CpG island methylator phenotype and Helicobacter pylori infection associated with gastric cancer

AIM:To investigate the association between the CpG island methylator phenotype(CIMP) and serum Helicobacter pylori(H.pylori) levels for clinical prediction of gastric cancer(GC) progression.METHODS:We analyzed the serum CIMP status of 75 patients with GC using a methylation marker panel and a methylation-specific polymerase chain reaction.Serum samples from 40 healthy persons were examined at the same time.The genes examined were APC,WIF-1,RUNX-3,DLC-1,SFRP-1,DKK and E-cad.H.pylori infection in serum was assayed with an anti-H.pylori immunoglobulin G antibody test and a rapid urease test.RESULTS:The frequencies of high-level methylation in GC tissues for the seven genes were:48% for APC,57.33% for WIF-1,56% for RUNX-3,50.67% for DLC-1,52% for SFRP-1,54.67% for DKK,and 48% for E-cad.The frequencies in GC serum were 30.67% for APC,34.67% for WIF-1,37.33% for RUNX-3,29.33% for DLC-1,33.33% for SFRP-1,32% for DKK,and 26.67% for E-cad.CIMP+(defined as ≥ 3 methylated genes) was associated with 47(62.67%) GC tissue samples and 44(58.67%) GC serum samples.CIMP+ was not associated with non-neoplastic mucosal tissues or the serum of healthy persons.Of the 75 GC cases,51(68%) were H.pylori +,and 24(32%) were H.pylori-.Of the 51 H.pylori + cases,36 were CIMP+ and 15 were CIMP-.In contrast,for the 24 H.pylori-cases,11 were CIMP+,and 13 were CIMP-.The difference was significant between the H.pylori + and H.pylori-groups(χ 2 = 4.27,P < 0.05).Of the 51 H.pylori + GC patients,34 were CIMP+ and 17 were CIMP-,while among the 24 H.pylori-GC cases,10 were CIMP+ and 14 were CIMP-.The difference was significant between the H.pylori+ and H.pylori-groups(χ 2 = 4.21,P < 0.05).A 2-year follow-up showed significant difference in the rates of metastasis and recurrence between H.pylori +/CIMP+ cases and the H.pylori +/CIMP-cases or CIMP-cases associated with H.pylori assayed in serum(P < 0.05).However,there were no significant differences in survival rates between the two groups.CONCLUSION:H.pylori +/CIMP+ cases are associated with higher rates of metastasis and recurrence than H.pylori +/CIMP-cases.Serum may be useful for examining CIMP status.

Contrast-enhanced ultrasound in association with serum biomarkers for differentiating combined hepatocellular-cholangiocarcinoma from hepatocellular carcinoma and intrahepatic cholangiocarcinoma

BACKGROUND Combined hepatocellular-cholangiocarcinoma(CHC)is a rare type of primary liver cancer.Due to its complex histopathological characteristics,the imaging features of CHC can overlap with those of hepatocellular carcinoma(HCC)and intrahepatic cholangiocarcinoma(ICC).AIM To investigate the possibility and efficacy of differentiating CHC from HCC and ICC by using contrast-enhanced ultrasound(CEUS)Liver Imaging Reporting and Data System(LI-RADS)and tumor biomarkers.METHODS Between January 2016 and December 2019,patients with histologically confirmed CHC,ICC and HCC with chronic liver disease were enrolled.The diagnostic formula for CHC was as follows:(1)LR-5 or LR-M with elevated alphafetoprotein(AFP)and carbohydrate antigen 19-9(CA19-9);(2)LR-M with elevated AFP and normal CA19-9;or(3)LR-5 with elevated CA19-9 and normal AFP.The sensitivity,specificity,accuracy and area under the receiver operating characteristic curve were calculated to determine the diagnostic value of the criteria.RESULTS After propensity score matching,134 patients(mean age of 51.4±9.4 years,108 men)were enrolled,including 35 CHC,29 ICC and 70 HCC patients.Based on CEUS LI-RADS classification,74.3%(26/35)and 25.7%(9/35)of CHC lesions were assessed as LR-M and LR-5,respectively.The rates of elevated AFP and CA19-9 in CHC patients were 51.4%and 11.4%,respectively,and simultaneous elevations of AFP and CA19-9 were found in 8.6%(3/35)of CHC patients.The sensitivity,specificity,positive predictive value,negative predictive value,accuracy and area under the receiver operating characteristic curve of the aforementioned diagnostic criteria for discriminating CHC from HCC and ICC were 40.0%,89.9%,58.3%,80.9%,76.9%and 0.649,respectively.When considering the reported prevalence of CHC(0.4%-14.2%),the positive predictive value and NPV were revised to 1.6%-39.6%and 90.1%-99.7%,respectively.CONCLUSION CHCs are more likely to be classified as LR-M than LR-5 by CEUS LI-RADS.The combination of the CEUS LI-RADS classification with serum tumor markers shows high specificity but low sensitivity for the diagnosis of CHC.Moreover,CHC could be confidently excluded with high NPV.

