Clinical features and prognosis of patients with extrahepatic metastases from hepatocellular carcinoma

AIM: To assess the clinical features and prognosis of 151 patients with extrahepatic metastases from primary hepatocellular carcinoma (HCC), and describe the treatment strategy for such patients. METHODS: After the diagnosis of HCC, all 995 consecutive HCC patients were followed up at regular intervals and 151 (15.2%) patients were found to have extrahepatic metastases at the initial diagnosis of primary HCC or developed such tumors during the follow-up period. We assessed their clinical features, prognosis, and treatment strategies. RESULTS: The most frequent site of extrahepatic metastases was the lungs (47%), followed by lymph nodes (45%), bones (37%), and adrenal glands (12%). The cumulative survival rates after the initial diagnosis of extrahepatic metastases at 6, 12, 24, and 36 mo were 44.1%, 21.7%, 14.2%, 7.1%, respectively. The median survival time was 4.9 mo (range, 0-37 mo). Fourteen patients (11%) died of extrahepatic HCC, others died of primary HCC or liver failure. CONCLUSION: The prognosis of HCC patients with extrahepatic metastases is poor. With regard to the cause of death, many patients would die of intrahepatic HCC and few of extrahepatic metastases. Although most of HCC patients with extrahepatic metastases should undergo treatment for the primary HCC mainly, treatment of extrahepatic metastases in selected HCC patients who have good hepatic reserve, intrahepatictumor stage (T0-T2), and are free of portal venous invasion may improve survival.

Immunohistochemical molecular markers as predictors of curability of endoscopically resected submucosal colorectal cancer

AIM: To clarify the usefulness of immunohistochemical molecular markers in predicting lymph node metastasis of submucosal colorectal cancer. METHODS: We examined microvessel density, lymphatic vessel density, the Ki-67 labeling index, expression of MUC1 and Matrix metalloproteinase-7 (MMP-7) in tumor cells, and expression of cathepsin D in stromal cells at the invasive front by immunostaining of samples resected from 214 patients with submucosal colorectal cancer. Pathologic features were assessed on hematoxylin-eosin- stained samples. We evaluated the relations between clinicopathologic/immunohistochemical features and lymph node metastasis. RESULTS: Lesions of the superficial type, with an unfavorable histologic grade, budding, lymphatic involvement, high microvessel density (≥ 40), high lymphatic vessel density (≥ 9), high Ki-67 labeling index (≥ 42), and positivity of MUC1, cathepsin D, and MMP-7 showed a significantly high incidence of lymph node metastasis. Multivariate analysis revealed that high microvessel density, unfavorable histologic grade, cathepsin D positivity, high lymphatic vessel density, superficial type, budding, and MUC1 positivity were independent risk factors for lymph node metastasis.A combined examination with four independent immunohistochemical markers (microvessel density, cathepsin D, lymphatic vessel density, and MUC1) revealed that all lesions that were negative for all markers or positive for only one marker were negative for lymph node metastasis. CONCLUSION: Analysis of a combination of immuno- histochemical molecular markers in endoscopically resected specimens of submucosal colorectal cancer allows prediction of curability regardless of the pathologic features visible of hematoxylin-eosin-stained sections.

Clinical significance of type V_I pit pattern subclassification in determining the depth of invasion of colorectal neoplasms

AIM: To clarify whether subclassification of the type VI pit pattern on the basis of magnifying colonoscopy findings is useful in determining the type and depth of invasion of colorectal neoplasms.METHODS: We retrospectively analyzed 272 colorectal neoplasms (117 dysplasias and 155 submucosal invasive carcinomas; 228 patients) with a type V pit pattern [type VI, n = 202; type VN, n = 70 (Kudo and Tsuruta classification system)]. We divided lesions with a type VI pit pattern into two subclasses, mildly irregular lesions and severely irregular lesions, according to the prominent and detailed magnifying colonoscopy findings. We examined the relation between these two subclasses and histology/invasion depth.RESULTS: One hundred and four lesions (51.5%) were judged to be mildly irregular, and 98 lesions (48.5%) were judged to be severely irregular. Ninety-seven (93.3%) mildly irregular lesions showed dysplasias or submucosal invasion of less than 1000 μm (SM < 1000 μm). Fifty-five (56.1%) severely irregular lesions showed submucosal invasion equal to or deeper than 1000 μm (SM ≥ 1000 μm). Mild irregularity was found significantly more often in dysplasias or lesions with SM < 1000 μm than in lesions with SM ≥ 1000 μm (P < 0.01).CONCLUSION: Subclassification of the type VI pit pattern is useful for identifying dysplasias or lesions with SM < 1000 μm.

