Identification of a 1p21 independent functional variant for abdominal obesity.

OBJECTIVES: Aiming to uncover the genetic basis of abdominal obesity, we performed a genome-wide association study (GWAS) meta-analysis of trunk fat mass adjusted by trunk lean mass (TFMadj) and followed by a series of functional investigations. SUBJECTS: A total of 11,569 subjects from six samples were included into the GWAS meta-analysis. METHODS: Meta-analysis was performed by a weighted fixed-effects model. In silico replication analysis was performed in the UK-Biobank (UKB) sample (N = 331,093) and in the GIANT study (N up to 110,204). Cis-expression QTL (cis-eQTL) analysis, dual-luciferase reporter assay and electrophoresis mobility shift assay (EMSA) were conducted to examine the functional relevance of the identified SNPs. At last, differential gene expression analysis (DGEA) was performed. RESULTS: We identified an independent SNP rs12409479 at 1p21 (MAF = 0.07, p = 7.26 × 10-10), whose association was replicated by the analysis of TFM in the UKB sample (one-sided p = 3.39 × 10-3), and was cross-validated by the analyses of BMI (one-sided p = 0.03) and WHRadj (one-sided p = 0.04) in the GIANT study. Cis-eQTL analysis demonstrated that allele A at rs12409479 was positively associated with PTBP2 expression level in subcutaneous adipose tissue (N = 385, p = 4.15 × 10-3). Dual-luciferase reporter assay showed that the region repressed PTBP2 gene expression by downregulating PTBP2 promoter activity (p < 0.001), and allele A at rs12409479 induced higher luciferase activity than allele G did (p = 4.15 × 10-3). EMSA experiment implied that allele A was more capable of binding to unknown transcription factors than allele G. Lastly, DGEA showed that the level of PTBP2 expression was higher in individuals with obesity than in individuals without obesity (N = 20 and 11, p = 0.04 and 9.22 × 10-3), suggesting a regulatory role in obesity development. CONCLUSIONS: Taken together, we hypothesize a regulating path from rs12409479 to trunk fat mass development through its allelic specific regulation of PTBP2 gene expression, thus providing some novel insight into the genetic basis of abdominal obesity.

Joint Association Analysis Identified 18 New Loci for Bone Mineral Density.

Bone mineral density (BMD) at various skeletal sites have shared genetic determinants. In the present study, aiming to identify shared loci associated with BMD, we conducted a joint association study of a genomewide association study (GWAS) and a meta-analysis of BMD at different skeletal sites: (i) a single GWAS of heel BMD in 142,487 individuals from the UK Biobank, and (ii) a meta-analysis of 30 GWASs of total body (TB) BMD in 66,628 individuals from the Genetic Factors for Osteoporosis (GEFOS) Consortium. The genetic correlation coefficient of the two traits was estimated to be 0.57. We performed joint association analysis with a recently developed statistical method multi-trait analysis of GWAS (MTAG) to account for trait heterogeneity and sample overlap. The joint association analysis combining samples of up to 209,115 individuals identified 18 novel loci associated with BMD at the genomewide significance level (α = 5.0 × 10-8 ), explaining an additional 0.43% and 0.60% of heel-BMD and TB-BMD heritability, respectively. The vast majority of the identified lead SNPs or their proxies exerted local expression quantitative trait loci (cis-eQTL) activity. Credible risk variants, defined as those SNPs located within 500 kilobases (kb) of the lead SNP and with p values within two orders of magnitude of the lead SNP, were enriched in transcription factor binding sites (p = 3.58 × 10-4 ) and coding regions (p = 5.71 × 10-4 ). Fifty-six candidate genes were prioritized at these novel loci using multiple sources of information, including several genes being previously reported to play a role in bone biology but not reported in previous GWASs (PPARG, FBN2, DEF6, TNFRSF19, and NFE2L1). One newly identified gene, SCMH1, was shown to upregulate the expression of several bone biomarkers, including alkaline phosphatase (ALP), collagen type 1 (COL-I), osteocalcin (OCN), osteopontin (OPN), and runt-related transcription factor 2 (RUNX2), in mouse osteoblastic MC3T3-E1 cells, highlighting its regulatory role in bone formation. Our results may provide useful candidate genes for future functional investigations. © 2019 American Society for Bone and Mineral Research.

PEG-conjugated PAMAM dendrimers mediate efficient intramuscular gene expression.

Generations 5 and 6 (G5 and G6) poly(amidoamine) (PAMAM) dendrimers have been shown to be highly efficient nonviral carriers in in vitro gene delivery. However, their high toxicity and unsatisfied in vivo efficacy limit their applications. In this study, to improve their characteristics as gene delivery carriers, polyethylene glycol (PEG, molecular weight 5,000) was conjugated to G5 and G6 PAMAM dendrimers (PEG-PAMAM) at three different molar ratios of 4%, 8%, and 15% (PEG to surface amine per PAMAM dendrimer molecular). Compared with unconjugated PAMAM dendrimers, PEG conjugation significantly decreased the in vitro and in vivo cytotoxicities and hemolysis of G5 and G6 dendrimers, especially at higher PEG molar ratios. Among all of the PEG-PAMAM dendrimers, 8% PEG-conjugated G5 and G6 dendrimers (G5-8% PEG, G6-8% PEG) resulted in the most efficient muscular gene expression when polyplexes were injected intramuscularly to the quadriceps of neonatal mice. Consistent with the in vivo results, these two 8% PEG-conjugated PAMAM dendrimers could also mediate the highest in vitro transfection in 293A cells. Therefore, G5-8% PEG and G6-8% PEG possess a great potential for gene delivery both in vivo and in vitro.