First report on establishment and characterization of the extrahepatic cholangiocarcinoma sarcoma cell line CBC2T-2

BACKGROUND Extrahepatic cholangiocarcinoma sarcoma is extremely rare in clinical practice.These cells consist of both epithelial and mesenchymal cells.Patient-derived cell lines that maintain tumor characteristics are valuable tools for studying the molecular mechanisms associated with carcinosarcoma.However,cholangiocarcinoma sarcoma cell lines are not available in cell banks.AIM To establish and characterize a new extrahepatic cholangiocarcinoma sarcoma cell line,namely CBC2T-2.METHODS We conducted a short tandem repeat(STR)test to confirm the identity of the CBC2T-2 cell line.Furthermore,we assessed the migratory and invasive properties of the cells and performed clonogenicity assay to evaluate the ability of individual cells to form colonies.The tumorigenic potential of CBC2T-2 cells was tested in vivo using nonobese diabetic/severe combined immunodeficient(NOD/SCID)mice.The cells were injected subcutaneously and tumor formation was observed.In addition,immunohistochemical analysis was carried out to examine the expression of epithelial marker CK19 and mesenchymal marker vimentin in both CBC2T-2 cells and xenografts.The CBC2T-2 cell line was used to screen the potential therapeutic effects of various clinical agents in patients with cholangiocarcinoma sarcoma.Lastly,whole-exome sequencing was performed to identify genetic alterations and screen for somatic mutations in the CBC2T-2 cell line.RESULTS The STR test showed that there was no cross-contamination and the results were identical to those of the original tissue.The cells showed round or oval-shaped epithelioid cells and mesenchymal cells with spindle-shaped or elongated morphology.The cells exhibited a high proliferation ratio with a doubling time of 47.11 h.This cell line has migratory,invasive,and clonogenic abilities.The chromosomes in the CBC2T-2 cells were polyploidy,with numbers ranging from 69 to 79.The subcutaneous tumorigenic assay confirmed the in vivo tumorigenic ability of CBC2T-2 cells in NOD/SCID mice.CBC2T-2 cells and xenografts were positive for both the epithelial marker,CK19,and the mesenchymal marker,vimentin.These results suggest that CBC2T-2 cells may have both epithelial and mesenchymal characteristics.The cells were also used to screen clinical agents in patients with cholangiocarcinoma sarcoma,and a combination of paclitaxel and gemcitabine was found to be the most effective treatment option.CONCLUSION We established the first human cholangiocarcinoma sarcoma cell line,CBC2T-2,with stable biogenetic traits.This cell line,as a research model,has a high clinical value and would facilitate the understanding of the pathogenesis of cholangiocarcinoma sarcoma.

内镜逆行胰胆管造影术在肝包虫病胆道并发症治疗中的应用

目的探讨内镜逆行胰胆管造影术(ERCP)在肝包虫胆道并发症治疗中的价值。方法回顾分析该院2010年10月-2016年10月ERCP参与治疗的肝包虫病27例,总结患者的临床表现、实验室检验、影像学检查、手术方式及治疗效果。结果该组有27例应用ERCP参与治疗的肝包虫胆道并发症病例。其中,12例因肝包虫囊肿破入胆道引起急性化脓性胆管炎及梗阻性黄疸,7例因肝包虫囊肿压迫胆总管引起重度黄疸,先行ERCP通畅胆道引流,择期行开腹手术;6例因肝包虫内囊摘除术后残腔胆瘘,2例因肝包虫术后胆道狭窄,术后行ERCP放置胆道支架,该组27例均取得良好的治疗效果;比较患者术前24 h内与术后48 h白细胞计数(WBC)、谷草转氨酶(AST)、谷丙转氨酶(ALT)、总胆红素(TBIL)、碱性磷酸酶(ALP)和γ-转肽酶(GGT)6项指标,术后均有改善,差异有统计学意义(P<0.05)。结论对于肝包虫病引起的急性化脓性胆管炎、胆瘘和胆道狭窄等胆道并发症,在术前或术后采用ERCP处理具有重要的诊治价值。

Plasma microRNAs as potential new biomarkers for early detection of early gastric cancer