Patients with early recurrence of hepatocellular carcinoma have poor prognosis

BACKGROUND： Early recurrence （ER） after hepatic resection （HR） is a poor prognostic factor for patients with hepatocellular carcinoma （HCC）. This study aimed to identify the clinico- pathological features, outcomes, and risk factors for ER after HR for small HCC in order to clarify the reasons why ER is a worse recurrence pattern. METHODS： We retrospectively examined 130 patients who underwent HR for small HCC （___30 mm）. Recurrence was clas- sifted into ER （〈2 years） and late recurrence （LR） （_〉2 years）. The clinicopathological features, outcomes, and risk factors for ER were analyzed by multivariate analysis. RESULTS： ER was observed in 39 patients （30.0%）. The sur- vival rate of the ER group was significantly lower than that of the LR group （P〈0.005）, and ER was an independent prognos- tic factor for poor survival （P=0.0001）. The ER group had a significantly higher frequency （P=0.0039） and shorter interval （P=0.027） of development to carcinoma beyond the Milan criteria （DBMC） compared with the LR group, and ER was an independent risk factor for DBMC （P〈0.0001）. Multi-nodularity, non-simple nodular type, and microvascular invasion were independent predictors for ER （P=0.012, 0.010, and 0.019, respectively）.CONCLUSIONS： ER was a highly malignant recurrence pattern associated with DBMC and subsequent poor survival after HR for small HCC. Multi-nodularity, non-simple nodular type, and microvascular invasion predict ER, and taking these factors into consideration may be useful for the decision of the treatment strategy for small HCC after HR.

Defensive medicine practices among gastroenterologists in Japan

AIM： To clarify the prevalence of defensive medicine and the specific defensive medicine practices among gastroenterologists in Japan. METHODS： A survey of gastroenterologists in Hiroshima, Japan, was conducted by mail in March 2006. The number of gastroenterologists reporting defensive medicine behaviors or changes in their scope of practice and the reported defensive medicine practices, i.e., assurance and avoidance behaviors, were examined. RESULTS： A total of 131 （77%） out of 171 gastroenterologists completed the survey. Three （2%） respondents were sued, and most respondents （96%） had liability insurance. Nearly all respondents （98%） reported practicing defensive medicine. Avoidance behaviors, such as avoiding certain procedures or interventions and avoiding caring for high-risk patients, were very common （96%）. Seventy-five percent of respondents reported often avoiding certain procedures or interventions. However, seasoned gastroenterologists （those in practice for more than 20 years） adopted avoidance behaviors significantly less often than those in practice for less than 10 years. Assurance behaviors, i.e., supplying additional services of marginal or no medical value, were also widespread （91%）. Sixty-eight percent of respondents reported that they sometimes or often referred patients to other specialists unnecessarily. CONCLUSION： Defensive medicine may be highly prevalent among gastroenterologists throughout 3apan, with potentially serious implications regarding costs, access, and both technical and interpersonal quality of care.

Strategy for treatment of nonerosive reflux disease in Asia

The paper is to review the clinical and pathophysiologic differences between of nonerosive reflux disease (NERD) and reflux esophagitis (RE), and to propose a treatment strategy for NERD, especially for patients in Asia. A Medline search was performed regarding the clinical and pathophysiologic differences between NERD and RE, and treatment of NERD and RE. The characteristics of NERD patients in Asia are as follows: (1) high proportion of female patients, (2) low frequency of hiatal hernia, (3) high frequency of H pylori infection, (4) severe glandular atrophy of the gastric mucosa, and (5) frequent resistance to proton pump inhibitor (PPI) therapy. In Asian NERD patients, exposure of the esophagus to acid is not increased, and esophageal motility is normal. These characteristics are similar to those of patients in Western countries. Our recommended first-choice treatment is administration of PPI in combination with a prokinetic agent. However, at present, because there is limited evidence regarding effective treatments for NERD, it is best to try several different treatment strategies to find the best treatment for each patient.

