Maik Brune

The CAG repeat polymorphism in the androgen receptor gene affects bone density and bone metabolism in healthy males

OBJECTIVE Bone metabolism and bone density (BD) are influenced by sex hormones. Testosterone (T) action is exerted through the androgen receptor (AR). We investigated the potential impact of the CAG repeat (CAGR) polymorphism within the AR gene on BD and bone metabolism in healthy younger males.

Monitoring bone density in hypogonadal men by quantitative phalangeal ultrasound

Monitoring bone density (BMD) in hypogonadal and testosterone (T) substituted men is a major component of andrological therapy and is performed by methods that are cost-intensive and use radiation, such as quantitative computer tomography (QCT). Therefore, we assessed the feasibility of a more practical and inexpensive approach through application of phalangeal quantitative ultrasound (pQUS; IGEA DBM BP Sonic 1200, Sensweiler, Germany) in a cross-sectional study of 521 men, aged 18-91 years (224 healthy controls, 156 newly diagnosed hypogonadal, and 141 T substituted men). The method was compared with QCT of the lumbar spine in the first 80 patients. We evaluated longitudinal changes of amplitude-dependent speed-of-sound (AdSoS) in 54 hypogonadal men from the beginning of T substitution. AdSoS decreased with age (p < 0.0001) and with declining total T concentration, with a four to fivefold larger reduction in AdSoS for each nanomole-per-liter decrement in total T in the hypogonadal range (<12 nmol/L) compared with the eugonadal range (p < 0.0001). AdSoS was higher in eugonadal and substituted men than in hypogonadal patients (p < 0.0001, by analysis of covariance [ANCOVA]). Substituted men <50 years of age showed lower AdSoS than eugonadal men (p = 0.004) and untreated men with secondary hypogonadism had lower values than men with primary hypogonadism (p = 0.005). Therapeutic effects were seen regardless of age, diagnosis, or T substitution modality. In the longitudinal approach, AdSoS increased from 1986 +/- 93 to 2035 +/- 77 m/sec over 237 +/- 57 days with the highest gain in those men with initially the lowest values (p < 0.0001, by ANCOVA for repeated measurements). In comparison to QCT, patients with a lumbar content of hydroxylapatite of <100 mg/cm(3) were reliably identified by pQUS (cutoff level 1965 m/sec, T score -3.5 based on eugonadal subjects; receiver operating characteristics: area under the curve [AUC] 0.94, sensitivity 94.1, specificity 92%, p < 0.0001), but specific values of lumbar BMD could not be predicted by pQUS. pQUS represents a feasible, sensitive, and inexpensive method for assessing bone tissue in hypogonadal men over the full age range and also for monitoring the effects of T substitution.

A Frequent PNPLA3 Variant Is a Sex Specific Disease Modifier in PSC Patients with Bile Duct Stenosis

Background & Aims Primary sclerosing cholangitis predominantly affects males and is an important indication for liver transplantation. The rs738409 variant (I148M) of the PNPLA3 gene is associated with alcoholic and non-alcoholic liver disease and we evaluated its impact on the disease course of PSC. Methods The I148M polymorphism was genotyped in 121 German PSC patients of a long-term prospective cohort and 347 Norwegian PSC patients. Results In the prospective German cohort, actuarial survival free of liver transplantation was significantly reduced for I148M carriers (p = 0.011) compared to wildtype patients. This effect was restricted to patients with severe disease, as defined by development of dominant stenosis (DS) requiring endoscopic intervention. DS patients showed markedly decreased survival (p = 0.004) when carrying the I148M variant (I148M: mean 13.8 years; 95% confidence interval: 11.6–16.0 vs. wildtype: mean 18.6 years; 95% confidence interval: 16.3–20.9) while there was no impact on survival in patients without a DS (p = 0.87). In line with previous observations of sex specific effects of the I148M polymorphism, the effect on survival was further restricted to male patients (mean survival 11.9 years; 95% confidence interval: 10.0–14.0 in I148M carriers vs. 18.8 years; 95% confidence interval: 16.2–21.5 in wildtype; p<0.001) while female patients were unaffected by the polymorphism (p = 0.65). These sex specific findings were validated in the Norwegian cohort (p = 0.013). Conclusions In male PSC patients with severe disease with bile duct stenosis requiring intervention, the common I148M variant of the PNPLA3 gene is a risk factor for reduced survival.

PNPLA3 in end-stage liver disease: Alcohol consumption, hepatocellular carcinoma development, and transplantation-free survival

The rs738409 variant (I148M) of the patatin‐like phospholipase domain‐containing protein 3 (PNPLA3) gene is associated with several liver malfunctions. Its impact on end‐stage liver disease has not been addressed yet.

