A mathematical model of the rat kidney IV: whole-kidney response to hyperkalemia

The renal response to acute hyperkalemia is mediated by increased K secretion within connecting tubule (CNT), flux that is modulated by tubular effects (e.g. aldosterone) in conjunction with increased luminal flow. There is ample evidence that peritubular K blunts Na reabsorption in proximal tubule, thick ascending Henle limb, and distal convoluted tubule (DCT). While any such reduction may augment CNT delivery, the relative contribution of each is uncertain. The kidney model of this lab was recently advanced with representation of cortical labyrinth and medullary ray. Model tubules capture the impact of hyperkalemia to blunt Na reabsorption within each upstream segment. However, this forces the question of the extent to which increased Na delivery is transmitted past macula densa and its tubuloglomerular feedback (TGF) signal. Beyond increasing macula densa Na delivery, peritubular K is predicted to raise cytosolic Cl and depolarize macula densa cells, which may also activate TGF. Thus, although upstream reduction in Na transport may be larger, it appears that the DCT effect is critical to increasing CNT delivery. Beyond the flow effect, hyperkalemia reduces ammoniagenesis and reduced ammoniagenesis enhances K excretion. What this model provides is a possible mechanism. When cortical NH4 is taken up via peritubular Na,K(NH4)-ATPase, it acidifies principal cells. Consequently, reduced ammoniagenesis increases principal cell pH, thereby increasing conductance of both ENaC and ROMK, enhancing K excretion. In this model, aldosterone's effect on principal cells, diminished DCT Na reabsorption, and reduced ammoniagenesis, all provide relatively equal and additive contributions to renal K excretion.

Mechanisms of Flow-Mediated Dilation of Pial Collaterals and the Effect of Hypertension

Leptomeningeal anastomoses are small distal anastomotic vessels also known as pial collaterals in the brain. These vessels redirect blood flow during an occlusion and are important for stroke treatment and outcome. Pial collaterals have unique hemodynamic forces and experience significantly increased luminal flow and shear stress after the onset of ischemic stroke. However, there is limited knowledge of how pial collaterals respond to flow and shear stress, and whether this response is altered in chronic hypertension. Using an in vitro system, pial collaterals from normotensive and hypertensive rats (n=6–8/group) were isolated and luminal flow was induced with intravascular pressure maintained at 40 mm Hg. Collateral lumen diameter was measured following each flow rate in the absence or presence of pharmacological inhibitors and activators. Collaterals from male and female Wistar rats dilated similarly to increased flow (2 µL/minute: 58.4±18.7% versus 67.9±7.4%; P =0.275), and this response was prevented by inhibition of the transient receptor potential vanilloid type 4 channel, as well as inhibitors of nitric oxide and intermediate-conductance calcium-activated potassium channels, suggesting shear stress-induced activation of this pathway was involved. However, the vasodilation was significantly impaired in hypertensive rats (2 µL/minute: 17.7±7.7%), which was restored by inhibitors of reactive oxygen species and mimicked by angiotensin II. Thus, flow- and shear stress-induced vasodilation of pial collaterals appears to be an important stimulus for increasing collateral flow during large vessel occlusion. Impairment of this response during chronic hypertension may be related to poorly engaged pial collaterals during ischemic stroke in hypertensive subjects.

Luminal Fluid Motion Inside an In Vitro Dissolution Model of the Human Ascending Colon Assessed Using Magnetic Resonance Imaging

Knowledge of luminal flow inside the human colon remains elusive, despite its importance for the design of new colon-targeted drug delivery systems and physiologically relevant in silico models of dissolution mechanics within the colon. This study uses magnetic resonance imaging (MRI) techniques to visualise, measure and differentiate between different motility patterns within an anatomically representative in vitro dissolution model of the human ascending colon: the dynamic colon model (DCM). The segmented architecture and peristalsis-like contractile activity of the DCM generated flow profiles that were distinct from compendial dissolution apparatuses. MRI enabled different motility patterns to be classified by the degree of mixing-related motion using a new tagging method. Different media viscosities could also be differentiated, which is important for an understanding of colonic pathophysiology, the conditions that a colon-targeted dosage form may be subjected to and the effectiveness of treatments. The tagged MRI data showed that the DCM effectively mimicked wall motion, luminal flow patterns and the velocities of the contents of the human ascending colon. Accurate reproduction of in vivo hydrodynamics is an essential capability for a biorelevant mechanical model of the colon to make it suitable for in vitro data generation for in vitro in vivo evaluation (IVIVE) or in vitro in vivo correlation (IVIVC). This work illustrates how the DCM provides new insight into how motion of the colonic walls may control luminal hydrodynamics, driving erosion of a dosage form and subsequent drug release, compared to traditional pharmacopeial methods.

