Role of lipids in pathophysiology, diagnosis and therapy of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is an aggressive and widespread cancer. Patients with liver cirrhosis of different aetiologies are at a risk to develop HCC. It is important to know that in approximately 20% of cases primary liver tumors arise in a non-cirrhotic liver. Lipid metabolism is variable in patients with chronic liver diseases, and lipid metabolites involved therein do play a role in the development of HCC. Of note, lipid composition of carcinogenic tissues differs from non-affected liver tissues. High cholesterol and low ceramide levels in the tumors protect the cells from oxidative stress and apoptosis, and do also promote cell proliferation. So far, detailed characterization of the mechanisms by which lipids enable the development of HCC has received little attention. Evaluation of the complex roles of lipids in HCC is needed to better understand the pathophysiology of HCC, the later being of paramount importance for the development of urgently needed therapeutic interventions. Disturbed hepatic lipid homeostasis has systemic consequences and lipid species may emerge as promising biomarkers for early diagnosis of HCC. The challenge is to distinguish lipids specifically related to HCC from changes simply related to the underlying liver disease. This review article discusses aberrant lipid metabolism in patients with HCC.

Haploidentical CD3 or α/ß T-cell depleted HSCT in advanced stage sickle cell disease.

Despite significant improvements in the supportive care, sickle cell disease (SCD) leads to significant morbidity and mortality. Allogeneic hematopoietic stem cell transplantation (HSCT), the only curative option, is limited due to matched donor availability. This could be met with T-cell-depleted haploidentical HSCT. Twenty advanced-stage SCD patients, median age 15 years, and 9 patients, median age 14 years, were transplanted with CD3/CD19- or TCRαß/CD19-depleted grafts and from matched sibling donors (MSDs). The conditioning consisted of ATG, thiotepa, fludarabine, and treosulfan. The median follow-up in the T-haplo-HSCT and the MSD patients was 21 (9-62) and 25 (7-60) months, respectively. The OS in the T-haplo-HSCT and MSD was 90% and 100%, respectively. In the T-haplo-HSCT group, two patient succumbed to a CMV pneumonitis and a macrophage activation syndrome (MAS). One patient in the T-haplo-HSCT group requires renal replacement therapy because of BK virus nephritis. None developed grade III-IV acute GvHD. In the T-haplo-HSCT and in the MSD, 20% and 22%, respectively, developed a mild or moderate chronic GvHD. These results demonstrate the feasibility, safety, and efficacy of T-haplo-HSCT also for adult advanced stage SCD patients.

Chemerin Isoforms and Activity in Obesity.

Overweight and adiposity are risk factors for several diseases, like type 2 diabetes and cancer. White adipose tissue is a major source for adipokines, comprising a diverse group of proteins exerting various functions. Chemerin is one of these proteins whose systemic levels are increased in obesity. Chemerin is involved in different physiological and pathophysiological processes and it regulates adipogenesis, insulin sensitivity, and immune response, suggesting a vital role in metabolic health. The majority of serum chemerin is biologically inert. Different proteases are involved in the C-terminal processing of chemerin and generate diverse isoforms that vary in their activity. Distribution of chemerin variants was analyzed in adipose tissues and plasma of lean and obese humans and mice. The Tango bioassay, which is suitable to monitor the activation of the beta-arrestin 2 pathway, was used to determine the ex-vivo activation of chemerin receptors by systemic chemerin. Further, the expression of the chemerin receptors was analyzed in adipose tissue, liver, and skeletal muscle. Present investigations assume that increased systemic chemerin in human obesity is not accompanied by higher biologic activity. More research is needed to fully understand the pathways that control chemerin processing and chemerin signaling.

Adipokines in Liver Cirrhosis.

