Hyposulfatemia, growth retardation, reduced fertility, and seizures in mice lacking a functional NaS _i -1 gene

Abstract

Inorganic sulfate is required for numerous functions in mammalian physiology, and its circulating levels are proposed to be maintained by the Na ؉ -SO 4 2؊ cotransporter, (NaSi-1). To determine the role of NaSi-1 in sulfate homeostasis and the physiological consequences in its absence, we have generated a mouse lacking a functional NaSi-1 gene, Nas1. Serum sulfate concentration was reduced by >75% in Nas1 ؊/؊ mice when compared with Nas1 ؉/؉ mice. Nas1 ؊/؊ mice exhibit increased urinary sulfate excretion, reduced renal and intestinal Na ؉ -SO 4 2؊ cotransport, and a general growth retardation. Nas1 ؊/؊ mouse body weight was reduced by >20% when compared with Nas1 ؉/؉ and Nas1 ؉/؊ littermates at 2 weeks of age and remained so throughout adulthood. Nas1 ؊/؊ females had a lowered fertility, with a 60% reduction in litter size. Spontaneous clonic seizures were observed in Nas1 ؊/؊ mice from 8 months of age. These data demonstrate NaSi-1 is essential for maintaining sulfate homeostasis, and its expression is necessary for a wide range of physiological functions.I norganic sulfate (SO 4 2Ϫ ) is an abundant anion in mammalian plasma and is essential for numerous physiological functions (1). SO 4 2Ϫ conjugation is an important step in the biotransformation of xenobiotics such as steroids, antiinflammatory agents, adrenergic stimulants and blockers, analgesics, and, in most cases, leads to an increase in their urinary excretion (2). SO 4 2Ϫ is also required for the activation of many endogenous compounds such as heparin, heparan sulfate, dermatan sulfate, and bile acids (3). In addition, sulfation of structural components such as glycosaminoglycans and cerebroside sulfate is essential for the maintenance of normal structure and function of tissues (4). Disturbances of SO 4 2Ϫ metabolism and transport have been associated with human syndromes and diseases, including metachromatic leukodystrophy, Hunter's syndrome, Morquio's syndrome, Maroteaux-Lamy syndrome, multiple-sulfohydrolase deficiency, and osteochondrodysplasias (5, 6). However, despite its importance in the body, SO 4 2Ϫ levels are rarely measured in a clinical setting and there remains areas in which knowledge of its significance is still lacking, or is at most, minimal.In humans, SO 4 2Ϫ absorption is initiated in the small intestine and its homeostasis is proposed to be maintained through renal tubular mechanisms (7). SO 4 2Ϫ is freely filtered in the glomerulus and is actively reabsorbed in the proximal tubule. Renal proximal tubular SO 4 2Ϫ reabsorption is mediated by entry through the brush-border membrane (BBM) by a Na ϩ -SO 4 2Ϫ cotransporter (NaS i -1), and exit through the basolateral membrane by an anion exchanger, Sat-1 (1). The NaS i -1 transporter is expressed primarily in the kidney and intestine (8) and has been proposed to play a major role in maintaining serum SO 4 2Ϫ concentrations within the normal physiological range of 0.33-0.47 mmol͞liter in humans (9-11). In recent years, a growing body of experimental evidence has demonstrated that variou...

Disruption of NaS1 sulfate transport function in mice leads to enhanced acetaminophen-induced hepatotoxicity

“…Because APAP administration has been shown to cause plasma sulfate depletion in humans and rodents, 9,10 we aimed to characterize APAP detoxification in a hyposulfatemic mouse model with reduced NaS1 function. 6 After a single dose of APAP (250 mg/kg), no change in serum sulfate levels was observed in Nas1 Ϫ/Ϫ mice, whereas in Nas1 ϩ/ϩ mice serum sulfate levels were significantly reduced, by about 30%, after 2 hours and were restored to basal levels after 12 hours ( Fig. 2A).…”

“…Because NaS1 mRNA is not expressed in the liver, 15 we propose that the observed differences in hepatotoxicity in the Nas1 Ϫ/Ϫ mice is most likely a result of reduced sulfate availability in the liver as a consequence of the hyposulfatemia found in the Nas1 Ϫ/Ϫ mice. 6 Renal Histology and Urinary Protein Levels. APAP nephrotoxicity has been observed in humans, 16 and large doses of APAP (1,200 mg/kg) cause renal injury in mice.…”

“…We have also shown increased hepatic phenol sulfotransferase activity in Nas1 Ϫ/Ϫ mice under basal conditions. 6 However, despite elevated sat-1 expression and sulfotransferase activity 6 in Nas1 Ϫ/Ϫ mice, it appears these two factors may not be sufficient to compensate for the low blood sulfate levels in order to overcome the increased APAP-induced hepatotoxicity in Nas1 Ϫ/Ϫ mice.…”

“…5 Recently, we generated NaS1-null (Nas1 Ϫ/Ϫ ) mice, which have very low (Ϸ0.2 mmol/L) serum sulfate levels compared to those (Ϸ1 mmol/L) in wild-type (Nas1 ϩ/ϩ ) mice. 6 In this study, we identified two loss-of-function polymorphisms in the human NaS1 gene and characterized the Nas1 Ϫ/Ϫ mouse showing increased APAP hepatotoxicity, suggesting disturbed NaS1 function produces hyposulfatemia that leads to increased drug-induced liver damage. From the…”

The Biological Roles of Steroid Sulfonation

“…The NaS1 protein is expressed on the apical membrane of epithelial cells in the proximal tubule where it mediates the first step of sulfate reabsorption [10], and SAT1 mediates the second step across the basolateral membrane [11] ( Figure 1A). Mice lacking the NaS1 or SAT1 genes have sulfate wasting into the urine which leads to low blood sulfate levels (hyposulfataemia) [12,13]. Humans with loss of function mutations (R12X and N174S) in the NaS1 gene also exhibit renal sulfate wasting and hyposulfataemia [14].…”