Biology subjects

Verma, G.

Publications and source records attributed to Verma, G..

12 recordsLinked to original sources

bioRxiv2026

Frequency Shift Imaging Using an Adiabatic SLR Self-Refocused Pulse for Positive-Contrast SPION Imaging at 7T

Abstract: Purpose: Superparamagnetic iron oxide nanoparticles (SPION) are widely used as MRI contrast agents for cell tracking and molecular imaging. However, their detection using conventional T2/T2* weighted and susceptibility-weighted MRI relies on negative contrast (signal voids), which lacks specificity because similar hypointense signals can arise from other sources of magnetic susceptibility. This study introduces Frequency Shift Imaging (FSI), a positive-contrast technique based on an adiabatic Shinnar-Le Roux (SLR) self-refocused pulse that addresses the limited specificity of conventional SPION imaging. Methods: An adiabatic self-refocused RF pulse was designed using the SLR algorithm for frequency-selective positive-contrast imaging. The optimized pulse had a spectral bandwidth of 370 Hz, a peak RF amplitude of 14 T, and a duration of 15 ms. FSI data were acquired on a 7T MRI scanner. Phantom experiments used agarose-embedded 30 nm cell-labeled SPION at 0%, 25%, 75%, and 100% concentrations (pure). In vivo validation was performed following intramuscular SPION injection into the hind limb of a mouse. FSI images were compared with conventional acquisitions. Results: Quantitative phantom analysis showed normalized FSI signal intensities of 25.02% and 75.16% for samples containing 25% and 75% labeled cells, respectively, demonstrating an approximately proportional relationship between FSI signal and labeled-cell concentration. In vivo, FSI generated localized hyperintense positive contrast at the SPIO injection site, providing improved conspicuity and localization compared with the corresponding negative-contrast signal void. Discussion and Conclusion: FSI enables robust, B1-insensitive, positive-contrast SPION imaging at 7T with reduced echo time and SAR compared to paired adiabatic approaches, supporting its potential for quantitative cell tracking and molecular imaging applications.

bioengineering↗

bioRxiv2026

Tuning T-cell immunological synapse by modular DNA-Nanobody engagers for precision immunotherapy

Bispecific T-cell engagers (TCEs) are a promising class of cancer immunotherapies, but their clinical use is limited by toxicity and insufficient specificity. Tuning the T cell- tumor interface through engager architecture may address these drawbacks. To this end, we engineered hybrid constructs composed of two nanobodies targeting CD3 and the model tumor antigen HER2, respectively, connected by rigid DNA linkers of variable length. Using cytotoxicity assays and hybrid biophysical platforms, we demonstrate a linker-length dependence of cell spreading on antigen, target killing and cytokine release, revealing a functional decoupling between killing and cytokine secretion, and implicating the glycocalyx as a key player. Through the addition of EGFR targeting, we also generate trispecific constructs implementing an "OR-gate" logic to address tumor heterogeneity and reduce resistance due to antigen loss. Overall, these versatile constructs show great therapeutic promise, and at the same time serve as platforms to test hypotheses on biophysical mechanisms.

synthetic biology↗

bioRxiv2026

Rheb membrane orientation dynamics and functional consequences elucidated by molecular simulations, single-molecule-FRET and signaling assays

Membrane binding and dynamics of prenylated Ras family small GTPases are being increasingly recognized as key regulators of their cellular signaling. However, previous studies have focused primarily on the plasma membrane-associated Ras proteins. The Ras homolog enriched in brain (Rheb) is predominantly found in endo-membranes, such as the endoplasmic reticulum and lysosomes where it directly activates mTORC1 to control cell growth and proliferation. Little is known about the molecular details and functional consequences of Rheb membrane engagement, interaction with lipids, and orientational dynamics. In this work, we combined molecular dynamics simulations, hidden Markov modeling and single-molecule FRET measurements to map the modes of membrane binding and orientation landscape of Rheb. We also assessed the functional consequence of these processes on mTORC1 activation through simulation-guided mutagenesis and cell signaling assays. We found that Rheb interconverts between two dominant orientation states through two intermediates, and that mutationally destabilizing some of these orientation states modulates mTORC1 activity. Together, these results establish membrane orientation dynamics as a functionally relevant phenomenon in the endo-membrane localized Rheb and suggest a broader role of orientational dynamics for the regulation of functional in lipid-modified small GTPases.