Liver fibrosis can progress to cirrhosis, which is considered a serious disease. The Child-Pugh score and the model of end-stage liver disease score have been established to assess residual liver function in patients with liver cirrhosis. The development of portal hypertension contributes to ascites, variceal bleeding and further complications in these patients. A transjugular intrahepatic portosystemic shunt (TIPS) is used to lower portal pressure, which represents a major improvement in the treatment of patients. Adipokines are proteins released from adipose tissue and modulate hepatic fibrogenesis. These proteins affect various biological processes that are involved in liver function, including angiogenesis, vasodilation, inflammation and deposition of extracellular matrix proteins. The best studied adipokines are adiponectin and leptin. Adiponectin protects against hepatic inflammation and fibrogenesis, and leptin functions as a profibrogenic factor. These and other adipokines are supposed to modulate disease severity in patients with liver cirrhosis. Consequently, circulating levels of these proteins have been analyzed to identify associations with parameters of hepatic function, portal hypertension and its associated complications in patients with liver cirrhosis. This review article briefly addresses the role of adipokines in hepatitis and liver fibrosis. Here, studies having analyzed these proteins in systemic blood in cirrhotic patients are listed to identify adipokines that are comparably changed in the different cohorts of patients with liver cirrhosis. Some studies measured these proteins in systemic, hepatic and portal vein blood or after TIPS to specify the tissues contributing to circulating levels of these proteins and the effect of portal hypertension, respectively.

Lung disease caused by ABCA3 mutations.

BACKGROUND: Knowledge about the clinical spectrum of lung disease caused by variations in the ATP binding cassette subfamily A member 3 (ABCA3) gene is limited. Here we describe genotype-phenotype correlations in a European cohort. METHODS: We retrospectively analysed baseline and outcome characteristics of 40 patients with two disease-causing ABCA3 mutations collected between 2001 and 2015. RESULTS: Of 22 homozygous (15 male) and 18 compound heterozygous patients (3 male), 37 presented with neonatal respiratory distress syndrome as term babies. At follow-up, two major phenotypes are documented: patients with (1) early lethal mutations subdivided into (1a) dying within the first 6 months or (1b) before the age of 5 years, and (2) patients with prolonged survival into childhood, adolescence or adulthood. Patients with null/null mutations predicting complete ABCA3 deficiency died within the 1st weeks to months of life, while those with null/other or other/other mutations had a more variable presentation and outcome. Treatment with exogenous surfactant, systemic steroids, hydroxychloroquine and whole lung lavages had apparent but many times transient effects in individual subjects. CONCLUSIONS: Overall long-term (>5 years) survival of subjects with two disease-causing ABCA3 mutations was <20%. Response to therapies needs to be ascertained in randomised controlled trials.

Hepatic chemerin mRNA expression is reduced in human nonalcoholic steatohepatitis.

BACKGROUND: Chemerin is associated with insulin resistance and is expressed in the liver. Nonalcoholic fatty liver disease (NAFLD) is related to impaired insulin sensitivity, but studies evaluating hepatic and serum chemerin in NAFLD resulted in discordant data. MATERIALS AND METHODS: Chemerin mRNA was determined in the liver tissue obtained from 33 controls and 76 NAFLD patients. Chemerin serum levels were measured in a different cohort of patients with ultrasound-diagnosed NAFLD and the respective controls. Hepatic stellate cells and hepatocytes were exposed to selected metabolites and nuclear receptor agonists to study the regulation of chemerin. Effect of recombinant chemerin on hepatocyte released proteins was analysed. RESULTS: Hepatic chemerin expression was not related to BMI, gender, type 2 diabetes and hypertension. Chemerin mRNA did not correlate with steatosis and was negatively associated with inflammation, fibrosis and nonalcoholic steatohepatitis (NASH) score. Patients with NASH had lower chemerin mRNA compared to those with borderline NASH and controls. Factors with a role in NASH mostly did not regulate chemerin in the liver cells. Of note, liver X receptor agonist reduced chemerin protein. Serum chemerin was not changed in NAFLD. Levels positively correlated with age, waist-to-hip ratio, systolic blood pressure, serum FGF21 and lipocalin 2, and negatively with transferrin saturation. Chemerin induced FGF21 in supernatants of primary human hepatocytes. Hepcidin, a major regulator of iron homoeostasis and lipocalin 2, were not regulated by chemerin. CONCLUSION: Chemerin mRNA is reduced in the liver of NASH patients, and liver X receptor seems to have a role herein.

