Fine-needle aspiration technique under endoscopic ultrasound guidance:A technical approach for RNA profiling of pancreatic neoplasms

BACKGROUND Early diagnosis of pancreatic ductal adenocarcinoma(PDAC)has been a longstanding challenge.The prognosis of patients with PDAC depends on the stage at diagnosis.It is necessary to identify biomarkers for the detection and differentiation of pancreatic tumors and optimize PDAC sample preparation procedures for DNA and RNA analysis.Most molecular studies are done using paraffin-embedded blocks;however,the integrity of DNA and RNA is often compromised in this format.Moreover,RNA isolated from human pancreatic tissue samples is generally of low quality,in part,because of the high concentration of endogenous pancreatic RNAse activity present.AIM To assess the potential of endoscopic ultrasound-guided fine-needle aspiration(EUS-FNA)to obtain specimens from pancreatic neoplasms for subsequent RNA molecular profiling,including next-generation sequencing(NGS).METHODS Thirty-four EUS-FNA samples were included in this study:PDAC(n=15),chronic pancreatitis(n=5),pancreatic cysts(n=14),mucinous cysts(mucinous cystic neoplasia/intraductal papillary mucinous neoplasia)n=7,serous cystic neoplasms n=5,and pseudocysts n=2.Cyst material consisted of cyst fluid and cyst wall samples obtained by through-the-needle biopsy(TTNB).Samples were stored at -80℃ until analysis.RNA purity(A260/230,A260/280 ratios),concentration,and integrity(RIN)were assessed.Real-time polymerase chain reaction was conducted on all samples,and small RNA libraries were prepared from solid mass samples.RESULTS RNA was successfully extracted from 29/34(85%)EUS-FNA samples:100% pancreatic adenocarcinoma samples,100% chronic pancreatitis samples,70% pancreatic fluid cyst samples,and 50%TTNB samples.The relative expression of GAPDH and HPRT were obtained for all successfully extracted RNA samples(n=29)including lowquality RNA specimens.Low concentration and nonoptimal RIN values(no less than 3)of RNA extracted from EUS-FNA samples did not prevent NGS library preparation.The suitability of cyst fluid samples for RNA profiling varied.The quality of RNA extracted from mucinous cyst fluid had a median RIN of 7.7(5.0-8.2),which was compatible with that from solid neoplasms[6.2(0-7.8)],whereas the quality of the RNA extracted from all fluids of serous cystic neoplasms and TTNB samples had a RIN of 0.CONCLUSION The results demonstrate the high potential of EUS-FNA material for RNA profiling of various pancreatic lesions,including low-quality RNA specimens.

Endoscopic ultrasound-guided fine-needle aspiration biopsy-Recent topics and technical tips

Endoscopic ultrasound-guided fine-needle aspiration biopsy (EUS-FNA) is a useful procedure that enables reliable pathological diagnoses of pancreatobiliary diseases, subepithelial lesions, and swollen lymph nodes. In recent years, a pathological diagnosis based on EUS-FNA has made it possible to provide accurate treatment methods not only in these fields, but also in respiratory organs and otorhinolaryngology. This review discusses the latest topics pertaining to EUS-FNA as well as procedural tips.

Endoscopic ultrasound-guided tissue acquisition for the diagnosis of focal liver lesion

In patients with liver tumors,the histopathology examination can assist in diagnosis,staging,prognosis,and therapeutic management strategy.Endoscopic ultrasound(EUS)-guided tissue acquisition using fine needle aspiration(FNA)or more newly fine needle biopsy(FNB)is a well-developed technique in order to evaluate and differentiate the liver masses.The goal of the EUS-FNA or EUS-FNB is to provide an accurate sample for a histopathology examination.Therefore,malignant tumors such as hepatocarcinoma,cholangiocarcinoma and liver metastasis or benign tumors such as liver adenoma,focal hyperplastic nodular tumors and cystic lesions can be accurately diagnosed using EUS-guided tissue acquisition.EUS-FNB using 19 or 22 Ga needle provide longer samples and a higher diagnostic accuracy in patients with liver masses when compared with EUS-FNA.Few data are available on the diagnostic accuracy of EUS-FNB when compared with percutaneously,ultrasound,computer tomography or transjugulary-guided liver biopsies.This review will discuss the EUS-guided tissue acquisition options in patients with liver tumors and its efficacy and safety in providing accurate samples.The results of the last studies comparing EUS-guided liver biopsy with other conventional techniques are presented.The EUS-guided tissue acquisition using FNB can be a suitable technique in suspected liver lesions in order to provide an accurate histopathology diagnosis,especially for those who require endoscopy.

