Shear stress and oxygen availability drive differential changes in opossum kidney proximal tubule cell metabolism and endocytosis

Abstract

Kidney proximal tubule (PT) cells have high-metabolic demands to drive the extraordinary ion and solute transport, water reabsorption, and endocytic uptake that occur in this nephron segment. Increases in renal blood flow alter glomerular filtration rate and lead to rapid mechanosensitive adaptations in PT transport, impacting metabolic demand. Although the PT reabsorbs essentially all of the filtered glucose, PT cells rely primarily on oxidative metabolism rather than glycolysis to meet their energy demands. We lack an understanding of how PT functions are impacted by changes in O 2 availability via cortical capillaries and mechanosensitive signaling in response to alterations in luminal flow. Previously, we found that opossum kidney (OK) cells recapitulate key features of PT cells in vivo, including enhanced endocytic uptake and ion transport, when exposed to mechanical stimulation by culture on an orbital shaker.We hypothesized that increased oxygenation resulting from orbital shaking also contributes to this more physiologic phenotype. RNA seq of OK cells maintained under static conditions or exposed to orbital shaking for up to 96 hours showed significant time-and culture-dependent changes in gene expression. Transcriptional and metabolomics data were consistent with a decrease in glycolytic flux and with an increased utilization of aerobic metabolic pathways in cells exposed to orbital shaking. Moreover, we found spatial differences in the pattern of mitogenesis vs development of ion transport and endocytic capacities in our culture system that highlight the complexity of O 2 -dependent and mechanosensitive crosstalk to regulate PT cell function.

Endocytic adaptation to functional demand by the kidney proximal tubule

“…Subsequent studies demonstrated that growing OK cells under continuous shear stress and with increased O 2 availability dramatically improved their baseline endocytic capacity by roughly 5-fold compared with cells maintained under static conditions (Long et al 2017). In addition to increased expression of megalin and cubilin, cells cultured in this manner acquire morphological, transport, metabolic and transcriptional adaptations characteristic of PTs in vivo, including dense microvilli, greater sodium transport capacity, and oxidative metabolism (Long et al 2017;Ren et al 2019;Park et al 2020). Similar to the observations using cells cultured under static conditions and exposed acutely to flow, endocytic capacity in this more differentiated cell model is rapidly and reversibly regulated by changes in FSS (Long et al 2017).…”

Differential kidney proximal tubule cell responses to protein overload by albumin and its ligands

“…We recently developed an opossum kidney (OK) cell model cultured under continuous shear stress that recapitulates key features of PT function, including well-differentiated apical and basolateral plasma membrane domains, increased energy utilization and ion transport, and dramatically expanded apical endocytic capacity (27). Similarly to the PT in vivo, these cells rely on oxidative metabolism for energy production, thus enabling us to better address the specific role of fatty acid ligands carried by albumin in overload responses (34). Using this model, we asked how exposure to nephrotic concentrations of human serum (HuSer), to FAF-BSA, or to BSA complexed with different ligands affects cellular outcomes known to be associated with proteinuria.…”

Drug Repurposing of potential drug targets for treatment of COVID-19

“…To establish a link between SARS-CoV-2-related genes to drugs, we used a commercial drug database of Metacore version 20.3 build 70200 from Clarivate Analytics [15] and IPA [14]. To highlight the most promising drugs that might be repurposed for COVID-19 treatment, only drug targets that were either FDA approved or had been examined in clinical trials were selected.…”

“…To establish a link between SARS-CoV-2-related genes to drugs, we used a commercial drug database of Metacore version 20.3 build 70200 from Clarivate Analytics [15] and IPA [14].…”