Ceramide and polyunsaturated phospholipids are strongly reduced in human hepatocellular carcinoma.

Lipid composition affects membrane function, cell proliferation and cell death and is changed in cancer tissues. Hepatocellular carcinoma (HCC) is an aggressive cancer and this study aimed at a comprehensive characterization of hepatic and serum lipids in human HCC. Cholesteryl ester were higher in tumorous tissues (TT) compared to adjacent non-tumorous tissues (NT). Free cholesterol exerting cytotoxic effects was not changed. Phosphatidylethanolamine, -serine (PS) and -inositol but not phosphatidylcholine (PC) and lysophosphatidylcholine (LPC) were reduced in HCC tissues. Saturated species mostly increased and polyunsaturated species were diminished in all of these phospholipids. Ceramide (Cer) was markedly reduced in HCC tissues and higher levels of sphingomyelin suggest impaired sphingomyelinase activity as one of the underlying mechanisms. Importantly, ceramide in NT increased in HCC stage T3. Ceramide released from hepatocytes attracts immune cells and a positive association of the macrophage specific receptor CD163 with NT ceramide was identified. HCC associated lipid changes did not differ in patients suffering from type 2 diabetes. Protein levels of p53 were induced in TT and negatively correlated with Cer d18:1/16:0 and PS 36:1. Of the lipid species changed in HCC tissues only TT Cer d18:1/16:0, Cer d18:1/24:1, PC 38:6 and LPC 22:6 correlated with the respective serum levels. Our study demonstrates a considerably altered hepatic lipidome in HCC tissues. Ceramide was markedly reduced in HCC tissues, and therefore, raising ceramide levels specifically in the tumor represents a reasonable therapeutic approach for the treatment of this malignancy.

Tools to explore ABCA3 mutations causing interstitial lung disease.

BACKGROUND: Interstitial lung diseases (ILD) comprise disorders of mostly unknown cause. Among the few molecularly defined entities, mutations in the gene encoding the ATP-binding cassette (ABC), subfamily A, member 3 (ABCA3) lipid transporter represent the main cause of inherited surfactant dysfunction disorders, a subgroup of ILD. Whereas many cases are reported, specific methods to functionally define such mutations are rarely presented. MATERIALS AND METHODS: In this study, we exemplarily utilized a set of molecular tools to characterize the mutation K1388N, which had been identified in a patient suffering from ILD with lethal outcome. We also aimed to correlate in vitro and ex vivo findings. RESULTS: We found that presence of the K1388N mutation did not affect protein expression, but resulted in an altered protein processing and a functional impairment of ABCA3. This was demonstrated by decreased dipalmitoyl-phosphatidylcholine (PC 32:0) content and malformed lamellar bodies in cells transfected with the K1388N variant as compared to controls. CONCLUSIONS: Here we present a set of tools useful for categorizing different ABCA3 mutations according to their impact upon ABCA3 activity. Knowledge of the molecular defects and close correlation of in vitro and ex vivo data will allow us to define groups of mutations that can be targeted by small molecule correctors for restoring impaired ABCA3 transporter in the future. Pediatr Pulmonol. 2016;51:1284-1294. © 2016 Wiley Periodicals, Inc.

Surfactant proteins in pediatric interstitial lung disease.

