The m6A methyltransferase METTL5 promotes neutrophil extracellular trap network release to regulate hepatocellular carcinoma progression.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common malignant tumors worldwide, it has a poor prognosis due to its highly invasive and metastatic nature. Consequently, identifying effective prognostic markers and potential therapeutic targets has been extensively investigated. METTL5, an 18S rRNA methyltransferase, is abnormally high in HCC. But its biological function and prognostic significance in HCC remain largely unelucidated. This study aimed to investigate the role of METTL5 in HCC progression, and elucidate its possible molecular mechanisms in HCC via transcriptome sequencing, providing new insights for identifying new HCC prognostic markers and therapeutic targets. METHODS: The METTL5 expression in HCC and paracancerous tissues was analyzed using HCC immunohistochemical microarrays and bioinformatic retrieval methods to correlate METTL5 with clinicopathological features and survival prognosis. We constructed a METTL5 knockdown hepatocellular carcinoma cell line model and an animal model to determine the effect of METTL5 on hepatocellular carcinoma progression. Subsequently, RNA sequencing was performed to analyze the molecular mechanism of METTL5 in HCC based on the sequencing results, and relevant experiments were performed to verify it. RESULTS: We found that METTL5 expression was elevated in hepatocellular carcinoma tissues and correlated with poor patient prognosis, and in the analysis of clinicopathological features showed a correlation with TNM staging. In hepatocellular carcinoma cell lines with knockdown of METTL5, the malignant biological behavior was significantly reduced both in vitro and in vivo. Based on the sequencing results as well as the results of GO functional enrichment analysis and KEGG pathway enrichment analysis, we found that METTL5 could promote the generation and release of neutrophil extracellular capture network (NETs) and might further accelerate the progression of HCC. CONCLUSION: The m6A methyltransferase METTL5 is overexpressed in hepatocellular carcinoma (HCC) and correlates with poor prognosis. METTL5 accelerates malignant progression of HCC by promoting generation and release of the neutrophil extracellular traps (NETs) network, providing new insights for clinical biomarkers and immunotherapeutic targets in HCC prognosis.

Overexpression of POLA2 in hepatocellular carcinoma is involved in immune infiltration and predicts a poor prognosis.

BACKGROUND: Hepatocellular carcinoma (HCC) is the second malignancy worldwide. POLA2 initiates DNA replication, regulates cell cycle and gene repair that promote tumorigenesis and disease progression. However, the prognostic and biological function roles of POLA2 in HCC had not been conclusively determined. METHODS: The expression levels and prognosis role of POLA1 and POLA2 in HCC were analyzed based on TCGA-LIHC database and recruited 24 HCC patients. Gene mutations were analyzed using "maftools" package. POLA2 and immune cells correlations were analyzed by TIMER. POLA2 co-expressed genes functional enrichment were evaluated using Metascape. The mRNA and protein level of POLA2 was detected in HCC cells and tissues. Cell migration, invasion, proliferation, cell cycle and HCC cell lines derived xenograft model were performed to investigate POLA2 biological function. RESULTS: POLA2 was significantly high expressed in HCC than in normal liver tissue in both TCGA-LIHC and our collected HCC samples. In validation cohort, POLA2 significantly related to tumor differentiation, tumor size and Ki-67 (p < 0.05). In TCGA-LIHC cohort, overexpression of POLA2 predicted a low OS and associated with different clinical stages. Multivariate Cox regression showed overexpression of POLA2 effectively distinguished the prognosis at different T, N, M, stages and grades of HCC. POLA2 expression correlated with mutation burden, immune cells infiltration and immune-associated genes expression of HCC. Functional enrichment revealed that POLA2 co-expressed genes were linked to cellular activity, plasma membrane protein complex and leukocyte activity, immune response-regulated cell surface receptor signaling pathway, and immune response-regulated signaling pathway. Moreover, POLA2 was also positively co-expressed with some immune checkpoints (CD274, CTL-4, HAVCR2, PDCD1, PDCD1LG2, TIGIT, and LAG3) (p < 0.001). Gene knockdown revealed that POLA2 promoted proliferation, migration, invasion, and cell cycle of SMMC-7721 and HepG2. The HCC xenograft tumor model also demonstrated remarkably tumor size inhibition, tumor proliferation inhibtion and tumor necrosis promotion when POLA2 knockdown. CONCLUSIONS: POLA2 influenced immune microenvironment and tumor progression of HCC indicated that it might be a potential molecular marker for prognostic evaluation or a therapeutic target for HCC.

