suggest a HF diet induces differential forms of neuronal dysfunction with age and sex-specific outcomes, characterised by enhanced oxidative stress and cell death which impacts on neuronal and behavioural functions.

neuroscience↗

bioRxiv2023

Developmental Mouse Brain Common Coordinate Framework

3D standard reference brains serve as key resources to understand the spatial organization of the brain and promote interoperability across different studies. However, unlike the adult mouse brain, the lack of standard 3D reference atlases for developing mouse brains has hindered advancement of our understanding of brain development. Here, we present a multimodal 3D developmental common coordinate framework (DevCCF) spanning mouse embryonic day (E) 11.5, E13.5, E15.5, E18.5, and postnatal day (P) 4, P14, and P56 with anatomical segmentations defined by a developmental ontology. At each age, the DevCCF features undistorted morphologically averaged atlas templates created from Magnetic Resonance Imaging and co-registered high-resolution templates from light sheet fluorescence microscopy. Expert-curated 3D anatomical segmentations at each age adhere to an updated prosomeric model and can be explored via an interactive 3D web-visualizer. As a use case, we employed the DevCCF to unveil the emergence of GABAergic neurons in embryonic brains. Moreover, we integrated the Allen CCFv3 into the P56 template with stereotaxic coordinates and mapped spatial transcriptome cell-type data with the developmental ontology. In summary, the DevCCF is an openly accessible resource that can be used for large-scale data integration to gain a comprehensive understanding of brain development.

neuroscience↗

bioRxiv2023

Development and Characterization of Co-crystals Assisted with In-silico Screening for Solubility and Permeability Enhancement of Curcumin

Despite being a promising phytochemical, Curcumins potential applications are limited due to its classification in BCS class IV, which is associated with low water solubility and permeability. Enhancing the bioavailability of BCS class IV drugs presents a significant challenge, but crystal chemistry provides a hopeful avenue for overcoming this hurdle. In this research, co-crystals of Curcumin were developed to improve both solubility and permeability. Unlike traditional methods that require extensive trial-based lab work and time-consuming screening of co-formers, the use of molecular docking in In-silico co-former screening offers a scientific and rational approach to selecting suitable partners. In this study, two distinct co-crystals were synthesized using a solvent evaporation technique with methanol as the solvent, employing a 1:1 molar ratio. L-proline and piperine were chosen as co-formers to enhance solubility and permeability, respectively. The co-crystals underwent optimization and characterization through Design of Experiments (DOE). Comparing the dissolution study results for the same curcumin concentration, the cumulative drug release (CDR) after 8 hours was 20% for pure curcumin and an impressive 71% for curcumin-L-proline co-crystals. The permeability study, conducted over four hours using the everted gut sac method in phosphate buffer pH 6.8, revealed curcumins permeability to be less than 0.05 mg/mL, while curcumin-piperine co-crystals exhibited a five-fold increase (0.2545 mg/mL) in permeability. The co-crystals formed through a molecular ratio of 1:1 for curcumin-L-proline to enhance solubility and 1:1 for curcumin-piperine to enhance permeability, both demonstrated positive outcomes with support from optimization analysis, FTIR, DSC, SEM, PXRD analysis, and dissolution studies.

pharmacology and toxicology↗

bioRxiv2022

Diurnal stomatal apertures and density ratios affect whole-canopy stomatal conductance, water-use efficiency and yield

