Double knockout of pendrin and Na-Cl cotransporter (NCC) causes severe salt wasting, volume depletion, and renal failure

Abstract

The Na-Cl cotransporter (NCC), which is the target of inhibition by thiazides, is located in close proximity to the chloride-absorbing transporter pendrin in the kidney distal nephron. Single deletion of pendrin or NCC does not cause salt wasting or excessive diuresis under basal conditions, raising the possibility that these transporters are predominantly active during salt depletion or in response to excess aldosterone. We hypothesized that pendrin and NCC compensate for loss of function of the other under basal conditions, thereby masking the role that each plays in salt absorption. To test our hypothesis, we generated pendrin/NCC double knockout (KO) mice by crossing pendrin KO mice with NCC KO mice. Pendrin/NCC double KO mice displayed severe salt wasting and sharp increase in urine output under basal conditions. As a result, animals developed profound volume depletion, renal failure, and metabolic alkalosis without hypokalemia, which were all corrected with salt replacement. We propose that the combined inhibition of pendrin and NCC can provide a strong diuretic regimen without causing hypokalemia for patients with fluid overload, including patients with congestive heart failure, nephrotic syndrome, diuretic resistance, or generalized edema.diuretics | kidney tubules | nephrogenic diabetes insipidus T he thiazide-sensitive Na-Cl cotransporter (NCC) (SLC12A3) and the Cl − /HCO 3 − exchanger pendrin (SLC26A4) are expressed on apical membranes of distal cortical nephron segments and mediate salt absorption, with pendrin working in tandem with the epithelial Na channel and NCC working by itself (1-6). Pendrin is expressed on the apical membrane of intercalated cells in late distal convoluted tubule (DCT), connecting tubule (CNT), and the cortical collecting duct (CCD) (7-9). The thiazide-sensitive NCC is primarily expressed on the apical membrane of DCT cells (10,11).Single deletion of pendrin or NCC does not cause salt wasting or excessive diuresis under basal conditions (5,(12)(13)(14)(15). Indeed, even a mild degree of salt wasting has not been demonstrated in these two genetically engineered mouse models at steady state. Kidney functions, including sodium and chloride excretion, urine output, and blood urea nitrogen (BUN) levels in mutant mice are comparable to wild-type (WT) animals (12-15). Both pendrin KO and NCC KO mice, however, show signs of volume depletion or develop hypotension during salt restriction (12,14). These findings have led investigators to conclude that pendrin and NCC are predominantly active during salt depletion (and or in response to increased aldosterone levels), and their contribution to salt reabsorption at baseline conditions is small.We hypothesized that NCC and pendrin may compensate for loss of the other under basal conditions, thereby masking the role that each plays in salt reabsorption. To test this hypothesis, we generated double knockout of pendrin and NCC mice by crossing animals with single deletion for NCC and pendrin. The double KO mice show significant salt and flui...

Protein Phosphatase 1 Inhibitor-1 Deficiency Reduces Phosphorylation of Renal NaCl Cotransporter and Causes Arterial Hypotension

“…51,52 Moreover, NCC 2/2 mice exhibit an enhanced abundance of the bicarbonatechloride cotransporter pendrin, 53 which when deleted in NCC 2/2 mice, led to lethal salt wasting. 54 Consistent with the unchanged aldosterone levels in I-1 2/2 mice, we did not detect any evidence for an altered ENaC regulation. However, the pendrin protein abundance was found to be upregulated, which could have contributed to the compensated phenotype.…”

Integrated compensatory network is activated in the absence of NCC phosphorylation

“…Because both β-and non-α/non-β ICs express the Cl/HCO 3 exchanger pendrin (39-41), we refer to them collectively herein as pendrin-positive ICs (PP-ICs) for simplicity. In addition to verifying the transcript abundance increase of pendrin (Slc264a4), which is activated in NCC KO mice (13,42), the profile revealed an increase in AE4 (Slc4a9), numerous V-type proton ATPase subunits (V0 subunit d2 and the V1 subunits B1, C2, and G3), and 2 different isoforms of carbonic anhydrase (CA2 and CA15). A comprehensive qPCR survey of IC transporters (Figtranscript levels was performed on kidney cortex samples from WT and SPAK KO mice using the Affymetrix whole-genome GeneChip Mouse Gene 1.0 ST Array, which represents all mouse genes and contains more than 28,000 total transcripts.…”

“…The nature of this compensation is poorly understood. In NCC KO mice, activation of pendrin-mediated chloride absorption (13), ENaC (11,14), and NHE3-mediated sodium reabsorption (14) have been shown to contribute, but an integrated molecular understanding of adaptive responses to NCC inhibition has been wanting. To develop a better appreciation of the response, we sought to identify the gene networks in the kidney that are activated to drive NaCl reabsorption when NCC phosphorylation is compromised, as occurs in the absence of the STE20/SPS-1-related proline-alanine-rich protein kinase (15,16).…”

Overexpression of Pendrin in Intercalated Cells Produces Chloride-Sensitive Hypertension

“…This possibility is further supported by a recent study showing that double deletion of NCC and pendrin leads to massive sodium wasting, hypovolemia, and hypotension. 24 From the study of two independent mouse models of pseudohypoaldosteronism type II, 25,26 it has been concluded that increased electroneutral NaCl absorption by NCC is the main mechanism leading to this syndrome. Functional coupling of pendrin and Ndcbe mediates NCC-like activity.…”