biophysics↗

bioRxiv2025

Suppression and modulation mechanism underlying the flavonoid-induced inhibition of fibrillation of α-synuclein

The accumulation of -synuclein (Syn) aggregates is a hallmark of synucleinopathies, including Parkinsons disease (PD). However, the progression from harmless monomeric Syn to misfolded oligomers and fibrillar species remains largely unclear. In this study, we examined the effect of Cyanidin, a naturally occurring neuroprotective compound, on the aggregation properties of -syn using a combination of various biophysical tools. Thioflavin T fluorescence measurements revealed the inhibition of -syn fibrillation in the presence of sub-stoichiometric concentration of Cyanidin, indicating its effect on -syn nucleation. Ultracentrifugation and size exclusion chromatographic analyses demonstrated that increasing concentrations of Cyanidin reduced the conversion of monomers to aggregated forms of -syn. Analysis of the aggregation reaction of -syn based on monomer concentration suggested that Cyanidin reduces the in-vitro conversion of monomers to amyloid nuclei at sub-stoichiometric micromolar concentrations. Cyanidin was observed to bind weakly with the unstructured, monomeric -syn, limiting the nucleation step and modulating the pathway to form conformationally restrained, SDS-resistant -helical higher-order oligomers ([~]670 kDa), which are less-hydrophobic. AFM and TEM images show that Cyanidin also possesses strong fibril disaggregation activity. Furthermore, seeding studies reveal that the higher-order oligomers, which are otherwise stable, destabilise upon sonication and attain seeding capability. Steady-state fluorescence spectroscopy and isothermal titration calorimetry (ITC) reveal weak interactions between Cyanidin and Syn, with a dissociation constant in the mM range. Interestingly, Cyanidin-generated oligomers increase the viability of neuroblastoma (SH-SY5Y) cells, thereby protecting the neuronal cells from degeneration. Our study suggests that stabilization of structured oligomers by small molecule modulators like Cyanidin provides a viable strategy to interfere with Syn fibrillization.

biophysics↗

bioRxiv2024

Assessing the efficacy of therapeutically promising combination of polymyxin B and triclosan against colistin resistant Klebsiella pneumoniae

Antimicrobial resistance (AMR) is one of the greatest public health challenges of the current time, and it is primarily associated with Gram-negative bacterial infections. Among these infections, Klebsiella pneumoniae is the most notorious bacterial pathogen in the context of AMR dissemination. In this dire circumstance, clinicians explored old regimes such as colistin to treat these multidrug-resistant infections; however, unfortunately, the resistance towards these last-resort drugs is also emerging rapidly. In this study, we examined the occurrence of colistin resistance in K. pneumoniae concerning its mechanism of resistance, chronology, and geographical distribution. We found that resistance towards these last-resort drugs, colistin and polymyxin B, is emerging rapidly, with evolving multiple mechanisms of resistance. While efforts continue to develop new antibacterial drugs, a combination of approved antibacterial drugs may be one of the most suitable strategies to fight against AMR in the current time-ticking situation. In this series, we evaluated the efficacy of a therapeutically viable combination of polymyxin B and triclosan using a set of standard antibacterial assays, used for in vitro preclinical efficacy. We found that the combination is highly effective in colistin-resistant clinical strains of K. pneumoniae, as triclosan sensitizes the pathogen towards polymyxin B. Furthermore, the results of additional complementary experiments, such as time-kill kinetics, biofilm inhibition, frequency of resistance, cell viability, and cytotoxicity assays, show very encouraging results for the feasibility and validation of this combination. Finally, we examined the presence of mutations in field isolates of K. pneumoniae FabI enzyme, as it has been hypothesized previously that triclosan may cause mutation in its binding target, FabI. Interestingly, we could not detect any mutation in the triclosan binding site of FabI in the genome of field isolates. Altogether, this study provides an overview of the current situation on colistin resistance, a promising solution for colistin resistance comprising a combination of two already being used antibacterial ingredients based on in vitro efficacy studies.