Shear wave elastography results correlate with liver fibrosis histology and liver function reserve

AIM: To evaluate the correlation of shear wave elastography(SWE) results with liver fibrosis histology and quantitative function reserve.METHODS: Weekly subcutaneous injection of 60% carbon tetrachloride(1.5 m L/kg) was given to 12 canines for 24 wk to induce experimental liver fibrosis, with olive oil given to 2 control canines. At 24 wk, liver condition was evaluated using clinical biochemistry assays, SWE imaging, lidocaine metabolite monoethylglycine-xylidide(MEGX) test, and histologic fibrosis grading. Clinical biochemistry assays were performed at the institutional central laboratory for routine liver function evaluation. Liver stiffness was measured in triplicate from three different intercostal spaces and expressed as mean liver stiffness modulus(LSM). Plasma concentrations of lidocaine and its metabolite MEGX were determined using high-performance liquid chromatography repeated in duplicate. Liver biopsy samples were fixed in 10% formaldehyde, and liver fibrosis was graded using the modified histological activity index Knodell score(F0-F4). Correlations among histologic grading, LSM, and MEGX measures were analyzed with the Pearson linear correlation coefficient.RESULTS: At 24 wk liver fibrosis histologic grading was as follows: F0, n = 2(control); F1, n = 0; F2, n = 3; F3, n = 7; and F4, n = 2. SWE LSM was positively correlated with histologic grading(r = 0.835, P < 0.001). Specifically, the F4 group had a significantly higher elastic modulus than the F3, F2, and F0 groups(P = 0.002, P = 0.003, and P = 0.006, respectively), and the F3 group also had a significantly higher modulus than the control F0 group(P = 0.039). LSM was negatively associated with plasma MEGX concentrations at 30 min(r =-0.642; P = 0.013) and 60 min(r =-0.651; P = 0.012), time to ? of the maximum concentration(r =-0.538; P = 0.047), and the area under the curve(r =-0.636; P = 0.014). Multiple comparisons showed identical differences in these three measures: significantly lower with F4(P = 0.037) and F3(P = 0.032) as compared to F0 and significantly lower with F4 as compared to F2(P = 0.032).CONCLUSION: SWE LSM shows a good correlation with histologic fibrosis grading and pharmacologic quantitative liver function reserve in experimental severe fibrosis and cirrhosis.

Can contrast enhanced ultrasound differentiate intrahepatic cholangiocarcinoma from hepatocellular carcinoma?

