Novel Genetic Loci Control Calcium Absorption and Femur Bone Mass as Well as Their Response to Low Calcium Intake in Male BXD Recombinant Inbred Mice.

Low dietary calcium (Ca) intake during growth limits peak bone mass but physiological adaptation can prevent this adverse effect. To assess the genetic control on the physiologic response to dietary Ca restriction (RCR), we conducted a study in 51 BXD lines fed either 0.5% (basal) or 0.25% (low) Ca diets from ages 4 to 12 weeks (n = 8/line/diet). Ca absorption (CaAbs), femur bone mineral density (BMD), and bone mineral content (BMC) were examined. ANCOVA with body size as covariate was used to detect significant line and diet main effects, and line-by-diet interactions. Body size-corrected residuals were used for linkage mapping and to estimate heritability (h(2) ). Loci controlling the phenotypes were identified using composite interval mapping on each diet and for the RCR. h(2) of basal phenotypes (0.37-0.43) and their RCR (0.32-0.38) was moderate. For each phenotype, we identified multiple quantitative trait loci (QTL) on each diet and for the RCR. Several loci affected multiple traits: Chr 1 (88.3-90.6 cM, CaAbs, BMC), Chr 4 (45.8-49.2 cM, CaAbs, BMD, BMC), Chr 8 (28.6-31.6 cM, CaAbs, BMD, RCR), and Chr 15 (13.6-24 cM, BMD, BMC; 32.3-36 cM, CaAbs RCR, BMD). This suggests that gene clusters may regulate interdependent bone-related phenotypes. Using in silico expression QTL (eQTL) mapping and bioinformatic tools, we identified novel candidates for the regulation of bone under Ca stress (Ext1, Deptor), and for the first time, we report genes modulating Ca absorption (Inadl, Sc4mol, Sh3rf1, and Dennd3), and both Ca and bone metabolism (Tceanc2, Tll1, and Aadat). Our data reveal gene-by-diet interactions and the existence of novel relationships between bone and Ca metabolism during growth. © 2015 American Society for Bone and Mineral Research.

Gene-by-Diet Interactions Affect Serum 1,25-Dihydroxyvitamin D Levels in Male BXD Recombinant Inbred Mice.

1,25-Dihydroxyvitamin D (1,25[OH]2D) regulates calcium (Ca), phosphate, and bone metabolism. Serum 1,25(OH)2D levels are reduced by low vitamin D status and high fibroblast growth factor 23 (FGF23) levels and increased by low Ca intake and high PTH levels. Natural genetic variation controls serum 25-hydroxyvitamin D (25[OH]D) levels, but it is unclear how it controls serum 1,25(OH)2D or the response of serum 1,25(OH)2D levels to dietary Ca restriction (RCR). Male mice from 11 inbred lines and from 51 BXD recombinant inbred lines were fed diets with either 0.5% (basal) or 0.25% Ca from 4 to 12 weeks of age (n = 8 per line per diet). Significant variation among the lines was found in basal serum 1,25(OH)2D and in the RCR as well as basal serum 25(OH)D and FGF23 levels. 1,25(OH)2D was not correlated to 25(OH)D but was negatively correlated to FGF23 (r = -0.5). Narrow sense heritability of 1,25(OH)2D was 0.67 on the 0.5% Ca diet, 0.66 on the 0.25% Ca diet, and 0.59 for the RCR, indicating a strong genetic control of serum 1,25(OH)2D. Genetic mapping revealed many loci controlling 1,25(OH)2D (seven loci) and the RCR (three loci) as well as 25(OH)D (four loci) and FGF23 (two loci); a locus on chromosome 18 controlled both 1,25(OH)2D and FGF23. Candidate genes underlying loci include the following: Ets1 (1,25[OH]2D), Elac1 (FGF23 and 1,25[OH]2D), Tbc1d15 (RCR), Plekha8 and Lyplal1 (25[OH]D), and Trim35 (FGF23). This report is the first to reveal that serum 1,25(OH)2D levels are controlled by multiple genetic factors and that some of these genetic loci interact with the dietary environment.

Gene-by-diet interactions influence calcium absorption and bone density in mice.

Dietary calcium (Ca) intake is needed to attain peak bone mineral density (BMD). Habitual low Ca intake increases intestinal Ca absorption efficiency to protect bone mass, but the mechanism controlling, and the impact of genetics on, this adaptive response is not clear. We fed 11 genetically diverse inbred mouse lines a normal (0.5%) or low (0.25%) Ca diet from 4 to 12 weeks of age (n = 8 per diet per line) and studied the independent and interacting effects of diet and genetics on Ca and bone metabolism. Significant genetic variation was observed in all bone, renal, and intestinal phenotypes measured including Ca absorption. Also, adaptation of Ca absorption and bone parameters to low dietary Ca was significantly different among the lines. Ca absorption was positively correlated to femur BMD (r = 0.17, p = 0.02), and distal femur bone volume/tissue volume (BV/TV) (r = 0.34, p < 0.0001). Although Ca absorption was correlated to 1,25 dihydroxyvitamin D (1,25(OH)2 D) (r = 0.35, p < 0.0001), the adaptation of Ca absorption to low Ca intake did not correlate to diet-induced adaptation of 1,25(OH)2 D across the 11 lines. Several intestinal proteins have been proposed to mediate Ca absorption: claudins 2 and 12, voltage gated Ca channel v1.3 (Cav1.3), plasma membrane Ca ATPase 1b (PMCA1b), transient receptor potential vanilloid member 6 (TRPV6), and calbindin D9k (CaBPD9k). Only the mRNA levels for TRPV6, CaBPD9k, and PMCA1b were related to Ca absorption (r = 0.42, 0.43, and 0.21, respectively). However, a significant amount of the variation in Ca absorption is not explained by the current model and suggests that novel mechanisms remain to be determined. These observations lay the groundwork for discovery-focused initiatives to identify novel genetic factors controlling gene-by-diet interactions affecting Ca/bone metabolism.