microbiology↗

bioRxiv2024

Deep brain stimulation induces white matter remodeling and functional changes to brain-wide networks

Deep brain stimulation (DBS) is an emerging therapeutic option for treatment resistant neurological and psychiatric disorders, most notably depression. Despite this, little is known about the anatomical and functional mechanisms that underlie this therapy. Here we targeted stimulation to the white matter adjacent to the subcallosal anterior cingulate cortex (SCC-DBS) in macaques, modeling the location in the brain proven effective for depression. We demonstrate that SCC-DBS has a selective effect on white matter macro- and micro-structure in the cingulum bundle distant to where stimulation was delivered. SCC-DBS also decreased functional connectivity between subcallosal and posterior cingulate cortex, two areas linked by the cingulum bundle and implicated in depression. Our data reveal that white matter remodeling as well as functional effects contribute to DBSs therapeutic efficacy.

neuroscience↗

bioRxiv2023

Epigenetic mediated functional reprogramming of immune cells leads to HBsAg seroconversion in Hepatitis B Virus Reactivation patients

BackgroundHepatitis B virus (HBV) modulates epigenetic landscape by epigenetic regulators. HBsAg seroconversion is possible with immune activation, therefore we aimed to investigate epigenetic modulation in HBV reactivation (rHBV) for viral clearance and seroconversion. MethodsSixteen retrospectively collected rHBV patients [Seroconverters (SC, n=7, HBsAg loss and anti-HBs>10 IU/ml), non- seroconverters (NSC, n=9)], chronic hepatitis B treatment naive (nCHBV, n=7) patients and healthy controls (HC, n=7) were included in this study. Genome methylation, gene expression, plasma-cytokines, and immune cell profiling was analysed by Reduced Representation Bisulfite Sequencing (RRBS), QRT-PCR, multiplex-cytokine-bead array and flow-cytometry. ResultsrHBV patients having high HBV DNA and ALT showed epigenetic remodellers; KDM2B, NCOR2 and GATA6, immune and metabolic genes; TGF-{beta}, IL-6, IRF8, RPTOR, HK3 significantly (p<0.05) hypomethylated at specific CpG islands compared to nCHBV. TOX was hypomethylated in nCHBV suggesting immune-exhaustion. At-baseline, seroconverters showed hypomethylation of KDM2B, COX19, IRF8, TLR5 and hypermethylation of LAG3 compared to non-seroconverters. Further, in seroconverters at week-24, IL17RA, IFN-{gamma}, TGF-{beta}, and STAT5B (p<0.05) were additionally hypomethylated at specific CpG islands suggesting immune activation. Cytokine-bead analysis revealed increased IL-6 (p=0.009) and decreased LAG3 plasma levels (p=0.01) also imply on significantly differentiated HBV specific CD8, Tfh and Th1/17 cells in seroconverters at baseline and week-24. However, both nCHBV and non-seroconverters had consistent hypomethylation of LAG3 and TOX, which leads to immune exhaustion. ConclusionIn rHBV, seroconversion is driven by position specific CpG islands methylation in epigenetic remodellers, immune and metabolic genes. Immune metabolic reprograming is reflected by Th1/17 differentiation, extensive interleukin production for HBsAg seroconversion. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=116 SRC="FIGDIR/small/554133v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@48e206org.highwire.dtl.DTLVardef@ef0fa7org.highwire.dtl.DTLVardef@ecb710org.highwire.dtl.DTLVardef@e6b34e_HPS_FORMAT_FIGEXP M_FIG C_FIG Lay summaryEpigenetic landscape in nCHBV depicts exhaustion and immune dysfunction. Out of many hypermethylated CpG islands of nCHBV, few become hypomethylated in rHBV and drives immune and metabolic reprogramming. This study provides insights into the cellular and molecular basis of epigenomic programs that regulate the differentiation and activation of immune cells leading to viral clearance and seroconversion. Targeting epigenetic mechanism could be promising strategy for the treatment of nCHBV and non-seroconverters.