Causal role of Helicobacter pylori infection and eradication therapy in gastric carcinogenesis

Many epidemiological reports indicate that Helicobacter pylori （H pylori） infection plays an important role in gastric carcinogenesis. Several genetic and epigenetic alterations contribute to the initiation, promotion, and progression of the cancer cells in a multi-step manner. H pylori is known to induce chronic inflammation in the gastric mucosa. Its products, including superoxides, participate in the DNA damage followed by initiation, and the inflammation-derived cytokines and growth factors contribute to the promotion of gastric carcinogenesis. By eradicating H pylori, gastric inflammation can be cured; the therapy diminishes the levels not only of inflammatory cell infiltration, but also atrophy/ intestinal metaplasia in part. A randomized controlled trial revealed that the eradication therapy diminished the gastric cancer prevalence in cases without precancerous conditions. In addition, recent epidemiological studies from Japanese groups demonstrated that the development of gastric cancer, especially of the intestinal type, was decreased by successful eradication therapy, although these were designed in a non- randomized manner. However, it should be mentioned that endoscopic detection is the only way to evaluate the degree of gastric carcinogenesis. We have reported that the endoscopic and histological morphologies could be modified by eradication therapy and it might contribute to the prevalence of gastric cancer development. Considering the biological nature of cancer cell proliferation, it is considered that a sufficiently long-term follow-up would be essential to discuss the anticancer effect of eradication therapy.

Selective decrease in colonic CD56^+ T and CD161^+ T cells in the inflamed mucosa of patients with ulcerative colitis

AIM： To investigate the role of local colonic mucosal NK receptor-positive T （NKR＋ T） cells in the regulation of intestinal inflammation, we analyzed the population and function of these cells in ulcerative colitis （UC）. METHODS： Colonic mucosal tissues were obtained from colonoscopic biopsies of the descending colon from 96 patients with UC （51 endoscopically uninflamed, 45 inflamed） and 18 normal controls. Endoscopic appearance and histologic score at the biopsied site were determined by MaLts＇ classification. A single cell suspension was prepared from each biopsy by collagenase digestion. Two NKR^＋ T cell subsets, CD56^＋ （CD56^＋CD3^＋） T cells and CD161＋ （CD161^＋CD3^＋） T cells, were detected by flow cytometric analysis. Intracellular cytokine analysis for anti-inflammatory cytokine interleukin-10 （IL-10） was performed by in vitro stimulation with phorbol-myristateacetate （PMA） and ionomycin. RESULTS： CD56^＋ T cells and CD161^＋ T cells are present in the normal human colon and account for 6.7% and 21.3% of all mononuclear cells, respectively. The populations of both CD56＋ T cells and CD161^＋ T cells were decreased significantly in the inflamed mucosa of UC. In contrast, the frequency of conventional T cells （CD56 CD3^＋ cells and CD161CD3^＋ cells） was similar among the patient and control groups. The populations of NKR^＋ T cells were correlated inversely with the severity of inflammation, which was classified according to the endoscopic and histologic Marts＇ criteria. Interestingly, approximately 4% of mucosal NKR＋ T cells expressing IL-10 were detected by in vitro stimulation with PMA and ionomycin.CONCLUSION： Selective reduction in the population of colonic mucosal NKR＋T cells may contribute to the development of intestinal inflammation in UC.

Outcome of patients who have undergone total enteroscopy for obscure gastrointestinal bleeding

AIM： To assess the diagnostic success and outcome among patients with obscure gastrointestinal bleeding who underwent total enteroscopy with double-balloon endoscopy.