BACKGROUND Hepatocellular carcinoma(HCC)and intrahepatic cholangiocarcinoma(ICC)differ in treatment and prognosis,warranting an effective differential diagnosis between them.The LR-M category in the contrast-enhanced ultrasound(CEUS)liver imaging reporting and data system(LI-RADS)was set up for lesions that are malignant but not specific to HCC.However,a substantial number of HCC cases in this category elevated the diagnostic challenge.AIM To investigate the possibility and efficacy of differentiating ICC from HCC classified in the LR-M category according to the CEUS LI-RADS.METHODS Patients with complete CEUS records together with pathologically confirmed ICC and LR-M HCC(HCC classified in the CEUS LI-RADS LR-M category)between January 2015 and October 2018 were included in this retrospective study.Each ICC was assigned a category as per the CEUS LI-RADS.The enhancement pattern,washout timing,and washout degree between the ICC and LR-M HCC were compared using theχ2 test.Logistic regression analysis was used for prediction of ICC.Receiver operating characteristic(ROC)curve analysis was used to investigate the possibility of LR-M criteria and serum tumor markers in differentiating ICC from LR-M HCC.RESULTS A total of 228 nodules(99 ICCs and 129 LR-M HCCs)in 228 patients were included.The mean sizes of ICC and LR-M HCC were 6.3±2.8 cm and 5.5±3.5 cm,respectively(P=0.03).Peripheral rim-like arterial phase hyperenhancement(APHE)was detected in 50.5%(50/99)of ICCs vs 16.3%(21/129)of LR-M HCCs(P<0.001).Early washout was found in 93.4%(93/99)of ICCs vs 96.1%(124/129)of LR-M HCCs(P>0.05).Marked washout was observed in 23.2%(23/99)of ICCs and 7.8%(10/129)of LR-M HCCs(P=0.002),while this feature did not show up alone either in ICC or LR-M HCC.Homogeneous hyperenhancement was detected in 15.2%(15/99)of ICCs and 37.2%(48/129)of LR-M HCCs(P<0.001).The logistic regression showed that rim APHE,carbohydrate antigen 19-9(CA 19-9),and alpha fetoprotein(AFP)had significant correlations with ICC(r=1.251,3.074,and-2.767,respectively;P<0.01).Rim APHE presented the best enhancement pattern for diagnosing ICC,with an area under the ROC curve(AUC)of 0.70,sensitivity of 70.4%,and specificity of 68.8%.When rim hyperenhancement was coupled with elevated CA 19-9 and normal AFP,the AUC and sensitivity improved to 0.82 and 100%,respectively,with specificity decreasing to 63.9%.CONCLUSION Rim APHE is a key predictor for differentiating ICC from LR-M HCC.Rim APHE plus elevated CA 19-9 and normal AFP is a strong predictor of ICC rather than LR-M HCC.Early washout and marked washout have limited value for the differentiation between the two entities.

Diagnostic value of two dimensional shear wave elastography combined with texture analysis in early liver fibrosis

