Utility of serum CA19-9 in diagnosis of cholangiocarcinoma:In comparison with CEA

AIM:The diagnosis of cholangiocarcinoma is often difficult,making management approaches problematic. A reliable serum marker for cholangiocarcinoma would be a useful diagnostic test. The aims of our study were to evaluate the usefulness of a serum CA19-9 determination in the diagnosis of cholangiocarcinoma.METHODS: We prospectively measured serum CA19-9 and CEA concentrations in patients with cholangiocarcinoma (n=35), benign biliary diseases (n=92), and healthy individuals (n=15). Serum CA19-9 and CEA concentrations were measured by an immunoradiometric assay without knowledge of the clinical diagnosis.RESULTS:The sensitivity of a CA19-9 value>37KU·L^-1 and a CEA value >22μg·L^-1 in diagnosing cholangiocarcinoma were 77.14% and 68.57%, respectively. When compared with the benign biliary diseases group,the true negative rates of serum CA19-9 and CEA were 84.78% and 81.52%,respectively. The false positive rates of serum CA19-9 and CEA were 15.22% and 18.48%, whereas the accuracy of serum CA19-9 and CEA were 82.68% and 77.95%,respectively. Serum CA19-9 and CEA concentrations were significantly elevated (P<0.001 and P<0.05) in patients with cholangiocarcinoma (290.31±5.34KU·L^-1 and 36.46±18.03μg·L^-1) compared with patients with benign biliary diseases (13.38±2.59KU·L^-1 and 13.84±3.85μg·L^-1) and healthy individuals (12.78±3.69KU·L^-1 and 11.48±3.37μg·L^-1). In 15 patients undergoing curative resection of cholangiocarcinoma,the mean serum CA19-9 concentration was decreased from a preoperative level of 286.41±4.36KU·L^-1 to a postoperative level of 62.01±17.43KU·L^-1 (P<0.001), and the mean serum CEA concentration from 39.41±24.35μg·L^-1 to 28.69±11.03μg·L6-1(P<0.05). In patients with cholangiocarcinoma,however, no correlation was found between serum CEA and CA19-9 concentrations (r=-0.036).CONCLUSION:These data suggest that the serum CA19-9 determination is a useful addition to the available tests for the differential diagnosis of cholangiocarcinoma. Serum CA19-9 is an effective tumor marker in diagnosing cholangiocarcinoma,deciding whether the tumor has been radically resected and monitoring effect of treatment.

Effects of herbal compound 861 on human hepatic stellate cell proliferation and activation

AIM: To investigate the effects of herbal compound 861 (Cpd 861) on cell proliferation in human hepatic stellate cells (LX-2) and human hepatocellular liver carcinoma cells (HepG2), and expression of α-smooth muscle actin (α-SMA) in LX-2 cells.METHODS: LX-2 and HepG2 cells were incubated with various concentrations of Cpd 861 (0.1-0.003 mg/mL)for 1, 2, 3, 5 and 7 d. Cell proliferation was analyzed by 3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay. Effects of Cpd861on the expression of cc-SMA mRNA in LX-2 cells weremeasured by real-time quantitative PCR method using SYBRGreen I technology.RESULTS: Cpd 861, at 0.1 mg/mL, significantly inhibitedLX-2 cell proliferation (15% decrease relative to control,P<0.05) after 3 d of incubation. The inhibitory effects seemedto increase with the treatment time (25% decrease after5 d of incubation and 35% decrease after 7 d of incubation,P<0.01). However, Cpd 861 did not affect HepG2 cellproliferation at the same concentration used for I_X-2 cells.The expression levels of ^-SMA mRNA decreased significantlywhen LX-2 cells were exposed to Cpd 861 for 48 h (59%decrease relative to control, P<0.05) or 72 h (60% decreaserelative to control, P<0.01).CONCLUSION: Cpd 861 can significantly inhibit LX-2 cellproliferation in a dose-dependant manner, and reduce theexpression levels of o^-SMA mRNA in LX-2 cells. Since hepaticcell proliferation and high level of ^-SMA are associatedwith liver fibrosis, the results suggest that Cpd 861 may beuseful in the treatment of this disease.

