Active intestinal calcium transport in the absence of transient receptor potential vanilloid type 6 and calbindin-D9k

To study the role of the epithelial calcium channel transient receptor potential vanilloid type 6 (TRPV6) and the calcium-binding protein calbindin-D9k in intestinal calcium absorption, TRPV6 knockout (KO), calbindin-D9k KO, and TRPV6/calbindin-D(9k) double-KO (DKO) mice were generated. TRPV6 KO, calbindin-D9k KO, and TRPV6/calbindin-D9k DKO mice have serum calcium levels similar to those of wild-type (WT) mice ( approximately 10 mg Ca2+/dl). In the TRPV6 KO and the DKO mice, however, there is a 1.8-fold increase in serum PTH levels (P < 0.05 compared with WT). Active intestinal calcium transport was measured using the everted gut sac method. Under low dietary calcium conditions there was a 4.1-, 2.9-, and 3.9-fold increase in calcium transport in the duodenum of WT, TRPV6 KO, and calbindin-D9k KO mice, respectively (n = 8-22 per group; P > 0.1, WT vs. calbindin-D9k KO, and P < 0.05, WT vs. TRPV6 KO on the low-calcium diet). Duodenal calcium transport was increased 2.1-fold in the TRPV6/calbindin-D9k DKO mice fed the low-calcium diet (P < 0.05, WT vs. DKO). Active calcium transport was not stimulated by low dietary calcium in the ileum of the WT or KO mice. 1,25-Dihydroxyvitamin D3 administration to vitamin D-deficient null mutant and WT mice also resulted in a significant increase in duodenal calcium transport (1.4- to 2.0-fold, P < 0.05 compared with vitamin D-deficient mice). This study provides evidence for the first time using null mutant mice that significant active intestinal calcium transport occurs in the absence of TRPV6 and calbindin-D9k, thus challenging the dogma that TRPV6 and calbindin-D9k are essential for vitamin D-induced active intestinal calcium transport.

Effect of disodium monofluorophosphate, calcium and vitamin D supplementation on bone mineral density in patients chronically treated with glucocorticosteroids: a prospective, randomized, double-blind study

To study the effect of fluoride on bone mineral density (BMD) in patients treated chronically with glucocorticosteroids, 15 subjects (renal grafted, n = 12; skin disease, n = 1; broncho pulmonary disorder, n = 1; Crohn's disease, n = 1) were prospectively studied in a double-blinded manner and randomly allocated either to group 1 (n = 8) receiving 13.2 mg/day fluoride given as disodium monofluorophosphate (MFP) supplemented with calcium (1,000 mg/day) and 25-hydroxyvitamin D (calcifediol) (50 micrograms/day), or to group 2 (n = 7) receiving Cas+ calcifediol alone. An additional group of 14 renal transplant patients treated chronically with glucocorticosteroids but exempt of specific therapeutic intervention for bone disease was set up as historical controls. BMD was measured by dual-energy X-ray absorptiometry (DXA, Hologic QDR 1000) performed at months 0, 6 and 12 for groups 1 and 2 (lumbar spine, total upper femur, diaphysis and epiphysis of distal tibia), or 11-31 months apart with calculation of linear yearly changes for the historical cohort. Lumbar BMD tended to rise in groups 1 and 2, and to fall in group 3, the change reaching statistical significance (p < 0.05) in group 1, thus leading to a significant difference between groups 1 and 3 (p < 0.05). At upper femur, tibial diaphysis and tibial epiphysis, no significant change in BMD occurred in any of the groups. In conclusion, lumbar BMD rises more after a mild dosis of fluoride given as MFP and combined to calcium and calcifediol than on Ca+ calcifediol alone, without changes in BMD at the upper femur or distal tibia.

Bicyclo[3.2.1]amide-DNA: a chiral, non-chiroselective base-pairing system

The design, synthesis and base-pairing properties of bicyclo[3.2.1]amide-(bca)DNA, a novel phosphodiester based DNA analogue, is reported. This analogue consists of a conformationally constrained backbone entity which emulates a B-DNA geometry, to which the nucleobases were attached via an extended, acyclic amide linker. Homobasic adenine-containing bca-decamers form duplexes with complementary oligonucleotides containing the bca-, the DNA the RNA and, surprisingly, also the L-RNA backbone. UV- and CD-spectroscopic investigations revealed the duplexes with D- or L-complement to be of similar stability and enantiomorphic in structure. Bca-oligonucleotides containing all four bases form strictly antiparallel, left-handed complementary duplexes with itself and complementary DNA but not with RNA. Base-mismatch discrimination is comparable to that of DNA while the overall thermal stabilities of bca-oligonucleotide duplexes are inferior relative to that of DNA or RNA. A detailed molecular modeling study of left- and right-handed bca-DNA containing duplexes showed only minor changes in the backbone structure and revealed a structural switch around the base-linker unit to be responsible for the generation of enantiomorphic duplex structures. The obtained data are discussed with respect to the structural and energetic role of the ribofuranose entities in DNA and RNA association