MACC1 and Gasdermin-E (GSDME) regulate the resistance of colorectal cancer cells to irinotecan.

Metastasis-associated in colon cancer 1 (MACC1) is an oncogene associated with the progression and metastasis of many solid cancer entities. High expression of MACC1 is found in colorectal cancer (CRC) tissues. So far, the role of MACC1 in CRC cell pyroptosis and resistance to irinotecan is unclear. The cleavage of Gasdermin-E (GSDME) is the main executors of activated pyroptosis. We found that GSDME enhanced CRC cell pyroptosis and reduced their resistance to irinotecan, while MACC1 inhibited the cleavage of GSDME and CRC cell pyroptosis, promoted CRC cell proliferation, and enhanced the resistance of CRC cells to irinotecan. Therefore, CRC cells with high MACC1 expression and low GSDME expression had higher resistance to irinotecan, while CRC cells with low MACC1 expression and high GSDME expression had lower resistance to irinotecan. Consistently, by analyzing CRC patients who received FOLFIRI (Fluorouracil + Irinotecan + Leucovorin) in combination with chemotherapy in the GEO database, we found that CRC patients with low MACC1 expression and high GSDME expression had higher survival rate. Our study suggests that the expression of MACC1 and GSDME can be used as detection markers to divide CRC patients into irinotecan resistant and sensitive groups, helping to determine the treatment strategy of patients.

Hepatocyte-derived extracellular vesicles miR-122-5p promotes hepatic ischemia reperfusion injury by regulating Kupffer cell polarization.

Ischemia reperfusion injury remains a major barrier to liver transplantation, especially using grafts from donation after circulatory death, and it is also a pressing issue to be solved in clinical practice. Kupffer cell polarization toward a proinflammatory M1 phenotype is an early trigger of liver ischemia-reperfusion injury. However, the molecular mechanism regulating Kupffer cell polarization has not yet been fully elucidated. We induced liver ischemia reperfusion injury in mice and obtained samples from patients undergoing liver transplantation, serum and hepatocytes-derived extracellular vesicles were isolated by differential ultracentrifugation. Kupffer cell polarization was examined by flow cytometry and immunofluorescence histochemistry. RNA-seq was conducted to detect the differentially expressed miRNAs in extracellular vesicles. The role and mechanism of exosomal miR-122-5p in liver ischemia-reperfusion injury were determined both in vitro and in vivo. We identified ischemia reperfusion induced extracellular vesicles as a major cause of hepatic inflammation and tissue damage using adoptive transfer and release inhibition. The study also demonstrated that hepatocyte-derived exosomal miR-122-5p mediates liver ischemia reperfusion injury by polarizing Kupffer cell via PPARδ down-regulation and NF-κB pathway activation using profiling and functional analysis. Moreover, inhibiting miR-122-5p with antagomir suppressed Kupffer cell M1 polarization and attenuated liver ischemia reperfusion injury. Overall, our study demonstrated that hepatocyte-derived exosomal miR-122-5p played a critical role in promoting hepatic ischemia reperfusion injury through modulating PPARδ signaling and NF-κB pathway to introduce M1 polarization of Kupffer cell. Inhibition of miR-122-5p exhibited a protective effect against liver ischemia reperfusion injury, suggesting a potential therapeutic target for liver transplantation.

Establishment and validation of nomogram model for the diagnosis of AFP-negative hepatocellular carcinoma.

Introduction: As one of the most common malignant tumors in clinical practice, hepatocellular carcinoma (HCC) is a major threat to human health, where alpha-fetoprotein (AFP) is widely used for early screening and diagnoses. However, the level of AFP would not elevate in about 30-40% of HCC patients, which is clinically referred to as AFP-negative HCC, with small tumors at an early stage and atypical imaging features, making it difficult to distinguish benign from malignant by imaging alone. Methods: A total of 798 patients, with the majority being HBV-positive, were enrolled in the study and were randomized 2:1 to the training and validation groups. Univariate and multivariate binary logistic regression analyses were used to determine the ability of each parameter to predict HCC. A nomogram model was constructed based on the independent predictors. Results: A unordered multicategorical logistic regression analyses showed that the age, TBIL, ALT, ALB, PT, GGT and GPR help identify non-hepatic disease, hepatitis, cirrhosis, and hepatocellular carcinoma. A multivariate logistic regression analyses showed that the gender, age, TBIL, GAR, and GPR were independent predictors for the diagnosis of AFP-negative HCC. And an efficient and reliable nomogram model (AUC=0.837) was constructed based on independent predictors. Discussion: Serum parameters help reveal intrinsic differences between non-hepatic disease, hepatitis, cirrhosis, and HCC. The nomogram based on clinical and serum parameters could be used as a marker for the diagnosis of AFP-negative HCC, providing an objective basis for the early diagnosis and individualized treatment of hepatocellular carcinoma patients.

