Hypertension may develop before or after the onset of diabetes and it is known to increase the risk of developing diabetic nephropathy. Alpha-1 antitrypsin (AAT) is a multi-functional protein with beneficial effects in various diseases but its role in reducing blood pressure in the diabetic kidney has not been thoroughly studied. Like blood pressure, epithelial sodium channels (ENaC) and its adaptor protein myristoylated alanine-rich C-kinase substrate (MARCKS) are regulated by circadian rhythms. Our hypothesis is that administration of human AAT (hAAT) reduces blood pressure in hypertensive diabetic mice by attenuating membrane expression of ENaC and its association with the actin cytoskeleton. First, we show hAAT administration results in reduced blood pressure in diabetic db/db mice compared to vehicle treatment in both the inactive and active cycles. Western blotting and immunohistochemistry analyses showed a reduction of ENaC and the actin cytoskeleton protein, MARCKS in the kidneys of diabetic db/db mice treated with hAAT compared to vehicle. hAAT treatment resulted in elevated amounts of extracellular vesicles present in the urine of diabetic db/db mice compared to vehicle treatment both in the inactive and active cycles. Multiple hexosylceramides, among other lipid classes increased in urinary EVs released from hAAT treated hypertensive diabetic mice compared to vehicle treated mice. Taken together, these data suggest hAAT treatment could normalize blood pressure in the diabetic kidney in a mechanism involving attenuation of renal ENaC and MARCKS protein expression and possibly ceramide metabolism to hexosylceramide in kidney cells.

Space travel may have promising implications for innovative approaches in biomedical research. But there are potential challenges and health concerns associated with exposures from space travel that warrant the need for mechanistic studies to determine the effects on various organ systems including the cardiovascular system. Radiation exposure associated with space flight is known to adversely affect the heart and vascular system. However, less is known about the effects of microgravity on the cardiovascular system. Various functions of the cardiovascular system may be affected by microgravity. Studies have investigated changes in the cytoskeleton of various cell types in response to simulated microgravity. Other studies investigated the effect of microgravity on the permeability and migration of endothelial cells and myocardial atrophy associated with endothelial dysfunction. In addition, coagulation, vasoconstriction, blood volume, and cardiac dimensions were outcome measures of studies aimed at understanding the implications of microgravity on the cardiovascular system. This comprehensive review summarizes the effect of microgravity on various aspects of the cardiovascular system.

The C-type natriuretic peptide receptor (NPRC) is expressed in many cell types and binds all natriuretic peptides with high affinity. Ligand binding results in the activation or inhibition of various intracellular signaling pathways. Although NPRC ligand binding has been shown to regulate various ion channels, the regulation of endothelial sodium channel (EnNaC) activity by NPRC activation has not been studied. The objective of this study was to investigate mechanisms of EnNaC regulation associated with NPRC activation in human aortic endothelial cells (hAoEC). EnNaC protein expression and activity was attenuated after treating hAoEC with the NPRC agonist cANF compared to vehicle, as demonstrated by Western blotting and patch clamping studies, respectively. NPRC knockdown studies using siRNA’s corroborated the specificity of EnNaC regulation by NPRC activation mediated by ligand binding. The concentration of multiple diacylglycerols (DAG) and the activity of protein kinase C (PKC) was augmented after treating hAoEC with cANF compared to vehicle, suggesting EnNaC activity is down-regulated upon NPRC ligand binding in a DAG-PKC dependent manner. The reciprocal cross-talk between NPRC activation and EnNaC inhibition represents a feedback mechanism that presumably is involved in the regulation of endothelial function and aortic stiffness.

The expression of the myristoylated alanine-rich C-kinase substrate (MARCKS) family of proteins in the kidneys plays an important role in the regulation of the renal epithelial sodium channel (ENaC) and hence overall blood pressure regulation. The function of MARCKS is regulated by post-translational modifications including myristoylation, phosphorylation, and proteolysis. Proteases known to cleave both ENaC and MARCKS have been shown to contribute to the development of high blood pressure, or hypertension. Here, we investigated protein expression and proteolysis of MARCKS, protein expression of multiple protein kinase C (PKC) isoforms, and protein expression and activity of several different proteases in the kidneys of diabetic db/db mice compared to wild-type littermate mice. In addition, MARCKS protein expression was assessed in cultured mouse cortical collecting duct (mpkCCD) cells treated with normal glucose and high glucose concentrations. Western blot and densitometric analysis showed less abundance of the unprocessed form of MARCKS and increased expression of a proteolytically cleaved form of MARCKS in the kidneys of diabetic db/db mice compared to wild-type mice. The protein expression levels of PKC delta and PKC epsilon were increased, while cathepsin B, cathepsin S, and cathepsin D were augmented in diabetic db/db kidneys compared to those of wild-type mice. An increase in the cleaved form of MARCKS was observed in mpkCCD cells cultured in high glucose compared to normal glucose concentrations. Taken together, these results suggest that high glucose may contribute to an increase in the proteolysis of renal MARCKS, while the upregulation of the cathepsin proteolytic pathway positively correlates with increased proteolysis of MARCKS in diabetic kidneys, where PKC expression is augmented.

Cathpesin B is a multi-functional protease that plays numerous roles in physiology and pathophysiology. We hypothesized that actin cytoskeleton proteins that are substrates of cathepsin B, various lipids, and kinases that are regulated by lipids would be down-regulated in the kidney of cathepsin B knockout mice. Here, we show by Western blot and densitometric analysis that the expression and proteolysis of the actin cytoskeleton proteins myristoylated alanine-rich C-kinase substrate (MARCKS) and spectrin are significantly reduced in kidney cortex membrane fractions of cathepsin B knockout mice compared to C57B6 wild-type control mice. Lipidomic results show that specific lipids are increased while other lipids, including lysophosphatidylcholine (LPC) species LPC (16:0), LPC (18:0), LPC (18:1), and LPC (18:2), are significantly decreased in membrane fractions of the kidney cortex from Cathepsin B null mice. Protein Kinase C (PKC) activity is significantly lower in the kidney cortex of cathepsin B knockout mice compared to wild-type mice, while calcium/calmodulin-dependent protein kinase II (CaMKII) activity and phospholipase D (PLD) activity are comparable between the two groups. Together, these results provide the first evidence of altered actin cytoskeleton organization, membrane lipid composition, and PKC activity in the kidneys of mice lacking cathepsin B.

Background: The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is responsible for causing the Coronavirus disease 2019 (COVID-19) outbreak. While mutations cause the emergence of new variants, the ancestral SARS-CoV-2 strain is unique among other strains. Methods: Various clinical parameters, the activity of cathepsin proteases, and the concentration of various proteins were measured in urine samples from COVID-19-negative participants and COVID-19-positive participants. Urinary extracellular vesicles (uEVs) were isolated from urine samples from the two groups and used for proteomic analysis and subsequent pathway analyses. Results: Activity levels of cathepsin S and L were greater in the urine of COVID-19-positive participants. The concentration of C-reactive protein, transmembrane serine protease 2, and klotho protein were significantly greater in the urine of COVID-19-positive participants. There was a greater amount of uEVs in the COVID-19 group and klotho protein was found to be enriched in uEVs from the COVID-19 group. Pathway analyses of the proteomics data showed most of the identified proteins were involved in signal transduction, stress response, protein metabolism, and transport. The identified proteins were predominantly associated with cellular membranes and with function of the cytoskeleton, enzyme regulation, and signal transduction. Conclusions: Taken together, our data identify novel urinary biomarkers that could be used to further investigate the long-term effects of SARS-CoV-2 infection.