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Singla, S.

Publications and source records attributed to Singla, S..

8 recordsLinked to original sources

From Scalp to Source: Precise Phase Retrieval of Intracerebral Epileptic Sources Based on Surface EEG

Accurate phase tracking of deep-brain activity is critical for effective closed-loop and phase-locked neuromodulation therapies. However, direct access to deep neural phase through intracranial recordings remains clinically restrictive due to the invasiveness. Here we validate and clinically benchmark the Gabor-Nelson (GN) dipole estimation method for reconstructing deep-brain oscillatory phase from non-invasive scalp EEG. GN is a geometry-based, imaging-independent approach that offers computationally efficient dipole reconstruction and has rarely been applied to source-level phase estimation in human neuroscience. We compared GN with an established MRI-informed Inverse Solution (IS) method using a three-stage reconstruction pipeline consisting of dipole modeling, dimensionality reduction, and frequency-dependent phase-delay correction. Validation is performed using (i) cadaveric recordings, where known ground-truth seizure waveforms were replayed through implanted deep electrodes, and (ii) simultaneous scalp EEG and SEEG recordings in human patients, where pseudo-ground truth was approximated via the intracranial contacts. GN achieved phase accuracy and signal fidelity comparable to IS across both datasets despite requiring no anatomical imaging. In cadaver recordings, phase-corrected reconstruction correlations exceeded r > 0.91 and {Delta}{Phi} < 9{degrees} in mean phase error. In patient SEEG data, GN reached up to r {approx} 0.80 with phase offsets suitable for neuromodulatory timing. GN offers a viable, low-barrier, imaging-independent alternative to traditional inverse modeling for non-invasive seizure phase tracking. This framework opens pathways for scalable, phase-locked and closed-loop stimulation therapies in epilepsy and potentially other network-based brain disorders.

neuroscience↗

Developing in vitro patient derived CERvical Cancer OrganoidS (CERCOS) as a potential preclinical tool in cervical cancer research

Advanced cervical cancer remains a major cause of mortality in women worldwide as it has limited treatment options and recurrence is very common. This highlights the necessity to develop patient derived organoids (PDOs) as preclinical models, that can recapitulate the clinical heterogeneity of the cancer in terms of molecular features, and genetic background. The PDOs have potential for guiding personalized treatment in clinical practice. In this study, we have established patient-derived cervical cancer organoids or CERvical Cancer OrganoidS (CERCOS) from biopsy samples of five patients having two different subtypes (squamous cell carcinoma and adenocarcinoma) using a modified protocol. The CERCOS were characterized to assess their genetic and phenotypic similarity with the parent tissue. The organoids developed in vitro showed preservation of the histopathological and somatic mutational profile of the parent tissue. Expression of cervical cancer-related genes, including PIK3CA, MET, and LRP1B, was found to be comparable between the organoids and the parent tissue. Moreover, characterization of the PDOs post cryopreservation showed the maintenance of the morphological and histopathological features of the parent tissue. This study demonstrates that the CERvical Cancer OrganoidS (CERCOS), established using the current protocol, preserves genetic and phenotypic similarity and can serve as a platform for precision medicine and biobanking purposes.

cancer biology↗

An envelope stress response governs long-chain fatty acid metabolism via a small RNA to maintain redox homeostasis in Escherichia coli

Long-chain fatty acids (LCFAs) are a tremendous source of energy for several bacteria but are complex to use because they induce redox stress. We previously showed that LCFA degradation impedes oxidative protein folding in the Escherichia coli envelope, an issue that arises due to the insufficiency of ubiquinone, a lipid-soluble electron carrier in the electron transport chain (ETC). To maintain redox homeostasis, E. coli activates the CpxAR two-component system; however, the nature of feedback imparted by this envelope stress response (ESR) remained unknown. Here, we show that contrary to the well-recognized remedial mode of Cpx restoring envelope integrity by upregulating protein quality control factors, in LCFA-grown cells, it uses a preventive measure to maintain homeostasis. Cpx increases ubiquinone availability for oxidative protein folding by suppressing LCFA metabolism and directly increasing ubiquinone levels. Further, rather than using its conventional mode of imparting regulation via CpxR working as a transcriptional regulator, during LCFA metabolism, Cpx mainly uses its non-coding arm to counteract envelope redox stress. The Cpx-regulated small RNA CpxQ represses fad genes involved in LCFA transport and {beta}-oxidation, downregulates components of glyoxylate shunt, gluconeogenesis, and ETC, and increases ubiquinone content. Corroborating with its role in repressing LCFA metabolism and maintaining redox homeostasis, CpxQ overexpression impairs growth of E. coli in LCFAs and CpxQ deletion renders LCFA-grown E. coli hypersensitive to a thiol agent. Our foremost work studying the interconnection between LCFA metabolism, redox stress, and ESR in E. coli provides a rationale for investigating similar networks in other LCFA-utilizing bacteria. Significance StatementLong-chain fatty acids (LCFAs) are energy-rich nutrients for Escherichia coli; however, their utilization hampers disulfide bond (DSB) formation in secreted proteins, an essential process that occurs in the envelope compartment. Here, we show that an envelope stress response manipulates LCFA metabolism in E. coli and uses a small RNA (sRNA) to restore homeostasis. Several bacteria with a huge impact on human health use host-derived LCFAs during infection. Because many virulence factors require DSB formation, the present study offers a basis to examine whether sRNAs play a role in governing envelope redox balance during LCFA metabolism in pathogens. The sRNA-mediated control is likely an ideal strategy both for rapid response to and quick recovery from LCFA-induced stress.