Lesion human leukocyte antigen-E is associated with favourable prognosis for patients with oesophageal squamous cell carcinoma.

OBJECTIVE: To investigate the clinical significance of human leukocyte antigen (HLA)-E levels in oesophageal squamous cell carcinoma (ESCC). METHODS: The levels of HLA-E immunostaining in ESCC lesions and 47 corresponding adjacent normal tissues were measured using immunohistochemistry. The correlation between the levels of immunostaining and clinical parameters was analysed. RESULTS: This study analysed 110 paraffin-embedded primary tumour lesions and 47 case-controlled paracancerous tissues that were surgically resected from 110 patients with ESCC. Positive immunostaining for HLA-E was observed in 88.2% (97 of 110) of ESCC lesions and 29.8% (14 of 47) of normal oesophageal tissues. There was no correlation between HLA-E immunostaining in ESCC lesions and clinicopathological characteristics such as lymph node metastasis, tumour-node-metastasis stage and differentiation grade. Kaplan-Meier survival analysis revealed a significantly better prognosis in patients with higher levels of HLA-E immunostaining than in those with lower levels of HLA-E immunostaining; overall survival was 28.6 months (95% confidence interval [CI], 23.2, 34.0) versus 15.3 months (95% CI, 11.5, 19.1), respectively. Furthermore, multivariate analysis showed that the HLA-E level was an independent prognostic factor in patients with ESCC. CONCLUSION: A higher level of HLA-E immunostaining was associated with favourable survival in patients with ESCC.

Laparoscopic cholecystectomy with two incisions: an improved, feasible and safe technique with superior cosmetic outcomes.

OBJECTIVE: Conventional laparoscopic cholecystectomy (CLC) is usually performed with four incisions. Minimally invasive surgery for gallbladder disease with less pain and smaller scars has become increasingly popular. This study reported a new, two-incision laparoscopic cholecystectomy (TILC) using conventional instruments. METHODS: In this prospective study, 43 patients were recruited to undergo TILC and were compared with 43 historical cases undergoing CLC. We evaluated operative time, postoperative pain, cosmesis and complications. RESULTS: There was no significant difference in gender, age, body mass index, bile duct damage, blood loss and postoperative hospital stay between the two groups. The mean operation time was longer with TILC than with CLC, but the difference was not statistically different. Postoperative pain scores were significantly lower with TILC than with CLC. The mean cosmetic satisfaction score was significantly higher with TILC than that with CLC. There was no significant difference in the incidence of complications between the two groups. CONCLUSION: Our work demonstrates that TILC generates less postoperative pain and significantly improved cosmesis for patients. TILC is a safe and feasible alternative to CLC.

The role of vesicular overexpressed in cancer pro-survival protein 1 in hepatocellular carcinoma proliferation.

BACKGROUND: Recently, hepatocellular carcinoma (HCC) has been ranked as the second leading cause of cancer-associated death. However, the underlying molecular mechanisms of HCC progression remain unclear. Vesicular overexpressed in cancer pro-survival protein 1 (VOPP1) could be upregulated in a quantity of human cancers, including squamous cell carcinoma (SCC), gastric cancer, and glioblastoma. However, the precise functional mechanism of VOPP1 in HCC remains poorly understood. The present study aimed to investigate the role of VOPP1 in HCC proliferation. METHODS: Immunohistochemistry (IHC), Western blot and Reverse-transcription polymerase chain reaction (RT-PCR) were used to analyze the protein and mRNA expressions of VOPP1, mitogen-activated protein kinase (MAPK) 14, ribosomal protein S6 kinase ß1 (RPS6KB1), cylindromatosis (CYLD) and Twist family bHLH transcription factor 1 (TWIST1). The cell proliferation and apoptosis were tested using Celigo cell imaging analyzer and annexin V-APC apoptosis detection kit respectively. Colony formation and tumor xenograft assays were performed to understand their roles in tumorigenicity. RESULTS: The expression of VOPP1 in HCC samples was higher than that in adjacent noncancerous tissues by immunohistochemistry. In addition, the down-regulation of VOPP1 using shRNA inhibited cell proliferation and tumour growth, and induced cell apoptosis in vitro and in vivo. Furthermore, VOPP1 silencing increased the expression of MAPK14 and RPS6KB1, indicating that the MAPK and mTOR signalling pathways might be involved in VOPP1-mediated cancer cell proliferation. CONCLUSION: The present data indicate that VOPP1 may play an important role in the progression of HCC by targeting the MAPK and mTOR signalling pathways, and that VOPP1 may potentially be a candidate as a novel molecular target for HCC therapy.