Systemic gemcitabine combined with intra-arterial low-dose cisplatin and 5-fluorouracil for advanced hepatocellular carcinoma: Seven cases

The combination of intra-arterial low-dose cisplatin and 5-fluorouracil (5-FU) is effective against advanced hepatocellular carcinoma (HCC). Systemic gemcitabine chemotherapy seems effective in many cancers. We report the results of combination therapy with systemic gemcitabine, intra-arterial low-dose cisplatin and 5-FU (GEMFP). Seven patients with non-resectable advanced HCC were treated with GEMFP. One course of chemotherapy consisted of daily intra-arterial cisplatin (20 mg/body weight/hour on d 1, 10 mg/body weight per 0.5 h on d 2-5 and 8-12), followed by 5-FU (250 mg/body weight per 5 h on d 1-5 and 8-12) via an injection port. Gemcitabine at 1000 mg/m2 was administered intravenously at 0.5 h on d 1 and 8. The objective response was 57%. The response to GEMFP was as follows: complete response (no patients), partial response (four patients), stable disease (three patients), and progressive disease (no patients). The median survival period was 8 mo (range, 5-55). With regard to the National Cancer Institute Common Toxicity Criteria (NCI-CTC) grade 3 or 4 adverse reactions, seven (100%), seven, six (86%) and one (14%) patients developed leukopenia, neutropenia, thrombocytopenia and anemia, respectively. GEMFP may potentially be effective for non- resectable advanced HCC, but it has severe hematologic toxicity.

Effects of a 24-week course of interferon-αtherapy after curative treatment of hepatitis C virus-associated hepatocellular carcinoma

AIM： To assess whether a 24-wk course of interferon （IFN） could prevent hepatocellular carcinoma （HCC） recurrence and worsening of liver function in patients with hepatitis C virus （HCV）-infected patients after receiving curative treatment for primary HCC.METHODS： Outcomes in 42 patients with HCV infection treated with IFN-α, after curative treatment for primary HCC （IFN group）, were compared with 42 matched curatively treated historical controls not given IFN （non- IFN group）.RESULTS： Although the rate of initial recurrence did not differ significantly between ]FN group and non-IFN group （0%, 44%, 61~, and 67% ys 4.8%, 53~, 81%, and 87% at 1, 3, 5, and 7 years, P = 0.153, respectively）, ]FN group showed a lower rate than the non-IFN group for second recurrence （0%, 10.4%, 28%, and 350/0 ys 0%, 30~ , 59%, and 66% at 1, 3, 5 and 7 years, P = 0.022, respectively）. Among the ]FN group, patients with sustained virologic response （SVR） were less likely to have a second HCC recurrence than ]FN patients without an SVR, or non-IFN patients. Multivariate analysis identified the lack of SVR as the only independent risk factor for a second recurrence, while SVR and Child-Pugh class A independently favored overall survival.CONCLUSION： Most intrahepatic recurrences of HCV-related HCC occurred during persistent viral infection. Eradication of HCV is essential for the prevention of HCC recurrence and improvement of survival.

Low-dose intermittent interferon-alpha therapy for HCV-related liver cirrhosis after curative treatment of hepatocellular carcinoma

AIM: To assess the efficacy of low-dose intermittent interferon (IFN) therapy in patients with hepatitis C virus (HCV)-related compensated cirrhosis who had received curative treatment for primary hepatocellular carcinoma (HCC). METHODS: We performed a prospective case controlled study. Sixteen patients received 3 MIU of natural IFN- alpha intramuscularly 3 times weekly for at least 48 wk (IFN group). They were compared with 16 matched historical controls (non-IFN group). RESULTS: The cumulative rate of first recurrence of HCC was not significantly different between the IFN group and the non-IFN group (0% vs 6.7% and 68.6% vs 80% at 1- and 3-year, P = 0.157, respectively). The cumulative rate of second recurrence was not also significantly different between the IFN group and the non-IFN group (0% vs 6.7% and 35.9% vs 67% at 1- and 3-year, P = 0.056, respectively). Although the difference in the Child-Pugh classification score between the groups at initial treatment of HCC was not signifi cant, the score was signifi cantly worse at the time of data analysis in the non-IFN group than IFN group (7.19 ± 1.42 vs 5.81 ± 0.75, P = 0.0008). The cumulative rate of deviation from objects of any treatment for recurrentHCC was also higher in the non-IFN group than IFN group (6.7% and 27% vs 0 and 0% at 1- and 3-year, P = 0.048, respectively). CONCLUSION: Low-dose intermittent IFN-alpha therapy for patients with HCV-related compensated cirrhosis after curative HCC treatment was effective by making patients tolerant to medical or surgical treatment for recurrent HCC in the later period of observation.