Mechanisms of decreased tubular flow-induced nitric oxide in Dahl salt-sensitive rat thick ascending limbs

Dahl salt-sensitive rat (SS) kidneys produce less nitric oxide (NO) than those of salt-resistant rats (SR). Thick ascending limb (THAL) NO synthase 3 (NOS3) is a major source of renal NO, and luminal flow enhances its activity. We hypothesized that flow-induced NO is reduced in SS THALs primarily due to NOS uncoupling and diminished NOS3 expression rather than scavenging. Rats were fed normal (NS) or high-salt (HS) diets. We measured flow-induced NO and superoxide in perfused THALs, and performed Western blots of renal outer medullas. For rats on NS, flow-induced NO was 35±6 arbitrary units (AU)/min in SR but only 11±2 AU/min in SS THALs (p<0.008). The superoxide scavenger tempol decreased the difference in flow-induced NO between strains by about 36% (p<0.020). The NOS inhibitor L-NAME decreased flow-induced superoxide 36 ± 8% in SS (p<0.02) but had no effect in SR THALs. NOS3 expression was not different between strains on NS. For rats on HS, the difference in flow-induced NO between strains was enhanced (SR: 44±10 vs SS: 9±2 AU/min; p<0.005). Tempol decreased the difference in flow-induced NO between strains by about 37% (p<0.012). L-NAME did not significantly reduce flow-induced superoxide in either strain. HS increased NOS3 expression in SR but not in SS THALs (p<0.003). We conclude: 1) on NS, flow-induced NO is diminished in SS THALs mainly due to NOS3 uncoupling such that it produces superoxide; and 2) on HS, the difference is enhanced due to failure of SS THALs to increase NOS3 expression.

Luminal Flow Actuation Generates Coupled Shear and Strain in a Microvessel-on-Chip

In the microvasculature, blood flow-derived forces are key regulators of vascular structure and function. Consequently, the development of hydrogel-based microvessel-on-chip systems that strive to mimic the in vivo cellular organization and mechanical environment has received great attention in recent years. However, despite intensive efforts, current microvessel- on-chip systems suffer from several limitations, most notably failure to produce physiologically relevant wall strain levels. In this study, a novel microvessel-on-chip based on the templating technique and using luminal flow actuation to generate physiologically relevant levels of wall shear stress and circumferential stretch is presented. Normal forces induced by the luminal pressure compress the surrounding soft collagen hydrogel, dilate the channel, and create large circumferential strain. The fluid pressure gradient in the system drives flow forward and generates realistic pulsatile wall shear stresses. Rigorous characterization of the system reveals the crucial role played by the poroelastic behavior of the hydrogel in determining the magnitudes of the wall shear stress and strain. The experimental measurements are combined with an analytical model of flow in both the lumen and the porous hydrogel to provide an exceptionally versatile user manual for an application-based choice of parameters in microvessels-on-chip. This unique strategy of flow actuation adds a dimension to the capabilities of microvessel-on-chip systems and provides a more general framework for improving hydrogel-based in vitro engineered platforms.Abstract Figure

The effects of luminal and trans-endothelial fluid flows on the extravasation and tissue invasion of tumor cells in a 3D in vitro microvascular platform

ABSTRACTThroughout the process of metastatic dissemination, tumor cells are continuously subjected to mechanical forces resulting from complex fluid flows due to changes in pressures in their local microenvironments. While these forces have been associated with invasive phenotypes in 3D matrices, their role in key steps of the metastatic cascade, namely extravasation and subsequent interstitial migration, remains poorly understood. In this study, an in vitro model of the human microvasculature was employed to subject tumor cells to physiological luminal, trans-endothelial, and interstitial flows to evaluate their effects on those key steps of metastasis. Luminal flow promoted the extravasation potential of tumor cells, possibly as a result of their increased intravascular migration speed. Trans-endothelial flow increased the speed with which tumor cells transmigrated across the endothelium as well as their migration speed in the matrix following extravasation. In addition, tumor cells possessed a greater propensity to migrate in close proximity to the endothelium when subjected to physiological flows, which may promote the successful formation of metastatic foci. These results show important roles of fluid flow during extravasation and invasion, which could determine the local metastatic potential of tumor cells.