Accuracy of endoscopic ultrasound-guided needle aspiration specimens for molecular diagnosis of non-small-cell lung carcinoma

BACKGROUND Endoscopic ultrasonography-guided fine-needle aspiration(EUS-FNA)and endobronchial ultrasound-guided transbronchial needle aspiration(EBUS-TBNA)are highly sensitive for diagnosing and staging lung cancer.In recent years,targeted therapy has shown great significance in the treatment of non-small cell lung carcinoma(NSCLC).Using these minimally invasive techniques to obtain specimens for molecular testing will provide patients with a more convenient diagnostic approach.AIM To evaluate the feasibility and accuracy of tissue samples obtained using EUSFNA and EBUS-TBNA for molecular diagnosis of NSCLC.METHODS A total of 83 patients with NSCLC underwent molecular testing using tissues obtained from EUS-FNA or EBUS-TBNA at the Tianjin Medical University Cancer Hospital from January 2017 to June 2019.All enrolled patients underwent chest computed tomography or positron emission tomography/computed tomography prior to puncture.We detected abnormal expression of EGFR,KRAS,MET,HER2,ROS1 and anaplastic lymphoma kinase protein.Two patients failed to complete molecular testing due to insufficient tumor tissue.The clinical features,puncture records,molecular testing results and targeted treatment in the remaining 81 patients were summarized.RESULTS In a total of 99 tissue samples obtained from 83 patients,molecular testing was successfully completed in 93 samples with a sample adequacy ratio of 93.9%(93/99).Biopsy samples from two patients failed to provide test results due to insufficient tumor tissue.In the remaining 81 patients,62 cases(76.5%)were found to have adenocarcinoma,11 cases(13.6%)had squamous cell carcinoma,3 cases(3.7%)had adenosquamous carcinoma and 5 cases(6.2%)had NSCLC-not otherwise specified.The results of molecular testing showed EGFR mutations in 21 cases(25.9%),KRAS mutations in 9 cases(11.1%),ROS-1 rearrangement in 1 case(1.2%)and anaplastic lymphoma kinase-positive in 5 cases(6.2%).Twentyfour patients with positive results received targeted therapy.The total effectiveness rate of targeted therapy was 66.7%(16/24),and the disease control rate was 83.3%(20/24).CONCLUSION Tissue samples obtained by EUS-FNA or EBUS-TBNA are feasible for the molecular diagnosis of NSCLC and can provide reliable evidence for clinical diagnosis and treatment.

Endoscopic ultrasound-guided through-the-needle microforceps biopsy and needle-based confocal laser-endomicroscopy increase detection of potentially malignant pancreatic cystic lesions:A single-center study

