Klotho Lacks a Vitamin D Independent Physiological Role in Glucose Homeostasis, Bone Turnover, and Steady-State PTH Secretion In Vivo

Apart from its function as co-receptor for fibroblast growth factor-23 (FGF23), Klotho is thought to regulate insulin signaling, intracellular oxidative stress, and parathyroid hormone (PTH) secretion in an FGF23 independent fashion. Here, we crossed Klotho deficient (Kl−/−) mice with vitamin D receptor (VDR) mutant mice to examine further vitamin D independent functions of Klotho. All mice were fed a rescue diet enriched with calcium, phosphorus, and lactose to prevent hyperparathyroidism in VDR mutants, and were killed at 4 weeks of age after double fluorochrome labeling. Kl−/− mice displayed hypercalcemia, hyperphosphatemia, dwarfism, organ atrophy, azotemia, pulmonary emphysema, and osteomalacia. In addition, glucose and insulin tolerance tests revealed hypoglycemia and profoundly increased peripheral insulin sensitivity in Kl−/− mice. Compound mutants were normocalcemic and normophosphatemic, did not show premature aging or organ atrophy, and were phenocopies of VDR mutant mice in terms of body weight, bone mineral density, bone metabolism, serum calcium, serum phosphate, serum PTH, gene expression in parathyroid glands, as well as urinary calcium and phosphate excretion. Furthermore, ablation of vitamin D signaling in double mutants completely normalized glucose and insulin tolerance, indicating that Klotho has no vitamin D independent effects on insulin signaling. Histomorphometry of pancreas islets showed similar beta cell volume per body weight in all groups of animals. In conclusion, our findings cast doubt on a physiologically relevant vitamin D and Fgf23 independent function of Klotho in the regulation of glucose metabolism, bone turnover, and steady-state PTH secretion in vivo

Nucleotide sequence of the na+/h+ exchanger-8 in patients with congenital sodium diarrhea.

Sodium absorption by the intestine is mediated by brush border Na(+)/H(+) exchangers, which include the NHE3 and NHE8 isoforms. We demonstrated a maturational decrease in NHE8 and increase in NHE3 in mouse intestine mRNA abundance and brush border membrane protein abundance, indicating a developmental switch of isoforms. Congenital sodium diarrhea is a rare autosomal recessive disorder characterized by polyhydramnios, hyponatremia, metabolic acidosis, and diarrhea with a high sodium content. Previous studies using intestinal brush border membrane vesicles from patients with this disorder have demonstrated a decrease in Na(+)/H(+) exchanger activity. Because some patients with congenital sodium diarrhea improve with age and knowing the developmental switch from NHE8 to NHE3, NHE8 may be a candidate gene for this disorder. We sequenced NHE8 from 5 patients with this disorder and found no disease-causing homozygous mutations. Although brush border membrane Na(+)/H(+) exchange activity may be decreased, exonic mutations in NHE8 cannot account for this disorder in these subjects

Effect of metabolic acidosis on neonatal proximal tubule acidification

The serum bicarbonate in neonates is lower than adults due in large part to a lower rate of proximal tubule acidification. It is unclear if the neonatal proximal tubule is functioning at maximal capacity or if the proximal tubule can respond to metabolic acidosis as has been described in adult proximal tubules. We find that neonatal mouse brush-border membranes have a lower Na+/H+ exchanger (NHE) 3 protein abundance (neonate 0.11 ± 0.05 vs. adult 0.64 ± 0.07; P < 0.05) and a higher NHE8 protein abundance (neonate 1.0 ± 0.01 vs. adult 0.13 ± 0.09; P < 0.001) compared with adults. To examine if neonates can adapt to acidosis, neonatal mice were gavaged with either acid or vehicle for 4 days, resulting in a drop in serum bicarbonate from 19.5 ± 1.0 to 8.9 ± 0.6 meq/l (P < 0.001). Proximal convoluted tubule Na+/H+ exchanger activity (dpHi/dt) was 1.68 ± 0.19 pH units/min in control tubules and 2.49 ± 0.60 pH units/min in acidemic neonatal mice (P < 0.05), indicating that the neonatal proximal tubule can respond to metabolic acidosis with an increase in Na+/H+ exchanger activity. Similarly, brush-border membrane vesicles from neonatal rats had an increase in Na+/H+ exchanger activity with acidemia that was almost totally inhibited by 10−6 M 5-(N-ethyl-n-isopropyl)-amiloride, a dose that has little effect on NHE3 but inhibits NHE8. There was a significant increase in both NHE3 (vehicle 0.35 ± 0.07 vs. acid 0.73 ± 0.07; P < 0.003) and NHE8 brush-border membrane protein abundance (vehicle 0.41 ± 0.05 vs. acid 0.73 ± 0.06; P < 0.001) in acidemic mouse neonates compared with controls. A comparable increase in NHE3 and NHE8 was found in neonatal rats with acidosis. In conclusion, the neonatal proximal tubule can adapt to metabolic acidosis with an increase in Na+/H+ exchanger activity