Assessment of clinical outcomes of advanced hilar cholangiocarcinoma.

BACKGROUND: Low resectability and poor survival outcome are common for hilar cholangiocarcinoma (HCCA), especially in advanced stages. The present study was to assess the clinical outcome of advanced HCCA, focusing on therapeutic modalities, survival analysis and prognostic assessment. METHODS: Clinical data of 176 advanced HCCA patients who had been treated in our hospital between January 2013 and December 2015 were analyzed retrospectively. Prognostic effects of clinicopathological factors were explored by univariate and multivariate analysis. Survival predictors were evaluated by the receiver operating characteristic (ROC) curve. RESULTS: The 3-year overall survival rate was 13% for patients with advanced HCCA. Preoperative total bilirubin (P = 0.009), hepatic artery invasion (P = 0.014) and treatment modalities (P = 0.020) were independent prognostic factors on overall survival. A model combining these independent prognostic factors (area under ROC curve: 0.748; 95% CI: 0.678-0.811; sensitivity: 82.3%, specificity: 53.5%) was highly predictive of tumor death. After R0 resection, the 3-year overall survival was up to 38%. Preoperative total bilirubin was still an independent negative factor, but not for hepatic artery invasion. CONCLUSIONS: Surgery is still the best treatment for advanced HCCA. Preoperative biliary drainage should be performed in highly-jaundiced patients to improve survival. Prediction of survival is improved significantly by a model that incorporates preoperative total bilirubin, hepatic artery invasion and treatment modalities.

MIR196A2 rs11614913 C > T polymorphism correlates with an increased risk of hepatopulmonary syndrome in liver cirrhosis: a case-control study in China.

OBJECTIVE: This case-control study is designed to explore the relationship between microRNA-196a2 (MIR196A2) rs11614913 C > T polymorphism and the risk of hepatopulmonary syndrome (HPS) in liver cirrhosis. METHODS: From January 2013 to January 2015, 163 liver cirrhosis patients with HPS (case group), 264 liver cirrhosis patients without HPS (control group), and 195 healthy people (normal group) were selected. A DNA extraction kit was used to extract plasma DNA from peripheral blood. Polymerase chain reaction-restriction fragment length polymorphism was used to detect the allele and genotype frequencies of MIR196A2 C > T polymorphism. Real-time quantitative polymerase chain reaction was adopted to detect the relative expression of MIR196A. RESULTS: The frequencies of C allele in the case group were higher than those in the control and normal groups (all P < 0.05), whereas no significant difference was found between the control and normal groups, which indicated that MIR196A2 C > T polymorphism was closely associated with an increased risk of HPS in patients with liver cirrhosis. Compared with the normal group, the relative expression of MIR196A in the case group was significantly increased (P < 0.05), but there was no significant difference in the control group (P > 0.05). In the case group, compared with patients carrying the TT genotype, the relative expression of MIR196A of patients carrying the C allele (CT + CC) evidently increased (P < 0.05). CONCLUSIONS: The MIR196A2 rs11614913 C > T polymorphism may contribute to an increased risk of HPS in liver cirrhosis patients.

Substrate uptake and subcellular compartmentation of anoxic cholesterol catabolism in Sterolibacterium denitrificans.

Cholesterol catabolism by actinobacteria has been extensively studied. In contrast, the uptake and catabolism of cholesterol by Gram-negative species are poorly understood. Here, we investigated microbial cholesterol catabolism at the subcellular level. (13)C metabolomic analysis revealed that anaerobically grown Sterolibacterium denitrificans, a ß-proteobacterium, adopts an oxygenase-independent pathway to degrade cholesterol. S. denitrificans cells did not produce biosurfactants upon growth on cholesterol and exhibited high cell surface hydrophobicity. Moreover, S. denitrificans did not produce extracellular catabolic enzymes to transform cholesterol. Accordingly, S. denitrificans accessed cholesterol by direction adhesion. Cholesterol is imported through the outer membrane via a putative FadL-like transport system, which is induced by neutral sterols. The outer membrane steroid transporter is able to selectively import various C27 sterols into the periplasm. S. denitrificans spheroplasts exhibited a significantly higher efficiency in cholest-4-en-3-one-26-oic acid uptake than in cholesterol uptake. We separated S. denitrificans proteins into four fractions, namely the outer membrane, periplasm, inner membrane, and cytoplasm, and we observed the individual catabolic reactions within them. Our data indicated that, in the periplasm, various periplasmic and peripheral membrane enzymes transform cholesterol into cholest-4-en-3-one-26-oic acid. The C27 acidic steroid is then transported into the cytoplasm, in which side-chain degradation and the subsequent sterane cleavage occur. This study sheds light into microbial cholesterol metabolism under anoxic conditions.

