Association of FXR gene variants with cholelithiasis.

BACKGROUND AND AIM: Impairment of bile acid homeostasis is the most important risk factor of gallstone disease. Thereby the bile acid sensor farnesoid X receptor (FXR) plays a pivotal role in hepatic and intestinal bile acid metabolism. In this explorative study, the FXR gene was investigated to identify gene variants, associated with gallstone formation in a Caucasian population. METHODS: Sequencing of the FXR gene was conducted in a randomly selected cohort of gallstone carriers (n=30) and control subjects (n=16) from Stuttgart, Germany. Genomic DNA was obtained from blood leukocytes. Genotype frequencies were established in the total cohort (controls: n=133, gallstone carriers: n=74). For expression analysis, total RNA and protein were isolated from ileal biopsies. RESULTS: The sequencing showed the sole appearance of 10 SNPs in gallstone carriers. Further genotype analysis revealed significant gender- and weight-dependent frequency differences of 3 SNPs between gallstone carriers and controls in males (rs35724: OR=4.73, P=0.022) and normal weight subjects (rs11110385: OR=3.67, P=0.027; rs11110386: OR=3.67, P=0.027) applying the 11+12<>22 allele model. Furthermore, rs11110385 carriers showed a significantly decreased FXR protein expression (11+12<>22: P=0.003). Significant mRNA expression differences between lean rs11110385 carriers and non-carriers were observed in FXR target genes (decrease: ILBP: P=0.042, OSTalpha: P=0.071, FGF19: P=0.011. Increase: LRH1: P=0.044). CONCLUSIONS: Three FXR gene variants (rs35724, rs11110385, rs11110386) were identified as potential susceptibility factors for cholelithiasis in a German cohort in gender- and weight-dependent manners. Thereby the tag SNP rs11110385 seemed to influence the activation of the FXR gene.

Role of the ABCG8 19H risk allele in cholesterol absorption and gallstone disease.

BACKGROUND: Gallstone disease is associated with p.D19H of ABCG8 as well as alterations of cholesterol and bile acid metabolism. However, molecular mechanisms have not been fully elucidated. It is important to understand the link between the sterol transporters ABCG5/8 and NPC1L1 and intestinal cholesterol absorption as well as de novo synthesis in gallstone patients stratified according to 19H risk allele. Moreover, the functional importance of the 19H variant on intestinal ABCG8 feature remains to be clarified. METHODS: Measurements of serum surrogate markers of cholesterol absorption (plant sterols: sitosterol, campesterol) and synthesis (cholesterol precursor: lathosterol) were carried out by gas chromatography/mass spectrometry (GC/MS). For expression studies, total RNA was isolated from 168 ileal biopsies of study participants with (34) and without gallstone disease (134). Messenger RNA was measured by LightCycler real-time PCR. Genomic DNA was obtained from blood leukocytes. Genotype frequencies of p.D19H were established using MALDI-TOF mass spectrometry. RESULTS: Compared to controls, cholesterol absorption but not synthesis in gallstone carriers was diminished by about 21% based on low serum sitosterol (P = 0.0269) and campesterol (P = 0.0231) to cholesterol ratios. D19H was found to be significantly associated with gallstones (odds ratio [OR] = 2.9, P = 0.0220, 95% confidence interval [CI]:1.22-6.89), particularly in the overweight cohort (OR = 3.2, P = 0.0430, 95% CI:1.07-9.26). Cholesterol absorption was about 24% lower in individuals carrying p.D19H compared to wild type (Psitosterol = 0.0080, Pcampesterol = 0.0206). Moreover, irrespective of phenotype, carriers of p.D19H displayed a significant lower absorption than carriers of the major allele. The most pronounced effect on cholesterol absorption ratio was observed for serum campesterol levels (wild type controls to mutated controls 28%, P = 0.0347 and wild type controls to gallstone carriers with 19H allele 37%, P = 0.0030). Notably, ABCG5/8 and NPC1L1 expression was similar in gallstone carriers and controls regardless of p.D19H presence. CONCLUSIONS: Both gallstone disease and p.D19H of ABCG8 are associated with diminished cholesterol absorption. However, p.D19H is not responsible for the differences in small intestinal sterol transporter expression.

