The anion exchanger pendrin (SLC26A4) and renal acid-base homeostasis

The anion exchanger pendrin (Pds, SLC26A4) transports various anions including bicarbonate, chloride and iodide. In the kidney, pendrin is exclusively expressed on the luminal pole of bicarbonate-secretory type B intercalated cells. Genetic ablation of pendrin in mice abolishes luminal chloride-bicarbonate exchanger activity from type B intercalated cells suggesting that pendrin is the apical bicarbonate extruding pathway. The renal expression of pendrin is developmentally adapted and pendrin positive cells originate from both the uretric bud and mesenchyme. In adult kidney, pendrin expression and activity is regulated by systemic acid-base status, dietary electrolyte intake (mostly chloride), and hormones such as angiotensin II and aldosterone which can affect subcellular localization, the relative number of pendrin expressing cells, and the overall abundance consistent with a role of pendrin in maintaining normal acid-base homeostasis. This review summarizes recent findings on the role and regulation of pendrin in the context of the kidneys role in acid-base homeostasis in health and disease

Renal Intercalated Cells Sense and Mediate Inflammation via the P2Y14 Receptor

Uncontrolled inflammation is one of the leading causes of kidney failure. Pro-inflammatory responses can occur in the absence of infection, a process called sterile inflammation. Here we show that the purinergic receptor P2Y14 (GPR105) is specifically and highly expressed in collecting duct intercalated cells (ICs) and mediates sterile inflammation in the kidney. P2Y14 is activated by UDP-glucose, a damage-associated molecular pattern molecule (DAMP) released by injured cells. We found that UDP-glucose increases pro-inflammatory chemokine expression in ICs as well as MDCK-C11 cells, and UDP-glucose activates the MEK1/2-ERK1/2 pathway in MDCK-C11 cells. These effects were prevented following inhibition of P2Y14 with the small molecule PPTN. Tail vein injection of mice with UDP-glucose induced the recruitment of neutrophils to the renal medulla. This study identifies ICs as novel sensors, mediators and effectors of inflammation in the kidney via P2Y14

Altered V-ATPase expression in renal intercalated cells isolated from B1-subunit deficient mice by fluorescence activated cell sorting

Unlike human patients with mutations in the 56-kDa B1 subunit isoform of the vacuolar proton-pumping ATPase (V-ATPase), B1-deficient mice (Atp6v1b1(-/-)) do not develop metabolic acidosis under baseline conditions. This is due to the insertion of V-ATPases containing the alternative B2 subunit isoform into the apical membrane of renal medullary collecting duct intercalated cells (ICs). We previously reported that quantitative Western blots (WBs) from whole kidneys showed similar B2 protein levels in Atp6v1b1(-/-) and wild type mice. However, WBs from renal medulla (including outer and inner medulla) membrane and cytosol fractions reveal a decrease in the levels of the ubiquitous V-ATPase E1 subunit. To compare V-ATPase expression specifically in ICs from wild type and Atp6v1b1(-/-) mice, we crossed mice in which EGFP expression is driven by the B1 subunit promoter (EGFP-B1(+/+) mice) with Atp6v1b1(-/-) mice to generate novel EGFP-B1(-/-) mice. We isolated pure IC populations by fluorescence-assisted cell sorting from EGFP-B1(+/+) and EGFP-B1(-/-) mice to compare their V-ATPase subunit protein levels. We report that V-ATPase A, E1, and H subunits are all significantly down-regulated in EGFP-B1(-/-) mice, while the B2 protein level is considerably increased in these animals. We conclude that under baseline conditions the B2 up-regulation compensates for the lack of B1, and is sufficient to maintain basal acid-base homeostasis, even when other V-ATPase subunits are down-regulated

Immunolocalization of neutrophils in kidney medulla of mice 48 h after injection with saline (Sham) or 100 μM UDP-glucose.

<p>Mosaic images of kidney medulla double-labeled for P2Y₁₄ (green) and the neutrophil marker Ly6G (red; white circles) from mice injected with saline (A) or 100 μM UDP-glucose (B). Individual neutrophils, delineated in the corresponding white circles in Panels A and B, are shown in the small panels A1-A5, and B1-B5, respectively. (C-D) High magnification images showing the presence of neutrophils (arrows) in proximity to medullary ICs after UDP-glucose injection (D, D', D"). In the sham animals (C, C', C"), the areas surrounding collecting ducts were often devoid of neutrophils. Scale bars = 200 μm in A and B, 25 μm in C-D.</p

Expression of P2Y₁₄ in EGFP(+) cells.

<p>(A) Representative immunoblot profile of P2Y₁₄ in two EGFP(+) cell samples isolated by FACS. (B) Binding of [³H]UDP-glucose to total membranes prepared from FACS isolated EGFP(+) and EGFP(−) cells in the presence or absence of a saturating concentration (10^–5 M) of unlabeled UDP-glucose or ATP. Data are represented as fold changes compared to the binding measured in the presence of unlabeled UDP-glucose. Each bar represent the average of 3 independent experiments each performed in triplicate. Values are expressed as mean ± SEM, * P<0.05.</p

Quantitative PCR detection of pro-inflammatory mediators in EGFP(+) cells.

<p>EGFP(+) cells were isolated by FACS from B1-EGFP mice 4h after an i.v. injection with saline (sham) or with saline containing 100 μM UDP-glucose (UDP-glu). All values are normalized to GAPDH. Data are represented as % changes relative to control. Values are mean ± SEM (n = 4), *P<0.05, ** P<0.001.</p

Flow cytometry analysis of immune cell infiltration in the kidneys of mice 48 h after injection with saline (sham) or 100 μM UDP-glucose (UDP-glu).

<p>Changes in kidney medulla (A) or cortex (B) infiltrated immune cell counts are represented as % changes relative to control. Values are means of percent of each cell population ± SEM from 4–6 animals. *P<0.05.</p