BACKGROUND Currently,there is insufficient data about the accuracy in the diagnosing of pancreatic cystic lesions(PCLs),especially with novel endoscopic techniques such as with direct intracystic micro-forceps biopsy(mFB)and needle-based confocal laser-endomicroscopy(nCLE).AIM To compare the accuracy of endoscopic ultrasound(EUS)and associated techniques for the detection of potentially malignant PCLs:EUS-guided fine needle aspiration(EUS-FNA),contrast-enhanced EUS(CE-EUS),EUS-guided fiberoptic probe cystoscopy(cystoscopy),mFB,and nCLE.METHODS This was a single-center,retrospective study.We identified patients who had undergone EUS,with or without additional diagnostic techniques,and had been diagnosed with PCLs.We determined agreement among malignancy after 24-mo follow-up findings with detection of potentially malignant PCLs via the EUSguided techniques and/or EUS-guided biopsy when available(EUS malignancy detection).RESULTS A total of 129 patients were included, with EUS performed alone in 47/129. In 82/129 patients,EUS procedures were performed with additional EUS-FNA (21/82), CE-EUS (20/82), cystoscopy(27/82), mFB (36/82), nCLE (44/82). Agreement between EUS malignancy detection and the 24-mo follow-up findings was higher when associated with additional diagnostic techniques thanEUS alone [62/82 (75.6%) vs 8/47 (17%);OR 4.35, 95%CI: 2.70-7.37;P < 0.001]. The highestmalignancy detection accuracy was reached when nCLE and direct intracystic mFB were bothperformed, with a sensitivity, specificity, positive predictive value, negative predictive value andobserved agreement of 100%, 89.4%, 77.8%, 100% and 92.3%, respectively (P < 0.001 comparedwith EUS-alone).CONCLUSIONThe combined use of EUS-guided mFB and nCLE improves detection of potentially malignantPCLs compared with EUS-alone, EUS-FNA, CE-EUS or cystoscopy.

Endoscopic ultrasound guided liver biopsy: Recent evidence

Liver biopsy(LB)is an essential tool in diagnosing,evaluating and managing various diseases of the liver.As such,histopathological results are critical as they establish or aid in diagnosis,provide information on prognosis,and guide the appropriate selection of medical therapy for patients.Indications for LB include evaluation of persistent elevation of liver chemistries of unclear etiology,diagnosis of chronic liver diseases such as Wilson's disease,autoimmune hepatitis,small duct primary sclerosing cholangitis,work up of fever of unknown origin,amyloidosis and more.Traditionally,methods of acquiring liver tissue have included percutaneous LB(PCLB),transjugular LB(TJLB)or biopsy taken surgically via laparotomy or laparoscopy.However,traditional methods of LB may be inferior to newer methods.Additionally,PCLB and TJLB carry higher risks of adverse events and complications.More recently,endoscopic ultrasound guided LB(EUS-LB)has evolved as an alternative method of tissue sampling that has proven to be safe and effective,with limited adverse events.Compared to PC and TJ routes,EUS-LB may also have a greater diagnostic yield of tissue,be superior for a targeted approach of focal lesions,provide higher quality images and allow for greater patient comfort.These advantages have contributed to the increased use of EUS-LB as a technique for obtaining liver tissue.Herein,we provide a review of the recent evidence of EUS-LB for liver disease.

Appropriate number of biliary biopsies and endoscopic retrograde cholangiopancreatography sessions for diagnosing biliary tract cancer

BACKGROUND Biliary ductal cancer(BDC) is a lethal disease; however, diagnosing BDC is challenging. Biliary biopsies are performed to pathologically diagnose BDC, but the appropriate parameters for biliary biopsy [number of biliary biopsies, number of endoscopic retrograde cholangiopancreatography(ERCP) sessions, etc.] are unknown.AIM To clarify what constitutes an adequate method for biliary biopsy.METHODS In total, 95 patients who underwent endoscopic biliary biopsy without choledochoscopy and who were pathologically diagnosed with BDC were enrolled in this study. The patients were divided into two groups. Seventy-six patients who were diagnosed by biliary biopsy were defined as the positive group(P group), and nineteen patients who were not diagnosed by biliary biopsy were defined as the negative group(N group). The patient characteristics and ERCP-related procedures were compared between the P and N groups.RESULTS The numbers of ERCP sessions and biliary biopsies were significantly different between the two groups [ERCP sessions(one/two), P group 72/4 vs N group15/4, P value = 0.048; number of biliary biopsies, P group 2(1-6) vs N group 2(1-7), P value = 0.039]. In a multivariate analysis, fewer than 2 ERCP sessions was an independent factor influencing the positivity of the biliary biopsies.CONCLUSION This study clarified that ERCP and biliary ductal biopsy should only be performed once. If biliary cancer is not pathologically diagnosed after the first ERCP session, other methods(Endoscopic ultrasonography-guided fine needle aspiration or choledochoscopy-guided biliary ductal biopsy) should be employed.