MALDI-TOF mass spectrometry screening of cholelithiasis risk markers in the gene of HNF1alpha.

In recent years MALDI-TOF MS gained importance for high-throughput DNA analysis. In the present study this technique was used for the pathogenetic analysis of gallstone disease. The intestinal apical sodium-dependent bile acid transporter (ASBT) shows a genetic association with gallstone disease. ASBT has 3 binding sites in its 5'UTR for hepatocyte nuclear factor 1alpha (HNF1alpha). We hypothesized that genetic alterations in the HNF1alpha gene could influence ASBT expression. The gene HNF1alpha was sequenced in 46 Stuttgart random samples, composed of 16 controls and 30 gallstone patients. Subsequently, two independent cohorts (Stuttgart: 67 gallstones carriers, 109 controls, Leutkirch: 112 gallstone carriers, 99 controls) were screened by MALDI-TOF MS. The subjects were further divided by gender and weight. 24 known polymorphisms and two novel SNPs in the 3'UTR of HNF1alpha were detected (c.*220G>A and c.*1151G>A). After gender-specific sub-division of the pooled cohorts, 4 SNPs resulted in significant differences between male gallstone carriers and male controls (Stuttgart/Leutkirch: rs2255531 OR=2.78; p=0.006, rs1169288 OR=2.13; p=0.032, rs7310409 OR=2.34; p=0.025 and rs1169294 OR=2.13; p=0.031). Two novel variants in the 3'UTR of HNF1alpha were detected and four SNPs of HNF1alpha show a significant association to cholelithiasis in male gallstone patients. This article is part of a Special Section entitled: Understanding genome regulation and genetic diversity by mass spectrometry.

Hip pain from impingement and dysplasia in patients aged 20-50 years. Workup and role for reconstruction.

In the 20-50-year age group, hip pain usually indicates dysplasia. Chronic mechanical pain is the usual pattern, although acute pain caused by avulsion or degeneration of the labrum may occur. The morphological characteristics of the dysplastic hip should be evaluated, and the link between the dysplasia and the osteoarthritis should be confirmed. Three factors indicate a favorable prognosis: joint space preservation, age younger than 40 years, and correctable femoral and acetabular abnormalities. Reconstruction is highly desirable, as it delays the need for joint replacement by 20 years. After 15 years, good outcomes are seen in 87% of patients after shelf arthroplasty and 85% after femoral varus osteotomy with or without shelf arthroplasty. Chiari acetabular osteotomy can be performed in patients with osteoarthritis but is followed by prolonged limping. Periacetabular osteotomy should be reserved for patients with moderate dysplasia and no evidence of osteoarthritis. Shelf arthroplasty and femoral osteotomy require 5-8 months off work (compared to 5 months after hip replacement surgery) but subsequently permits a far more active lifestyle. Hip replacement, which is required 20 years or more after biologic reconstruction, carries the same prognosis as first-line hip replacement (good results in 80% of patients after 15 years). Acute sharp pain related to anterior hip derangement also occurs in primary femoroacetabular impingement (FAI). The most common pattern is cam impingement, which is due to a decrease in head-neck offset and manifests as pain during flexion and adduction of the hip. Cam impingement can be corrected by anterolateral osteoplasty, which is often performed arthroscopically. Pincer-type impingement is contact between the anterior acetabular rim and the femoral neck due to retroversion of the proximal acetabulum. The imaging study strategy is discussed. Coxometry, computed tomography, and arthrography can be used. Primary FAI, which occurs as a result of geometric abnormalities, should be distinguished from secondary impingement. Causes of secondary impingement include exaggerated lumbar lordosis with pelvic tilt and to hip osteophytosis (sports or posterior hip osteoarthritis). Osteoplasty is rarely appropriate in patients with secondary impingement. The features of acute anterior hip derangement are now better defined. They can be used to guide palliative treatment, which is effective, in the medium term at least. Experience acquired over the last two decades has established the efficacy of surgery for hip dysplasia.