Simple scoring system for predicting cirrhosis in nonalcoholic fatty liver disease

AIM: To investigate a simple noninvasive scoring system for predicting liver cirrhosis in nonalcoholic fatty liver disease (NAFLD) patients.

Alpha-fetoprotein and des-gamma-carboxy-prothrombin at twenty-four weeks after interferon-based therapy predict hepatocellular carcinoma development

AIM: To investigate risk factors for development of hepatocellular carcinoma(HCC) in patients with hepatitis C virus-related liver cirrhosis(LC-C).METHODS: To evaluate the relationship between clinical factors including virological response and the development of HCC in patients with LC-C treated with interferon(IFN) and ribavirin, we conducted a multicenter, retrospective study in 14 hospitals in Japan. All patients had compensated LC-C with clinical or histological data available. HCC was diagnosed by the presence of typical hypervascular characteristics on computed tomography and/or magnetic resonance imaging.RESULTS: HCC was diagnosis in 50(21.6%) of 231 LC-C patients during a median observation period of 3.8 years after IFN and ribavirin therapy. Patients who developed HCC were older(P = 0.018) and had higher serum levels of pretreatment alpha-fetoprotein(AFP)(P = 0.038). Multivariate analysis revealed the following independent risk factors for HCC development: history of treatment for HCC [P < 0.001, odds ratio(OR) = 15.27, 95%CI: 4.98-59.51], AFP levels of ≥ 10 ng/m L(P = 0.009, OR = 3.89, 95%CI: 1.38-11.94), and des-γ-carboxy prothrombin(DCP) levels of ≥ 40 m AU/mL at 24 wk after the completion of IFN and ribavirin therapy(P < 0.001, OR = 24.43, 95%CI: 4.11-238.67).CONCLUSION: We suggested that the elevation of AFP and DCP levels at 24 wk after the completion of IFN and ribavirin therapy were strongly associated with the incidence of HCC irrespective of virological response among Japanese LC-C patients.

Impaired coronary microvascular endothelial function in men with metabolic syndrome

AIM: To assess coronary endothelial function of conduit and resistance vessels in patients with metabolic syndrome (MS). METHODS: Seventy-eight men (mean age, 57 years) with chest pain and angiographically normal coronary arteries were included in the study. Patients with coronary spastic angina were excluded. Changes in coronary artery diameter and coronary blood flow (CBF) in response to acetylcholine (ACh) were determined using quantitative coronary angiography and Doppler velocity measurements. Coronary flow reserve was calculated as the ratio of coronary blood velocity after adenosine triphosphate infusion relative to baseline values. Patients were divided into two groups based on the presence or absence of MS. RESULTS: There were 24 patients in the MS group (31%). The increase in CBF in response to ACh infusion was impaired in the MS group (P < 0.0001) compared to the non-MS group, whereas changes in coronary artery diameter in response to ACh infusion did not differ between the two groups. Multivariate regression analysis revealed that MS was a significant factor associated with the lesser change in CBF induced by ACh infusion at 30 μg/min (P < 0.0001, r 2 = 0.46). CONCLUSION: Coronary endothelial dysfunction was present at the level of resistance vessels but not conduit vessels in the MS patients included in our study.