BACKGROUND: Children's interstitial lung diseases (chILD) comprise a broad spectrum of diseases. Besides the genetically defined surfactant dysfunction disorders, most entities pathologically involve the alveolar surfactant region, possibly affecting the surfactant proteins SP-B and SP-C. Therefore, our objective was to determine the value of quantitation of SP-B and SP-C levels in bronchoalveolar lavage fluid (BALF) for the diagnosis of chILD. METHODS: Levels of SP-B and SP-C in BALF from 302 children with chILD and in controls were quantified using western blotting. In a subset, single-nucleotide polymorphisms (SNPs) in the SFTPC promoter were genotyped by direct sequencing. RESULTS: While a lack of dimeric SP-B was found only in the sole subject with hereditary SP-B deficiency, low or absent SP-C was observed not only in surfactant dysfunction disorders but also in patients with other diffuse parenchymal lung diseases pathogenetically related to the alveolar surfactant region. Genetic analysis of the SFTPC promoter showed association of a single SNP with SP-C level. CONCLUSION: SP-B levels may be used for screening for SP-B deficiency, while low SP-C levels may point out diseases caused by mutations in TTF1, SFTPC, ABCA3, and likely in other genes involved in surfactant metabolism that remain to be identified. We conclude that measurement of levels of SP-B and SP-C was useful for the differential diagnosis of chILD, and for the precise molecular diagnosis, sequencing of the genes is necessary.

Identification and genotype/phenotype correlation of mutations in a large German cohort with hearing loss.

The prevalence of hearing impairment is estimated as approximately 1 on 1,000 newborn children. To assess a higher mutation detection rate in individuals with hearing loss a three-step mutation screening program consisting of GJB2 in first line, then GJB1, GJB3 and GJB6 (second step) and if tested negative or heterozygote, testing of GJA1, GJB4, SLC26A4 and PJVK (third) was performed. Audiograms were derived from all patients to characterize audiological features of GJB2 mutations especially. In 59 patients (31.3%) of the 188 probands, the hearing impairment was due to GJB2 mutations, 45 (23.9%) of these being homozygous for 35delG mutation and 14 (7.4%) compound heterozygous for GJB2 mutations in the coding region of exon 2 whereas no significant sequence variation was found in exon 1. In 22 (11.7%) additional patients a single recessive mutation in GJB2, GJB3, GJB6 and SLC26A4 without a second mutation on the other allele was identified, making genetic counseling difficult. Our study showed significant difference in hearing loss degree in the patients with GJB2-mutations. Forty-five (45.5%) GJB2-cases were identified in 99 individuals diagnosed with severe to profound hearing loss, 14 (17.7%) GJB2-cases were identified in 79 individuals with moderate deafness whereas no clear GJB2 mutation was found in 10 patients with mild hearing loss (p < 0.001). Revealing a high variability of hearing levels in identical genotypes (even intrafamilial), a significant genotype-phenotype correlation could not be established. Based on the identified mutations spectrum and frequencies, speaking mostly of GJB2, a step by step screening for mutations can be devised and in addition may lead to a better stratification of patients for specific therapeutical approaches.

Ultrastructure of skin from Refsum disease with emphasis on epidermal lamellar bodies and stratum corneum barrier lipid organization.

Classic Refsum disease (RD) is a rare, autosomal recessively-inherited disorder of peroxisome metabolism due to a defect in the initial step in the alpha oxidation of phytanic acid (PA), a C16 saturated fatty acid with four methyl side groups, which accumulates in plasma and lipid enriched tissues (please see van den Brink and Wanders, Cell Mol Life Sci 63:1752-1765, 2006). It has been proposed that the disease complex in RD is in part due to the high affinity of phytanic acid for retinoid X receptors and peroxisome proliferator-activated receptors. Structurally, epidermal hyperplasia, increased numbers of cornified cell layers, presence of cells with lipid droplets in stratum basale and reduction of granular layer to a single layer have been reported by Blanchet-Bardon et al. (The ichthyoses, SP Medical & Scientific Books, New York, pp 65-69, 1978). However, lamellar body (LB) density and secretion were reportedly normal. We recently examined biopsies from four unrelated patients, using both OsO4 and RuO4 post-fixation to evaluate the barrier lipid structural organization. Although lamellar body density appeared normal, individual organelles often had distorted shape, or had non-lamellar domains interspersed with lamellar structures. Some of the organelles seemed to lack lamellar contents altogether, showing instead uniformly electron-dense contents. In addition, we also observed mitochondrial abnormalities in the nucleated epidermis. Stratum granulosum-stratum corneum junctions also showed co-existence of non-lamellar and lamellar domains, indicative of lipid phase separation. Also, partial detachment or complete absence of corneocyte lipid envelopes (CLE) was seen in the stratum corneum of all RD patients. In conclusion, abnormal LB contents, resulting in defective lamellar bilayers, as well as reduced CLEs, likely lead to impaired barrier function in RD.