Surgical treatment of an old avulsion fracture of the ischial tuberosity and ischial ramus: A case report.

BACKGROUND: Avulsion fracture of the ischial tuberosity is a relatively clinically rare type of trauma that is mainly incurred by adolescents during competitive sports activities. According to previous literature, the most commonly involved sports are soccer, sprinting, and gymnastics, in descending order. Dance-induced avulsion fracture of the ischial tuberosity and ischial ramus is extremely clinically rare. CASE SUMMARY: A case of a neglected avulsion fracture of the ischial tuberosity and ischial ramus was diagnosed in a young female dancer who complained of pain and restricted movement of her right hip. She stated that she had suffered the injury while performing a split leap during a dance performance 9 mo prior. Eventually, she underwent surgery and obtained satisfactory treatment results. CONCLUSION: Early diagnosis of these fractures is important to ensuring early proper treatment towards a quicker recovery. For old fractures with nonunion and chronic buttock pain, surgery is a preferred therapeutic choice with good treatment outcomes.

Risk factors and incidence of acute pyogenic cholangitis.

BACKGROUND: Acute cholangitis varies from mild to severe form. Acute suppurative cholangitis (ASC), the severe form of acute cholangitis, is a fatal disease and requires urgent biliary decompression. Which patients are at a high risk of ASC and need emergency drainage is still unclear. The present study aimed to identify the factors for determining early-stage ASC and distinguishing ASC from acute cholangitis. METHODS: We analyzed 359 consecutive patients with acute cholangitis who had been admitted to the First Affiliated Hospital, Zhejiang University School of Medicine from January 2004 to May 2011. Emergency endoscopic retrograde cholangiopancreatography (ERCP) was carried out in all patients to decompress or clear the stones by experienced endoscopists. Clinical and therapeutic data were collected, and univariate and multivariate analyses were performed to identify the potential risk factors of ASC. RESULTS: Of the 359 patients, 1 was excluded because of failure of ERCP drainage. Of the remaining 358 patients with an average age of 62.7 years (range 17-90), 162 were diagnosed with ASC, and 196 with non-ASC. ENBD catheters were placed in 343 patients (95.8%), of whom 182 patients had stones removed at the same time, and plastic stent was placed in 25 patients (7.0%). Clinical conditions were improved quickly after emergency biliary drainage in all patients. Complications were identified in 11 patients (3.1%): mild pancreatitis occurred in 8 patients and hemorrhage in 3 patients. There was no mortality. Univariate analysis showed that several variables were associated with ASC: age, fever, decreased urine output, hypotension, tachycardia, abnormal white blood cell count (WBC), low platelet, high C reactive protein (CRP), and duration of the disease. Multivariate analysis revealed that advanced age, hypotension, abnormal WBC, high CRP, and duration of the disease were independent risk factors for ASC. CONCLUSIONS: This study demonstrates that advanced age, hypotension, abnormal WBC, high CRP, and long duration of antibiotic therapy are significantly associated with ASC. We recommend decompression by ERCP should be carried out in patients as early as possible.

Isolated cryptococcal osteomyelitis of the ulna in an immunocompetent patient: A case report.