BACKGROUND Staging diagnosis of liver fibrosis is a prerequisite for timely diagnosis and therapy in patients with chronic hepatitis B.In recent years,ultrasound elastography has become an important method for clinical noninvasive assessment of liver fibrosis stage,but its diagnostic value for early liver fibrosis still needs to be further improved.In this study,the texture analysis was carried out on the basis of two dimensional shear wave elastography(2D-SWE),and the feasibility of 2D-SWE plus texture analysis in the diagnosis of early liver fibrosis was discussed.AIM To assess the diagnostic value of 2D-SWE combined with textural analysis in liver fibrosis staging.METHODS This study recruited 46 patients with chronic hepatitis B.Patients underwent 2DSWE and texture analysis;Young's modulus values and textural patterns were obtained,respectively.Textural pattern was analyzed with regard to contrast,correlation,angular second moment(ASM),and homogeneity.Pathological results of biopsy specimens were the gold standard;comparison and assessment of the diagnosis efficiency were conducted for 2D-SWE,texture analysis and their combination.RESULTS 2D-SWE displayed diagnosis efficiency in early fibrosis,significant fibrosis,severe fibrosis,and early cirrhosis(AUC>0.7,P<0.05)with respective AUC values of 0.823(0.678-0.921),0.808(0.662-0.911),0.920(0.798-0.980),and 0.855(0.716-0.943).Contrast and homogeneity displayed independent diagnosis efficiency in liver fibrosis stage(AUC>0.7,P<0.05),whereas correlation and ASM showed limited values.AUC of contrast and homogeneity were respectively 0.906(0.779-0.973),0.835(0.693-0.930),0.807(0.660-0.910)and 0.925(0.805-0.983),0.789(0.639-0.897),0.736(0.582-0.858),0.705(0.549-0.883)and 0.798(0.650-0.904)in four liver fibrosis stages,which exhibited equivalence to 2D-SWE in diagnostic efficiency(P>0.05).Combined diagnosis(PRE)displayed diagnostic efficiency(AUC>0.7,P<0.01)for all fibrosis stages with respective AUC of 0.952(0.841-0.994),0.896(0.766-0.967),0.978(0.881-0.999),0.947(0.835-0.992).The combined diagnosis showed higher diagnosis efficiency over 2D-SWE in early liver fibrosis(P<0.05),whereas no significant differences were observed in other comparisons(P>0.05).CONCLUSION Texture analysis was capable of diagnosing liver fibrosis stage,combined diagnosis had obvious advantages in early liver fibrosis,liver fibrosis stage might be related to the hepatic tissue hardness distribution.

Effect of a poloxamer 407-based thermosensitive gel on minimization of thermal injury to diaphragm during microwave ablation of the liver

AIM To assess the insulating effect of a poloxamer 407(P407)-based gel during microwave ablation of liver adjacent to the diaphragm.METHODS We prepared serial dilutions of P407, and 22.5%(w/w) concentration was identified as suitable for ablation procedures. Subsequently, microwave ablations were performed on the livers of 24 rabbits(gel, saline, control groups, n = 8 in each). The P407 solution and 0.9% normal saline were injected into the potential space between the diaphragm and liver in experimental groups. No barriers were applied to the controls. After microwave ablations, the frequency, size and degree of thermal injury were compared histologically among the three groups. Subsequently, another 8 rabbits were injected with the P407 solution and microwave ablation was performed. The levels of alanine aminotransferase(ALT), aspartate aminotransferase(AST), blood urea nitrogen(BUN) and creatinine(Cr) in serum were tested at 1 d before microwave ablation and 3 and 7 d after operation. RESULTS In vivo ablation thermal injury to the adjacent diaphragm was evaluated in the control, saline and 22.5% P407 gel groups(P = 0.001-0.040). However, there was no significant difference in the volume of ablation zone among the three groups(P > 0.05). Moreover, there were no statistical differences among the preoperative and postoperative gel groups according to the levels of ALT, AST, BUN and Cr in serum(all P > 0.05).CONCLUSION Twenty-two point five percent P407 gel could be a more effective choice during microwave ablation of hepatic tumors adjacent to the diaphragm. Further studies for clinical translation are warranted.

Contrast-enhanced ultrasound for quantitative assessment of portal pressure in canine liver fibrosis

