RESUMEN
BACKGROUND: Phosphate is absorbed in the small intestine via passive flow and active transport.NaPi-IIb, a type II sodium-dependent phosphate transporter, is considered to mediate active phosphate transport in rodents. To study the regulation of intestinal phosphate transport in chronic kidney disease (CKD), we analyzed the expression levels of NaPi-IIb, pituitary-specific transcription factor 1 (PiT-1) and PiT-2 and the kinetics of intestinal phosphate transport using two CKD models. METHODS: CKD was induced in rats via adenine orThy1 antibody injection. Phosphate uptake by intestinal brush border membrane vesicles (BBMV) and the messenger RNA (mRNA) expression of NaPi-IIb, PiT-1 and PiT-2 were analyzed. The protein expression level of NaPi-IIb was measured by mass spectrometry (e.g. liquid chromatography tandem mass spectrometry). RESULTS: In normal rats, phosphate uptake into BBMV consisted of a single saturable component and its Michaelis constant (Km) was comparable to that of NaPi-IIb. The maximum velocity (Vmax) correlated with mRNA and protein levels of NaPi-IIb. In the CKD models, intestinal phosphate uptake consisted of two saturable components. The Vmax of the higher-affinity transport, which is thought to be responsible for NaPi-IIb, significantly decreased and the decrease correlated with reduced NaPi-IIb expression. The Km of the lower-affinity transport was comparable to that of PiT-1 and -2. PiT-1 mRNA expression was much higher than that of PiT-2, suggesting that PiT-1 was mostly responsible for phosphate transport. CONCLUSIONS: This study suggests that the contribution of NaPi-IIb to intestinal phosphate absorption dramatically decreases in rats with CKD and that a low-affinity alternative to NaPi-IIb, in particular PiT-1, is upregulated in a compensatory manner in CKD.
Asunto(s)
Intestinos/fisiología , Proteínas de Transporte de Fosfato/metabolismo , Fosfatos/metabolismo , Insuficiencia Renal Crónica/metabolismo , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo IIb/metabolismo , Sodio/metabolismo , Adenina/toxicidad , Animales , Masculino , Ratas , Ratas Endogámicas F344 , Ratas Wistar , Insuficiencia Renal Crónica/inducido químicamente , Insuficiencia Renal Crónica/patología , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo III/genética , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo III/metabolismo , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo IIb/clasificación , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo IIb/genética , Factor de Transcripción Pit-1/genética , Factor de Transcripción Pit-1/metabolismoRESUMEN
Chronic kidney disease is characterized as impaired renal function along with the imbalance and dysregulation of mineral metabolism; recognized as chronic kidney disease-mineral and bone disorder. Hyperphosphatemia, characterized by altered phosphate homeostasis along with elevated fibroblast growth factor-23 and intact parathyroid hormone, is such an alteration of mineral metabolism. We discovered a novel inhibitor, EOS789, that interacts with several sodium-dependent phosphate transporters (NaPi-IIb, PiT-1, and PiT-2) known to contribute to intestinal phosphate absorption. This inhibitor dose-dependently increased the fecal phosphorus excretion rate and inversely decreased the urinary phosphorus excretion rate in normal rats, suggesting inhibition of intestinal phosphorus absorption. In rats with adenine-induced hyperphosphatemia, EOS789 markedly decreased the serum phosphate, fibroblast growth factor-23, and intact parathyroid hormone below values found in normal control rats. Notably, this pan-phosphate transporter inhibitor exhibited a more potent effect on serum phosphate than a NaPi-IIb-selective inhibitor in rats with hyperphosphatemia indicating that PiT-1 and PiT-2 play important roles in intestinal phosphate absorption. Moreover, in a long-term study, EOS789 sustained the suppression of serum phosphorus in parallel with fibroblast growth factor-23 and intact parathyroid hormone and ameliorated ectopic calcification of the thoracic aorta. Additionally, EOS789 treatment also ameliorated kidney deterioration in rats with progressive kidney injury, probably due to the strict phosphate control. Thus, EOS789 has potent efficacy against hyperphosphatemia and its complications and could provide a significant benefit to patients who are ineffectively treated with phosphate binders.