immunology↗

bioRxiv2023

A broadly protective CHO cell expressed recombinant spike protein subunit based vaccine (IMT-CVAX) against SARS-CoV-2

Protective immunity induced by COVID-19 vaccines is mediated mainly by spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, we report the development of a recombinant prefusion stabilized SARS-CoV-2 spike protein-subunit-based COVID-19 vaccine produced in the mammalian cell line. The gene encoding ectodomain (ECD) of the spike protein was engineered and cloned into Freedom pCHO 1.0, a mammalian expression vector, and subsequently expressed in the Chinese Hamster Ovary suspension cell line (CHO-S). The recombinant S protein ectodomain (hereafter referred to as IMT-CVAX) was purified using a combination of tangential flow filtration and liquid chromatography. Biochemical and biophysical characterization of IMT-CVAX was done to ensure its vital quality attributes. Intramuscular immunization of mice with two doses of adjuvanted IMT-CVAX elicited a strong anti-Spike IgG response. In pseudovirus-based assays, IMT-CVAX- immune mice sera exhibited a broad-spectrum neutralization of several SARS-CoV-2 variants of concern (VoCs). Golden Syrian Hamster immunized with IMT-CVAX provided excellent protection against SARS-CoV-2 infection, and, hamster immune sera neutralized the live SARS-CoV-2 virus. The adjuvanted IMT-CVAX induced robust Tfh-cells response and germinal center (GC) reaction in human ACE2 receptor-expressing transgenic mice. The findings of this study may pave the way for developing next-generation protein subunit-based vaccines to combat the existing SARS-CoV-2 and its emerging VoCs. The IMT-CVAX is produced using a scalable process and can be used for large-scale vaccine production in an industrial setup.

microbiology↗

bioRxiv2023

Differential patterns of automatic segmentation of 3T and 7T MRI: Implications in neuropsychiatric disease

Automatic segmentation was performed on T1-MPRAGE structural MRI data acquired at 3T and 7T from 37 and 69 distinct healthy controls, respectively. Additionally, segmentation was performed on imaging acquired from 215 major depressive disorder (MDD) patients at 3T and 40 MDD patients at 7T. Of 259 segmentation-derived imaging features evaluated, 120 showed significant 3T vs. 7T differences among controls, and 153 among patients. 7T imaging metrics showed consistently lower cortical thickness and cortical gray/white matter ratios. Subcortical and cortical volumes measured at 7T were more mixed, with 7T images showing greater frontal lobe volume, but lower cortical volumes elsewhere.

bioengineering↗

bioRxiv2020

Perivascular Space Semi-Automatic Segmentation (PVSSAS): A Tool for Segmenting, Viewing and Editing Perivascular Spaces

ObjectiveIn this study, we validate and describe a user-friendly tool for PVS tracing that uses a Frangi-based detection algorithm; which will be made freely available to aid in future clinical and research applications. All PVS detected by the semi-automated method had a match with the manual dataset and 94% of the manual PVS had a match within the semi-automated dataset. MethodsWe deployed a Frangi-based filter using a pre-existing Matlab toolbox. The PVSSAS tool pre-processes the images and is optimized for maximum effectiveness in this application. A user-friendly GUI was developed to aid the speed and ease in marking large numbers of PVS across the entire brain at once. ResultsUsing a tolerance of 0.7 cm, 83% of all PVSs detected by the semi-automated method had a match with the manual dataset and 94% of the manual PVS had a match within the semi-automated dataset. As shown in figure 3, there was generally excellent agreement between the manual and semi-automated markings in any given slice. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=80 SRC="FIGDIR/small/385336v1_fig3.gif" ALT="Figure 3"> View larger version (19K): org.highwire.dtl.DTLVardef@35f651org.highwire.dtl.DTLVardef@be6b08org.highwire.dtl.DTLVardef@164ec0aorg.highwire.dtl.DTLVardef@c48643_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 3&4C_FLOATNO Rendered 3-D view of total PVS found across all slices for one patient. C_FIG SignificanceThe primary benefit of PVSSAS will be time saved marking PVS. Clinical MRI use is likely to become more widespread in the diagnosis, treatment, and study of MS and other degenerative neurological conditions in the coming years. Tools like the one presented here will be invaluable in ensuring that the tracing and quantitative analysis of these PVS does not act as a bottle neck to treatment and further research.