Hepatocellular HO-1 mediated iNOS-induced hepatoprotection against liver ischemia reperfusion injury.

Hepatocyte-derived inducible nitric oxide synthase (iNOS) was proved to impart protection against liver ischemia reperfusion (I/R) injury in our prior analysis. However, the mechanism for this hepatoprotection remains incompletely understood. Bone marrow chimeric mice were generated using expression of iNOS in a hepatocyte-selective manner against an iNOS-knockout background. The function of heme oxygenase 1 (HO-1) in iNOS-stimulated hepatoprotection and the molecular mechanisms were explored in vitro and in vivo, respectively. Hepatocyte-derived iNOS conferred protection from I/R injury and anoxia/reoxygenation stimulation. Mechanistically, iNOS activated nuclear factor erythroid 2-related factor 2 (Nrf-2) and subsequently, stimulated the transcription of HO-1. Results from our study led to the conclusion that HO-1 is another potent mediator of iNOS-mediated protection after liver I/R.

LECT2, a Ligand for Tie1, Plays a Crucial Role in Liver Fibrogenesis.

Liver fibrosis is a very common condition seen in millions of patients with various liver diseases, and yet no effective treatments are available owing to poorly characterized molecular pathogenesis. Here, we show that leukocyte cell-derived chemotaxin 2 (LECT2) is a functional ligand of Tie1, a poorly characterized endothelial cell (EC)-specific orphan receptor. Upon binding to Tie1, LECT2 interrupts Tie1/Tie2 heterodimerization, facilitates Tie2/Tie2 homodimerization, activates PPAR signaling, and inhibits the migration and tube formations of EC. In vivo studies showed that LECT2 overexpression inhibits portal angiogenesis, promotes sinusoid capillarization, and worsens fibrosis, whereas these changes were reversed in Lect2-KO mice. Adeno-associated viral vector serotype 9 (AAV9)-LECT2 small hairpin RNA (shRNA) treatment significantly attenuates fibrosis. Upregulation of LECT2 is associated with advanced human liver fibrosis staging. We concluded that targeting LECT2/Tie1 signaling may represent a potential therapeutic target for liver fibrosis, and serum LECT2 level may be a potential biomarker for the screening and diagnosis of liver fibrosis.

Hepatocellular iNOS protects liver from NASH through Nrf2-dependent activation of HO-1.

Multiple molecular events are involved in non-alcoholic steatohepatitis (NASH). There is no consensus on the role of inducible nitric oxide synthase (iNOS) in the progression of NASH. The present study therefore investigated the role of iNOS in NASH pathogenesis using bone marrow-transplanted iNOS chimeric mice under high-fat diet (HFD) conditions. The chimeric mice were fed a HFD for 16 wk, and primary hepatocytes were stimulated with oleic acid (OA). The molecular mechanisms underlying the role of iNOS in NASH were investigated. Marked hepatic steatosis and injury observed in the HFD mice and OA-stimulated hepatocytes were reduced by hepatocyte-derived iNOS. Mechanistically, iNOS upregulated heme oxygenase 1 (HO-1) by augmenting nuclear factor erythroid 2-related factor 2 (Nrf-2) binding to the HO-1 gene promoter. In conclusion, hepatocyte-derived iNOS may play a protective role against the progression of NASH by upregulating HO-1 through Nrf-2. Upregulation of hepatocellular iNOS may represent a potentially new therapeutic paradigm to combat NASH.

Hepatocellular iNOS protects liver from ischemia/reperfusion injury through HSF1-dependent activation of HSP70.