BACKGROUND: Cryptococcal osteomyelitis is a bone infection caused by cryptococcus. As an opportunistic infection, bone cryptococcosis usually occurs in patients with immunodeficiency diseases or in those undergoing immunosuppressive therapy and often displays characteristics of disseminated disease. Isolated cryptococcal osteomyelitis is extremely unusual in immunocompetent person. The pathogenic fungus often invades vertebrae, femur, tibia, rib, clavicle, pelvis, and humerus, but the ulna is a rare target. CASE SUMMARY: A 79-year-old woman complaining of chronic pain, skin ulceration and a sinus on her right forearm was admitted, and soon after was diagnosed with cryptococcal osteomyelitis in the right ulna. Unexpectedly, she was also found to have apparently normal immunity. After treatment with antifungal therapy combined with surgery debridement, the patient's osteomyelitis healed with a satisfactory outcome. CONCLUSION: Although rare, cryptococcal osteomyelitis should be considered in the differential diagnosis of osteolytic lesions even in immunocompetent patients, and good outcomes can be expected if early definitive diagnosis and etiological treatment are established.

No therapeutic ERCP in anastomotic stricture without intrahepatic biliary dilation after liver transplantation.

BACKGROUND/AIMS: Differentiating surgical jaundice from non-surgical jaundice is of vital importance after liver transplantation (LT) and endoscopic retrograde cholangiopancreatography (ERCP) is not effective for all anastomotic stricture (AS) cases. In the present study, we aim to determine the optimal indication of ERCP treatment for AS after LT. METHODOLOGY: Twenty-eight jaundice patients who underwent successful ERCP treatments for post-transplant AS were classified into two groups: AS with intrahepatic biliary dilation (group 1, n=22) and AS without intrahepatic biliary dilation (group 2, n=6). The outcomes of the two groups were evaluated. RESULTS: The median time intervals from LT to the occurrence of AS were 38 days and 434 days for group 1 and group 2, respectively. The median total bilirubin significantly decreased from 142umol/L to 49umol/L (p<0.05) two weeks after ERCP treatment in group 1. Fourteen patients (63.6%) were cured and for the other 8 the treatment proved effective in group 1. But total bilirubin was not improved after the ERCP treatment in group 2 (p>0.05). CONCLUSIONS: Therapeutic ERCP is not effective in AS without intrahepatic biliary dilation after LT.

Effects of HGF gene polymorphisms and protein expression on transhepatic arterial chemotherapeutic embolism efficacy and prognosis in patients with primary liver cancer.

OBJECTIVE: To investigate the correlations of two hepatocyte growth factor (HGF) gene polymorphisms (rs5745652 and rs2074725) and their protein expression levels with the efficacy of transhepatic arterial chemotherapeutic embolism (TACE) and prognosis in patients with primary liver cancer (PLC). METHODS: From March 2011 to June 2012, 109 PLC patients (the case group) who chose TACE as primary treatment and 80 healthy people (the control group) who had undergone physical examination in The First Affiliated Hospital, Zhejiang University were selected during the same period. Gene polymorphisms of HGF rs5745652 and HGF rs2074725 were detected. Serum HGF level, treating efficacy, survival quality, and 3-year survival rate for PLC patients who received TACE were observed. RESULTS: There were significant differences in genotype and allele frequencies of HGF rs5745652 and HGF rs2074725, between the case and control groups (all P<0.05). Compared with CT+TT genotype of HGF rs5745652, patients carrying CC genotype had lower serum HGF levels, higher efficacy, better survival quality, and prolonged 3-year survival rate (all P<0.05). In rs2074725, patients carrying CA+AA genotype had lower serum HGF levels, higher efficacy, better survival quality, and prolonged 3-year survival rate compared with patients carrying rs2074725 CC genotype (all P<0.05). Gene polymorphisms of HGF rs5745652 and HGF rs2074725, tumor size, and Barcelona Clinic Liver Cancer stage were independent prognostic factors for PLC (P<0.05). CONCLUSION: Our results indicated that HGF gene polymorphisms affect TACE efficacy and survival quality of PLC patients. Patients with HGF CC genotype of rs5745652 and CA+AA genotype of rs2074725 had decreased HGF level, better curative effect, high survival quality, and a good prognosis after TACE treatment.