Asunto(s)
Trastorno Mineral y Óseo Asociado a la Enfermedad Renal Crónica , Hiperfosfatemia , Insuficiencia Renal Crónica , Animales , Humanos , Hiperfosfatemia/tratamiento farmacológico , Minerales , Proteínas de Transporte de Fosfato , Fosfatos/metabolismo , Ratas , Insuficiencia Renal Crónica/tratamiento farmacológicoRESUMEN
BACKGROUND: Hyperphosphatemia is common in chronic kidney disease and is associated with morbidity and mortality. The intestinal Na+-dependent phosphate transporter Npt2b is thought to be an important molecular target for the prevention of hyperphosphatemia. The role of Npt2b in the net absorption of inorganic phosphate (Pi), however, is controversial. METHODS: In the present study, we made tamoxifen-inducible Npt2b conditional knockout (CKO) mice to analyze systemic Pi metabolism, including intestinal Pi absorption. RESULTS: Although the Na+-dependent Pi transport in brush-border membrane vesicle uptake levels was significantly decreased in the distal intestine of Npt2b CKO mice compared with control mice, plasma Pi and fecal Pi excretion levels were not significantly different. Data obtained using the intestinal loop technique showed that Pi uptake in Npt2b CKO mice was not affected at a Pi concentration of 4 mM, which is considered the typical luminal Pi concentration after meals in mice. Claudin, which may be involved in paracellular pathways, as well as claudin-2, 12, and 15 protein levels were significantly decreased in the Npt2b CKO mice. Thus, Npt2b deficiency did not affect Pi absorption within the range of Pi concentrations that normally occurs after meals. CONCLUSION: These findings indicate that abnormal Pi metabolism may also be involved in tight junction molecules such as Cldns that are affected by Npt2b deficiency.
Asunto(s)
Absorción Intestinal , Riñón/metabolismo , Fosfatos/metabolismo , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo IIb/fisiología , Animales , Claudinas/metabolismo , Ratones Noqueados , Microvellosidades/metabolismoRESUMEN
BACKGROUND: Inorganic phosphate (Pi) binders are the only pharmacologic treatment approved for hyperphosphatemia. However, Pi binders induce the expression of intestinal Pi transporters and have limited effects on the inhibition of Pi transport. EOS789, a novel pan-Pi transporter inhibitor, reportedly has potent efficacy in treating hyperphosphatemia. We investigated the properties of EOS789 with comparison to a conventional Pi binder. METHODS: Protein and mRNA expression levels of Pi transporters were measured in intestinal and kidney tissues from male Wistar rats fed diets supplemented with EOS789 or lanthanum carbonate (LC). 32Pi permeability was measured in intestinal tissues from normal rats using a chamber. RESULTS: Increased protein levels of NaPi-2b, an intestinal Pi transporter, and luminal Pi removal were observed in rats treated with LC but not in rats treated with EOS789. EOS789 but not LC suppressed intestinal protein levels of the Pi transporter Pit-1 and sodium/hydrogen exchanger isoform 3. 32Pi flux experiments using small intestine tissues from rats demonstrated that EOS789 may affect transcellular Pi transport in addition to paracellular Pi transport. CONCLUSION: EOS789 has differing regulatory effects on Pi metabolism compared to LC. The properties of EOS789 may compensate for the limitations of LC therapy. The combined or selective use of EOS789 and conventional Pi binders may allow tighter control of hyperphosphatemia. J. Med. Invest. 70 : 260-270, February, 2023.
Asunto(s)
Hiperfosfatemia , Proteínas de Transporte de Fosfato , Ratas , Masculino , Animales , Proteínas de Transporte de Fosfato/metabolismo , Ratas Wistar , Hiperfosfatemia/tratamiento farmacológico , Absorción Intestinal , Fosfatos/metabolismoRESUMEN
Hyperphosphatemia associated with chronic kidney disease (CKD) not only dysregulates mineral metabolism and bone diseases, but also strongly contributes to the progression of kidney disease itself. We have identified a novel drug for hyperphosphatemia, EOS789, that interacts with several sodium-dependent phosphate transporters (NaPi-IIb, PiT-1, and PiT-2) known to contribute to intestinal phosphate absorption. In this study, we investigated whether EOS789 could ameliorate kidney disease progression in glomerulonephritis rats. Anti-glomerular basement membrane (GBM) nephritis was induced in rats by intravenously administering two types of anti-rat GBM antibodies. We evaluated the effect of EOS789 administered in food admixture on hyperphosphatemia and kidney disease progression. In an anti-GBM nephritis rats, which exhibit a significant increase in serum phosphate and a decline in renal function, EOS789 dose-dependently improved hyperphosphatemia and EOS789 at 0.3% food admixture significantly ameliorated kidney dysfunction as shown in the decline of serum creatinine and BUN. Renal histopathology analysis showed that EOS789 significantly decreased crescent formation in glomeruli. To elucidate the mechanism underlying glomerular disease progression, human mesangial cells were used. High phosphate concentration in media significantly increased the expression of Collagen 1A1, 3A1, and αSMA mRNA in human mesangial cells and EOS789 dose-dependently suppressed these fibrotic markers. These results indicate that EOS789 prevented glomerular crescent formation caused by mesangial fibrosis by ameliorating hyperphosphatemia. In conclusion, EOS789 would not only be useful against hyperphosphatemia but may also have the potential to relieve mesangial proliferative glomerulonephritis with crescent formation.