BACKGROUND Early gastric cancer(EGC), compared with advanced gastric cancer(AGC), has a higher 5-year survival rate. However, due to the lack of typical symptoms and the difficulty in diagnosing EGC, no effective biomarkers exist for the detection of EGC, and gastroscopy is the only detection method.AIM To provide new biomarkers with high specificity and sensitivity through analyzed the differentially expressed microRNAs(miRNAs) in EGC and AGC and compared them with those in benign gastritis(BG).METHODSWe examined the differentially expressed miRNAs in the plasma of 30 patients with EGC, AGC, and BG by miRNA chip analysis. Then, we analyzed and selected the significantly different miRNAs using bioinformatics. Reverse transcription quantitative real-time polymerase chain reaction(RT-qPCR)confirmed the relative transcription level of these miRNAs in another 122 patients, including patients with EGC, AGC, Helicobacter pylori(H. pylori)-negative gastritis(Control-1), and H. pylori-positive atrophic gastritis(Control-2).To establish a diagnostic model for the detection of plasma miRNA in EGC, we chose miRNAs that can be used to determine EGC and AGC from Control-1 and Control-2 and miRNAs in EGC from all other groups.RESULTS Among the expression profiles of the miRNA chips in the three groups in the discovery set, of 117 aberrantly expressed miRNAs, 30 confirmed target prediction, whereas 14 were included as potential miRNAs. The RT-qPCR results showed that 14 potential miRNAs expression profiles in the two groups exhibited no differences in terms of H. pylori-negative gastritis(Control-1) and H. pyloripositive atrophic gastritis(Control-2). Hence, these two groups were incorporated into the Control group. A combination of four types of miRNAs,miR-7641, miR-425-5 p, miR-1180-3 p and miR-122-5 p, were used to effectively distinguish the Cancer group(EGC + AGC) from the Control group [area under the curve(AUC) = 0.799, 95% confidence interval(CI): 0.691-0.908, P < 0.001].Additionally, miR-425-5 p, miR-24-3 p, miR-1180-3 p and miR-122-5 p were utilized to distinguish EGC from the Control group(AUC = 0.829, 95%CI: 0.657-1.000, P =0.001). Moreover, the miR-24-3 p expression level in EGC was lower than that in the AGC(AUC = 0.782, 95%CI: 0.571-0.993, P = 0.029), and the miR-4632-5 p expression level in EGC was significantly higher than that in AGC(AUC = 0.791,95%CI: 0.574-1.000, P = 0.024).CONCLUSION The differentially expressed circulatory plasma miR-425-5 p, miR-1180-3 p, miR-122-5 p, miR-24-3 p and miR-4632-5 p can be regarded as a new potential biomarker panel for the diagnosis of EGC. The prediction and early diagnosis of EGC can be considerably facilitated by combining gastroscopy with the use of these miRNA biomarkers, thereby optimizing the strategy for effective detection of EGC. Nevertheless, larger-scale human experiments are still required to confirm our findings.

Clinical significance of different periampullary diverticulum classifications for endoscopic retrograde cholangiopancreatography cannulation

BACKGROUND Different types of periampullary diverticulum(PAD) may differentially affect the success of endoscopic retrograde cholangiopancreatography(ERCP) cannulation,but the clinical significance of the two current PAD classifications for cannulation is limited.AIM To verify the clinical value of our newly proposed PAD classification.METHODS A new PAD classification(Li-Tanaka classification) was proposed at our center.All PAD patients with native papillae who underwent ERCP from January 2012 to December 2017 were classified according to three classification systems, and the effects of various types of PAD on ERCP cannulation were compared.RESULTS A total of 3564 patients with native papillae were enrolled, including 967(27.13%)PAD patients and 2597(72.87%) non-PAD patients. In the Li-Tanaka classification, type Ⅰ PAD patients exhibited the highest difficult cannulation rate(23.1%, P = 0.01), and type Ⅱ and Ⅳ patients had the highest cannulation success rates(99.4% in type Ⅱ and 99.3% in type Ⅳ, P < 0.001). In a multivariableadjusted logistic model, the overall successful cannulation rate in PAD patients was higher than that in non-PAD patients [odds ratio(OR) = 1.87, 95% confidence interval(CI): 1.04-3037, P = 0.037]. In addition, compared to the non-PAD group,the difficulty of cannulation in the type Ⅰ PAD group according to the Li-Tanaka classification was greater(OR = 2.04, 95%CI: 1.13-3.68, P = 0.004), and the successful cannulation rate was lower(OR = 0.27, 95%CI: 0.11-0.66, P < 0.001),while it was higher in the type Ⅱ PAD group(OR = 4.44, 95%CI: 1.61-12.29, P <0.01).CONCLUSION Among the three PAD classifications, the Li-Tanaka classification has an obvious clinical advantage for ERCP cannulation, and it is helpful for evaluating potentially difficult and successful cannulation cases among different types of PAD patients.

