Effect of Insulin on D_5 Dopamine Receptor Expression and Function in Renal Proximal Tubule Cells / 中华高血压杂志

Objective To investigate the effect of insulin on D_5 dopamine receptor expression and function in renal proximal tubule (RPT).Methods Immortalized RPT cells and D_5 receptor transfected HEK293 (HEK-D_5) cells were used in the study to investigate the effect of insulin on D_5 receptor expression and function,and those effects were compared in RPT cells from Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). The function of D_5 receptor was determined by measurement of the Na~+-K~+-ATPase activity in HEK-D_5 cells. Results Insulin increased D_5 receptor protein expression in a concentration and time-dependent manner in WKY RPT cells,but not in SHR.The basal level of D_5 receptor expression was higher in WKY cells than that in SHR cells. Stimulation with fenoldopam(D_1-like dopamine receptor agonist) inhibited the Na~+-K~+-ATPase activity;pretreat- ment with insulin increased the inhibitory effect of fenoldopam on Na~+-K~+-ATPase activity in HEK-D_5 cells. Conclusion The abnormal regulation of insulin on D_5 receptor expression and function might be involved in the path- ogenesis of essential hypertension.

Effects of bioartificial kidney on cardiovascular function and cytokine response in endotoxic shock pigs / 中华急诊医学杂志

Objective To test the role of a bioartificial kidney which consisted of a continuous vein-vein hemofiltration (CVVH) system and renal proximal tubule cells (PTCs)device, also called the renal tubule assist device (RAD),in endotoxin shock.Methods Female crossbred pigs 25 to 30 kg received with 3×1011 bacteria / kg body weight of E.coli intraperitoneally followed by treatment with a sham RAD without cells (n=7) or a RAD with cells (n=7). Data on blood pressure, cardiac output, renal blood flow and systemic marker of inflammation (IL-6) were measured.Results Cardiac outputs and renal blood flows were higher in the RAD group with cells than in the sham RAD group after E.coli infusion (P＜0.05). Plasma IL-6 levels were lower in the cell RAD group than in the sham RAD group after bacterial administration (P＜0.05). A significant difference on survival time was also observed between the two groups,with(9.07 ± 0.88) h in the cell RAD group vs. (5.10 ± 0.46)h in the sham RAD group, P＜ 0.01.Conclusion The bioartificial kidney containing PTCs can improve cardiovascular function,reduce cytokine levels,and prolong survival time.

Co-localization and interaction of organic anion transporter 1 with caveolin-2 in rat kidney

The organic anion transporters (OAT) have recently been identified. Although the some transport properties of OATs in the kidney have been verified, the regulatory mechanisms for OAT's functions are still not fully understood. The rat OAT1 (rOAT1) transports a number of negatively charged organic compounds between the cells and their extracellular milieu. Caveolin (Cav) also plays a role in membrane transport. Therefore, we investigated the protein-protein interactions between rOAT1 and caveolin-2. In the rat kidney, the expressions of rOAT1 mRNA and protein were observed in both the cortex and the outer medulla. With respect to Cav-2, the expressions of mRNA and protein were observed in all portions of the kidney (cortex < outer medulla = inner medulla). The results of Western blot analysis using the isolated caveolae-enriched membrane fractions or the immunoprecipitates by respective antibodies from the rat kidney showed that rOAT1 and Cav-2 co-localized in the same fractions and they formed complexes each other. These results were confirmed by performing confocal microscopy with immunocytochemistry using the primary cultured renal proximal tubular cells. When the synthesized cRNA of rOAT1 along with the antisense oligodeoxynucleotides of Xenopus Cav-2 were co-injected into Xenopus oocytes, the [14C]p-aminohippurate and [3H]methotrexate uptake was slightly, but significantly decreased. The similar results were also observed in rOAT1 over-expressed Chinese hamster ovary cells. These findings suggest that rOAT1 and caveolin-2 are co-expressed in the plasma membrane and rOAT1's function for organic compound transport is upregulated by Cav-2 in the normal physiological condition.

Effects of Dopamine on Intracellular pH in Opossum Kidney Cells

Na+/H+ exchanger (NHE) has a critical role in regulation of intracellular pH (pHi) in the renal proximal tubular cells. It has recently been shown that dopamine inhibits NHE in the renal proximal tubules. Nevertheless, there is a dearth of information on the effects of long-term (chronic) dopamine treatment on NHE activities. This study was performed to elucidate the pHi regulatory mechanisms during the chronic dopamine treatments in renal proximal tubular OK cells. The resting pHi was greatly decreased by chronic dopamine treatments. The initial rate and the amplitude of intracellular acidification by isosmotical Na+ removal from the bath medium in chronically dopamine-treated cells were much smaller than those in control. Although it seemed to be attenuated in Na+-dependent pH regulation system, Na+-dependent pHi recovery by NHE after intracelluar acid loading in the dopamine-treated groups was not significantly different from the control. The result is interpreted to be due to the balance between the stimulation effects of lower pHi on the NHE activity and counterbalance by dopamine. Our data strongly suggested that chronic dopamine treatment increased intrinsic intracellular buffer capacity, since higher buffer capacity was induced by lower resting pHi and this effect could attenuate pHi changes under extracellular Na+-free conditions in chronically dopamine-treated cells. Our study also demonstrated that intracellular acidification induced by chronic dopamine treatments was not mediated by changes in NHE activity.

Study of K~(+) channels at the apical membrane of mouse renal proximal tubule cell with patch-clamp / 第三军医大学学报

Objective To study the property of the K~(+) channels at the apical membrane of mouse renal proximal tubule cells.Methods Single-channel recording of the K~(+) channel currents in apical membrane of freshly isolated renal proximal tubule cells was performed using the cell-attached mode of the patch-clamp technique.Results This kind of potassium channel appeared to be an inward rectifier,and its limiting inward slope conductance was(11.4?0.6) pS.The channel activity increased with hyperpolarization.Conclusion The potassium channel with spontaneous current was observed in most of the cell-attached patches,which is thought to be involved in volume regulation,potassium recycling and stabilization of the apical potential.