Biology subjects

Jena, B. P.

Publications and source records attributed to Jena, B. P..

11 recordsLinked to original sources

bioRxiv2025

Porosome reconstitution reverses Alzheimers in human brain organoids

A central challenge in overcoming diseases resulting from defects in the cellular nanoscale multiprotein porosome complexes, is the consequent alteration of other proteins within the complex. By exploiting the distinct porosomes present in different tissues, Porosome Therapeutics, Inc., has developed a clinically relevant approach to incorporate (reconstitute) functional wild type porosomes into the plasma membrane of diseased cells to overcome secretory defects. In a combinatorial strategy of reprogramming the neuronal secretory and metabolic components, we established a dual-target therapeutic framework that repairs synaptic and metabolic defects to counteract neurodegeneration in Alzheimers. Utilizing iPSC derived human brain organoids, as recommended by the NIH and FDA, we demonstrate both the morphological and functional reversal of Alzheimers following porosome-apigenin treatment. Using the UHD-CMOS-MEA System developed jointly by Sony Semiconductor Solutions (Sony), SCREEN Holdings (SCREEN) and VitroVo, we were able to monitor thousands of neuronal firing in the brain organoids, helping in assessing the restoration of neuronal function in Alzheimers following porosome-apigenin therapy.

neuroscience↗

bioRxiv2025

Mechanism of clinically relevant neurosecretory and metabolic impairment in Alzheimers and identification of engineered nanotherapy

A central challenge in Alzheimers disease is understanding the mechanism of neuronal secretory dysfunction. By exploiting AI, we reveal how beta-amyloid disrupts key protein interactions within the neuronal secretory machinery. In a combinatorial strategy of reprogramming the neuronal secretory and metabolic components, we established a dual-target therapeutic framework that repairs synaptic and mitochondrial defects to counteract neurodegeneration in Alzheimers.

neuroscience↗

bioRxiv2024

Reprogramming the neuronal secretory and metabolic machinery using correctors for Alzheimer therapy

Alzheimer disease (AD) is a neurodegenerative disorder characterized by memory loss and personality changes, leading to dementia. The primary cause of the cognitive decline that characterizes AD is the loss of neurons, hypothesized as being due to secretory defects in neurotransmitter release and synaptic plasticity. There is growing evidence that in Alzheimer there is impaired neuronal metabolism due to an increase in free radicals and mitochondrial fission, leading to loss in ATP synthesis required for neurotransmission. Therefore, impaired metabolism in neurons may lead to the observed defects in neurotransmitter release, resulting in the loss of neuronal function and connections in regions of the brain involved in memory and reasoning. In our earlier studies, peptide inhibitor targeting oligomeric ATAD3A, a protein essential for mitochondrial fission and bioenergetics, were found to partially rescue from AD, both in AD neurons and AD mice. In the current study in AD neurons, neuronal porosome reconstitution combined with either a linear or circular peptide inhibitor of ATAD3A oligomers, was able to restore AD neurons to near normal levels of viability and greatly reduce oxidative stress in mitochondria. Additionally, the peptide and a small molecule considered safe by the FDA and used as a food additive, were also able to greatly increase viability in AD neurons and provide reduction to near normal levels of mitochondrial oxidative stress. Collectively, these results demonstrate great promise as secretory and metabolic correctors for AD therapy.

neuroscience↗

bioRxiv2024

Porosome reconstitution therapy: A biologic rescue from cystic fibrosis

Cystic fibrosis (CF) is a genetic disorder resulting from mutations in the CF Transmembrane Conductance Regulator (CFTR) gene that codes for a chloride transporting channel at the cell plasma membrane. In CF, highly viscous mucus is secreted in the airways preventing its clearance, leading to lung infections and respiratory failure. A major challenge in treating CF patients has been the presence of more than 2,000 different CFTR mutations or due to the absence of CFTR expression. CFTR is among the 34 major proteins composing the 100 nm porosome secretory machinery in the human airway epithelia, involved in mucin secretion. The airways is coated with a thin film of mucus, composed primarily of mucin MUC5AC and MUC5B. Sputum from patients with CF show a >70% decrease in MUC5B and MUC5AC secretion. Our studies using differentiated 3D cultures of human airway epithelial cell line, also demonstrate loss of both chloride and mucus secretion following exposure to CFTR inhibitors thiazolidinone 172 or the hydrazide GlyH101. Our studies show that human bronchial epithelial (HBE) cells with {Delta}F508 CFTR mutation, affects nearly a dozen porosome proteins including CFTR. Therefore, we hypothesized that the introduction of normal functional porosomes carrying wild type CFTR into the cell plasma membrane of CF cells would rescue from all forms of CF. Air liquid interface (ALI) 3D differentiated HBE WT-CFTR cells and {Delta}F508-CFTR CF HBE cell cultures mimicking normal lung physiology, responding to CFTR inhibitors and CF corrector and modulator drugs Tezacaftor, Ivacaftor and TRIKAFTA, was used in the study. Introduction of functional porosome complexes obtained from WT-CFTR HBE cells into the plasma membrane (PM) of {Delta}F508-CFTR CF cells, was demonstrated by an increase in PM-associated CFTR using Magnify expansion microscopy. Mucin secretion assays demonstrate porosome reconstitution to restore mucin secretion more than twice as effectively as TRIKAFTA. These results are further supported by preliminary nasal potential different studies in {Delta}F508 mice, where treatment of the nasal passage with porosome isolates from WT-CFTR HBE cells, restore chloride secretion in the nasal passage of mice, a further validation of the highly effective porosome reconstitution therapy for CF.