Could saline irrigation clear all residual common bile duct stones after lithotripsy?A self-controlled prospective cohort study

BACKGROUND A previous study showed that irrigation with 100 mL saline reduced residual common bile duct(CBD)stones,which potentially cause recurrent stones after endoscopic retrograde cholangiopancreatography.AIM To determine whether saline irrigation can improve CBD clearance after lithotripsy.METHODS This prospective self-controlled study enrolled patients receiving mechanical lithotripsy for large(>1.2 cm)CBD stones.After occlusion cholangiography confirmed CBD stone clearance,peroral cholangioscopy(POC)was performed to determine clearance scores based on the number of residual stones.The amounts of residual stones spotted via POC were graded on a 5-point scale(score 1,worst;score 5,best).Scores were documented after only stone removal(control)and after irrigation with 50 mL and 100 mL saline,respectively.The stone composition was analyzed using infrared spectroscopy.RESULTS Between October 2018 and January 2020,47 patients had CBD clearance scores of 2.4±1.1 without saline irrigation,3.5±0.7 with 50 mL irrigation,and 4.6±0.6 with 100 mL irrigation(P<0.001).Multivariate analysis showed that CBD diameter>15 mm[odds ratio(OR)=0.08,95%confidence interval(CI):0.01-0.49;P=0.007]and periampullary diverticula(PAD)(OR=6.51,95%CI:1.08-39.21;P=0.041)were independent risk factors for residual stones.Bilirubin pigment stones constituted the main residual stones found in patients with PAD(P=0.004).CONCLUSION Irrigation with 100 mL of saline may not clear all residual CBD stones after lithotripsy,especially in patients with PAD and/or a dilated(>15 mm)CBD.Pigment residual stones are soft and commonly found in patients with PAD.Additional saline irrigation may be required to remove retained stones.

Associations between serum uric acid and hepatobiliary-pancreatic cancer:A cohort study

BACKGROUND Uric acid is the end product of purine metabolism.Previous studies have found that serum uric acid(SUA)levels are associated with the total cancer risk.However,due to the dual effect of uric acid on cancer,the relationship between the SUA levels and most specific-site cancer remains unclear.AIM To investigate the associations between the SUA levels and incidence of hepatobiliary-pancreatic cancer.METHODS In this prospective cohort study,444462 participants free of cancer from the UK Biobank were included.The SUA levels were measured at baseline,and the incidence of hepatobiliary-pancreatic cancer was determined by contacting the cancer registry.The hazard ratios(HRs)and 95%confidence intervals(CIs)between the SUA levels and hepatobiliary-pancreatic cancer were investigated using multiple adjusted Cox regression models adjusted for potential confounders.RESULTS In total,920 participants developed liver,gallbladder,biliary tract or pancreatic cancer during a median of 6.6 yrs of follow-up.We found that the HR of pancreatic cancer in the highest SUA group was 1.77(95%CI:1.29-2.42)compared with that in the lowest group.After stratifying by gender,we further found that SUA was associated with an increased risk of pancreatic cancer only among the females(highest quartile vs lowest quartile HR 2.04,95%CI:1.35-3.08).Among the males,the SUA levels were positively associated with the gallbladder cancer risk(highest quartile vs lowest quartile HR 3.09,95%CI:1.28-7.46),but a U-shaped association with the liver cancer risk was observed(P-nonlinear=0.03).CONCLUSION SUA is likely to have gender-specific effects on hepatobiliary-pancreatic cancer.High SUA levels are a risk factor for pancreatic cancer in females and gallbladder cancer in males.A U-shaped association with the liver cancer risk was identified.

Cytokines predict virological response in chronic hepatitis B patients receiving peginterferon alfa-2a therapy