microbiology↗

Malaria parasite HOPS/CORVET complexes are critical for endocytosis and invasion organelles function

The tethering complexes HOPS/CORVET are central for vesicular fusion through the eukaryotic endolysosomal system, but the functions of these complexes in the intracellular development of malaria parasites are unknown. Here we show that early inactivation of core HOPS/CORVET complex subunits in Plasmodium falciparum leads to developmental arrest and accumulation of cytosolic vesicles, indicating a role of HOPS/CORVET in parasite endocytosis and fusion of endosomes to the digestive vacuole membrane. Late inactivation of the core HOPS/CORVET subunits led to the mislocalization of luminal rhoptry and microneme proteins, and to a severe defect in merozoite invasion. Ultra-expansion microscopy revealed a reduced rhoptry volume and the accumulation of numerous vesicles, further supporting a role of HOPS/CORVET in protein trafficking to the apical organelles. Malaria parasites have therefore repurposed HOPS/CORVET to perform dual functions consistent with a canonical endocytosis pathway for delivery of host cell material to the digestive vacuole in trophozoite stages and a parasite specific role in trafficking of protein cargo to the apical organelles required for invasion in schizont stages.

cell biology↗

Bibliometric and Temporal Trend Analysis of Nipah Virus- An Emerging Zoonotic Disease: What Do We Know So Far

BackgroundNipah virus (Genus Henipavirus) was first detected in 1999, during the Malaysia-Singapore outbreak, and is an emerging bat-borne pathogen. It causes rare but fatal disease, with a 40-75% fatality rate, and clinically ranging from asymptomatic infection to fatal encephalitis. MethodsPublicly available literature, including preclinical and clinical studies, have been retrieved from PubMed, Scopus, and Web of Science. Bibliometric analysis was done using embedded tools of these search engines along with VOSviewer. Results601 documents from PubMed, 955 from Web of Science, and 784 from Scopus were analyzed, and we found that maximum contributions are from the USA, India, Malaysia, Australia, and Bangladesh, and in the past five years, there has been an exponential surge in the publication rate. More research and high-throughput screening are needed to explore drugs against critical targets and receptors like ephrin-B2, non-structural protein C, F protein, L protein, G glycoprotein, nucleocapsid protein, V protein, P protein, and W protein. Research for possible vaccination is underway, but the rate is not significant. Clinical studies in this direction are minimal, undermining the fatality of this lethal disease and possible outbreak. ConclusionTo our knowledge, this is the first bibliometric analysis of Nipah virus-related documents. It urgently demands expedited research in this direction before it is too late. Author SummaryNipah virus, a rare but deadly bat-borne pathogen, has gained increased attention in recent years. Our pioneering bibliometric analysis reveals a surge in research interest. This study underscores the pressing need for accelerated research on drugs and vaccines to combat this lethal disease and prevent potential outbreaks. Urgency is paramount.

microbiology↗

Endothelial Knockdown of the Tumor Suppressor, WWOX, Increases Inflammation in Ventilator-Induced Lung Injury

BackgroundChronic cigarette smoke exposure downregulates lung expression of WWOX, an ARDS relevant tumor suppressor. Prior work has revealed a barrier protective function of WWOX during infectious models of ARDS. Proteomic analysis of WWOX-silenced lung endothelial cells suggest involvement of WWOX in protection against mechanical stretch-induced inflammation. MethodsProtein lysates from WWOX-silenced endothelial cells (ECs) were analyzed using tandem mass tag mass spectrometry (TMT-MS) to determine the differential expression status of the proteome compared to wild type ECs. WWOX-silenced ECs as well as those isolated from endothelial Wwox knockout (EC Wwox KO) mice were subjected to cyclic stretch (18% elongation, 0.5 Hz, 4 hours). Cellular lysates and media supernatant were harvested for assays of cellular signaling, protein expression, and cytokine release. Dual silencing of WWOX and zyxin was achieved to determine the role of zyxin upregulation in IL-8 production following mechanical stretch and during WWOX knockdown. Control and EC Wwox KO mice were subjected to high tidal volume ventilation (VILI, 40ml/kg, 65 breath/min, 4hours). Bronchoalveolar lavage fluid and mouse lung tissue were harvested for cellular signaling, cytokine secretion, and histologic assays. ResultsTMT-MS revealed upregulation of zyxin expression during WWOX knockdown which predicted a heightened inflammatory response to mechanical stretch. WWOX-silenced ECs and ECs isolated from EC Wwox mice displayed significantly increased cyclic stretch-mediated secretion of various cytokines (IL-6, KC/IL-8, IL-1{beta}, and MCP-1) relative to controls. This was associated with increased ERK and JNK phosphorylation but decreased p38 MAPK phosphorylation. EC Wwox KO mice subjected to VILI sustained a greater degree of injury than corresponding controls. Silencing of zyxin during WWOX knockdown abrogated stretch-induced increases in IL-8 secretion. ConclusionLoss of WWOX function in ECs is associated with a heightened inflammatory response during mechanical stretch that is associated with increased MAPK phosphorylation and appears to be dependent on upregulation of zyxin.