Overlooked risk for needle tract seeding following endoscopic ultrasound-guided minimally invasive tissue acquisition

Endoscopic ultrasound-guided minimally invasive tissue acquisition can be performed by two approaches as follows: Endoscopic ultrasound-guided fineneedle aspiration(EUS-FNA) and endoscopic ultrasound-guided fine-needle biopsy(EUS-FNB). These have been evolved into leading approaches and widely used for the histological diagnosis of tumors in the gastrointestinal tract and adjacent organs. However, the role of EUS-FNA and EUS-FNB in disease diagnosis and evaluation remains controversial. Although the incidence of surgery-associated complications remains low, the consequences of needle tract seeding can be serious or even life-threatening. Recently, increasing case reports of needle tract seeding are emerging, especially caused by EUS-FNA. This complication needs serious consideration. In the present work, we integrated these case reports and the related literature, and summarized the relevant cases and technical characteristics of needle tract seeding caused by EUS-FNA and EUSFNB. Collectively, our findings provided valuable insights into the prevention and reduction of such serious complication.

Evaluating the risk of adverse events with interventional endoscopic retrograde cholangiopancreatography and endoscopic ultrasound procedures in cirrhotic patients

BACKGROUND Hepatic cirrhosis is associated with greater adverse event rates following surgical procedures and is thought to have a higher risk of complications with interventional procedures in general.However,these same patients often require interventional gastrointestinal procedures such as endoscopic retrograde cholangiopancreatography(ERCP)and endoscopic ultrasound(EUS).While studies examining this scenario exist,the overall body of evidence for adverse event rates associated with ERCP/EUS procedures is more limited.We sought add to the literature by examining the incidence of adverse events after ERCP/EUS procedures in our safety-net hospital population with the hypothesis that severity of cirrhosis correlates with higher adverse event rates.AIM To examine whether increasing severity of cirrhosis is associated with greater incidence of adverse events after interventional ERCP/EUS procedures.METHODS We performed a retrospective study of patients diagnosed with hepatic cirrhosis who underwent ERCP and/or EUS-guided fine needle aspirations/fine needle biopsies from January 1,2016 to March 14,2019 at our safety net hospital.We recorded Child-Pugh and Model for End-stage Liver Disease(MELD-Na)scores at time of procedure,interventions completed,and 30-day post-procedural adverse events.Statistical analyses were done to assess whether Child-Pugh class and MELD-Na score were associated with greater adverse event rates and whether advanced techniques(single-operator cholangioscopy,electrohydraulic lithotripsy/laser lithotripsy,or needle-knife techniques)were associated with higher complication rates.RESULTS 77 procedures performed on 36 patients were included.The study population consisted primarily of middle-aged Hispanic males.30-d procedure-related adverse events included gastrointestinal bleeding(7.8%),infection(6.5%),and bile leak(2%).The effect of Child-Pugh class C vs class A and B significantly predicted adverse events(β=0.55,P<0.01).MELD-Na scores also significantly predicted adverse events(β=0.037,P<0.01).Presence of advanced techniques was not associated with higher adverse events(P>0.05).When MELD-Na scores were added as predictors with the effect of Child-Pugh class C,logistic regression showed MELD-Na scores were a significant predictor of adverse events(P<0.01).The findings held after controlling for age,gender,ethnicity and repeat cases.CONCLUSION Increasing cirrhosis severity predicted adverse events while the presence of advanced techniques did not.MELD-Na score may be more useful in predicting adverse events than Child-Pugh class.