neuroscience↗

bioRxiv2020

DIMT1, a regulator of ribosomal biogenesis, controls beta-cell protein synthesis, mitochondrial function and insulin secretion

We previously reported that transcription factor B1 mitochondrial (TFB1M) is involved in the pathogenesis of type 2 diabetes (T2D) owing to mitochondrial dysfunction. Here, we describe that dimethyladenosine transferase 1 homolog (DIMT1), a homologue of TFB1M, is expressed and active in pancreatic {beta}-cells. Like TFB1M, it has been implicated in control of ribosomal RNA (rRNA) but its role in {beta}-cells or T2D remains to be identified. Silencing of DIMT1 impacted mitochondrial function, leading to reduced expression of mitochondrial OXPHOS proteins, reduced oxygen consumption rate (OCR), dissipated mitochondrial membrane potential ({Delta}{Psi}m) and caused a lower rate of ATP production (mATP). In addition, DIMT1 knockdown slowed the rate of protein synthesis. In accordance with these findings, DIMT1-deficiency perturbed insulin secretion in rodent and human {beta}-cell lines. These effects are likely a result of destabilization of ribosomal assembly, involving NIN1 (RPN12) binding protein 1 homolog (NOB-1) and Pescadillo ribosomal biogenesis factor 1 (PES-1). These are two critical ribosomal subunits proteins, whose interactions were perturbed upon DIMT1-deficiency, thereby disturbing protein synthesis in {beta}-cells. Thus, we have here highlighted a role of DIMT1 in ribosomal biogenesis that perturbs protein synthesis, resulting in mitochondrial dysfunction and disrupted insulin secretion, both being potential pathogenetic factors in T2D.

molecular biology↗

bioRxiv2018

Ultra-high field MRI reveals mood-related circuit disturbances in depression: A systematic comparison between 3-Tesla and 7-Tesla

Ultra-high field 7-Tesla (7T) MRI has the potential to advance our understanding of neuropsychiatric disorders, including major depressive disorder (MDD). To date, few studies have quantified the advantage of resting state functional MRI (fMRI) at 7T compared to 3-Tesla (3T). We conducted a series of experiments that demonstrate the improvement in temporal signal-to-noise ratio (TSNR) of a multi-echo fMRI protocol with ultra-high field 7T, compared to 3T MRI in healthy controls (HC). We also directly tested the enhancement in ultra-high field 7T fMRI signal power by examining the ventral tegmental area (VTA), a small midbrain structure that is critical to the expected neuropathology of MDD but difficult to discern with standard 3T MRI. We demonstrate 200-300% improvement in TSNR and resting state functional connectivity coefficients provided by ultra-high field 7T fMRI compared to 3T, indicating enhanced power for detection of functional neural architecture. A multi-echo based acquisition protocol and signal denoising pipeline afforded greater gain in signal power at ultra-high field compared to classic acquisition and denoising pipelines. Furthermore, ultra-high field fMRI revealed mood-related neuro-circuit disturbances in patients with MDD compared to HC, which were not detectable with 3T fMRI. Ultra-high field 7T fMRI may provide an effective tool for studying functional neural architecture relevant to MDD and other neuropsychiatric disorders.

neuroscience↗