Expression of FOXO6 is Associated With Oxidative Stress Level and Predicts the Prognosis in Hepatocellular Cancer: A Comparative Study.

The aim of this study was to explore the association of Forkhead box O6 (FOXO6) expression with oxidative stress level and prognosis of hepatocellular cancer (HCC).The case group included tissues of HCC from 128 patients who were hospitalized in Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery of First Affiliated Hospital, School of Medicine, Zhejiang University. The control group included normal liver tissues from 74 patients. RT-PCR and Western blot were used to test expressions of FOXO6, heme oxygenase (HO)-1, glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT). Dihydroethidium (DHE) was dyed to observe reactive oxygen species (ROS) level. Immunohistochemistry was used to test FOXO6 expression. FOXO6 was silenced in HepG2 cells to detect cell proliferation and apoptosis. The expressions of ROS, HO-1, GPx, SOD, CAT, p27, and cyclin D1 were also detected to further explore the possible mechanism.The expressions of FOXO6, HO-1, GPx, SOD, and CAT in HCC tissue was significantly higher than those in normal and adjacent HCC tissues (P <0.05). The tumor size, TNM stage, Alpha-fetoprotein (AFP) level, the presence or absence of hepatitis B surface antigen (HbsAg), and differentiation degree were related to FOXO6 expression level (all P <0.05). COX analysis showed that high FOXO6 expression, male, positive HBsAg, advanced TNM staging, high expression of AFP, and low degree of differentiation were all risk factors for prognosis in HCC (P <0.05). Compared with the blank group (C group, without transfection) and the negative control (NC) group, the mRNA expressions of ROS, FOXO6, HO-1, SOD, GPx, and CAT were decreased (P <0.05). si-RNA group had significantly decreased proliferation speed during 24 to 72 hours (P <0.05), whereas si-FOXO6 group had remarkably increased G0/G1 staged cells and decreased S-staged cells (P <0.05). The si-FOXO6 group showed notably increased apoptosis rate (P <0.05) and p27 expressions as well as decreased cyclin D1 expressions (P <0.05).FOXO6 was highly expressed in HCC tissue and was related to oxidative stress levels. Furthermore, FOXO6 expression can be used as a biomarker for deterioration and prognosis of liver cancer, which may provide a novel treatment target for HCC therapy.

Identification of Comamonas testosteroni as an androgen degrader in sewage.

Numerous studies have reported the masculinization of freshwater wildlife exposed to androgens in polluted rivers. Microbial degradation is a crucial mechanism for eliminating steroid hormones from contaminated ecosystems. The aerobic degradation of testosterone was observed in various bacterial isolates. However, the ecophysiological relevance of androgen-degrading microorganisms in the environment is unclear. Here, we investigated the biochemical mechanisms and corresponding microorganisms of androgen degradation in aerobic sewage. Sewage samples collected from the Dihua Sewage Treatment Plant (Taipei, Taiwan) were aerobically incubated with testosterone (1 mM). Androgen metabolite analysis revealed that bacteria adopt the 9, 10-seco pathway to degrade testosterone. A metagenomic analysis indicated the apparent enrichment of Comamonas spp. (mainly C. testosteroni) and Pseudomonas spp. in sewage incubated with testosterone. We used the degenerate primers derived from the meta-cleavage dioxygenase gene (tesB) of various proteobacteria to track this essential catabolic gene in the sewage. The amplified sequences showed the highest similarity (87-96%) to tesB of C. testosteroni. Using quantitative PCR, we detected a remarkable increase of the 16S rRNA and catabolic genes of C. testosteroni in the testosterone-treated sewage. Together, our data suggest that C. testosteroni, the model microorganism for aerobic testosterone degradation, plays a role in androgen biodegradation in aerobic sewage.

Integrated multi-omics analyses reveal the biochemical mechanisms and phylogenetic relevance of anaerobic androgen biodegradation in the environment.