Mucosal-incision assisted biopsy for suspected gastric gastrointestinal stromal tumors

To evaluate the diagnostic yield of the procedure, mucosal-incision assisted biopsy (MIAB), for the histological diagnosis of gastric gastrointestinal stromal tumor (GIST), we performed a retrospective review of the 27 patients with suspected gastric GIST who underwent MIAB in our hospitals. Tissue samples obtained by MIAB were sufficient to make a histological diagnosis (diagnostic MIAB) in 23 out of the 27 patients, where the lesions had intraluminal growth patterns. Alternatively, the samples were insufficient (non-diagnosticMIAB) in remaining 4 patients, three of whom had gastric submucosal tumor with extraluminal growth patterns. Although endoscopic ultrasound and fine needle aspiration is the gold standard for obtaining tissue specimens for histological and cytological analysis of suspected gastric GISTs, MIAB can be used as an alternative method for obtaining biopsy specimens of lesions with an intraluminal growth pattern.

Clinical course of suspected small gastrointestinal stromal tumors in the stomach

BACKGROUND Gastric subepithelial lesions are frequently encountered during endoscopic examinations,and the majority of them are small and asymptomatic.Among these lesions,gastrointestinal stromal tumors(GISTs)are the major concern for patients and clinicians owing to their malignant potentials.Although previous guidelines suggested periodic surveillance for such small(≤20 mm)lesions,several patients and clinicians have still requested or prescribed repeated examinations or radical resection,posing extra medical burdens and risks.AIM To describe the clinical course of suspected small gastric GISTs and provide further evidence for surveillance strategy for tumor therapy.METHODS This single-center,retrospective study was conducted at West China Hospital,Sichuan University.Consecutive patients with suspected small gastric GISTs were reviewed from November 2004 to November 2018.GIST was suspected according to endoscopic ultrasonography features:hypoechoic lesions from muscularis propria or muscularis mucosa.Eligible patients with suspected small(≤20 mm)GISTs were included for analysis.Patients’demographic data,lesions’characteristics,and follow-up medical records were collected.RESULTS A total of 383 patients(male/female,121/262;mean age,54 years)with 410 suspected small gastric GISTs(1 lesion in 362 patients,2 lesions in 16,3 lesions in4,and 4 lesions in 1)were included for analysis.The most common location was gastric fundus(56.6%),followed by body(29.0%),cardia(12.2%),and antrum(2.2%).After a median follow-up of 28 mo(interquartile range,16-48;range,3-156),402 lesions(98.0%)showed no changes in size,and size of 8 lesions(2.0%)was increased(mean increment,10 mm).Of the 8 lesions with size increment,endoscopic or surgical resection was performed in 6 patients(5 GISTs and 1 leiomyoma).For other 2 remaining patients,unroofing biopsy or endoscopic ultrasound-guided fine-needle aspiration was carried out(2 GISTs),while no further change in size was noted over a period of 62-64 mo.CONCLUSION The majority of suspected small(≤20 mm)gastric GISTs had no size increment during follow-up.Regular endoscopic follow-up without pathological diagnosis may be highly helpful for such small gastric subepithelial lesions.

Role of the preoperative usefulness of the pathological diagnosis of pancreatic diseases

Pancreatic cancer is the fifth leading cause of cancer death and has the lowest survival rate of any solid cancer.Endoscopic ultrasound-guided fine-needle aspiration biopsy(EUS-FNA) is currently capable of providing a cytopathological diagnosis of pancreatic malignancies with a higher diagnostic power,with a sensitivity and specificity of 85%-89% and 98%-99%,compared to pancreatic juice cytology(PJC),whose sensitivity and specificity are only 33.3%-93% and 83.3%-100%.However,EUS-FNA is not effective in the cases of carcinoma in situ and minimally invasive carcinoma because both are undetectable by endoscopic ultrasonography,although PJC is able to detect them.As for the frequency of complications such as post endoscopic retrograde cholangiopancreatography pancreatitis,EUS-FNA is safer than PJC.To diagnose pancreatic cancer appropriately,it is necessary for us to master both procedures so that we can select the best methods of sampling tissues while considering the patient's safety and condition.