Effect of luminal flow on doming of mpkCCD cells in a 3D perfusable kidney cortical collecting duct model

The cortical collecting duct (CCD) of the mammalian kidney plays a major role in the maintenance of total body electrolyte, acid/base, and fluid homeostasis by tubular reabsorption and excretion. The mammalian CCD is heterogeneous, composed of Na+-absorbing principal cells (PCs) and acid-base-transporting intercalated cells (ICs). Perturbations in luminal flow rate alter hydrodynamic forces to which these cells in the cylindrical tubules are exposed. However, most studies of tubular ion transport have been performed in cell monolayers grown on or epithelial sheets affixed to a flat support, since analysis of transepithelial transport in native tubules by in vitro microperfusion requires considerable expertise. Here, we report on the generation and characterization of an in vitro, perfusable three-dimensional kidney CCD model (3D CCD), in which immortalized mouse PC-like mpkCCD cells are seeded within a cylindrical channel embedded within an engineered extracellular matrix and subjected to luminal fluid flow. We find that a tight epithelial barrier composed of differentiated and polarized PCs forms within 1 wk. Immunofluorescence microscopy reveals the apical epithelial Na+ channel ENaC and basolateral Na+/K+-ATPase. On cessation of luminal flow, benzamil-inhibitable cell doming is observed within these 3D CCDs consistent with the presence of ENaC-mediated Na+ absorption. Our 3D CCD provides a geometrically and microphysiologically relevant platform for studying the development and physiology of renal tubule segments.

Numerical Analysis of Urine Flow with Multiple Sizes of Double-J Stents

This study investigated which sizes of double-J stents are more effective in achieving an acceptable urine flow through stenotic and stented ureters. Sixty four computational fluid dynamics models of the combinations of two different gauge ureters (4.57 mm and 5.39 mm in diameter) with four different levels of ureteral and four different sizes of double-J stents were developed for the numerical analysis of urine flow in the ureter. Luminal, extraluminal, and total flow rates along the ureter were measured, and the flow patterns around the ports and side holes were investigated. For the 4.57-mm ureter, the total flow rate for each gauge of stent was 23–63 mL/h (5 Fr), 20–47 mL/h (6 Fr), 17–35 mL/h (7 Fr), and 16–26 mL/h (8 Fr) and for the 5.39-mm ureter, the total flow rate for each gauge of stent was 43–147 mL/h (5 Fr), 36–116 mL/h (6 Fr), 29–92 mL/h (7 Fr), and 26–71 mL/h (8 Fr). With a 74% stenosis, all stents allowed a low flow rate, and the differences in flow rates between the stents were small. At the other levels of stenosis, 5 Fr stents allowed greater flow rates than the 8 Fr stents. The luminal flow rate increased just before the area of stenosis and decreased after the stenosis because of the increase and decrease in the luminal flow through the side holes before and after the stenosis. Therefore, a larger double-J stent is not favorable in achieving an acceptable urine flow through the stenotic and stented ureters. The results in this study could not be necessarily correlated with clinical situation because peristalsis, viscosity of the urine and real format of the ureter were not considered in our model. In vivo experiments are necessary for confirmation of our findings. Double J stents are commonly used in the ureteral stenosis or occlusion, especially due to ureter stones which obstruct the flow of urine. Clinicians choose the size of double J stent on the basis of their clinical experience. Here, we tried to know which sizes of double J stents are better for sufficient urine flow. According to various documents that try to determine the optimal shape of double J stents to increase the urine flow through the ureter, mostly bigger stent is recommended to occur maximum urine flow. However, in case of ureter with stenosis or occlusion, the right size of the double J stent may vary depending on the degree of stenosis in the ureter. To find appropriate stent size for the ureter with stenosis, computational fluid dynamics was conducted. This study shows that smaller diameter stents are more appropriate than larger diameter stents depending on the situation.

A Computer Model of Oxygen Dynamics in the Cortex of the Rat Kidney at the Cell-Tissue Level

The renal cortex drives renal function. Hypoxia/reoxygenation are primary factors in ischemia-reperfusion (IR) injuries, but renal oxygenation per se is complex and awaits full elucidation. Few mathematical models address this issue: none captures cortical tissue heterogeneity. Using agent-based modeling, we develop the first model of cortical oxygenation at the cell-tissue level (RCM), based on first principles and careful bibliographical analysis. Entirely parameterized with Rat data, RCM is a morphometrically equivalent 2D-slice of cortical tissue, featuring peritubular capillaries (PTC), tubules and interstitium. It implements hemoglobin/O2 binding-release, oxygen diffusion, and consumption, as well as capillary and tubular flows. Inputs are renal blood flow RBF and PO2 feeds; output is average tissue PO2 (tPO2). After verification and sensitivity analysis, RCM was validated at steady-state (tPO2 37.7 ± 2.2 vs. 36.9 ± 6 mmHg) and under transients (ischemic oxygen half-time: 4.5 ± 2.5 vs. 2.3 ± 0.5 s in situ). Simulations confirm that PO2 is largely independent of RBF, except at low values. They suggest that, at least in the proximal tubule, the luminal flow dominantly contributes to oxygen delivery, while the contribution of capillaries increases under partial ischemia. Before addressing IR-induced injuries, upcoming developments include ATP production, adaptation to minutes–hours scale, and segmental and regional specification.