Free fatty acids, lipopolysaccharide and IL-1α induce adipocyte manganese superoxide dismutase which is increased in visceral adipose tissues of obese rodents.

Excess fat storage in adipocytes is associated with increased generation of reactive oxygen species (ROS) and impaired activity of antioxidant mechanisms. Manganese superoxide dismutase (MnSOD) is a mitochondrial enzyme involved in detoxification of ROS, and objective of the current study is to analyze expression and regulation of MnSOD in obesity. MnSOD is increased in visceral but not subcutaneous fat depots of rodents kept on high fat diets (HFD) and ob/ob mice. MnSOD is elevated in visceral adipocytes of fat fed mice and exposure of differentiating 3T3-L1 cells to lipopolysaccharide, IL-1α, saturated, monounsaturated and polyunsaturated free fatty acids (FFA) upregulates its level. FFA do not alter cytochrome oxidase 4 arguing against overall induction of mitochondrial enzymes. Upregulation of MnSOD in fat loaded cells is not mediated by IL-6, TNF or sterol regulatory element binding protein 2 which are induced in these cells. MnSOD is similarly abundant in perirenal fat of Zucker diabetic rats and non-diabetic animals with similar body weight and glucose has no effect on MnSOD in 3T3-L1 cells. To evaluate whether MnSOD affects adipocyte fat storage, MnSOD was knocked-down in adipocytes for the last three days of differentiation and in mature adipocytes. Knock-down of MnSOD does neither alter lipid storage nor viability of these cells. Heme oxygenase-1 which is induced upon oxidative stress is not altered while antioxidative capacity of the cells is modestly reduced. Current data show that inflammation and excess triglyceride storage raise adipocyte MnSOD which is induced in epididymal adipocytes in obesity.

Low incidence of oncogenic EGFR, HRAS, and KRAS mutations in seborrheic keratosis.

Seborrheic keratosis (SK) represents a frequent epidermal skin tumor. Although lacking a malignant potential, these tumors reveal multiple oncogenic mutations. A previous study identified activating mutations in 89% of SK, particularly in FGFR3 and PIK3CA genes. The aim of this study was to identify further oncogenic mutations in human SK. Therefore, we screened for mutations in EGFR, FGFR2, PIK3R1, HRAS, KRAS, and NRAS genes using both Sanger sequencing of selected exons and a multiplex SNaPshot assay in 58 SK of 14 patients. We identified a somatic EGFR p.L858R mutation in 1 SK. Furthermore, the HRAS mutations p.G13R (2/58 SK) and p.Q61L (2/58 SK) were found. These mutations have not been described in human SK yet. In addition, 1 SK revealed the KRAS p.G12V mutation, which has already been reported in SK. No mutations were detected in FGFR2, PIK3R1, and NRAS genes. The results of this study suggest that activating mutations of EGFR, HRAS, and KRAS contribute to the pathogenesis of human SK, although at a lower frequency than FGFR3 and PIK3CA mutations. FGFR2, PIK3R1, and NRAS mutations obviously do not have a significant role in the development of SK.

Sterol regulatory element-binding protein 2 (SREBP2) activation after excess triglyceride storage induces chemerin in hypertrophic adipocytes.