Although the role of inducible nitric oxide synthase (iNOS) in hepatic ischemia/reperfusion (I/R) injury remains controversial and confusing, with both harmful and beneficial effects in animal studies, the mechanism of these incongruous actions remains unclear. In the current study, we generated bone marrow chimeric mice with hepatocyte-restricted expression of iNOS. Chimeric mice and primary hepatocytes were subjected to I/R or anoxia/reoxygenation stimulation, respectively. The role of iNOS in liver I/R injury and the underlying molecular mechanisms were investigated. Hepatocyte-derived iNOS resulted in hepatoprotection from I/R injury, as well as in vitro experiments. Mechanistically, iNOS upregulates Heat shock protein (HSP) 70 by augmenting heat shock factor 1 (HSF1) binding to the HSP70 gene promoter. Importantly, inhibition of HSP70 partly reversed the iNOS overexpression-mediated hepatoprotection. The present findings demonstrate that hepatocellular iNOS protects from hepatic I/R injury through the HSF1-dependent activation of the HSP70. The upregulation of hepatocellular iNOS may offer a promising strategy for protecting against I/R injury.

A clinical study of choledochoscopic holmium laser lithotripsy for multiple intrahepatic calculi within ERAS programs.

OBJECTIVES: Multiple intrahepatic calculi, especially calculi in both sides of the liver, cannot be completely resolved by traditional surgery. In addition, morbidity after liver resection remains high. ERAS programs have been suggested that could relieve surgical stress and accelerate postoperative recovery. This study aimed to evaluate the safety and efficacy of choledocholithotomy combined with holmium laser lithotripsy in the treatment of multiple intrahepatic calculi within ERAS programs. METHODS: In all, 109 patients with multiple intrahepatic calculi were enrolled between January 2012 and September 2016, 42 of whom received choledocholithotomy combined with holmium laser lithotripsy. The remaining 67 patients underwent choledocholithotomy combined with choledochoscopic mechanical lithotripsy. Perioperative outcomes were compared and analyzed. RESULTS: Patient characteristics and preoperative details were similar between the groups (P > 0.05). The implementation of holmium laser lithotripsy could reduce the calculi residual rate (7.1% vs. 22.4%, P = 0.037), and even the liver resection rate (16.7% vs. 35.8%, P = 0.031). Additionally, holmium laser lithotripsy did not result in a higher morbidity (11.9% vs. 16.4%, P = 0.517), readmission rate (0% vs. 6%, P = 0.158), hospital stay (P = 0.189), hospital cost (P = 0.998), transfusion rate (P = 0.576), or operative time (P = 0.638). CONCLUSIONS: Holmium laser lithotripsy is feasible and efficient for treating multiple intrahepatic calculi within ERAS programs, which could reduce the liver resection rate and render refractory hepatic calculi easy to eliminate. In addition, holmium laser lithotripsy could be well coupled to the ERAS program to relieve surgical stress and accelerate postoperative recovery. Lasers Surg. Med. 51:161-166, 2019. © 2018 Wiley Periodicals, Inc.

Laparoscopic hepatectomy combined with endoscopic papillary balloon dilation for complex bile duct stones.

BACKGROUND: We evaluated the feasibility and efficacy of laparoscopic hepatectomy (LH) combined with endoscopic papillary balloon dilation (EPBD) for removing intra- and extrahepatic bile duct stones. METHODS: A total of 26 patients with intra- and extrahepatic bile duct stones underwent LH and EPBD. Selective hemi-hepatic vascular occlusion was used to complete the LH. EPBD was performed under the guidance of a guidewire, and stones were removed with a stone basket or balloon. RESULTS: A one-stage LH with EPBD lithotomy was successfully performed in 26 cases. No residual bile duct stones, intestinal or bile duct perforations were found. In addition, no severe post-operative bleeding, severe pancreatitis or mortality occurred. Post-operative hyperamylasemia was observed in five cases and bile leakage in one case. Post-operative hospital stays lasted 7-11 days. CONCLUSION: LH combined with EPBD applied to intra- and extrahepatic bile duct stones was feasible, effective and safe, resulting in rapid recovery and few post-operative complications.

HGF/R-spondin1 rescues liver dysfunction through the induction of Lgr5+ liver stem cells.