Real-time determination of human telomerase reverse transcriptase mRNA in gastric cancer

AIM: To set up a real±time fluorescent quantitative reverse transcription±polymerase chain reaction (RT±PCR) assay,to detect human telomerase reverse transcriptase (hTERT) messenger RNA in gastric carcinomas, and to evaluate quantitative determination of hTERT mRNA in the diagnostic value of gastric carcinomas, and to analyze the correlation between the expression level of hTERT mRNA and clinicopathological parameters in patients with gastric cancer.METHODS: A real±time quantitative RT±PCR (RQ±PCR) based on TaqMan fluorescence methodology and the LightCyder system was used to quantify the full range of hTERT mRNA copy numbers in 35 samples of gastric carcinomas and ccrrespongding adjacent non±cancerous tissues. The normalized hTERT (NhTERT) was standardized by quantifying the number of GAPDH transcripts as intemal control and expressed as 100x (hTERT/GAPDH) ratio. Variables were analyzed bythe Student's t±test, X^2 test and Fisher's exact test.P,FESULIS: NhTERT from gastric cardnomas and corresponding adjacent non±cancerous tissues was 6.27±0.89 and 0.934±0.18,respectively (t = 12.76, P<0.001). There was no significant associaUon between gastric cancer hTERT mRNA expression level and patient's age, gender, tumor size, location and stage (pTNM), but a significant correlation was found between hTERT mRNA expression level in gastric carcinomas and the degree of differentiation.CONCLUSION: Quantitative determination of hTERT mRNA by RQ±PCR is a rapid and sensitive method, hTERT might be a potential biomarker for the early detection of gastric cancer.

Relationship between trefoil factor 1 expression and gastric mucosa injuries and gastric cancer

AIM: To determine whether trefoil factor 1 (TFF1) is associated with mucosa healing and carcinoma suppression,we assess the expression of trefoil factor 1 in normal and pathologic gastric mucosa.METHODS: TFFI in normal and pathologic gastric mucosa was assessed by immunohistochemical method, and the average positive A was estimated by Motic Images Advanced 3.0 software.RESULTS: Increased TFF1 was detected in gastritis,gastric ulcer and duodenal ulcer compared with normal mucosa. The same result could be seen in multiple and compound ulcer compared with simple ulcer. There was no significant difference between gastric ulcer and duodenal ulcer, gastritis and simple ulcer respectively.Increased TFF1 was detected in the peripheral mucosa of the gastric adenocarcinoma compared with normal mucosa. The expression of TFF1 in gastric adenocarcinoma was related to the differentiation of adenocarcinoma. The lower the differentiation of adenocarcinoma, the weaker the expression of TFF1. There was no TFF1 expressed in low-differentiated adenocarcinoma. The expression of TFF1 in middle and highly differentiated adenocarcinoma was a little lower than that in normal mucosa. But there was no significant difference. No TFF1 was assessed in esophageal squamous carcinoma and peripheral tissue. There was no significant difference between male and female.CONCLUSION: The expression of TFF1 was higher in gastritis and peptic ulcer than that in normal mucosa,and was also higher in multiple and compound ulcer than in simple ulcer. It seems that TFF1 plays a role in gastric mucosa protection and epithelial restitution. Increased expression of TFF1 in peripheral tissue suggests that TFF1 is associated with mechanism of carcinoma suppression and differentiation. Decreased expression of TFFI in carcinoma and its relativity to the differentiation suggests that TFF1 is related to gland and cell destruction of carcinoma.

Genes transactivated by hepatitis C virus core protein, a microarray assay

AIM: To explore the new target genes transactivated by hepatitis C virus (HCV) core protein and to elucidate the pathogenesis of HCV infection.METHODS: Reverse transcribed cDNA was subjected tomicroarray assay. The coding gene transactivated by HCV core protein was cloned and analyzed with bioinformatics methods.RESULTS: The expressive vector of pcDNA3.1(-)-core was constructed and confirmed by restriction enzyme digestion and DNA sequencing and approved correct. mRNA was purified from HepG2 and HepG2 cells transfected with pcDNA3.1(-)-core, respectively. The cDNA derived was subjected to microarray assay. A new gene namedHCTP4 was cloned with molecular biological method in combination with bioinformatics method.CONCLUSION: HCV core is a potential transactivator.Microarray is an efficient and convenient method for analysis of differentially expressed genes.