Chemerin is an adipokine whose systemic concentration and adipose tissue expression is increased in obesity. Chemerin is highly abundant in adipocytes, yet the molecular mechanisms mediating its further induction in obesity have not been clarified. Adipocyte hypertrophy contributes to dysregulated adipokine synthesis, and we hypothesized that excess loading with free fatty acids (FFA) stimulates chemerin synthesis. Chemerin was expressed in mature adipocytes, and differentiation of 3T3-L1 cells in the presence of FFA further increased its level. TNF and IL-6 were induced by FFA, but concentrations were too low to up-regulate chemerin. Sterol regulatory element-binding protein 2 (SREBP2) was activated in these cells, indicative for cholesterol shortage. Suppression of cholesterol synthesis by lovastatin led to activation of SREBP2 and increased chemerin, and supplementation with mevalonate reversed this effect. Knockdown of SREBP2 reduced basal and FFA-induced chemerin. EMSA confirmed binding of 3T3-L1 adipocyte nuclear proteins to a SREBP site in the chemerin promotor. SREBP2 was activated and chemerin was induced in adipose tissue of mice fed a high-fat diet, and higher systemic levels seem to be derived from adipocytes. Lipopolysaccharide-mediated elevation of chemerin was similarly effective as induction by FFA, indicating that both mechanisms are equally important. Chemokine-like receptor 1 was not altered by the incubations mentioned above, and higher expression in fat of mice fed a high-fat diet may reflect increased number of adipose tissue-resident macrophages in obesity. In conclusion, the current data show that adipocyte hypertrophy and chronic inflammation are equally important in inducing chemerin synthesis.

Serum galectin-3 is elevated in obesity and negatively correlates with glycosylated hemoglobin in type 2 diabetes.

CONTEXT: Adipocytes synthesize galectin-3 whose deficiency protects from inflammation associated with metabolic diseases. We aimed to study circulating galectin-3 in obesity and type 2 diabetes (T2D). STUDY DESIGN: Galectin-3 was measured by ELISA in the serum of male normal-weight and overweight controls and T2D patients and in T2D patients of both sexes. Because visceral fat contributes to systemic inflammation, galectin-3 was analyzed in paired samples of human and rodent sc and visceral adipose tissue. Visceral adipose tissue adipokines are released to the portal vein, and galectin-3 was analyzed in portal, hepatic, and systemic venous serum (PVS, HVS, and SVS, respectively) of patients with liver cirrhosis and in patients who underwent surgery for nonhepatic diseases. The effect of metformin on adipocyte galectin-3 was analyzed by immunoblot. RESULTS: Circulating galectin-3 was similarly elevated in T2D and obesity compared with normal-weight individuals and revealed a body mass index-dependent positive correlation with leptin, resistin, IL-6, and age. In T2D patients, galectin-3 was increased in serum of patients with elevated C-reactive protein and negatively correlated with glycated hemoglobin. Metformin treatment was associated with lower systemic galectin-3. Reduced galectin-3 in metformin-incubated human adipocytes indicated that low galectin-3 may be a direct effect of this drug. Galectin-3 was higher in PVS compared with HVS and SVS, suggesting that the splanchnic region is a major site of galectin-3 synthesis. Low galectin-3 in HVS compared with PVS demonstrated hepatic removal. CONCLUSIONS: Systemic galectin-3 is elevated in obesity and negatively correlates with glycated hemoglobin in T2D patients, pointing to a modifying function of galectin-3 in human metabolic diseases.

Effects of the new adiponectin paralogous protein CTRP-3 and of LPS on cytokine release from monocytes of patients with type 2 diabetes mellitus.