AIM: To explore the feasibility of non-invasive quantitative estimation of portal venous pressure by contrast-enhanced ultrasound(CEUS) in a canine model.METHODS: Liver fibrosis was established in adult canines(Beagles; n = 14) by subcutaneous injection of carbon tetrachloride(CCl4). CEUS parameters, including the area under the time-intensity curve and intensity at portal/arterial phases(Qp/Qa and Ip/Ia, respectively), were used to quantitatively assess the blood flow ratio of the portal vein/hepatic artery at multiple time points. The free portal venous pressures(FPP) were measured by a multi-channel baroreceptor using a percutaneous approach at baseline and 8, 16, and 24 wk after CCl4 injections in each canine. Liver biopsies were obtained at the end of 8, 16, and 24 wk from each animal, and the stage of the fibrosis was assessed according to the Metavir scoring system. A Pearson correlation test was performed to compare the FPP with Q p/Q a and I p/I a.RESULTS: Pathologic examination of 42 biopsies from the 14 canines at weeks 8, 16, and 24 revealed that liver fibrosis was induced by CCl4 and represented various stages of liver fibrosis, including F0(n = 3), F1(n = 12), F2(n = 14), F3(n = 11), and F4(n = 2). There were significant differences in the measurements of Qp/Qa(19.85 ± 3.30 vs 10.43 ± 1.21, 9.63 ± 1.03, and 8.77 ± 0.96) and Ip/Ia(1.77 ± 0.37 vs 1.03 ± 0.12, 0.83 ± 0.10, and 0.69 ± 0.13) between control and canine fibrosis at 8, 16, and 24 wk, respectively(all P < 0.001). There were statistically significant negative correlations between FPP and Q p/Q a(r =-0.707, P < 0.001), and between FPP and Ip/Ia(r =-0.759, P < 0.001) in the canine fibrosis model. Prediction of elevated FPP based on Q p/Q a and I p/I a was highly sensitive, as assessed by the area under the receiveroperating curve(0.866 and 0.895, respectively).CONCLUSION: C E U S i s a p o t e n t i a l m e t h o d t o accurately, but non-invasively, estimate portal venous pressure through measurement of Qp/Qa and Ip/Ia parameters.

Quantitative Assessment of Hepatic Fibrosis by Contrast-enhanced Ultrasonography

Objective To explore the contrast-enhanced ultrasonographic features for quantitative assessment of hepatic fibrosis. Methods 86 patients with chronic viral hepatitis B were enrolled in this study from March 2007 to August 2009. The patients were classified into 5 groups (S0-S4) according to fibrosis stage evaluated with ultrasound guided liver biopsy. New contrast-enhanced ultrasonography (CEUS) features including area under the time-intensity curve (TIC) of portal venous phase/hepatic arterial phase (Qp/Qa) and intensity of portal venous phase/hepatic arterial phase (Ip/Ia) were used to detect the blood supply ratio (portal vein/hepatic artery) in each group. Arrival time of portal vein trunk (Tp) and decreasing rate of TIC (β ) were also analyzed. Results Qp/Qa and Ip/Ia decreased from S0 to S4, while Tp and β increased. These 4 features were significantly correlated with the degree of fibrosis (P<0.001) and were significantly different among the five groups (P<0.001). Sensitivity and specificity of Ip/Ia were 80% and 86% for groups ≥S1, 75% and 86% for groups ≥ S2, 71% and 84% for groups ≥ S3, and 76% and 80% for group S4, respectively. Sensitivity and specificity of Qp/Qa were 70% and 88% for groups ≥ S1, 80% and 76% for groups ≥ S2, 74% and 70% for groups ≥ S3, and 81% and 95% for group S4, respectively. Conclusion Ip/Ia and Qp/Qa could be adopted as reliable, non-invasive features for quantitative assessment of hepatic fibrosis.

Ultrasound Assessraent of Intima-media Thickness and Diameter of Carotid Arteries in Patients Undergoing Hemodialysis or Renal Transplantation

Renal transplant （RT） recipients have a high risk of developing cardiovascular diseases. However, the effects of renal transplantation on the development of arteriosclerosis have been controversial. The carotid intima-media thickness （CIMT） and diameter （CD） are important indicators of vascular remodeling and arteriosclerosis. In this study, 31 patients with hemodialysis （HD）, 31 RT recipients and 84 age- and gender-matched control subjects were enrolled. Their CIMT and CD were measured by ultrasonic radiofrequency tracking, and the linear regression models and Z test were used to identify the progression of arteriosclerosis and the risk factors. Compared with HD group, RT group had significantly lower CIMT and CD. CIMT was found to be associated with age, body weight, resistance index and diastolic velocity, while CD was associated significantly with age, body weight, pulsatility index, end diastolic velocity and diastolic blood pressure （DBP）, respectively. The correlation curves between CIMT and age showed the slopes of curves were decreased successively in control, RT and HD groups, and the curves between CD and age showed the slopes were decreased in order of RT 〉 control 〉 HD groups. It was concluded that CIMT and CD were significantly correlated with age in RT and moderately with age in HD patients. RT could reduce the progress of arteriosclerosis in patients with end-stage renal disease.