molecular biology↗

A novel mouse model of focal limbic seizures that reproduces behavioral impairment associated with cortical slow wave activity

Patients with focal temporal lobe seizures often experience loss of consciousness. In humans, this loss of consciousness has been shown to be positively correlated with EEG neocortical slow waves, similar to those seen in non-REM sleep. Previous work in rat models of temporal lobe seizures suggests that decreased activity of subcortical arousal systems cause depressed cortical function during seizures. However, these studies were performed under light anesthesia, making it impossible to correlate behavior, and therefore consciousness, to electrophysiologic data. Further, the genetic and molecular toolkits allowing for precise study of the underlying neural circuitry are much more developed in mice than in rats. Here, we describe an awake-behaving, head-fixed mouse model of temporal lobe seizures with both spared and impaired behavior reflecting level of consciousness. Water-restricted mice were head-fixed on a running wheel and trained to associate an auditory stimulus to the delivery of a drop of water from a dispenser. To investigate the effect of seizures on behavior, seizures were electrically induced by stimulating either the left or right hippocampus via a chronically-implanted electrode, while mice were performing the task. Behavior was measured by monitoring lick responses to the auditory stimulus and running speed on the wheel. Further, local field potentials (LFP) signals were simultaneously recorded from hippocampus and orbitofrontal cortex (OFC). Induced focal seizures were 5-30s in duration, and repeatable for several weeks (n=20 animals). Behavioral responses showed a decrease in lick rate to auditory stimulus, and decreased running speed during seizures (p<0.01, n=20 animals). Interestingly, licking response to sound could vary from being impaired to normal during seizures. We found that behavioral impairment is correlated with large amplitude cortical slow-wave activity in frontal cortex, as seen in patients with temporal lobe seizures. These results suggest that induced focal limbic seizures in the mouse can impair consciousness and that the impaired consciousness is correlated with depressed cortical function resembling slow wave sleep. This novel mouse model has similar characteristics with previously studied rat models and human temporal lobe seizures. By leveraging the genetic and molecular techniques available in the mouse, this model can be used to further uncover fundamental mechanisms for loss of consciousness in focal seizures.

neuroscience↗

Cigarette Smoke and Nicotine-Containing E-cigarette Vapor Downregulate Lung WWOX Expression Which is Associated with Increased Severity of Murine ARDS

RATIONALEA history of chronic cigarette smoking is known to increase risk for ARDS, but the corresponding risks associated with chronic e-cigarette use are largely unknown. The chromosomal fragile site gene, WWOX, is highly susceptible to genotoxic stress from environmental exposures, and thus an interesting candidate gene for the study of exposure-related lung disease. METHODS AND RESULTSLungs harvested from current versus former/never smokers exhibited a 47% decrease in WWOX mRNA levels. Exposure to nicotine-containing e-cigarette vapor resulted in an average 57% decrease in WWOX mRNA levels relative to vehicle treated controls. In separate studies, endothelial (EC)-specific WWOX KO versus wild type mice were examined under ARDS-producing conditions. EC WWOX KO mice exhibited significantly greater levels of vascular leak and histologic lung injury. ECs were isolated from digested lungs of untreated EC WWOX KO mice using sorting by flow cytometry for CD31+CD45- cells. These were grown in culture, confirmed to be WWOX-deficient by RT-PCR and Western blotting, and analyzed by electric cell impedance sensing (ECIS) as well as a FITC dextran transwell assay for their barrier properties during MRSA or LPS exposure. WWOX KO ECs demonstrated significantly greater declines in barrier function relative to wild type cells during either MRSA or LPS treatment as measured by both ECIS and the transwell assay. CONCLUSIONThe increased risk for ARDS observed in chronic smokers may be mechanistically linked, at least in part, to lung WWOX downregulation, and this phenomenon may also manifest in the near future in chronic users of e-cigarettes.

molecular biology↗