AIMS/HYPOTHESIS: It was the aim to investigate the hypothesis that the new C1q/TNF-family member CTRP-3 (C1q/TNF-related protein-3) acts anti-inflammatory in human monocytes from healthy controls and patients with type 2 diabetes mellitus (T2D). METHODS: Monocytes were isolated from 20 healthy controls and 30 patients with T2D. IL-6 and TNF concentrations were measured by ELISA. CTRP-3 was expressed in insect cells and used for stimulation experiments. RESULTS: Basal IL-6 and TNF were not different in control and in T2D monocytes. LPS-stimulation (1 microg/ml) significantly (p<0.001) increased IL-6 and TNF in the supernatants of control and in T2D monocytes to a similar extent. CTRP-3 (1 microg/ml) significantly (p=0.03) inhibited LPS-induced IL-6 in control monocytes but not in T2D monocytes. TNF upon co-stimulation with LPS and CTRP-3 was significantly (p=0.012) lower in control than in T2D monocytes. LPS-induced TNF concentration was significantly and positively correlated with serum total cholesterol and LDL cholesterol in T2D patients. CONCLUSIONS: CTRP-3 inhibits LPS-induced IL-6 and TNF release. This anti-inflammatory effect is lost in T2D. Serum cholesterol concentration affects the pro-inflammatory potential of LPS to induce TNF release from T2D monocytes in the presence or absence of CTRP-3. CTRP-3 might partly account for the pro-inflammatory state in T2D.

Circulating levels of chemerin and adiponectin are higher in ulcerative colitis and chemerin is elevated in Crohn's disease.

BACKGROUND: Chemerin is an adipokine that stimulates chemotaxis of cells of the innate immune system. Inflammatory bowel disease (IBD) is linked to an impaired immune response and, therefore, we hypothesized that systemic chemerin may be altered in IBD patients. METHODS: Serum was collected from patients with Crohn's disease (CD, 230 patients), ulcerative colitis (UC, 80 patients), and healthy controls (HC, 80 probands). Chemerin and adiponectin, which has already been measured in the serum of similar cohorts by others, were determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: Chemerin was elevated in IBD compared to HC and was higher in male CD than UC patients. Female and male CD patients had lower adiponectin levels compared to UC, and adiponectin was lower in female CD patients compared to female HC. Adiponectin tended to be higher in female and male UC patients compared to HC and this difference became significant in the whole study group. Correlations with disease activity were only found in males. Here, chemerin was higher in CD patients on remission but was reduced in UC with nonactive disease. Adiponectin was higher in UC with inactive disease. Treatment with corticosteroids was linked to elevated adiponectin in male CD patients and higher chemerin in female UC patients. Unlike adiponectin, which was elevated in female serum in all cohorts, chemerin was only higher in female UC patients. CONCLUSIONS: These findings further indicate potential regulatory functions of adipokines in intestinal inflammation that are partly gender-dependent and that may even be associated with the distinct immunopathogenesis of UC and CD.

Adiponectin-stimulated CXCL8 release in primary human hepatocytes is regulated by ERK1/ERK2, p38 MAPK, NF-kappaB, and STAT3 signaling pathways.

Adiponectin is believed to exert hepatoprotective effects and induces CXCL8, a chemokine that functions as a survival factor, in vascular cells. In the current study, it is demonstrated that adiponectin also induces CXCL8 expression in primary human hepatocytes but not in hepatocellular carcinoma cell lines. Knock down of the adiponectin receptor (AdipoR) 1 or AdipoR2 by small-interfering RNA indicates that AdipoR1 is involved in adiponectin-stimulated CXCL8 release. Adiponectin activates nuclear factor (NF)-kappaB in primary hepatocytes and pharmacological inhibition of NF-kappaB, the p38 mitogen-activated protein kinase, and extracellular signal-regulated kinase (ERK) 1/ERK2 reduces adiponectin-mediated CXCL8 secretion. Furthermore, adiponectin also activates STAT3 involved in interleukin (IL)-6 and leptin-mediated CXCL8 induction in primary hepatocytes. Inhibition of JAK2 by AG-490 does not abolish adiponectin-stimulated CXCL8, indicating that this kinase is not involved. Pretreatment of primary cells with "STAT3 Inhibitor VI," however, elevates hepatocytic CXCL8 secretion, demonstrating that STAT3 is a negative regulator of CXCL8 in these cells. In accordance with this assumption, IL-6, a well-characterized activator of STAT3, reduces hepatocytic CXCL8. Therefore, adiponectin-stimulated induction of CXCL8 seems to be tightly controlled in primary human hepatocytes, whereas neither NF-kappaB, STAT3, nor CXCL8 are influenced in hepatocytic cell lines. CXCL8 is a survival factor, and its upregulation by adiponectin may contribute to the hepatoprotective effects of this adipokine.