Induction of endogenous adult stem cells by administering soluble molecules provides an advantageous approach for tissue damage repair, which could be a clinically applicable and cost-effective alternative to transplantation of embryonic or pluripotent stem cell-derived tissues for the treatment of acute organ failures. Here, we show that HGF/Rspo1 induce liver stem cells and rescue liver dysfunction. Carbon tetrachloride treatment promotes both fibrosis and Lgr5+ liver stem cell proliferation, whereas Lgr5 knockdown worsens fibrosis. Injection of HGF in combination with Rspo1 increases the number of Lgr5+ liver stem cells and improves liver function by attenuating fibrosis. We observe Lgr5+ liver stem cells in human liver fibrosis tissues, and once they are isolated, these cells are able to form organoids, and treatment with HGF/Rspo1 promotes their expansion. We suggest that Lgr5+ liver stem cells represent a valuable target for liver damage treatment, and that HGF/Rspo1 can be used to promote liver stem cell expansion.

Long non-coding RNA CCAT2 is associated with poor prognosis in hepatocellular carcinoma and promotes tumor metastasis by regulating Snail2-mediated epithelial-mesenchymal transition.

Increasing evidence has demonstrated that aberrant expressions of long non-coding RNAs (lncRNAs) are involved in various malignancies, including hepatocellular carcinoma (HCC). This study aimed to investigate the role of lncRNA colon cancer-associated transcript 2 (CCAT2) in the progression of HCC. Quantitative real-time polymerase chain reaction analysis confirmed that CCAT2 was upregulated in HCC cell lines and cancerous tissues compared with normal liver cell line and adjacent normal tissue samples. The level of CCAT2 was positively associated with tumor-node-metastasis stages and vessel invasion. Survival analyses revealed that high CCAT2 expression predicted poor prognostic outcomes, serving as an independent prognostic factor for HCC patients. Patients with high CCAT2 expression had a 1.849-fold increased risk of death compared with those with low CCAT2 expression. Moreover, we also found that knockdown of CCAT2 expression reduced cell migration and invasion in vitro. We further demonstrated that CCAT2 played a key role in enhancing the epithelial-mesenchymal transition (EMT) through the regulation of vimentin, E-cadherin and transcription factor snail2 expression. Taken together, our findings showed that high CCAT2 expression is associated with poor survival in HCC patients. CCAT2 promotes HCC progression by regulating Snail2-induced EMT. CCAT2 may be a prognostic biomarker and therapeutic target for HCC.

Enhanced Recovery after Surgery Programs for Liver Resection: a Meta-analysis.

BACKGROUND AND AIMS: Due to the limited number of high-quality randomized controlled trials on enhanced recovery after surgery for hepatectomy, previous reviews have not been sufficiently comprehensive. Our objectives were to evaluate and compare the safety and efficacy of enhanced recovery after surgery programs and traditional care in patients undergoing open or laparoscopic surgery and to assess the optimized items for hepatectomy. METHODS: We searched the PubMed, Embase, and the Cochrane Library databases for all the relevant studies regardless of study design. We assessed the methodological quality of the included studies and excluded studies of poor quality. We performed a meta-analysis using RevMan 5.3 software. RESULTS: In total, 19 original studies with 2575 patients, including four randomized controlled trials and 15 non-randomized controlled trials, were analyzed. The meta-analysis demonstrated that enhanced recovery after surgery programs could reduce morbidity, hospital stays and cost, blood loss, and time to bowel function recovery for both open and laparoscopic surgery without increasing mortality, readmission rate, or transfusion rate. Twelve items were essential for liver surgery. CONCLUSIONS: Enhanced recovery after surgery programs for hepatectomy are feasible and efficient. Further studies should optimize perioperative outcomes for liver surgery.

Choledochoscopic Holmium Laser Lithotripsy for Difficult Bile Duct Stones.

OBJECTIVE: The aim of this study was to evaluate the feasibility and efficacy of choledochoscopic holmium laser lithotripsy as a means of removing resistant extrahepatic and intrahepatic bile duct stones. METHODS: Clinical data on 28 patients who had undergone choledochoscopic holmium laser lithotripsy were analyzed. RESULTS: Complete stone clearance was obtained in 24 patients; small numbers of residual stones in the left or right hepatic duct were found in 4 patients. No severe complications such as hemobilia and bile duct injuries occurred. CONCLUSION: Choledochoscopic holmium laser lithotripsy is a simple, safe, and effective treatment method for patients with resistant bile duct stones.