Asunto(s)
Glomerulonefritis , Hiperfosfatemia , Proteínas de Transporte de Fosfato , Animales , Autoanticuerpos , Progresión de la Enfermedad , Fibrosis , Glomerulonefritis/inducido químicamente , Glomerulonefritis/tratamiento farmacológico , Hiperfosfatemia/tratamiento farmacológico , Hiperfosfatemia/patología , Riñón , Proteínas de Transporte de Fosfato/antagonistas & inhibidores , Fosfatos/farmacología , Ratas , Ratas Endogámicas WKYRESUMEN
A treatment for hyperphosphatemia would be expected to reduce mortality rates for CKD and dialysis patients. Although rodent studies have suggested sodium-dependent phosphate transporter type IIb (NaPi-IIb) as a potential target for hyperphosphatemia, NaPi-IIb selective inhibitors failed to achieve efficacy in human clinical trials. In this study, we analyzed phosphate metabolism in rats, dogs, and monkeys to confirm the species differences. Factors related to phosphate metabolism were measured and intestinal phosphate absorption rate was calculated from fecal excretion in each species. Phosphate uptake by intestinal brush border membrane vesicles (BBMV) and the mRNA expression of NaPi-IIb, PiT-1, and PiT-2 were analyzed. In addition, alkaline phosphatase (ALP) activity was evaluated. The intestinal phosphate absorption rate, including phosphate uptake by BBMV and NaPi-IIb expression, was the highest in dogs. Notably, urinary phosphate excretion was the lowest in monkeys, and their intestinal phosphate absorption rate was by far the lowest. Dogs and rats showed positive correlations between Vmax/Km of phosphate uptake in BBMV and NaPi-IIb expression. Although phosphate uptake was observed in the BBMV of monkeys, NaPi-IIb expression was not detected and ALP activity was low. This study revealed significant species differences in intestinal phosphate absorption. NaPi-IIb contributes to intestinal phosphate uptake in rats and dogs. However, in monkeys, phosphate is poorly absorbed due to the slight degradation of organic phosphate in the intestine.
Asunto(s)
Absorción Intestinal/fisiología , Microvellosidades/metabolismo , Fosfatos/metabolismo , Fosfatasa Alcalina/metabolismo , Animales , Perros , Haplorrinos , Ratas , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo IIb/análisis , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo IIb/genética , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo IIb/metabolismo , Especificidad de la EspecieRESUMEN
The neural basis of functional lateralization in language processing is a fundamental issue in systems neuroscience. We used functional MRI (fMRI) to examine hemispheric dominance during the processing of signed and spoken sentences. By using tasks involving comprehension of sentences (Sc) and sentential non-word detection (Sn), we compared different groups and stimulus conditions. Under the sign condition with sentence stimuli in Japanese Sign Language (JSL), we tested two groups of subjects: Deaf signers (Deaf) of JSL, and hearing bilinguals (children of Deaf adults, CODA) of JSL and Japanese (JPN). Under the speech condition, we tested hearing monolinguals (Mono) of JPN with auditory JPN stimuli alone (AUD), or with an audiovisual presentation of JPN and JSL stimuli (A&V). We found that the overall bilateral activation patterns under the four experimental conditions of Deaf, CODA, AUD and A&V were almost identical, despite differences in stimuli (JSL and JPN) and groups (Deaf, CODA and Mono). Moreover, consistently left-dominant activations involving frontal and temporo-parietal regions were observed across all four conditions. Furthermore, irrespective of the modalities of sign and speech, the main effects of task (Sc-Sn) were found primarily in the left regions: the ventral part of the inferior frontal gyrus (F3t/F3O), the precentral sulcus, the superior frontal gyrus, the middle temporal gyrus, the angular gyrus and the inferior parietal gyrus. Among these regions, only the left F3t/F3O showed no main effects of modality condition. These results demonstrate amodal commonality in the functional dominance of the left cortical regions for comprehension of sentences, as well as the essential and universal role of the left F3t/F3O in processing linguistic information from both signed and spoken sentences.
Asunto(s)
Comprensión/fisiología , Dominancia Cerebral/fisiología , Lengua de Signos , Percepción del Habla/fisiología , Estimulación Acústica/métodos , Adulto , Análisis de Varianza , Mapeo Encefálico/métodos , Sordera/fisiopatología , Sordera/psicología , Femenino , Lóbulo Frontal/fisiología , Lóbulo Frontal/fisiopatología , Humanos , Imagen por Resonancia Magnética/métodos , Masculino , Estimulación Luminosa/métodos , PsicolingüísticaRESUMEN
We have previously mapped a diet-induced hypercholesterolemia locus (Dihc2) to chromosome 14 in the F2 generation cross of high-responsive exogenous hypercholesterolemia rats and low-responsive BN rats. To identify a causal gene within this locus, we constructed interval-specific congenic lines and carried out expression and sequencing analyses. Here we narrowed Dihc2 to a region including 33 genes and predicted transcripts and identified RGD1309450_predicted, a homologous gene of SMEK2, as a strong candidate for responsiveness to dietary cholesterol. Our finding provides new insights into the pathway underlying the individual responsiveness to dietary cholesterol in vivo.