Biology subjects

Rao, J. S.

Publications and source records attributed to Rao, J. S..

5 recordsLinked to original sources

bioRxiv2026

Functional recovery of vitrified and nanowarmed human-scale pig kidneys

Organ transplantation is critically constrained by the short preservation window for donor organs, limiting allocation and contributing to organ discard and waitlist mortality. Cryogenic organ banking could extend preservation from hours to months or years, but ice formation, thermal stress and cryoprotectant toxicity become increasingly difficult to control as organ size increases. Vitrification and nanowarming have enabled long-term preservation and transplantation of rodent kidneys but have not been demonstrated at the scale of clinically relevant organs. Here we show that vitrification and electromagnetic nanowarming enable long-term cryogenic storage and functional recovery of human-scale porcine kidneys. Kidneys 100-fold larger than previously nanowarmed kidneys were successfully recovered after storage at -150 {degrees}C for up to one year. A new cryoprotectant formulation and multithermic perfusion strategy enabled ice-free vitrification at cooling rates achievable in large kidneys, while controlled cooling with annealing and staged volumetric nanowarming limited thermal stress and enabled rapid rewarming without cracking. Rewarmed kidneys demonstrated vascular, metabolic and excretory function during normothermic perfusion, with limited histological injury, and reperfused homogeneously and produced urine immediately after transplantation. Functional recovery approached static cold storage outcomes across several measures and was similar to that of kidneys exposed to cryoprotectant alone, suggesting minimal additional functional injury from vitrification, cryogenic storage and nanowarming. These findings overcome major scale and storage-duration barriers to organ cryopreservation and establish a path toward organ banking on timescales far beyond those of conventional preservation.

bioengineering↗

bioRxiv2026

MS4A8B regulates Orai1-dependent Ca2+ influx to control motile cilia function in human nasal epithelial cells

Motile cilia demonstrate coordinated beating to propel fluids across epithelial tissues, and changes to their beating frequency are largely regulated by intracellular second messengers including Ca2+. In the airway epithelium, ciliary beating is essential to mucociliary clearance. Mucociliary clearance involves trapping inhaled pathogens and irritants in sticky mucus lining the airways for motile cilia to sweep away contaminated mucus, preventing infection and reducing general airway inflammation. Many chronic respiratory diseases, including chronic rhinosinusitis and asthma, are characterized by an acquired ciliary dysfunction. Despite the importance of Ca2+ signaling in cilia physiology, the identity and molecular mechanisms governing localized ciliary Ca2+ transport remain poorly understood. MS4A8B is an uncharacterized cilia-localized transmembrane protein. Other MS4A homologs have been indirectly linked to Ca2+ signaling via uncharacterized mechanisms. Using primary human nasal epithelial cells differentiated at air-liquid interface, we demonstrated that MS4A8B regulates motile cilia function. MS4A8B knockdown impairs ciliary beating and impacts cilia structure. Live-cell imaging combined with genetic analysis revealed that MS4A8B potentiates Orai1-mediated Ca2+ influx. Co-immunoprecipitation and FRET microscopy in ectopic expression systems demonstrated that MS4A8B interacts with Orai1 channels. Orai1 was further identified to reside in motile cilia of primary human nasal epithelial cells, allowing ciliary beat frequency to be stimulated by Orai1 agonists including arachidonic acid. MS4A8B functional coupling with Orai1 acts as an autonomous cilia signaling network. Targeting this compartmentalized signaling pathway offers a novel therapeutic approach to restore or enhance mucociliary clearance in airway diseases.

physiology↗

bioRxiv2026

Clinical grade cryopreservation unlocks transplant ready human pancreatic and stem cell derived islets for diabetes therapy

Pancreatic islet transplantation can restore endogenous insulin production and offers a potential cure for diabetes, but its clinical impact has been limited by the inability to preserve large numbers of functional islets for timely use. Cryopreservation could provide an on-demand supply, yet conventional methods cause ice formation, cell injury, and loss of insulin secretion. We developed CryoMesh, a vitrification platform that combines a low-toxicity cryoprotectant with a thermally conductive, biocompatible mesh to enable ultra-rapid cooling and rewarming for ice-free cryopreservation. This approach supports long-term, clinical-scale preservation of both human pancreatic and stem cell-derived islets while maintaining viability, architecture, mitochondrial integrity, and glucose-responsive insulin secretion. Human islets preserved for up to one year restored normoglycemia in diabetic mice, with complete recovery of function and no increase in immunogenicity or loss of potency. The combination of high viability, recovery, and functional preservation at clinical scale has not been achieved previously. By decoupling islet isolation and manufacture from transplantation, CryoMesh enables extended quality, potency, and safety testing, cost-effective batch production, and global banking and distribution. These capabilities remove a major barrier to curative cell therapy for diabetes and establish a generalizable strategy for preserving complex multicellular therapeutics. One Sentence SummaryA clinical scale vitrification platform enables long-term banking of human pancreatic and stem cell-derived islets for transplantation.