Steroid hormones, such as androgens, are common surface-water contaminants. However, literature on the ecophysiological relevance of steroid-degrading organisms in the environment, particularly in anoxic ecosystems, is extremely limited. We previously reported that Steroidobacter denitrificans anaerobically degrades androgens through the 2,3-seco pathway. In this study, the genome of Sdo. denitrificans was completely sequenced. Transcriptomic data revealed gene clusters that were distinctly expressed during anaerobic growth on testosterone. We isolated and characterized the bifunctional 1-testosterone hydratase/dehydrogenase, which is essential for anaerobic degradation of steroid A-ring. Because of apparent substrate preference of this molybdoenzyme, corresponding genes, along with the signature metabolites of the 2,3-seco pathway, were used as biomarkers to investigate androgen biodegradation in the largest sewage treatment plant in Taipei, Taiwan. Androgen metabolite analysis indicated that denitrifying bacteria in anoxic sewage use the 2,3-seco pathway to degrade androgens. Metagenomic analysis and PCR-based functional assays showed androgen degradation in anoxic sewage by Thauera spp. through the action of 1-testosterone hydratase/dehydrogenase. Our integrative 'omics' approach can be used for culture-independent investigations of the microbial degradation of structurally complex compounds where isotope-labeled substrates are not easily available.

Toll-Interacting Protein Suppresses HIV-1 Long-Terminal-Repeat-Driven Gene Expression and Silences the Post-Integrational Transcription of Viral Proviral DNA.

Toll-interacting protein (Tollip) is a host adaptor protein for negatively regulating Toll-like receptor 2-, 4-, and IL-1R (interleukin-1 receptor)-mediated signaling. We found that Tollip expression could be induced in MDDCs (monocyte-derived dendritic cells) by HIV-1 particles and recombinant gp120 glycoprotein. Hence, we investigated the role of Tollip in modulating HIV-1 infection. We found that Tollip expression suppressed NF-κB-dependent HIV-1 long terminal repeat (LTR)-driven transcription and thus inhibited HIV-1 infection. Our protein truncation experiments proved that the intact C-terminus of Tollip was required for inhibition of both NF-κB activity and HIV-1 LTR-driven gene expression. Intriguingly, Tollip silenced the post-integrational transcription of HIV-1 proviral DNA, indicating the potential role of Tollip in maintaining viral persistence. Our results reveal the novel role of host factor Tollip in modulating HIV-1 infection, and may suggest the hijacking of Tollip as the negative regulator of the TLR pathway and even the downstream signaling, by HIV-1 for maintaining persistent infection. Further elucidation of the mechanisms by which HIV-1 induces Tollip expression and identification of the role of Tollip in modulating HIV-1 latency will facilitate the understanding of host regulation in viral replication and benefit the exploration of novel strategies for combating HIV-1 infection.

A modified rat model for hepatocellular carcinoma.

BACKGROUND: Rat hepatocellular carcinoma (HCC) model which has a high analogy to clinical liver cancer is of great value in understanding the pathogenesis and evolution of liver cancer, in searching effective anti-cancer treatments (drug, hepatectomy and liver transplantation), and designing cancer prevention strategies. In this study we established a modified rat model of hepatocellular carcinoma to enhance rats' physique and surgical endurance. METHODS: Wistar rats were fed with diethylnitrosamine (DENA) by three methods for evaluation of general conditions for 130 days: Doppler ultrasonographic measurement, laparotomy and histopathological examination. RESULTS: No rat died in control group (group A) and modified DENA-induction-HCC group(group C), but 6 deaths in classical DENA-induction-HCC group (group B) (survival rate 80%). All survived rats in groups B and C developed diffusive hepatocellular carcinoma and liver cirrhosis. General appearance of rats in the group C was better than that in the group B. CONCLUSION: With good general conditions for surgery, the modified rat model for hepatocellular carcinoma has a high carcinogenic rate and a high survival rate.

