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.

Role of macroscopic on-site evaluation of endoscopic ultrasoundguided fine-needle aspiration/biopsy:Results of a multicentric prospective study

BACKGROUND The concept of macroscopic on-site evaluation(MOSE)was introduced in 2015 when the endoscopist observed better diagnostic yield when the macroscopically visible core on MOSE was superior to 4 mm.Recent studies suggest that MOSE by the endoscopist may be an excellent alternative to rapid on-site evaluation,and some classi-fications have been published.Few studies have assessed the adequacy of histologic cores in MOSE during endoscopic ultrasound-guided fine-needle aspiration/biopsy(EUS-FNA/FNB).AIM To evaluate the performance of MOSE during EUS-FNA/FNB.METHODS This multicentric prospective study was conducted in 16 centers in 3 countries(Egypt,Iraq,and Morocco)and included 1108 patients with pancreatic,biliary,or gastrointestinal pathology who were referred for EUS examination.We prospectively analyzed the MOSE in 1008 patients with available histopathological reports according to 2 classifications to determine the adequacy of the histological core samples.Data management and analysis were performed using a Statistical Package for Social Sciences(SPSS)version 27.RESULTS A total of 1074 solid lesions were biopsied in 1008 patients with available cytopathological reports.Mean age was 59 years,and 509 patients(50.5%)were male.The mean lesion size was 38 mm.The most frequently utilized needles were FNB-Franseen(74.5%)and 22 G(93.4%),with a median of 2 passes.According to 2 classifications,618 non-bloody cores(61.3%)and 964 good samples(95.6%)were adequate for histological evaluation.The overall diagnostic yield of cytopathology was 95.5%.The cytological examination confirmed the diagnosis of malignancy in 861 patients(85.4%),while 45 samples(4.5%)were inconclusive.Post-procedural adverse events occurred in 33 patients(3.3%).Statistical analysis showed a difference between needle types(P=0.035)with a high sensitivity of FNB(97%).The analysis of the relationship between the MOSE-score and the final diagnosis showed a significant difference between the different scores of the MOSE(P<0.001).CONCLUSION MOSE is a simple method that allows endoscopists to increase needle passes to improve sample quality.There is significantly higher FNB sensitivity and cytopathology diagnostic yield with good MOSE cores.

Influence of the safety and diagnostic accuracy of preoperative endoscopic ultrasound-guided fine-needle aspiration for resectable pancreatic cancer on clinical performance

AIM: To evaluate the safety and diagnostic accuracy of endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) in a cohort of pancreatic cancer patients.

Diagnostic utility of endoscopic ultrasound-guided fineneedle aspiration biopsy for glomus tumor of the stomach

A 52-year-old man was referred for further investigation of a gastric submucosal tumor on the greater curvature of the antrum. Endoscopic ultrasonography demonstrated a hypoechoic solid mass, which was primarily connected to the muscular layer of the stomach. We performed endoscopic ultrasoundguided fine-needle aspiration biopsy. The pathological examination showed proliferation of oval-shaped cells with nest formation, which stained strongly positive for muscle actin, and negative for c-kit, CD34, CD56,desmin, S-100, chromogranin, and neuron-specific enolase. Therefore, we performed laparoscopy and endoscopy cooperative surgery based on the preoperative diagnosis of glomus tumor of the stomach. The final histological diagnosis confirmed the preoperative diagnosis. Although preoperative diagnosis of glomus tumor of the stomach is difficult with conventional images and endoscopic biopsy, endoscopic ultrasoundguided fine-needle aspiration biopsy is an essential tool to gain histological evidence of glomus tumor of the stomach for early diagnosis.

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.

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.

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.

Accuracy of endoscopic ultrasound-guided tissue acquisition in the evaluation of lymph nodes enlargement in the absence of on-site pathologist

To evaluate factors that influence the diagnostic accuracy of endoscopic ultrasound (EUS)-guided tissue acquisition for lymph node enlargement in the absence of an on-site pathologist. METHODSA retrospective analysis of patients who underwent EUS-guided tissue acquisition for the pathological diagnosis of lymph node enlargement between April 2012 and June 2015 is reported. Tissue acquisition was performed with both cytology and biopsy needles of different calibers. The variables evaluated were lymph node location and size, number of passes and type of needle used. Final diagnosis was based on surgical histopathology or, in non-operated cases, on EUS-guided tissue acquisition and imaging assessment with a minimum clinical follow-up of 6 mo. RESULTSDuring the study period, 168 lymph nodes with a median size of 20.3 mm (range 12.5-27) were sampled from 152 patients. Ninety lymph nodes (53.6%) were located at mediastinum, and 105 (62.5%) were acquired with biopsy needles. The final diagnosis was benign/reactive origin in 87 cases (51.8%), malignant in 65 cases (38.7%), and lymphoma in 16 cases (9.5%). The sensitivity, specificity, positive predictive value and negative predictive value for the detection of malignancy were 74.1%, 100%, 100% and 80.6%, respectively. The overall accuracy was 87.5% (95%CI: 81.7-91.7). No variables were independently associated with a correct final diagnosis according to the multivariate analysis. CONCLUSIONEUS-guided tissue acquisition is a highly accurate technique for assessing lymph node enlargement. None of the variables evaluated were associated with diagnostic accuracy.

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.

An exclusive fine-needle biopsy approach to sampling solid lesions under EUS guidance:a prospective cohort study

Background Endoscopic ultrasound-guided fine-needle biopsy(EUS-FNB)is increasingly utilized to enhance the cytological yield of sampling solid lesions,but its superiority over existing fine-needle aspiration(FNA)platforms has not been clearly demonstrated.The aim of our study was to compare the diagnostic accuracy and procedural outcomes of FNB using a new Franseen-tip needle to that of a traditional FNA in sampling solid lesions under EUS guidance.Methods Consecutive patients with solid lesions referred for EUS-FNB sampling were included.Procedure-related outcomes were collected prospectively including patient demographics,number of passes performed,diagnostic sample adequacy,adverse events,and recovery time.The Acquire needle was used to sample all lesions in the study group.Consecutive EUSFNA procedures performed to sample solid lesions using the Expect needle were utilized as controls.Results There were 180 patients undergoing EUS-FNB compared to 183 patients undergoing EUS-FNA procedures for solidlesion sampling.The procedure time was significantly shorter in patients who underwent FNB compared to FNA(mean:37.4 vs 44.9 minutes,P<0.001).Significantly fewer passes were performed in the FNB cohort compared to the FNA group(mean:2.9 vs 3.8,P<0.001).The cytologic diagnostic yield was significantly higher in the FNB group compared to the FNA group(98.3%vs 90.2%,P=0.003).No significant difference in the incidence of adverse events was observed between the FNB and FNA groups(1.1%vs 0.5%,P=0.564).Conclusions An FNB-exclusive approach to sampling solid lesions under EUS guidance is safe and feasible,and may result in fewer overall passes,shorter procedure time,and improved diagnostic adequacy.FNB may replace FNA as the primary sampling modality of choice in all solid lesions.

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.