Key physiological traits of crop plants, such as transpiration, stomatal conductance, and photosynthesis are highly related to plant productivity. However, these traits are typically studied under steady-state conditions or modeled using only a few measured data points which do not reflect the dynamic behavior of the plant in response to field conditions. In this work, we hypothesized that the plastic behavior of whole-plant water balance regulation, as previously observed in tomato WT, was partially lost in the breeding process due to selective pressure towards productivity. We also hypothesized that this plastic behavior would be observed in some members of the tomato introgression lines (ILs) population, which was created by crossing the WT and M82 lines, particularly in ILs that demonstrate improved drought response. To overcome the steady-state bottleneck, and to test our hypothesis we used a gravimetric functional-phenotyping platform and a reverse-phenotyping method to examine the dynamic whole-plant water-regulation responses of tomato ILs and compared those responses with several years of yield performance in commercial fields. Indeed, our study enabled us to identify high plasticity in a few ideotypic ILs. We found that ideotype lines with highly plastic stomatal conductance and high abaxial-adaxial stomatal density ratios had stomatal apertures that peaked early in the day, even under water-deficit conditions. These traits resulted in dynamic daily water-use efficiency, along with rapid recovery of transpiration when irrigation was resumed after a period of imposed drought. Abaxial stomatal density was also found to be strongly correlated with the expression of the stomatal-development genes SPCH and ZEP. This study demonstrates how a reverse functional phenotyping approach based on field yield data, continuous and simultaneous whole-plant water-balance measurements and anatomical examination of individual leaves can help us to understand and identify dynamic and complex yield-related physiological traits.

plant biology↗

bioRxiv2021

Baseline cell proliferation rates and response to UV differ in Lymphoblastoid Cell Lines derived from healthy individuals of extreme constitution types

Differences in human phenotypes and susceptibility to complex diseases are an outcome of genetic and environmental interactions. This is evident in diseases that progress through a common set of intermediate patho-endophenotypes. Precision medicine aims to delineate the molecular players for individualized and early interventions. Functional studies in Lymphoblastoid Cell Line (LCL) model of phenotypically well characterized healthy individuals can help deconvolute and validate these molecular mechanisms. We developed LCLs from eight healthy individuals belonging to three extreme constitution types, deep phenotyped on the basis of Ayurveda. LCLs were characterized by karyotyping and immunophenotyping. Growth characteristics and response to UV was studied in these LCLs. We observed significant differences in cell proliferation rates between the contrasting groups such that one type (Kapha) proliferates significantly slower than the other two (Vata, Pitta). In response to UV, one fast growing group (Vata) shows higher cell death but recovers its numbers due to inherent higher rates of proliferation. The baseline differences in cell proliferation are key to understanding the survival of cells in UV stress. Variability in baseline cellular phenotypes not only explains the cellular basis of different constitutions types but can also help set priors during designing an individualized therapy with DNA damaging agents. This is the first study of its kind that shows variability of intermediate patho-phenotypes amongst healthy individuals that have implications in precision medicine.

cell biology↗

bioRxiv2020

Cocculus hirsutus -derived phytopharmaceutical drug has potent anti-dengue activity