Clinical significance of mucosal suppressors of cytokine signaling 3 expression in ulcerative colitis

AIM:To investigate the clinical significance of mucosal expression of suppressors of cytokine signaling 1 (SOCS1) and SOCS3 in human ulcerative colitis (UC). METHODS:Biopsy specimens for histological analysis and mRNA detection were obtained endoscopically from the rectum of 62 patients with UC (36 men; age 13-76 years). The patients were classified endoscopically according to Matts' grade (grade 1 to 4). Expression of SOCS1 and SOCS3 mRNAs was quantified in samples by competitive reverse transcription-polymerase chain reaction (RT-PCR). GAPDH was used as an internal control for efficiency of RT-PCR and amount of RNA. RESULTS:SOCS3 mRNA expression was significantly higher in inflamed mucosa of UC than in inactive mucosa. The level of expression was well correlated with the degree of both endoscopic and histologic inflammation. Interestingly,among the patients in remission,the group with relatively low expression of SOCS3 showed a higher rate of remission maintenance over a 12-mo period. In contrast,SOCS1 mRNA was expressed in both inflamed and non-inflamed colonic mucosa and was not correlated with the activity of colonic mucosa or prognosis. CONCLUSION:These observations suggest that increased expression of mucosal SOCS3,but not of SOCS1,may play a critical role in the development of the colonic inflammation of UC.

Medical malpractice litigation related to gastrointestinal endoscopy in Japan:A two-decade review of civil court cases

AIM: To examine the allegations in malpractice litigations related to gastrointestinal endoscopy in Japan. METHODS: A retrospective review of cases tried in the civil court system during the 21-year period from 1985 to 2005, identified in a computerized legal database, was undertaken. RESULTS: Eighteen malpractice litigations and a total of 30 allegations were identified. Of the 18 (44%) malpractice litigations, 8 (44%) were related to eso- phagogastroduodenoscopy, 4 (22%) to colonoscopy, 4 (22%) to endoscopic sphincterotomy, and 2 (11%) to endoscopic retrograde cholangiopancreatography. Seventeen (94%) cases pertained to complications, and the remaining (6%) case pertained to misdiagnosis. In 10 cases, the patient died of the complications. Allegations were categorized as: (1) performance error during the endoscopic procedure (n = 12, 40%); (2) lack of informed consent (n = 9, 30%); (3) performance error during the treatment after the endoscopic procedure (n = 4, 13%); (4) premedication error (n = 3, 10%); (5) diagnostic error (n = 1, 3%); and (6) indication error for the endoscopic procedure (n = 1, 3%). CONCLUSION: These data may aid in the design of risk prevention strategies to be used by gastrointestinal endoscopists.

Prevalence and impact of musculoskeletal pain in Japanese gastrointestinal endoscopists:A controlled study

AIM:To examine the frequency and prevention of musculoskeletal pain in Japanese gastrointestinal endoscopists and non-endoscopist physicians.METHODS:Questionnaires were sent to 275 endoscopists and 173 non-endoscopists working in Hiroshima University Hospital and its affiliated hospitals.RESULTS:The completed questionnaires were returned by 190(69%)endoscopists and 120(69%)non-endoscopists.The frequency of pain in the hand and wrist,and especially the left thumb,was significantly higher in endoscopists than in non-endoscopists(17%vs 6%,P=0.004).Using multivariate analysis,the only significant factor associated with this pain was the age of the endoscopist(odds ratio 2.77,95%confidence interval,1.23-6.71,P=0.018).Interestingly,endoscopists had made significantly fewer modifications to their endoscopic practices than non-endoscopists(12%vs 33%,P <0.0001)to prevent pain.CONCLUSION:Pain in the hand and wrist may be endoscopy-related.However,endoscopists made little modifications in practice to prevent such pain.More attention to prevention appears necessary.

Involvement of Krüppel-like factor 6 (KLF6) mutation in the development of nonpolypoid colorectal carcinoma