cell biology↗

bioRxiv2023

Reprogramming the cells secretory machinery: A cystic fibrosis rescue

Cystic fibrosis (CF) is a genetic disorder resulting in the secretion of highly viscous mucus in the airways, leading to lung infection, respiratory failure, morbidity and mortality. CF is attributed to mutation in the CF Transmembrane Conductance Regulator (CFTR) gene that codes for a chloride transporting channel at the cell plasma membrane. More than 2,000 mutations in the CF gene have been identified; however, the {Delta}F508 CFTR is the most common, accounting for approximately 70% of all CFTR mutations. Our earlier studies demonstrate the CFTR protein to be among the nearly 30 proteins constituting the porosome secretory machinery at the plasma membrane in human airway epithelial mucous-secreting cells. Additionally, our recent studies show that stimulated human airway epithelial cells pre-exposed to CFTR inhibitors result in loss of mucus secretion, suggesting the involvement of CFTR in porosome-mediated mucus secretion. To further test the hypothesis that CFTR is involved in porosome-mediated mucus secretion in the human airways, and to develop a therapeutic approach to overcome this defect in {Delta}F508 CFTR human bronchial epithelial cells, the current study was undertaken. Mass spectrometry and Western Blot analysis of porosomes isolated from WT-CFTR Human Bronchial Epithelial (HBE) Cells and {Delta}F508-CFTR CF HBE cells, demonstrate a varying loss or gain of several porosome proteins, including a decrease in the t-SNARE protein SNAP-23 and undetectable levels of the Ras GTPase activating like-protein IQGAP1 in the {Delta}F508-CFTR CF cells. This suggested that mutation in porosome-associated CFTR protein additionally affects other proteins within the porosome secretory machinery, negatively impacting mucus secretion. Hence, to ameliorate defects in mucus secretion in CF, the reconstitution of functional porosomes obtained from WT-CFTR HBE cells into the plasma membrane of {Delta}F508-CFTR mutant cells was performed. Results from the study demonstrate that porosome reconstitution rescues mucus secretion approximately four-fold more effectively than the currently available CF drugs Tezacaftor and Ivacaftor.

cell biology↗

bioRxiv2022

Possible Mechanism of Porosome-Mediated Exosome Release

Supramolecular cup-shaped lipoprotein structures called porosomes embedded in the cell plasma membrane mediate fractional release of intra-vesicular contents from cells during secretion. The presence of porosomes, have been documented in many cell types including neurons, acinar cells of the exocrine pancreas, growth hormone secreting cells of the pituitary, and mouse insulin-secreting beta cell insulinomas called Min6. Electron micrographs of the nerve terminal in rat brain and of Min6 cells, suggest that besides the docking, fusion and content release of neurotransmitter containing synaptic vesicles and insulin containing granules, multi-vesicular bodies and exosome-like vesicles are released at the porosome complex. To test this hypothesis, the porosome-associated calcium transporting ATPase 1 [SPCA1] encoded by the ATP2C1 gene, was knocked-out in Min6 cells using CRISPR, to study glucose-stimulated insulin and exosome release. In agreement with electron micrographs, results from the study demonstrate a loss of both glucose-stimulated insulin and exosome release in the ATP2C1 knockout Min6 cells. These results further confirm the role of the porosome complex in insulin secretion and establishes a new paradigm in porosome-mediated exosome release in beta cells of the endocrine pancreas.

cell biology↗

bioRxiv2020

Skeletal Muscle Remodeling in Immobilized Patients: Determined Using a Parameter Estimation Histomorphometric Approach