PREPARATION OF POLYELECTROLYTE MULTILAYER COATED MICROBUBBLES FOR USE AS ULTRASOUND CONTRAST AGENT

Objective To prepare and characterize polyelectrolyte multilayer film coated microbubbles for use as ultrasound contrast agent(UCA)and evaluate its effects in ultrasonic imaging on normal rabbit's liver parenchyma.Methods Perfluorocarbon(PFC)-containing microbubbles(ST68-PFC)were prepared by sonication based on surfactant(Span 60 and Tween 80).Subsequently,the resulting ST68-PFC microbubbles were coated using oppositely charged polyelectrolytes by microbubble-templated layer-by-layer self-assembly technique via electrostatic interaction.The enhancement effects in ultrasonic imaging on normal rabbit's liver parenchyma were assessed.Results The obtained microbubbles exhibited a narrow size distribution.The polyelectrolytes were successfully assembled onto the surface of ST68-PFC microbubbles.In vivo experiment showed that polyelectrolyte multilayer film coated UCA effectively enhanced the imaging of rabbit's liver parenchyma.Conclusions The novel microbubbles UCA coated with polyelectrolyte multilayer,when enabled more function,has no obvious difference in enhancement effects compared with the pre-modified microbubbles.The polymers with chemically active groups(such as amino group and carboxyl group)can be used as the outermost layer for attachment of targeting ligands onto microbubbles,allowing selective targeting of the microbubbles to combine with desired sites.

Inhibition of USP14 and UCH37 deubiquitinating activity by b-AP15 as a potential therapy for tumors with p53 deficiency

Dear Editor,Tumor-suppressor protein p53 is important for cell function and genome integrity.1 The decrease of p53 protein is a common feature of human malignant tumors,which leads to the deficiency of cell cycle detection point control and apoptosis induction.An enormous amount of research effort goes into small molecules that regulate p53,including wild-type repair of mutant p53 gene and interruption of the binding between p53 and an E3-ubiquitin ligase Mdm2 to prevent ubiquitination degradation of p53 and rescue the protein level of p53.2 However,it is not clear whether it is necessary to continue to inhibit the ubiquitin activity of proteasome in order to restore the p53 protein level.

Expression quantitative trait loci for PVT1contributes to the prognosis of hepatocellular carcinoma

Aim: Plasmacytoma variant translocation 1 (PVT1), a long intergenic non-coding RNA, was overexpressed in liver cancer. A single nucleotide polymorphism (SNP) rs4733586 was identified as an expression quantitative trait loci (eQTL) for PVT1 using bioinformatics analysis. This study was to assess the association of PVT1 eQTL with hepatocellular carcinoma (HCC) prognosis. Methods: A case-only study was performed to assess the association between SNP and HCC overall survival in 331 HCC patients with hepatitis B virus. Cox proportional hazard regression models were conducted for survival analysis with adjustment for age, gender, smoking status, drinking status, Barcelona-Clinic Liver Cancer stages, and chemotherapy or transcatheter hepatic arterial chemoembolization (TACE) status. Results: The variant genotype C allele of rs4733586 was significantly associated with a higher death risk compared with T allele (adjusted hazard ratio = 1.26, 95% confidence intervals = 1.05-1.51,P = 0.012 in the additive model). By stepwise Cox proportional hazard analysis, four variables (age, drinking status, chemotherapy or TACE status, PVT1 eQTL) were remained in the final regression model. In the stratified analysis, no heterogeneity was observed among different subgroups. Conclusion: These findings suggest that eQTL SNP for PVT1 may be susceptibility marker for the HCC overall survival.