AIM： To examine Kruppel-like factor 6 （KLF6） mutations in nonpolypoid-type tumors and alterations of K-ras, p53, and B-raf in relation between mutation and morphologic type, particularly nonpolypoid-type colorectal carcinomas. METHODS： Fifty-five early nonpolypoid colorectal carcinomas were analyzed. Loss of heterozygosity （LOH） of KLF6 and p53 was determined by microsatellite assay. Mutations of KLF6, K-ras, and B-raf were examined by polymerase chain reaction-single-strand conformation polymorphism followed by direct sequencing. In LOH- positive and/or mutation-positive tumors, multiple （4-7） samples in each tumor were microdissected and examined for genetic alterations, p53 expression was evaluated by immunohistochemistry. RESULTS： LOH of KLF6 and p53 was found in 14 of 29 （48.3%） and 14 of 31 （45.2%） tumors, respectively. In tO of the 14 （71.4%） KLF6 LOH-positive tumors and 9 of the 14 （64.3%） p53 LOH-positive tumors, LOH was found in all of the microdissected samples. In 1 of the tO （10.0%） KLF6 LOH-positive tumors, a single missense mutation was identified. K-ras and B-raf mutations were found in 5 of 55 （9.1%） and 6 of 55 （10.9%） tumors, respectively. However, these mutations were detected only in subsets of microdissected tumor samples. CONCLUSION： These data suggest that KLF6 and p53 mutations are involved in the development of nonpolypoid colorectal carcinoma, whereas K-ras and B-raf mutations are not.

Earthworm fibrinolytic enzyme:anti-tumor activity on human hepatoma cells in vitro and in vivo

Background The earthworm fibrinolytic enzyme （EFE） is a complex protein enzyme that is widely distributed in the earthworm＇s digestive cavity. Possessing strong protein hydrolysis activity, EFE not only has a direct effect on fibrin, but also can activate plasminogen. Its therapeutic and preventative effects on thrombosis-related disease have been confirmed clinically. Recently, there has been increased interest in the anti-tumor activity of EFE. In this study, the anti-tumor activity of EFE, isolated from Eisenia foetida, on human hepatoma cells was evaluated in vitro and in vivo. The potential mechanisms involved were also studied. Methods In vitro experiments were performed in four human hepatoma cell lines： HLE, Huh7, PLC/PRF/5 and HepG2. After treatment with EFE in various concentrations, the inhibition of the rate of cell proliferation was measured. For the in vivo studies, tumor-bearing models xenografted with Huh7 cells were developed in nude mice, and then the mice were fed with EFE once a day for 4 weeks, and the control group received only saline. An inhibitory effect on tumor growth was observed. Also, apoptosis was observed with flow cytometric assay and fluorescent dye staining with acridine orange and ethidium bromide （AO/EB）. The expression of matrix metalloproteinase 2 （MMP-2） were detected by Western blotting assay. Results After treatment with various concentrations of EFE, the proliferation of all hepatoma cell lines was suppressed to varying degrees in vitro. The IC50 for HLE, Huh7, PLC/PCF/5 and HepG2 were 2.11, 5.87, 25.29 and 17.30 uku/ml, respectively. After administration of EFE orally for 4 weeks, the growth of tumor xenograft of Huh7 cells in nude mice was significantly inhibited in vivo. The tumor inhibitory rates in the EFE 500 uku/（kg·d） and 1000 uku/（kg·d） groups were 46.08% （compared with control group, P=0.026） and 57.52% （compared with control group, P=0.002） respectively. Meanwhile, the average weight of body, spleen or thymus did not show any remarkable differences among the various groups. The population in sub-G1 stage was more in the EFE treated groups than in the control group according to flow cytometric assay. After treatment with EFE 0, 5, 10 uku/ml for 72 hours, the apoptotic rates were 3.5%, 10.9% and 12.3% in HLE cells, and 5.0%, 24.7% and 34.5% in Huh7 cells respectively. Under fluorescent staining with AO/EB, the apoptotic morphological changes could be detected more significantly in the EFE treated groups than in the untreated groups. After treatment with EFE in doses of 0, 5, 10 uku/ml for 72 hours, the apoptotic rates were 3.02%, 8.76%, 10.54% in HLE cells, and 3.95%, 18.27%, 30.89% in Huh7 cells respectively. The apoptosis-inducing effects of EFE occurred in a dose dependent manner. Western blotting assay showed that, after treatment with EFE, the secretions of MMP-2 were significantly inhibited in HLE and Huh7 cells. Conclusions EFE showed significant anti-tumor activity in hepatoma cells both in vitro and in vivo, which may be because EFE could induce apoptosis of hepatoma cells and inhibit the expression of MMP-2. This suggests that EFE has a potential role in the treatment of hepatoma.