Elsa Bello-Reuss

Expression of MDR1 (Multidrug Resistance) Gene and Its Protein in Normal Human Kidney

P-glycoprotein (Pgp), the product of the multidrug resistance (MDR) gene overexpressed in cancer cells, is present also in normal tissues. In the kidney, MDR1 Pgp has been found in the proximal tubule and in cultured mesangial cells. In situ hybridization and immunohistochemistry were used to determine the complete nephronal localization of MDR mRNA and its product, Pgp, in the human kidney. MDR mRNA expression was studied with the use of nonradioactive in situ MDR RNA probes. MDR1 Pgp was immunolocalized using the specific monoclonal antibody MRK16. The presence of MDR mRNA was confirmed in proximal tubules and mesangium, and demonstrated as well in thick limb of Henles loops and in collecting ducts. MDR1 Pgp colocalized in the same nephronal segments. This suggests that, in addition to secreting xenobiotics, Pgp may play a role in the transport of endogenous substrates or in the regulation of Cl- channels.

Electrophysiological identification of cell types in cortical collecting duct monolayers.

Double-barrel microelectrodes were used to determine membrane voltages and the intracellular pH (pHi) in primary cultures of cortical collecting duct cells (CCD) grown in the absence of aldosterone. Electrophysiologically, two main cell types were identified. In cell type 1, the apical membrane voltage (Va) was -60 +/- 5 mV. The fractional resistance of the apical membrane (fRa) was 0.40 +/- 0.03, and pHi was 7.21 +/- 0.04. Exposure to 50 mM K+ on the apical side depolarized Va by 21 +/- 4 mV. When Cl- was replaced by cyclamate two types of responses were observed: (a) depolarization of Va by 26 +/- 3 mV while pHi remained unchanged, and (b) no change in Va. In cell type 2, Va was -36 +/- 5 mV, fRa was 0.91 +/- 0.03 and increasing apical [K+] from 5 to 50 mM did not change Va. Two subpopulations were distinguished by the response of pHi to lowering apical [Cl-]. In one of them pHi increased from 6.99 +/- 0.05 to 7.11 +/- 0.07. In the other, pHi was significantly decreased from 7.16 +/- 0.08 to 7.03 +/- 0.07. These results are compatible with the conclusion that about 50% of the impaled cells type 2 have a Cl-/HCO-3 exchanger at the apical membrane. In summary, two different cell types can be identified electrophysiologically in CCD monolayers. Cell type 1 has the electrical characteristics of principal cells. Cell type 2 resembles the intercalated cells. The cell alkalinization observed in approximately 50% of the cells type 2 in response to Cl- removal suggests the presence of an apical Cl-/HCO-3 exchanger. Thus, these cells should be the bicarbonate-secreting cells. The remaining cells should correspond to the acid-secreting cells.

CHAPTER 346 – Kidney

The kidney is a complex, highly sophisticated organ containing diverse cells types responsible for specialized functions, both in the renal vasculature and nephron segments. The structure of the kidney and the process of urine formation allow for cell-to-cell influences along single nephron segments that are distant in space. In addition, the proximity of parallel structures permits lateral communication between tubules, capillaries, and interstitial cells. These two kinds of cell-to-cell communication are central for functional integration at the single-nephron and whole-organ levels. Communication is mediated by paracrine and autocrine agents that act extracellularly or intracellularly by turning on signaling systems, thus eventually unifying homeostatic regulation of renal hemodynamics, glomerular filtration rate, tubule-transport processes, and urinary excretion. Endothelial cells regulate renal vascularresistance and reabsorption of salt and water by the renal tubules via a variety of messenger molecules, including nitric oxide, prostaglandins, EETs, 20-HETE, and endothelin. The vascular and epithelial cells regulate their own functions via the production of the autocrine agent 20-HETE. Tubule cells produce dopamine, purine nucleotides, nitric oxide, prostanoids, and endothelin. These agents act locally to regulate the vasculature and transepithelial transport of salt and water. Despite a rapidly growing understanding of cell-cell signaling in the kidney, the wide variety of autocrine and paracrine systems and their actions on multiple cell types are not completely understood. Interactions with hormones and neural control systems add to this complexity. Current research is largely reductionist. Integrative studies are also needed, in particular, to discern the relative importance, cross-talk, redundancy, and compensatory effects of the various systems under physiological and pathophysiological conditions.