cell biology↗

bioRxiv2024

Vitrification and rapid rewarming of precision-cut liver slices for pharmacological and biomedical research

Background and AimsHigh-throughput in vitro pharmacological toxicity testing is essential for drug discovery. Precision-cut liver slices (PCLS) provide a robust system for screening that is more representative of the complex 3D structure of the whole liver than isolated hepatocytes. However, PCLS are not available as off-the-shelf products, significantly limiting their translational potential. Cryopreservation could solve this bottleneck by effectively preserving PCLS indefinitely until their time of use. Conventional cryopreservation (slow cooling in DMSO-forming ice) results in poor PCLS viability and function and, therefore, has proven unsuitable. Here, we explore an "ice-free" cryopreservation approach called vitrification and focus on culturing and assessing PCLS for 3 days post-vitrification and rewarming, given that most acute drug toxicity tests are conducted over 24h. MethodsRat liver slices were diffusively loaded with a cryoprotective agent (CPA) cocktail consisting of EG and Sucrose. The CPA-loaded PCLS were placed on a polymer cryomesh, vitrified in liquid nitrogen (LN2), and rapidly rewarmed in CPA. The vitrified and rewarmed PCLS were subsequently cultured in a controlled volume of serum-free, chemically defined media for 3 days. ResultsThe cryopreserved PCLS maintained high viability, morphology, function, enzymatic activity, and drug toxicity response. Results show that the vitrified PCLS perform comparably to untreated controls and significantly outperform conventionally cryopreserved PCLS in all assessments (p < 0.05). ConclusionsRapid vitrification and rewarming of PCLS using cryomesh enabled successful preservation and culture. This approach maintained high viability, function, enzymatic activity, and drug response for 3 days in culture, similar to controls. Impact and ImplicationsThe implications of using vitrification to store PCLS are extensive. This technology provides an exciting opportunity for the development of an "off-the-shelf" cold supply chain of human PCLS from organs declined for transplant, which are sliced, cryopreserved, and stored in a repository and available for on-demand shipping for industrial and academic biomedical research. This would also allow PCLS to become a scalable, reproducible, wide-ranging, and population-representative source of tissue that can accurately mimic in-vivo conditions of the human liver. These transformative technologies could revolutionize our practice in studying not just the metabolism of drugs but also increase our capacity to study the zonal progression of many liver diseases and conduct other exciting biomedical research.

bioengineering↗

bioRxiv2022

Lumped Parameter Liver Simulation to Predict Acute Hemodynamic Alterations Following Partial Resections

Partial liver resections are routinely performed in living donor liver transplantation and to debulk tumors in liver malignancies, but surgical decisions on vessel reconstruction for adequate inflow and outflow are challenging. Pre-operative evaluation is often limited to radiological imaging, which fails to account for post-resection hemodynamic alterations. Substantial evidence suggests post-surgical increase in local volume flow rate enhances shear stress, signaling hepatic regeneration, but excessive shear stress has been postulated to result in small for size syndrome and liver failure. Predicting hemodynamic alterations throughout the liver is particularly challenging due to the dendritic architecture the vasculature, spanning several orders of magnitude in diameter. Therefore, we developed a mathematical lumped parameter model with realistic heterogeneities capturing inflow/outflow of the human liver to simulate acute perfusion alterations following surgical resection. Our model is parameterized using clinical measurements, relies on a single free parameter, and accurately captures established perfusion characteristics. We quantify acute changes in volume flow rate, flow speed, and wall shear stress following variable, realistic liver resections and make comparisons to the intact liver. Our numerical model runs in minutes and can be adapted to patient-specific anatomy, providing a novel computational tool aimed at assisting pre- and intra-operative surgical decisions for liver resections.

bioengineering↗