BackgroundDengue is a serious public health concern worldwide, with ~3 billion people at risk of contracting dengue virus (DENV) infections. Currently, no effective vaccine or drug is available for the prevention or treatment of dengue, which leaves only anti-mosquito strategies to combat this disease. The present study was initiated to determine the in-vitro and in vivo protective effects of a plant-derived phytopharmaceutical drug for the treatment of dengue. Methodology/Principal FindingsIn our previous report, we had identified methanolic extract of the aerial parts of Cissampelos pareira to exhibit in vitro and in vivo anti-dengue activity against all the four DENV serotypes. In the current study, we have identified another Indian medicinal plant, Cocculus hirsutus, which has a more potent anti-dengue activity than C. pareira. The activity has been evaluated through flow-cytometry-based virus inhibition assay. Interestingly, the stem of C. hirsutus was found to be more potent than the aerial part irrespective of the extraction solvent used viz., denatured spirit, hydro-alcohol (50:50) and water. Hence, the aqueous extract of stem of C. hirsutus (AQCH) was further advanced for investigations because of greater regulatory acceptance. The AQCH exhibited dose-dependent inhibition of release of DENV and its secretory antigen, NS1. Five chemical markers viz. Sinococuline, 20-Hydroxyecdysone, Makisterone-A, Magnoflorine and Coniferyl alcohol were identified as the major chemical ingredients of the AQCH extract. These chemicals were subsequently used for extract standardisation. Importantly, AQCH completely protected AG129 mice at 25 mg/kg/dose body weight when fed 4 times a day post-infection with a lethal dose of DENV-2 S221 strain. Because of its potential as an effective phytopharmaceutical drug against dengue, AQCH, has been formulated into tablets for further pre-clinical and clinical developments. Conclusions/SignificanceWe provide evidence of the pan anti-dengue potential of C. hirsutus-based phytopharmaceutical drug as determined through in vitro and in vivo experiments. We have also characterized five chemical entities in the drug substance, which provides means for standardization of drug substance and drug product. Based on these findings, a program to develop a safe and effective C. hirsutus-derived phytopharmaceutical drug for the treatment of dengue has been initiated. Author summaryThere is an urgent need to develop a safe and effective drug against dengue, which is a rapidly expanding mosquito-borne viral disease. Half of the worlds population has been estimated to be at risk of contracting this disease and the situation remains grim due to lack of an approved drug. We aimed to develop an ethnopharmacological drug against dengue by exploring traditional Indian medicinal science, Ayurveda. This led us to identify a creeper, Cocculus hirsutus, as a more potent anti-dengue plant than Cissampelos pareira, reported in our earlier published study. The stem part of C. hirsutus was found to be more efficacious in inhibiting the propagation of dengue viruses (DENVs) in cell culture than its aerial part. Hence, we chose to advance aqueous extract of stem of C. hirsutus (AQCH) for further studies. Importantly, AQCH also protected immune-compromised mice from lethal DENV infection, which is suggestive of its potential clinical relevance. We have identified five chemical marker compounds in AQCH to gauge the quality and consistency of extract preparation and its formulation into stable tablets. Based on the findings of this study, we have undertaken the development of a safe and effective C. hirsutus-derived phytopharmaceutical drug for the treatment of dengue.

microbiology↗

bioRxiv2020

Genetic differences between extreme and composite constitution types from whole exome sequences reveal actionable variations

Personalized medicine relies on successful identification of genome-wide variations that governs inter-individual differences in phenotypes and system level outcomes. In Ayurveda, assessment of composite constitution types "Prakriti" forms the basis for risk stratification, predicting health and disease trajectories and personalized recommendations. Here, we report a novel method for identifying pleiotropic genes and variants that associate with healthy individuals of three extreme and contrasting "Prakriti" constitutions through exome sequencing and state-of-the-art computational methods. Exome Seq of three extreme Prakriti types from 108 healthy individuals 54 each from genetically homogeneous populations of North India (NI, Discovery cohort) and Western India (VADU, Replication cohort) were evaluated. Fishers Exact Test was applied between Prakriti types in both cohorts and further permutation based p-value was used for selection of exonic variants. To investigate the effect of sample size per genetic association test, we performed power analysis. Functional impact of differentiating genes and variations were inferred using diverse resources -Toppfun, GTEx, GWAS, PheWAS, UK Biobank and mouse knockdown/knockout phenotype (MGI). We also applied supervised machine learning approach to evaluate the association of exonic variants with multisystem phenotypes of Prakriti. Our targeted investigation into exome sequencing from NI (discovery) and VADU (validation) cohorts datasets provide ~7,000 differentiating SNPs. Closer inspection further identified a subset of SNPs (2407 (NI) and 2393 (VADU)), that mapped to an overlapping set of 1181 genes. This set can robustly stratify the Prakriti groups into three distinct clusters with distinct gene ontological (GO) enrichments. Functional analysis further strengthens the potential pleiotropic effects of these differentiating genes/variants and multisystem phenotypic consequences. Replicated SNPs map to some very prominent genes like FIG4, EDNRA, ANKLE1, BCKDHA, ATP5SL, EXOCS5, IFIT5, ZNF502, PNPLA3 and IL6R. Lastly, multivariate analysis using random forest uncovered rs7244213 within urea transporter SLC14A2, that associate with an ensemble of features linked to distinct constitutions. Our results reinforce the concept of integration of Prakriti based deep phenotypes for risk stratification of healthy individuals and provides markers for early actionable interventions.