Transactivating effect of hepatitis C virus core protein: A suppression subtractive hybridization study

AIM: To investigate the transactivating effect of hepatitis C virus (HCV) core protein and to screen genes transactivated by HCV core protein. METHODS: pcDNA3.1(-)-core containing full-length HCV core gene was constructed by insertion of HCV core gene into EcoRI/BarnHI site. HepG2 cells were cotransfected with pcDNA3.1(-)-core and pSV-lacZ. After 48 h, cells were collected and detected for the expression of β-gal by an enzyme-linked immunosorbent assay (ELISA) kit. HepG2 cell swere transiently transfected with pcDNA3.1(-)-core using Upofectamine reagent. Cells were collected and total mRNA was isolated. A subtracted cDNA library was generated and constructed into a pGEM-Teasy vector. The library was amplified with E. coil strain JM109. The cDNAs were sequenced and analyzed in GenBank with BLAST search after polymerase chain reaction (PCR). RESULTS: The core mRNA and protein could be detected in HepG2 cell lysate which was transfected by the pcDNA3.1(-)-core. The activity of β-galactosidase in HepG2 cells transfected by the pcDNA3.1(-)-core was 5.4 times higher than that of HepG2 cells transfected by control plasmid. The subtractive library of genes transactivated by HCV core protein was constructed successfully. The amplified library contained 233 positive clones. Colony PCR showed that 213 clones contained 100-1 000 bp inserts. Sequence analysis was performed in 63 clones. Six of the sequences were unknown genes. The full length sequences were obtained with bioinformatics method, accepted by Genl3ank. It was suggested that six novel cDNA sequences might be target genes transactivated by HCV core protein. CONCLUSION: The core protein of HCV has transactivating effects on SV40 early promoter/enhancer. A total of 63 clones from cDNA library were randomly chosen and sequenced. Using the BLAST program at the National Center for Biotechnology Information, six of the sequences were unknown genes. The other 57 sequences were highly similar to known genes.

Screening of hepatocyte proteins binding to complete S protein of hepatitis B virus by yeast-two hybrid system

AIM: To investigate the biological function of complete S protein and to look for proteins interacting with complete S protein in hepatocytes.METHODS: We constructed bait plasmid expressing complete S protein of HBV by cloning the gene of complete S protein into pGBKT7, then the recombinant plasmid DNA was transformed into yeast AH109 (a type). The transformed yeast AH109 was mated with yeast Y187 (α type) containing liver cDNA library plasmid in 2xYPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/Trp-Leu-His-Ade) containing X-α-gal for selection and screening. After extracting and sequencing of plasmids from positive (blue) colonies, we underwent sequence analysis by bioinformatics.RESULTS: Nineteen colonies were selected and sequenced.Among them, five colonies were Homo sapiens solute carrier family 25, member 23 (SLC25A23), one was Homo sapiens calreticulin, one was human serum albumin (ALB)gene, one was Homo sapiens metallothionein 2A, two were Homo sapiens betaine-homocysteine methyltransferase,three were Homo sapiensNa+ and H+ coupled amino acid transport system N, one was Homo sapiens CD81 antigen (target of anti-proliferative antibody 1) (CD81), three were Homo sapiens diazepam binding inhibitor, two colonies were new genes with unknown function.CONCLUSION: The yeast-two hybrid system is an effective method for identifying hepatocyte proteins interacting with complete S protein of HBV. The complete S protein may bind to different proteins i.e., its multiple functions in vivo.

