Journal: JCI insight

Article Title: Activation of the calcium sensing receptor attenuates TRPV6-dependent intestinal calcium absorption.

doi: 10.1172/jci.insight.128013

Figure Lengend Snippet: Figure 1. Relative intestinal mRNA expression of transcellular Ca2+ transport mediators under altered extracellular Ca2+ conditions. (A–C) Relative mRNA expression of transcellular Ca2+ transport mediators TRPV6 (Trpv6), CABP9K (S100g), NCX1 (Slc8a1), or PMCA1b (Atp2b1), normalized to 18S rRNA expression in mice on high-, normal- (Con), or low-Ca2+ diet for 21 days (n = 7 for each diet). (D–F) Relative mRNA expression in animals treated with 1,25- [OH]2 D3 (VD) or vehicle (Veh) (n = 8 for each). (G–I) Relative mRNA expression in animals treated with cinacalcet (Cin) or control (Veh) diet (n = 8 for each). All data are presented as the mean ± SEM, normalized to the mice on the normal/control diet. Asterisks indicate a statistically significant difference from the normal/control mice by 1-way ANOVA (all genes in A and Slc8a1 and Atp2b1 in B and C), Brown-Forsythe test (S100g in B), Kruskal-Wallis test (Trpv6 in B and C), or Student’s unpaired t tests (D–I); *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: Capped RNA of human TRPV6 (accession number NM_018646, generated using in vitro transcription with mMESSAGE mMACHINE kit by Ambion) and human CaSR cDNA (Origene; catalog RC211229) were injected into Xenopus oocytes.

Techniques: Expressing, Control

Journal: JCI insight

Article Title: Activation of the calcium sensing receptor attenuates TRPV6-dependent intestinal calcium absorption.

doi: 10.1172/jci.insight.128013

Figure Lengend Snippet: Figure 6. Effect of phospholipase C inhibition on CaSR-mediated inhibition of Ca2+ absorption in in vitro and ex vivo. (A) Effect of CaSR activation on Ca2+-induced currents (ICa) in TRPV6 expressing oocytes in the presence and absence of cinacalcet and/or U73122, a phospholipase C (PLC) inhibitor (n = 6 each). Mean ICa values obtained from TRPV6- and CaSR-expressing oocytes were normalized to vehicle ICa values from TRPV6-expressing oocytes ± SEM; asterisks indi- cate a statistically significant difference between the conditions (multiple-comparisons Kruskal-Wallis test; *P < 0.05; **P < 0.01). (B) Effect of cinacalcet and/or U73122 on the plasma membrane expression of TRPV6 and CaSR in oocytes determined by immunoblot. As a loading control, β-actin was blotted (bottom). (C) Quantification of surface TRPV6 expression, normalized to total TRPV6 (n = 3 each). (D) Effect of basolateral cinacalcet (10 μM) and vehicle (DMSO) or PLC inhibitor U73122 (10 μM) on mouse proximal colon (n = 6 each). Raw values are presented; asterisks indicate a statistical difference between the conditions (Student’s paired t tests; *P < 0.05).

Article Snippet: Capped RNA of human TRPV6 (accession number NM_018646, generated using in vitro transcription with mMESSAGE mMACHINE kit by Ambion) and human CaSR cDNA (Origene; catalog RC211229) were injected into Xenopus oocytes.

Techniques: Inhibition, In Vitro, Ex Vivo, Activation Assay, Expressing, Clinical Proteomics, Membrane, Western Blot, Control

Journal: JCI insight

Article Title: Activation of the calcium sensing receptor attenuates TRPV6-dependent intestinal calcium absorption.

doi: 10.1172/jci.insight.128013

Figure Lengend Snippet: Figure 7. Proposed model of CaSR-mediated inhibition of Ca2+ absorption. (A) Increased plasma Ca2+ is sensed by the CaSR expressed in kidneys, bone, intestine, and parathyroid glands. In response, the kidneys decrease Ca2+ reabsorption, there is decreased bone resorption, and parathyroid hormone (PTH) secretion is decreased. We show herein that intestinal Ca2+ absorption is inhibited. In concert, this reduces plasma Ca2+. Importantly, PTH acts to increase plasma Ca2+ by increasing bone Ca2+ resorption, renal tubular reabsorption, and indirectly by increasing intestinal Ca2+ absorption by increasing 1,25-[OH]2 D3 synthesis in the kidneys. (B) High plasma Ca2+ is detected by the intestinal epithelial basolateral CaSR, which inhibits TRPV6-mediated transcellular Ca2+ transport via PLC.

Article Snippet: Capped RNA of human TRPV6 (accession number NM_018646, generated using in vitro transcription with mMESSAGE mMACHINE kit by Ambion) and human CaSR cDNA (Origene; catalog RC211229) were injected into Xenopus oocytes.

Techniques: Inhibition, Clinical Proteomics