[Influence of combined cyclosporine A and tacrolimus with 5-fluorouracil on hepatocellular carcinoma rats].

OBJECTIVE: To investigate the effect of combined CsA and FK506 with 5-FU on hepatocellular carcinoma rats. METHODS: A syngeneic rat model of hepatocellular carcinoma was used. Control group (A) underwent 4 ml 5% GS. Treatment group was divided into 3 groups namely, group B: only 5-FU and 5% GS; group C: 5-FU, CsA and 5% GS; group D: 5-FU, FK506 and 5%GS. Cell cycle, apoptosis, necrosis and mitochondrial transmembrane potential were measured by flow cytometry, laser scanning confocal microscopy, and electron transmission microscopy. Statistical analysis was performed by SPSS 10.0 for Windows software. Statistical comparisons were made with ANOVA followed by Dunnett's T3 or LSD test. RESULTS: Compared to the control group, the percentage of apoptotic cells including trifle necrotic cells was significantly higher, and among the treatment group, group D was the highest, and group C was higher than group B. In the treatment group, cell cycle of hepatoma cells was mainly arrested at S phase, but in group D, G0/G1 phase cells were significantly decreased and S phase cells significantly increased. Compared to the control group, mitochondrial transmembrane potential was significantly decreased in the treatment group, among with, group B was the lowest, group C was higher than group D. Morphological changes demonstrated by electron microscopy included dispersed nuclear chromatin, loss of nucleoli, membrane bleeding, cell shrinkage, typical apoptotic bodies and marked swelling of mitochondria in the treatment group. In the control group, however, they were characterized by normal cell ultrastructure. CONCLUSIONS: The present study reveals that 5-FU combined with CsA or FK506 demonstrated a synergistic effect on hepatocellular carcinoma rats. For FK506, the powerful mutual effect is related to the increase of tumor cell's quantity in S phase. Both CsA and FK506 can provide protection on mitochondrial transmembrane potential reduction against hepatoma cells damage from 5-FU.

[Repair of segmental bone defects in rhesus monkeys' radius with allogeneic tissue engineered bones].

OBJECTIVE: To construct tissue engineering bone with bio-derived materials and bone marrow stromal cells (MSCs), and to investigate the effect of allogeneic engineering bone implants on healing of segmental bone defects. METHODS: MSCs being aspirated aseptically from tibial tuberosities of young rhesus monkeys were induced into osteoblasts in vitro and then were cultured and marked with 5-bromo-2-deoxyuridine (BrdU). Tissue engineering bones were constructed with these labeled osteoblasts being seeded onto bio-derived materials made from fresh human bones which were treated physically and chemically, Then the constructs were implanted in 15 allogeneic monkeys to bridge 2.5 cm segmental bone defects of left radius as experimental groups, bio-derived materials only were implanted to bridge same size defects of right radius as control group. and, 2.5 cm segmental bone defects of both sides of radius were left empty in two rhesus monkeys as blank group. Every 3 monkeys were sacrificed in the 1st, 2nd, 3rd, 6th and 12th weeks postoperatively and both sides of the implants samples were examined macroscopically, histologicaly, and immunohistochemicaly. The two monkeys in blank group were sacrificed in the 12th week postoperatively. RESULTS: Apparent inflammatory reactions were seen around both sides of the implants samples in the 1st, 2nd, 3rd weeks, but it weakened in the 6th week and disappeared at the 12th week. The labeled osteoblasts existed at the 6th week but disappeared at the 12th week. The bone defects in experimental group were repaired and the new bone formed in multipoint way, and osteoid tissue, cartilage, woven bone and lamellar bone occurred earlier when compared with control group in which the bone defects were repaired in 'creep substitution' way. The bone defects in blank group remained same size at the 12th week. CONCLUSIONS: Engineering bones constructed with bio-derived materials and MSCs were capable of repairing segmental bone defects in allogeneic monkeys beyond 'creep substitution' way and making it healed earlier. Bio-derived materials being constituted with allogeneic MSCs may be a good option in construction of bone tissue engineering.