Parathyroid hormone-related protein confers chemoresistance by blocking apoptosis signaling via death receptors and mitochondria

Parathyroid hormone (PTH) has a central role in the regulation of serum calcium and phosphate, whereas parathyroid hormone‐related peptide (PTHrP) has important developmental roles. In addition, PTHrP has been discovered as a causative agent of hypercalcemia of malignancy. PTHrP is also expressed in many tumors, and expression often correlates with unfavorable prognosis. We have investigated the effects of PTHrP on apoptosis signaling pathways initiated by DNA damaging chemotherapeutic drugs. Stimulation experiments of the CD95‐, the TNF‐R‐, and the TRAIL‐R‐death receptor systems in Saos human osteosarcoma cells revealed that PTHrP can block signaling via each of these death receptors. Furthermore, our findings demonstrate a link between PTHrP and the mitochondrial apoptosis pathway. PTHrP down‐regulates expression of pro‐apoptotic Bcl‐2 family members like Bax and PUMA and up‐regulates expression of antiapoptotic molecules like Bcl‐2 and Bcl‐xl. It is of clinical relevance that PTHrP and anticancer drugs show opposing interactions on death receptor‐triggered as well as on mitochondrial apoptosis pathways. In addition, PTHrP induces chemoresistance by interference with p53 family‐dependent apoptosis signaling pathways and p53‐mediated transactivation of apoptosis target genes. Inhibition of CD95‐ and Bax gene transactivation is a mechanism by which PTHrP reduced the apoptosis response and treatment sensitivity of tumor cells. Our data indicate that PTHrP inhibits major apoptosis signaling pathways by blocking signaling via p53, death receptors and mitochondria and, consequently, confers chemoresistance of cancer cells. Thus, beyond its importance in development and differentiation, we describe an important role for PTHrP in tumorigenesis.

TaqIA polymorphism in dopamine D2 receptor gene complicates weight maintenance in younger obese patients

OBJECTIVE The A1 allele of the TaqIA polymorphism in the dopamine D2 receptor gene (rs1800497) has been associated with obesity. However, the effect of the polymorphism on the success in weight loss and/or weight maintenance during weight-loss programs has not been evaluated thus far. METHODS The rs1800497 was genotyped in 202 (135 female, 67 male) severely obese individuals with an initial body mass index of 41.7 ± 0.5 kg/m² who participated in a weight-loss program consisting of a weight-loss phase with a formula diet (12 wk) and a weight-maintenance phase (40 wk). Measurements were collected at baseline, after the weight-loss phase, and at the end of the weight-maintenance phase at 1 y. RESULTS Genotyping revealed 4 A1A1, 67 A1A2, and 131 A2A2 genotype carriers. Of the 202 subjects in the program, 66.8% completed the program and 33.2% terminated prematurely. Neither the attrition rate (P = 0.44) nor the overall weight loss was influenced by the different genotypes (P = 0.96). However, younger A1⁺ participants (A1A1 and A1A2) had a higher body mass index at all time points (baseline, P = 0.04; after weight loss, P = 0.05; after weight maintenance, P = 0.02). They also showed less overall weight loss (P = 0.05), which derived mainly from a greater weight regain during the maintenance phase (P = 0.02). CONCLUSION In this program, younger A1⁺ participants exhibited problems in maintaining weight loss during a weight-loss program.

Depletion of the receptor for advanced glycation end products (RAGE) sensitizes towards apoptosis via p53 and p73 posttranslational regulation

The receptor for advanced glycation endproduct (RAGE) is involved in diabetic complications and chronic inflammation, conditions known to affect the sensitivity towards apoptosis. Here, we studied the effect of genetically depleting RAGE on the susceptibility towards apoptosis. In murine osteoblastic cells, RAGE knockout increased both spontaneous and induced apoptosis. Decreased levels of B-cell lymphoma 2 protein and increased intrinsic apoptosis were observed in Rage−/− cells. Furthermore, loss of RAGE increased expression of the death receptor CD95 (Fas, Apo-1), CD95-dependent caspase activation and extrinsic apoptosis, whereas NF-kB-p65 nuclear translocation was diminished. Importantly, depletion of RAGE reduced the ubiquitination and degradation of p53 and p73 and increased their nuclear translocation. The increase of p53 and p73 transactivational activity was essential for the RAGE-dependent regulation of apoptosis, because knockdown of p53 and p73 significantly decreased apoptosis in RAGE-deficient but not in RAGE-expressing cells. Thus, the RAGE-mediated posttranslational regulation of p53 and p73 orchestrates a sequence of events culminating in control of intrinsic and extrinsic apoptosis signaling pathways.

CD14 is associated with biliary stricture formation.

