Association between a variation in LRCH1 and knee osteoarthritis: a genome-wide single-nucleotide polymorphism association study using DNA pooling.

OBJECTIVE: To perform a large-scale association analysis of single-nucleotide polymorphisms (SNPs) in patients with radiographically defined osteoarthritis (OA) of the knee. METHODS: We examined >25,000 SNPs located within approximately 14,000 genes for associations with radiographically defined knee OA, using polymerase chain reaction and MassExtend amplification techniques. Allele frequencies were estimated initially in DNA pools from 335 female patients with knee OA and 335 asymptomatic and radiographically negative female control subjects. All were of northern European ancestry. Significant allele frequency differences were validated by genotyping of individual DNA samples. Confirmed significant findings were verified in 2 additional case-control samples from the UK (443 cases and 303 controls) and Newfoundland (346 cases and 264 controls). Chondrosarcoma cell lines were used to test for potential differences in gene expression. RESULTS: The marker most strongly associated with the risk of knee OA was rs912428, a C/T polymorphism in intron 1 of LRCH1, a gene on chromosome 13q14 that encodes a novel protein of as-yet-unknown function. The frequency of the T allele compared with controls was consistently increased by 40% across all 3 case-control groups. Additional subanalyses in case-control samples with hip OA and hand OA suggested similar trends, but did not reach statistical significance. Association fine-mapping using 10 additional SNPs in LRCH1 confirmed intron 1 as the region of highest association but failed to reveal variations with significance stronger than the marker SNP, as did the haplotype analysis. LRCH1 was not up-regulated or overexpressed in chondrosarcoma cell lines exposed to inflammatory stimuli, suggesting a possible structural role. CONCLUSION: A genetic variant in LRCH1 was consistently associated with knee OA in 3 samples from 2 populations. Our results also suggest that the same association with OA may exist at other sites. Additional genetic and experimental work is needed to elucidate the precise mechanism by which the LRCH1 gene influences OA risk.

Linkage and potential association of obesity-related phenotypes with two genes on chromosome 12q24 in a female dizygous twin cohort.

Obesity is a multifactorial disorder with a complex phenotype. It is a significant risk factor for diabetes and hypertension. We assessed obesity-related traits in a large cohort of twins and performed a genome-wide linkage scan and positional candidate analysis to identify genes that play a role in regulating fat mass and distribution in women. Dizygous female twin pairs from 1,094 pedigrees were studied (mean age 47.0+/-11.5 years (range 18-79 years)). Nonparametric multipoint linkage analyses showed linkage for central fat mass to 12q24 (141 cM) with LOD 2.2 and body mass index to 8q11 (67 cM) with LOD 1.3, supporting previously established linkage data. Novel areas of suggestive linkage were for total fat percentage at 6q12 (LOD 2.4) and for total lean mass at 2q37 (LOD 2.4). Data from follow-up fine mapping in an expanded cohort of 1243 twin pairs reinforced the linkage for central fat mass to 12q24 (LOD 2.6; 143 cM) and narrowed the -1 LOD support interval to 22 cM. In all, 45 single-nucleotide polymorphisms (SNPs) from 26 positional candidate genes within the 12q24 interval were then tested for association in a cohort of 1102 twins. Single-point Monks-Kaplan analysis provided evidence of association between central fat mass and SNPs in two genes - PLA2G1B (P = 0.0067) and P2RX4 (P = 0.017). These data provide replication and refinement of the 12q24 obesity locus and suggest that genes involved in phospholipase and purinoreceptor pathways may regulate fat accumulation and distribution.

Effects of prenatal glucocorticoid exposure on cardiac calreticulin and calsequestrin protein expression during early development and in adulthood.

Overexpression of the conserved Ca(2+)-binding proteins calreticulin and calsequestrin impairs cardiac function, leading to premature death. Calreticulin is vital for embryonic development, but also impairs glucocorticoid action. Glucocorticoid overexposure during late fetal life causes intra-uterine growth retardation and programmed hypertension in adulthood. To determine whether intra-uterine growth retardation or programmed hypertension was associated with altered calreticulin or calsequestrin expression, effects of prenatal glucocorticoid overexposure (maternal dexamethasone treatment on days 15-21 of pregnancy) were examined during fetal life and postnatal development until adulthood (24 weeks). Dexamethasone (100 or 200 microg/kg of maternal body weight) was administered via osmotic pump. Calreticulin was detected as a 55 kDa band and calsequestrin as 55 and 63 kDa bands in 21 day fetal hearts. Only the 55 kDa calsequestrin band was detected postnatally. Prenatal glucocorticoid overexposure at the higher dose decreased calreticulin protein expression (26%; P <0.05) but increased calsequestrin protein expression, both 55 and 63 kDa bands, by 87% ( P <0.01) and 78% ( P <0.01); only the 55 kDa calsequestrin band was increased at the lower dose (66%; P <0.05). Offspring of dams treated at the lower dexamethasone dose were studied further. In control offspring, cardiac calreticulin protein expression declined between 2 and 3 weeks of age, and remained suppressed until adulthood. Cardiac calsequestrin protein expression increased 2-fold between fetal day 21 and postnatal day 1 and continued to increase until adulthood, at which time it was 3.4-fold higher ( P <0.001). Prenatal dexamethasone exposure minimally affected postnatal calsequestrin protein expression, but the postnatal decline in calreticulin protein expression was abrogated and calreticulin protein expression in adulthood was 2.2-fold increased ( P <0.001) compared with adult controls. In view of the known associations between cardiac calreticulin overexpression and impaired cardiac function, targeted up-regulation of calreticulin may contribute to the increased risk of adult heart disease introduced as a result of prenatal overexposure to glucocorticoids.