Comparison of smear cytology with liquid-based cytology in pancreatic lesions: A systematic review and meta-analysis

BACKGROUND Endoscopic ultrasonography-guided fine-needle aspiration(EUS-FNA)is a safe and accurate technique to confirm the diagnosis of pancreatic cancers.Recently,numerous studies comparing the diagnostic efficacy of smear cytology(SC)and liquid-based cytology(LBC)for pancreatic lesions yielded mixed results.AIM To compare and identify the better cytology method for EUS-FNA in pancreatic lesions.METHODS A comprehensive search of PubMed,Embase,and Cochrane was undertaken through July 18,2020.The primary endpoint was diagnostic accuracy(sensitivity and specificity).Secondary outcomes included sample adequacy and post procedure complications.In addition,factors affecting diagnostic efficacy were discussed.RESULTS Data on a total of 1121 comparisons from 10 studies met the inclusion criteria.Pooled rates of sensitivity for SC and LBC were 78%(67%-87%)vs 75%(67%-81%),respectively.In any case,both SC and LBC exhibited a high specificity close to 100%.Inadequate samples more often appeared in LBC compared with SC.However,the LBC samples exhibited a better visual field than SC.Very few post procedure complications were observed.CONCLUSION Our data suggested that for EUS-FNA in pancreatic lesions(particularly solid lesions),SC with Rapid On-Site Evaluation represents a superior diagnostic technique.If Rapid On-Site Evaluation is unavailable,LBC may replace smears.The diagnostic accuracy of LBC depends on different LBC techniques.

Nonfunctional pancreatic neuroendocrine tumours misdiagnosed as autoimmune pancreatitis:A case report and review of literature

BACKGROUND Nonfunctional pancreatic neuroendocrine tumours are difficult to diagnose in the early stage of disease due to a lack of clinical symptoms,but they can rarely manifest as autoimmune pancreatitis.Autoimmune pancreatitis is an uncommon disease that may cause recurrent acute pancreatitis and is therefore often regarded as a special type of chronic pancreatitis.CASE SUMMARY We report a case of a 42-year-old female who had nonspecific upper abdominal pain for 4 years and radiological abnormalities of the pancreas that mimicked autoimmune pancreatitis.The symptoms and pancreatic imaging did not improve following 1 year of steroid therapy.Finally,pancreatic biopsy was performed through endoscopic ultrasonography-guided fine-needle aspiration biopsy,and nonfunctional pancreatic neuroendocrine tumours were ultimately diagnosed.Pancreatectomy has resolved her symptoms.CONCLUSION Therefore,the differentiation of nonfunctional pancreatic neuroendocrine tumours from autoimmune pancreatitis is very important,although it is rare.We propose that endoscopic ultrasonography-guided fine-needle aspiration biopsy should be performed if imaging characteristics are equivocal or the diagnosis is in question.

内镜超声引导下细针穿刺活检用于实质病变的标本获取:一项前瞻性队列研究

背景:内镜超声引导下细针穿刺活检(EUS-FNB)的应用日渐增多以提高实质病变的细胞学检出率,但其相较于目前的细针穿刺抽吸(FNA)的优越性并未获明确证实。本研究旨在比较在EUS引导下采用新型Franseen-tip穿刺针的FNB与传统FNA的诊断准确性及操作相关指标。方法:入组接受EUS-FNB检查的连续实质病变患者,前瞻性记录其操作相关指标,包括患者人口统计资料、穿刺次数、诊断样本的充分性、不良反应和恢复时间。研究组采用Acquire针抽取样本。同时纳入行EUSFNA检查的实质病变患者作为对照组,对照组采用Expect针抽取样本。结果:180例行EUS-FNB和183例行EUS-FNA进行实质病变样本采集的患者纳入研究。FNB组较FNA组平均操作时间显著缩短(37.4 min vs 44.9 min,P<0.001),平均穿刺次数明显减少(2.9次vs 3.8次,P<0.001),细胞学检出率明显提高(98.3%vs 90.2%,P=0.003)。两组不良事件发生率的差异无统计学意义(1.1%vs 0.5%,P=0.564)。结论:EUS-FNB用于实质病变的病理活检是安全可行的,可减少穿刺次数,缩短操作时间,提高病变检出率。FNB或可替代FNA作为所有实质病变的首选标本获取方法。