The pathogenesis of intrahepatic biliary stricture formation in patients with primary sclerosing cholangitis (PSC) or after liver transplantation (LTx) remains elusive. CD14 receptor signaling is a key mediator of the innate immune system; its common genetic variant is associated with alcoholic liver disease. PSC and LTx cohort patients and primary biliary cirrhosis (PBC) control patients were genotyped for the CD14 ‐260C>T (rs2569190) polymorphism, and genotypes were correlated with long‐term clinical outcome. Biliary tissue, bile, and whole blood of PSC patients and healthy controls were screened for markers of the innate immune system and bacterial infection. In 121 PSC patients, the CD14 ‐260C>T genotype was associated with development of dominant bile duct strictures (P = 0.02). In 365 LTx patients, TT carriers (4.1%) were protected against the formation of nonanastomotic biliary strictures versus CC/CT patients (12.6%; P = 0.01). Chemokine ligand 8 (P = 0.04) and chemokine receptor 6 (P = 0.004) were up‐regulated in biliary tissue of PSC patients with the TT versus the CC/CT genotype. Lipopolysaccharide whole‐blood stimulation resulted in a significant change in interleukin (IL)‐8 (P = 0.05) and IL‐12p40 levels (P = 0.04) in healthy control subjects carrying the TT genotype. TT PSC patients were protected against Gram‐negative bacterial biliary infection (TT: 0% vs. CC/CT: 22.5%; P = 0.02). Serum‐soluble CD14 levels correlated with the CD14 ‐260C>T genotype (P = 0.02), representing an independent risk indicator of survival in PSC patients (hazard ratio, 0.40; 95% confidence interval, 0.19‐0.86; P =0.01). Conclusions: The function of the innate immune response by CD14 is crucial during biliary infection and stricture formation. The benefits of CD14 signaling modification should be addressed in future studies. (Hepatology 2016;64:843‐852)

A common genetic variant of fucosyltransferase 2 correlates with serum carcinoembryonic antigen levels and affects cancer screening in patients with primary sclerosing cholangitis.

Background Primary sclerosing cholangitis (PSC) patients are at increased risk of biliary tract cancer, and carcinoembryonic antigen (CEA) serum levels might be used for screening. Objective To examine cancer screening with CEA in PSC patients and analyse how serum CEA levels are affected by genetic variants of fucosyltransferase (FUT) 2 and 3. Methods In a retrospective cohort analysis we evaluated CEA levels in 226 PSC patients, including 19 with biliary malignancy, and investigated how FUT2 and FUT3 SNPs affected CEA levels. Receiver-operating-characteristic (ROC) analysis was performed and cut-off values were determined based on Youden’s index. A control cohort contained 240 patients, including 28 with biliary malignancy. Results Median CEA concentration was lower in cancer-free patients (1.4 ng/mL) than in cancer patients (2.0 ng/mL, P = 0.014). ROC analysis revealed an area under the curve (AUC) of 0.671, the optimal cut-off was 3.2 ng/mL. The FUT2 variant rs601338 (G428A) correlated with CEA levels, and the effect was most prominent in a subgroup of patients genetically incapable of expressing CA19-9. The AUC improved if ROC analysis was performed separately for wild-type (AUC: 0.731) and homozygous mutant (AUC: 0.816) G428A. The influence of FUT2 on CEA was confirmed in the control cohort. Conclusions CEA is interesting for biliary-malignancy screening in PSC patients, especially in patients who do not express CA19-9. This is the first study to show that the combined use of CEA measurement and FUT genotyping is clinically beneficial and that it might enhance the early detection of biliary malignancy in clinical practice. This approach could also be effective when screening for other common gastrointestinal malignancies.

Effects of Increased Von Willebrand Factor Levels on Primary Hemostasis in Thrombocytopenic Patients with Liver Cirrhosis

In patients with liver cirrhosis procoagulant and anticoagulant changes occur simultaneously. During primary hemostasis, platelets adhere to subendothelial structures, via von Willebrand factor (vWF). We aimed to investigate the influence of vWF on primary hemostasis in patients with liver cirrhosis. Therefore we assessed in-vitro bleeding time as marker of primary hemostasis in cirrhotic patients, measuring the Platelet Function Analyzer (PFA-100) closure times with collagen and epinephrine (Col-Epi, upper limit of normal ≤165 s) or collagen and ADP (Col-ADP, upper limit of normal ≤118 s). If Col-Epi and Col-ADP were prolonged, the PFA-100 was considered to be pathological. Effects of vWF on primary hemostasis in thrombocytopenic patients were analyzed and plasma vWF levels were modified by adding recombinant vWF or anti-vWF antibody. Of the 72 included cirrhotic patients, 32 (44.4%) showed a pathological result for the PFA-100. They had mean closure times (± SD) of 180±62 s with Col-Epi and 160±70 s with Col-ADP. Multivariate analysis revealed that hematocrit (P = 0.027) and vWF-antigen levels (P = 0.010) are the predictors of a pathological PFA-100 test in cirrhotic patients. In 21.4% of cirrhotic patients with platelet count ≥150/nL and hematocrit ≥27.0%, pathological PFA-100 results were found. In thrombocytopenic (<150/nL) patients with cirrhosis, normal PFA-100 results were associated with higher vWF-antigen levels (462.3±235.9% vs. 338.7±151.6%, P = 0.021). These results were confirmed by multivariate analysis in these patients as well as by adding recombinant vWF or polyclonal anti-vWF antibody that significantly shortened or prolonged closure times, respectively. In conclusion, primary hemostasis is impaired in cirrhotic patients. The effect of reduced platelet count in cirrhotic patients can at least be partly compensated by increased vWF levels. Recombinant vWF could be an alternative to platelet transfusions in the future.