Innate immunity and adipocyte function: ligand-specific activation of multiple Toll-like receptors modulates cytokine, adipokine, and chemokine secretion in adipocytes.

The aim of this study was to analyze Toll-like receptor (TLR) expression in preadipocytes and mature adipocytes and to investigate whether TLR ligands influence the release of cytokines, chemokines, and adipokines. Murine 3T3-L1 preadipocytes and mature adipocytes were used for stimulation experiments. The effects of lipopolysaccharide (LPS), flagellin, Poly (U), Poly (I:C), macrophage-activating lipopeptide-2 (MALP2), Pam3Cys, and CpG on the release of interleukin-6 (IL-6), resistin, and monocyte chemoattractant protein-1 (MCP-1) were determined by enzyme-linked immunosorbent assay (ELISA). Nuclear translocation and promoter binding of NFkappaB were analyzed by electrophoretic mobility shift assays. TLR expression was investigated by reverse-transcriptase (RT-PCR). All TLRs except TLR5 and TRL7 are expressed in the stromal vascular cell (SVC) fraction and in mature adipocytes of different fat stores. Whereas basal and LPS-induced IL-6 release is higher in preadipocytes, basal and LPS-induced MCP-1 release is higher in mature adipocytes. Mature adipocytes respond to corticosterone regarding MCP-1 and resistin release. The ligands for TLRs influence IL-6, MCP-1, and resistin release differentially. Some of these ligands induce nuclear translocation and promoter binding of NFkappaB. Besides TLR5, that is not expressed in mature adipocytes, all TLR family members are involved. There exists a functional TRL pathway in adipocytes that connects innate immunity with adipocyte function. As a consequence, the role of the adipose tissue in both immunity and metabolism has to be investigated in future studies. The results of this approach will help to explain the metabolic changes such as insulin resistance observed during infection and the immunological phenomena such as macrophage infiltration of adipose tissue seen in obesity.

KIT, PDGFRalpha and EGFR analysis in nephroblastoma.

Nephroblastoma prognosis has dramatically improved, but an unfavourable prognostic subgroup warrants development of novel therapeutic strategies. Selective KIT, PDGFRalpha and epidermal growth factor receptor (EGFR) tyrosine kinase inhibition evolved as powerful targeted therapy for gastrointestinal stromal tumours and non-small-cell lung cancer. To investigate a potential role for tyrosine kinase inhibition, we analyzed 209 nephroblastomas for immunohistochemical KIT and EGFR expression, 63 nephroblastomas for mutations in KIT exons 9, 11, 13, EGFR exons 18, 19, 20 and 21, and all 209 nephroblastomas for PDGFRalpha exons 12, 14 and 18. Twenty-two tumours (10.5%) expressed KIT, 31 (14.8%) EGFR, and 10 (4.8%) both KIT and EGFR, respectively. KIT expression was relatively more common among high-risk tumours (6/27; 22.3%) compared to low-/intermediate-risk tumours (26/181; 14.4%). Nine patients deceased, four of which had high-risk tumours with KIT expression in two of four and EGFR expression in one of four. There were no KIT, PDGFRalpha or EGFR mutations. Our results suggest no significant contribution of KIT, EGFR or PDGFRalpha mutations to nephroblastoma pathogenesis. Despite a trend towards association of immunohistochemical KIT and EGFR expression with poor outcome in high-risk nephroblastomas, statistical analysis did not yield significant correlations in this subgroup. Therefore, it remains open if KIT, PDGFRalpha or EGFR tyrosine kinase inhibition constitute a therapeutic target in nephroblastoma in the absence of KIT, PDGFRalpha or EGFR mutations.