genomics↗

bioRxiv2019

In Vivo Monitoring of Rare Circulating Tumor Cell and Cluster Dissemination in a Multiple Myeloma Xenograft Model

We recently developed Diffuse in vivo Flow Cytometry (DiFC), a new pre-clinical research tool for enumerating extremely rare fluorescently-labeled circulating cells directly in vivo. In this paper, we developed a green fluorescent protein (GFP) compatible version of DiFC, and used it to non-invasively monitor the circulating tumor cell (CTC) burden over time in a multiple myeloma disseminated xenograft model. We show that DiFC allowed counting of CTCs at estimated concentrations below 1 cell per mL in peripheral blood with a negligible false alarm rate. DiFC also revealed the presence of CTC clusters in circulation to our knowledge for the first time in this model, and allowed us to calculate their size, kinetics, and frequency of shedding. We anticipate that the unique capabilities of DiFC will have many applications in the study of hematogenous metastasis, and as a powerful complementary methodology to liquid biopsy assays.

bioengineering↗

bioRxiv2018

In Vivo Flow Cytometry of Extremely Rare Circulating Cells

Circulating tumor cells (CTCs) are of great interest in cancer research, but methods for their enumeration remain far from optimal. We developed a new small animal research tool called \"Diffuse in vivo Flow Cytometry\" (DiFC) for detecting extremely rare fluorescently-labeled circulating cells directly in the bloodstream. The technique exploits near-infrared diffuse photons to detect and count cells flowing in large superficial arteries and veins without drawing blood samples. DiFC uses custom-designed, dual fiber optic probes that are placed in contact with the skin surface approximately above a major vascular bundle. In combination with a novel signal processing, algorithm DiFC allows counting of individual cells moving in arterial or venous directions, as well as measurement of their speed and depth. We show that DiFC allows sampling of the entire circulating blood volume of a mouse in under 10 minutes, while maintaining a false alarm rate of 0.014 per minute. Hence, the unique capabilities of DiFC are highly suited to biological applications involving very rare cell types such as the study of hematogenic cancer metastasis.

bioengineering↗

bioRxiv2018

Nano immunoconjugates crossing blood-brain barrier activate local brain tumor immune system for glioma treatment

Treatment of brain gliomas with checkpoint inhibitor antibodies to cytotoxic T-lymphocyte-associated antigen 4 (a-CTLA-4) and programmed cell death-1 (a-PD-1) was largely unsuccessful due to their inability to cross the blood-brain barrier (BBB). We describe a new generation of nano immunoconjugates (NICs) developed on natural biopolymer scaffold, poly({beta}-L-malic acid), with covalently attached a-CTLA-4 and/or a-PD-1 for delivery across the BBB and activation of local brain anti-tumor immune response in glioma-bearing mice. NIC treatment of mice bearing intracranial GL261 glioblastoma (GBM) resulted in an increase of CD8+ T-cells with a decrease of T regulatory cells (Tregs) in the brain tumor area. Survival of GBM-bearing mice treated with combination of NICs was significantly longer compared to animals treated by single checkpoint inhibitor-bearing NICs or free a-CTLA-4 and a-PD-1. Our study demonstrates trans-BBB delivery of nanopolymer-conjugated checkpoint inhibitors as an effective treatment of GBM via activation of both systemic and local brain tumor immune response.

immunology↗

bioRxiv2018

Sequencing and de novo assembly reveals genomic variations associated with differential responses of Candida albicans ATCC 10231 towards fluconazole, pH and non-invasive growth

The whole genome sequencing generated 8.09 million paired-end reads, which were assembled into 2,262 scaffolds totaling 17,113,050 bp in length. We predict 7,654 coding regions and 7,647 genes, and annotated the coding regions with gene ontology terms based on similarity to other annotated genomes. Genome comparisons revealed variations (including SNP and indels in genes involved in pH response, fluconazole resistance and invasive growth) compared to C. albicans SC5314 and WO1.

genomics↗