Skeletal muscle biopsy commonly used for light microscopic, electron microscopic and biochemical and transcriptional evaluation remains the gold standard for establishing the etiology of a myopathy. While most myopathies exhibit one or more phenotypes, early stages or several metabolic myopathies often exhibit normal muscle morphology, making diagnosis difficult. In such cases where standard staining techniques fail to offer definitive diagnostic information, a combination of expensive and time-consuming electron microscopy and biochemical testing is required to provide definitive diagnosis. As a step toward overcoming these limitations in diagnostic pathology of skeletal muscle tissue, here we report the application of parameter estimation machine learning approaches on immunofluorescent images of human skeletal muscle tissue acquired using fluorescent microscopy. The machine learning morphometric approach enables the recognition of fine cellular changes in skeletal muscle tissue, allowing determination of skeletal muscle remodeling as a consequence of immobilization.

cell biology↗

bioRxiv2020

Cystic fibrosis transmembrane conductance regulator (CFTR) inhibition results in mucus accumulation in human airway epithelia Calu-3 cells: Experimental and Machine Learning Studies

Porosomes are cup-shaped lipoprotein structures at the cell plasma membrane involved in fractional release of intra-vesicular contents during secretion. At the base of the porosome facing the cell cytoplasm, secretory vesicles dock, fuse and swell, to release intra-vesicular content during secretion. Earlier studies demonstrate the cystic fibrosis trans-membrane conductance regulator (CFTR) associated with the porosome in human airways epithelial Calu-3 mucous-secreting cells, suggesting its possible involvement in porosome-mediated mucus secretion. The current study was undertaken to test this hypothesis. Electron microscopy followed by morphometric analysis using manual and computational machine learning approaches were used to assess changes in secretory vesicle size and content, following stimulation of secretion in the absence and presence of CFTR inhibitors. Results from the study demonstrate that stimulated Calu-3 cells pre-exposed to CFTR inhibitors, demonstrate attenuation of secretory vesicle swelling and the release of mucin. Consequently, accumulation of intracellular mucin is observed in cells exposed to CFTR inhibitors. These results further suggest that mucin secretion from Calu-3 cells involve CFTR both at the secretory vesicle membrane to regulate vesicle volume and hydration, and at the porosome to facilitate mucin hydration and secretion. These new findings progress our understanding of the involvement of CFTR on mucus hydration and secretion, providing critical insights into the etiology of CF disease.

cell biology↗

bioRxiv2019

Deep Learning Strategies for Differential Expansion Microscopy

Differential expansion microscopy (DiExM) achieves greater than 500-fold volumetric expansion of biological specimens without loss of cellular antigens. The anisotropic character of this expansion, which affects tissues, cells, organelles and even sub-organelle features, requires the application of novel machine learning approaches to extract accurate and meaningful morphological and biochemical information from DiExM images. Here we describe current strategies to achieve this goal.

cell biology↗

bioRxiv2019

vH+-ATPase-induced intracellular acidification is critical to glucose-stimulated insulin secretion

Swelling of secretory vesicles is critical for the regulated expulsion of intra-vesicular contents from cells during secretion. At the secretory vesicle membrane of the exocrine pancreas and neurons, GTP-binding G proteins, vH+-ATPase, potassium channels and AQP water channels, are among the players implicated in vesicle volume regulation. Here we report in insulin secreting MIN6 cells, the requirement of vH+-ATPase-mediated intracellular acidification, on glucose-stimulated insulin release. MIN6 cells exposed to the vH+-ATPase inhibitor Bafilomycin A show decreased acidification of the cytosolic compartment that include insulin-carrying granules. Additionally, a loss of insulin granule association with the cell plasma membrane is demonstrated and results in a decrease in glucose-stimulated insulin secretion and accumulation of intracellular insulin. These results suggest that vH+-ATPase-mediated intracellular acidification is required both at the level of secretory vesicles and the cell plasma membrane for cell secretion.

cell biology↗

bioRxiv2019

Differential expansion microscopy

Expansion microscopy (ExM) involves the use of hydration-competent polymers to physically expand biological specimens approximately 4-fold linear increase to achieve 70 nanometer resolution using an ordinary diffraction limited optical microscope. Optimal conditions however for antigen retention during the expansion process and the relative expansion between organelles within cells has remained unclear. It is reported that different tissues expand to different extents, suggesting that although isotropic expansion is believed to occur, different subcellular compartments with different composition would undergo anisotropic or differential expansion (DiEx). Consequently, there would be distortion of the native shape and size of subcellular compartments upon expansion, parameters which are critical in assessing cellular states in health and disease. Here we report optimal fixation and expansion conditions that retain structural integrity of cells while exhibiting up to 8-fold linear and therefore 512-fold volumetric expansion. Anisotropic expansion is observed not just between tissues, but between different subcellular compartments and even within subcellular compartments. Combining image analysis and machine learning, we provide an approach for the rapid and precise measurement of cellular and subcellular structures in expanded tissue. Using both manual and computation assessment of morphometric parameters, we demonstrate expansion to be anisotropic and name this method differential expansion microscopy (DiExM).

cell biology↗