BACKGROUND Chronic hepatitis B virus infection remains a major global public health problem.Peginterferon-alpha-2a(PEG-IFN)has direct antiviral and immunoregulatory effects,and it has become one of the first choice drugs for the treatment of chronic hepatitis B(CHB).Cytokines play an important role in immunity,and they directly inhibit viral replication and indirectly determine the predominant pattern of the host immune response.AIM To determine the correlation between cytokine/chemokine expression levels and response to PEG-IFN treatment in patients with CHB.METHODS Forty-six kinds of cytokines were analyzed before PEG-IFN therapy and at 24 wk during therapy in 26 CHB patients.RESULTS The monokine induced by INF-γ(CXCL9)and serum interferon-inducible protein 10(IP-10)levels at baseline were higher in virological responders than in nonvirological responders(NRs)and decreased during treatment,whereas the NRs did not exhibit significant changes.The macrophage inflammatory protein 1d(MIP-1d)levels at baseline and during treatment were significantly higher in the virological responders than in the NRs,while thymus and activation-regulated chemokine(TARC)levels at baseline and during treatment were significantly lower in the virological responders than in the NRs.The CXCL9,IP-10,MIP-1d,and TARC baseline levels exhibited the expected effects for interferon treatment.The area under the receiver operating characteristic curve values of CXCL9,IP-10,MIP-1d,and TARC for predicting virological responses were 0.787,0.799,0.787,and 0.77(P=0.01,0.013,0.01,and 0.021),respectively.CONCLUSION We found that cytokine levels before and during treatment may represent potential biomarkers to select CHB patients who can respond to PEG-IFN.Therefore,cytokines can be used as an indicator of antiviral drug selection before CHB treatment.

Three initial diets for management of mild acute pancreatitis:A meta-analysis

AIM:To compare non-liquid and clear-liquid diets,and to assess whether the latter is the optimal treatment for mild acute pancreatitis.METHODS:The Cochrane Library,PUBMED,EMBASE,EBM review databases,Science Citation Index Expanded,and several Chinese databases were searched up to March 2011.Randomized controlled trials(RCTs) that compared non-liquid with clear-liquid diets in patients with mild acute pancreatitis were included.A meta-analysis was performed using available evidence from RCTs.RESULTS:Three RCTs of adequate quality involving a total of 362 participants were included in the final analysis.Compared to liquid diet,non-liquid diet significantly decreased the length of hospitalization [mean difference(MD):1.18,95% CI:0.82-1.55;P﹤0.00001] and total length of hospitalization(MD:1.31,95% CI:0.45-2.17;P = 0.003).The subgroup analysis showed solid diet was more favorable than clear liquid diet in the length of hospitalization,with a pooled MD being-1.05(95% CI:-1.43 to-0.66;P﹤0.00001).However,compared with clear liquid diet,both soft and solid diets did not show any significant differences for recurrence of pain after re-feeding,either alone [relative risk(RR):0.95;95% CI:0.51-1.87;P = 0.88] and(RR:1.22;95% CI:0.69-2.16;P = 0.49),respectively,or analyzed together as non-liquid diet(RR:0.80;95% CI:0.47-1.36;P = 0.41).CONCLUSION:The non-liquid soft or solid diet did not increase pain recurrence after re-feeding,compared with the clear-liquid diet.The non-liquid diet reduced hospitalization.

Choledocholithiasis characteristics with periampullary diverticulum and endoscopic retrograde cholangiopancreatography procedures:Comparison between two centers from Lanzhou and Kyoto

BACKGROUND Most of study regarding periampullary diverticulum(PAD)impact on endoscopic retrograde cholangiopancreatography(ERCP)therapy for choledocholithiasis based on data from one endoscopy center and lacked to compare the clinical characteristic of choledocholithiasis with PAD from different geographical patients.AIM To compare the choledocholithiasis clinical characteristics between two regional endoscopy centers and analyze impacts of clinical characteristics on ERCP methods for choledocholithiasis patients with PAD.METHODS Patients seen in two endoscopy centers(The First Hospital of Lanzhou University,Lanzhou,Gansu Province,China,and Kyoto Second Red Cross Hospital,Kyoto,Japan)underwent ERCP treatment for the first time between January 2012 and December 2017.The characteristics of choledocholithiasis with PAD were compared between the two centers,and their ERCP procedures and therapeutic outcomes were analyzed.RESULTS A total of 829 out of 3608 patients in the Lanzhou center and 241 out of 1198 in the Kyoto center had choledocholithiasis with PAD.Lots of clinical characteristics were significantly different between the two centers.The common bile duct(CBD)diameter was wider,choledocholithiasis size was lager and multiple CBD stones were more in the Lanzhou center patients than those in the Kyoto center patients(14.8±5.2 mm vs 11.6±4.2 mm,12.2±6.5 mm vs 8.2±5.3 mm,45.3%vs 20.3%,P<0.001 for all).In addition,concomitant diseases,such as acute cholangitis,gallbladder stones,obstructive jaundice,cholecystectomy,and acute pancreatitis,were significantly different between the two centers(P=0.03 to<0.001).In the Lanzhou center,CBD diameter and choledocholithiasis size were lower,and multiple CBD stones and acute cholangitis were less in non-PAD patients than those in PAD patients(13.4±5.1 mm vs 14.8±5.2 mm,10.3±5.4 mm vs 12.2±6.5,39%vs 45.3%,13.9%vs 18.5%,P=0.002 to<0.001).But all these characteristics were not significantly different in the Kyoto center.The proportions of endoscopic sphincterotomy(EST),endoscopic balloon dilatation(EPBD),and EST+EPBD were 50.5%,1.7%,and 42.5%in the Lanzhou center and 90.0%,0.0%,and 0.4%in the Kyoto center,respectively.However,the overall post-ERCP complication rate was not significantly different between the two centers(8.9%in the Lanzhou and 5.8%in the Kyoto.P=0.12).In the Lanzhou center,the difficulty rate in removing CBD stones in PAD was higher than in non-PAD group(35.3%vs 26.0%,P<0.001).But the rate was no significant difference between the two groups in Kyoto center.The residual rates of choledocholithiasis were not significantly different between the two groups in both centers.Post-ERCP complications occurred in 8.9%of the PAD patients and 8.1%of the non-PAD patients in the Lanzhou Center,and it occurred in 5.8%in PAD patients and 10.0%in non-PAD patients in the Kyoto center,all P>0.05.CONCLUSION Many clinical characteristics of choledocholithiasis patients with PAD were significantly different between the Lanzhou and Kyoto centers.The patients had larger and multiple stones,wider CBD diameter,and more possibility of acute cholangitis and obstructive jaundice in the Lanzhou center than those in the Kyoto center.The ERCP procedures to manage native duodenal papilla were different depending on the different clinical characteristics while the overall post-ERCP complications were not significantly different between the two centers.The stone residual rate and post-ERCP complications were not significantly different between choledocholithiasis patients with PAD and without PAD in each center.