Standard pancreatoduodenectomy versus extended pancreatoduodenectomy with modified retroperitoneal nerve resection in patients with pancreatic head cancer:a multicenter randomized controlled trial

Background:The extent of pancreatoduodenectomy for pancreatic head cancer remains controversial,and more high-level clinical evidence is needed.This study aimed to evaluate the outcome of extended pancreatoduodenectomy(EPD)with retroperitoneal nerve resection in pancreatic head cancer.Methods:This multicenter randomized trial was performed at 6 Chinese highvolume hospitals that enrolled patients between October 3,2012,and September 21,2017.Four hundred patients with stage I or II pancreatic head cancer and without specific pancreatic cancer treatments(preoperative chemotherapy or chemoradiation)within three months were randomly assigned to undergo standard pancreatoduodenectomy(SPD)or EPD,with the latter followed by dissection of additional lymph nodes(LNs),nerves and soft tissues 270◦on the right side surrounding the superior mesenteric artery and celiac axis.The primary endpoint was overall survival(OS)by intention-to-treat(ITT).The secondary endpoints were disease-free survival(DFS),mortality,morbidity,and postoperative pain intensity.Results:TheR1 ratewas slightly lower with EPD(8.46%)thanwith SPD(12.56%).The morbidity and mortality rates were similar between the two groups.The median OS was similar in the EPD and SPD groups by ITT in the whole study cohort(23.0 vs.20.2 months,P=0.100),while the median DFS was superior in the EPD group(16.1 vs.13.2 months,P=0.031).Patients with preoperative CA19–9<200.0 U/mL had significantly improved OS and DFS with EPD(EPD vs.SPD,30.8 vs.20.9 months,P=0.009;23.4 vs.13.5 months,P<0.001).The EPD group exhibited significantly lower locoregional(16.48%vs.35.20%,P<0.001)andmesenteric LNrecurrence rates(3.98%vs.10.06%,P=0.022).The EPD group exhibited less back pain 6 months postoperation than the SPD group.Conclusions:EPD for pancreatic head cancer did not significantly improve OS,but patients with EPD treatment had significantly improved DFS.In the subgroup analysis,improvements in bothOS and DFS in the EPD armwere observed in patients with preoperative CA19–9<200.0 U/mL.EPD could be used as an effective surgical procedure for patients with pancreatic head cancer,especially those with preoperative CA19–9<200.0 U/mL.

Soil Organic Matter Dynamics Along a Vertical Vegetation Gradient in the Gongga Mountain on the Tibetan Plateau

Abstract: Our knowledge about soil organic matter (SOM) dynamics is limited although this is an important issue in the study of responses of ecosystems to global climate changes. Twelve sampling plots were set up every 200 m from 1 700 to 3 900 m along the vertical vegetation gradient along the east slope of Gongga Mountain. Samples were taken from all 12 plots for SOM content measurement, although only 5 of the 12 plots were subjected to radiocarbon measurements. A radiocarbon isotope method and a time-dependent model were used to quantify the SOM dynamics and SOM turnover rates along the vertical vegetation gradient. The results showed that the SOM turnover rate decreased and turnover time increased with soil depth for all vegetation types. The litter layer turnover rates presented a clear trend along the gradient. The litter layer turnover rates decreased with an increase in elevation, except that the litter layer turnover rate of mixed forest was higher than that of evergreen forest. Climatic factors, such as temperature and precipitation, were the main factors influencing the surface soil carbon dynamics. The turnover rates of the subsoil (including the A, B, and C horizons in the soil profiles) along the vertical gradient had no clear trends. The SOM of subalpine shrub and meadow turned over more slowly than that of the forest types in almost all soil horizons. The characteristic of short roots distributing in the upper part of the soil profile leads to different SOM dynamics of shrub and meadow compared with the forest types. Coniferous and mixed forests were susceptible to carbon loss from the young carbon pool, but their long and big roots resulted in high Δ14C values of the deep soil profiles and increased the input of young carbon to the deep soil. In evergreen forest, the carbon cumulative ability from the B horizon to the C horizon was weak. The different vegetation types, together with their different modes of nutrient and carbon intake, may be the mechanism conditioning the subsoil organic matter dynamics.