Chronic metabolic acidosis results in a negative calcium balance as a result of bone resorption and renal calcium loss. However, reports on the changes in intestinal calcium transport have been controversial. The present investigation therefore aimed to study the effects of chronic metabolic acidosis induced by 1.5% NH₄Cl administration on the three components of duodenal calcium transport, namely, solvent drag-induced, transcellular active, and passive paracellular components, in rats using an in vitro Ussing chamber technique. The relative mRNA expression of genes related to duodenal calcium transport was also determined. We found that 21-day chronic metabolic acidosis stimulated solvent drag-induced and transcellular active duodenal calcium transport but not passive paracellular calcium transport. Our results further demonstrated that an acute direct exposure to serosal acidic pH, in contrast, decreased solvent drag-induced calcium transport in a pH-dependent fashion but had no effect on transcellular active calcium transport. Neither the transepithelial resistance nor duodenal permeability to Na⁺, Cl⁻, and Ca²⁺via the passive paracellular pathway were altered by chronic metabolic acidosis, suggesting that widening of the tight junction and changes in the charge-selective property of the tight junction did not occur. Thus the enhanced duodenal calcium transport observed in chronic metabolic acidosis could have resulted from a long-term adaptation, possibly at the molecular level. RT-PCR study revealed that chronic metabolic acidosis significantly increased the relative mRNA expression of duodenal genes associated with solvent drag-induced transport, i.e., the β₁-subunit of Na⁺-K⁺-ATPase, zonula occludens-1, occludin, and claudin-3, and with transcellular active transport, i.e., transient receptor potential vanilloid family Ca²⁺channels 5 and 6 and plasma membrane Ca²⁺-ATPase isoform 1b. Total plasma calcium and free ionized calcium and magnesium concentrations were also increased, whereas serum parathyroid hormone and 1α,25-dihydroxyvitamin D₃levels were not changed. The results indicated that 21-day chronic metabolic acidosis affected the calcium metabolism in rats partly through enhancing the mRNA expression of crucial duodenal genes involved in calcium absorption, thereby stimulating solvent drag-induced and transcellular active calcium transport in the duodenum.

Studying the molecular regulation of the thiazide-sensitive Na⁺-Cl⁻ cotransporter (NCC) is important for understanding how the kidney contributes to blood pressure regulation. Until now, a native mammalian cell model to investigate this transporter remained unknown. Our aim here is to establish, for the first time, a primary distal convoluted tubule (DCT) cell culture exhibiting transcellular thiazide-sensitive Na⁺ transport. Because parvalbumin (PV) is primarily expressed in the DCT, where it colocalizes with NCC, kidneys from mice expressing enhanced green-fluorescent protein (eGFP) under the PV gene promoter (PV-eGFP-mice) were employed. The Complex Object Parametric Analyzer and Sorter (COPAS) was used to sort fluorescent PV-positive tubules from these kidneys, which were then seeded onto permeable supports. After 6 days, DCT cell monolayers developed transepithelial resistance values of 630 ± 33 Ω·cm². The monolayers also established opposing transcellular concentration gradients of Na⁺ and K⁺. Radioactive ²²Na⁺ flux experiments showed a net apical-to-basolateral thiazide-sensitive Na⁺ transport across the monolayers. Both hypotonic low-chloride medium and 1 μM angiotensin II increased this ²²Na⁺ transport significantly by four times, which could be totally blocked by 100 μM hydrochlorothiazide. Angiotensin II-stimulated ²²Na⁺ transport was also inhibited by 1 μM losartan. Furthermore, NCC present in the DCT monolayers was detected by immunoblot and immunocytochemistry studies. In conclusion, a murine primary DCT culture was established which expresses functional thiazide-sensitive Na⁺-Cl⁻ transport.

A calcium-regulating hormone 1alpha,25-dihydroxyvitamin D(3) (1,25-[OH](2)D(3)) has been known to rapidly stimulate the transcellular active calcium transport in the chick duodenum. However, its effects on the solvent drag-induced paracellular calcium transport, which normally contributes approximately 70% of the total active calcium transport, and the underlying mechanism were unknown. The present study aimed to investigate the rapid nongenomic actions of physiological concentrations of 1,25-(OH)(2)D(3), i.e., 1, 10, and 100 nmol/l, on the duodenal calcium absorption in female rats. Quantitative real-time PCR revealed strong expressions of the classical vitamin D receptor (VDR) and the membrane-associated rapid response steroid binding receptors (MARRS) in both small and large intestines. By using the Ussing chamber technique, we found that duodenal epithelia acutely exposed to 10 and 100 nmol/l 1,25-(OH)(2)D(3) rapidly increased the solvent drag-induced calcium transport, but not the transcellular calcium transport, in a dose-response manner. On the other hand, 3-day daily injections of 1,25-(OH)(2)D(3) enhanced the transcellular active duodenal calcium transport. The 1,25-(OH)(2)D(3)-stimulated solvent drag-induced transport was abolished by the phosphatidylinositol 3-kinase (PI3K) inhibitors, 200 nmol/l wortmannin and 75 micromol/l LY294002, as well as PKC (1 micromol/l GF109203X) and MEK inhibitors (10 micromol/l U0126). Although 100 nmol/l 1,25-(OH)(2)D(3) did not alter the transepithelial mannitol flux, indicating no widening of the tight junction, it decreased the transepithelial resistance and increased both sodium and chloride permeability through the paracellular channel. We conclude that 1,25-(OH)(2)D(3) uses the nongenomic signaling pathways involving PI3K, PKC, and MEK to rapidly enhance the solvent drag-induced calcium transport, partly by altering the charge-selective property of the duodenal epithelium at least for the pathways involving PI3K and MEK.

Pigmented rice varieties are abundant in phenolic compounds. Antioxidant activity and bioaccessibility of phenolic compounds are modified in the gastrointestinal tract. After in vitro simulated digestion, changes in antioxidant activity and bioaccessibility of phenolic compounds (phenolic acids, flavonoids, and anthocyanins) in purple rice brans (Hom Nil and Riceberry) were compared with undigested crude extracts. The digestion method was conducted following the INFOGEST protocol. Antioxidant activity was determined using the ferric-reducing antioxidant power (FRAP) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity assays. The bioaccessibility index (BI) was calculated from the ratio of digested to undigested soluble phenolic content. Overall results showed that the in vitro simulated digested rice brans had lower antioxidant activity and lower total phenolic, flavonoid, and anthocyanin contents. However, the concentration of sinapic acid was stable, while other phenolic acids (gallic, protocatechuic, vanillic, ρ-coumaric, and ferulic acids) degraded after the oral, gastric, and intestinal phases. The BI of sinapic, gallic, vanillic, and ferulic acids remained stable, and the BI of quercetin was resistant to digestion. Conversely, anthocyanins degraded during the intestinal phase. In conclusion, selective phenolic compounds are lost along the gastrointestinal tract, suggesting that controlled food delivery is of further interest.