A novel endoscopic retrograde cholangiopancreatography technique to reduce stone size in type IV Mirizzi syndrome: avoiding cholangiojejunostomy

To the Editor:Mirizzi syndrome(MS)is a rare type of cholelithiasis with an incidence of 0.1%in all gallstone.[1]cases.Its clinical manifestations and imaging features can sometimes be diffcult to distinguish from those of choledocholithiasis and biliary tumors.s.The traditional treatment for MS is usually surgery,including cholecys-tectomy,exploration of common bile.duct plus T-tube drainage,and cholangiojejunostomy.[2] There have even been reports of endoscopic therapies for MS in recent years,but it is technically difficult and non-feasible in cases where the stones get stuck in the Hartmann pouch.[3]Here we report a case where the MS was downstaged via endoscopic modalities to avoid cholangiojejunostomy.

Emergent Endoscopic Retrograde Cholangiopancreatography with Placement of Biliary Double Stents to Salvage Endoscopic Retrograde Cholangiopancreatography-lnduced Stapfer＇s Type Ⅱ Perforation

To the Editor：Endoscopic retrograde cholangiopancreatography （ERCP） plays a vital role in the management ofpancreaticobiliary diseases in recent years and it concomitantly carries a risk of complications including post-ERCP pancreatitis,cholangitis,bleeding,and perforation.The incidence of primary post-ERCP complications ranges from 5.4% to 23.0%,and ERCP-induced perforation can occur in 0.3-1.0% of cases,but the associated mortality is high ranging from 8% to 23%.[1] Because of the confluence of the bile duct and pancreatic duct meet at the papilla in proximity to the site of perforation in the duodenum,there is a potential risk of leakage of bile and pancreatic juice into the retroperitoneal space or peritoneum.Patients with Stapfer＇s Type Ⅱ perforation （perivaterian perforation） generally suffered from systemic inflammatory response syndrome,which could progress rapidly to acute lung injury and acute renal insufficiency and even multiple organ failure.All of these contribute to the significantly high mortality.A timely recognition and appropriate treatment are crucial to the management of ERCP-induced perforation to reduce the overall mortality.Conventionally,surgery remains the primary treatment for iatrogenic perforations.With the improvement in endoscopic technique and development of new accessories,nonsurgical management with endoscopic treatment of perforation is increasingly being reported.[2] We report a retrospective analysis of ERCP-induced Stapfer＇s Type Ⅱ perforations over a 5-year period managed with nonsurgical approach combined with salvage ERCP.

Repair of Large Diaphragmatic Defect Using Artificial Patch in Hydatid Disease

To the Editor： Hydatid is an endemic zoonosis disease which brings great harm to the human. It is popular in developing countries especially in North Western China where animal husbandry is the primary industry. For liver hydatid, a few cases may be associated with diaphragmatic muscle invasion, and even invasion into the chest and the right lung, which become more difficult in clinical treatment.