AIM: To investigate the effects of agkis-trodon halys venom anti-tumor component (AHVAC-) on the biological behavior of gastric cancer MKN-28 cells. METHODS: Gastric cancer MKN-28 cells were treated with the experimental concentrations (5, 10, 15 μg/mL) of AHAVC- for 24 h. Cell proliferation and toxicity assay (cell counting kit-8, CCK-8) was used to detect the inhibition rates of the cells in different concentrations of AHVAC-. The migration ability of the cells was evaluated by wound-healing and Transwell assay. The apoptosis were observed by laser confocal microscopy with annexin V-mCherry/DAPI double staining, and the apoptosis rates were analyzed by flow cytometry with annexin V-FITC/PI double fluorescence staining. The protein level of Caspease-3 was determined by Western blot. RESULTS: Compared with normal control group, the results of AHVAC- concentration groups showed that with the increase of AHVAC- concentration, the proliferative activity of MN-28 cells decreased gradually (P<0.01), the cell migration ability decreased gradually (P<0.01), and the cell apoptosis rate increased (P<0.05). The expression of apoptosis-related protein Caspease-3 was up-regulated (P<0.01). CONCLUSION: AHVAC- inhibits proliferation and migration of gastric cancer MSN-28 cells and induces apoptosis.

Objective To investigate the expression of the ubiquitination enzyme UBE2S in different cell types in hepatocellular carcinoma(HCC)microenvironment and its impact on proliferation and stemness of HCC cells.Methods TCGA and CPTAC database were used to analyze the transcriptional and promoter methylation levels and protein expressions of UBE2S in HCC.Specific expression patterns of UBE2S,intercellular communication and key transcription factors in different cell types were analyzed based on single-cell sequencing data from TISCH website.We further examined UBE2S expressions in clinical samples of HCC tissues,HCC cells and T cells using immunohistochemistry and immunofluorescence staining.We also tested the effects of UBE2S knockdown on stemness of HCC-LM3 and HepG2 cells using clone formation experiments and sphere formation assay.Results Analysis based on TCGA database suggested significant overexpression of UBE2S in both paired and non-paired tumor tissues(P<0.001),and its transcriptional level increased with tumor grades.The methylation level of UBE2S promoter was significantly decreased in HCC(P<0.001),and its transcription level increased obviously in HCC with TP53 mutation(P<0.001).Analysis of CPTAC database also demonstrated overexpression of UBE2S protein in HCC tissues(P<0.001).Three prognostic models suggested that HCC patients with high UBE2S expression had poorer prognosis(P<0.001).Single-cell sequencing data analysis revealed high expressions of UBE2S in T cells and high intensities of interaction between endothelial cells,epithelial cells and fibroblasts in HCC microenvironment.Immunohistochemistry and immunofluorescence staining demonstrated high UBE2S expressions in clinical samples of HCC tissues,HCC cells and T cells.In HCC-LM3 and HepG2 cells,UBE2S knockdown significantly inhibited cell clone formation and tumor sphere formation(P<0.05).Conclusion UBE2S is highly expressed in T cells in HCC microenvironment in close correlation with a poor prognosis.High UBE2S expression promotes the stemness of HCC cells.

Objective:To investigate the effect of decarbromodiphenyl ether(BDE-209)exposure on the migration ability of triple-negative breast cancer(TNBC)cells and to explore the underlying mechanism.Methods:Human TNBC MDA-MB-231 cells were divided into blank control group and BDE-209 exposure groups(treated with 0.02,0.20,2.00,20.00 and 200.00 ng/mL BDE-209 in high glucose DMEM).Extracellular vehicles(EVs)secreted by MDA-MB-231 cells were isolated by differential ultracentrifugation.Transmission electron microscopy(SEM),nanoparticle tracking analysis(NTA)and Western blotting were performed to characterize the EVs.The effect of the EVs induced by BDE-209 exposure(EVs-BDE-209)on the migration and invasion of MDA-MB-231 cells was detected by wound-healing assay and Transwell test.qRT-PCR was used to measure the miR-221 level in EVs-BDE-209.The expression of MMP9 in MDA-MB-231 cells was determined by Western blotting.Results:Compared with the blank control,BDE-209 exposure increased the tumor cell-derived EVs in dose-dependent manner.The MDA-MB-231 cells co-cultured with EVs released by 200.00 ng/mL BDE-209 exposure showed an 86%increase in cell migration rate,a 1.32-fold higher number of membrane-penetrating cells,a 2.71-fold higher expression level of miR-221,and a 1.62-fold higher expression level of MMP9 compared with the blank control group(all P<0.05).While transfection with anti-miR-221 antibody to decrease miR-221 level in EVs significantly reversed the increased invasion ability of the MDA-MB-231 cells treated with EVs-BDE-209.Conclusion:BDE-209 exposure may promote metastasis potential of MDA-MB-231 cells via EVs-BDE-209 transmitted miR-221.

Objective To investigate the expression of the ubiquitination enzyme UBE2S in different cell types in hepatocellular carcinoma(HCC)microenvironment and its impact on proliferation and stemness of HCC cells.Methods TCGA and CPTAC database were used to analyze the transcriptional and promoter methylation levels and protein expressions of UBE2S in HCC.Specific expression patterns of UBE2S,intercellular communication and key transcription factors in different cell types were analyzed based on single-cell sequencing data from TISCH website.We further examined UBE2S expressions in clinical samples of HCC tissues,HCC cells and T cells using immunohistochemistry and immunofluorescence staining.We also tested the effects of UBE2S knockdown on stemness of HCC-LM3 and HepG2 cells using clone formation experiments and sphere formation assay.Results Analysis based on TCGA database suggested significant overexpression of UBE2S in both paired and non-paired tumor tissues(P<0.001),and its transcriptional level increased with tumor grades.The methylation level of UBE2S promoter was significantly decreased in HCC(P<0.001),and its transcription level increased obviously in HCC with TP53 mutation(P<0.001).Analysis of CPTAC database also demonstrated overexpression of UBE2S protein in HCC tissues(P<0.001).Three prognostic models suggested that HCC patients with high UBE2S expression had poorer prognosis(P<0.001).Single-cell sequencing data analysis revealed high expressions of UBE2S in T cells and high intensities of interaction between endothelial cells,epithelial cells and fibroblasts in HCC microenvironment.Immunohistochemistry and immunofluorescence staining demonstrated high UBE2S expressions in clinical samples of HCC tissues,HCC cells and T cells.In HCC-LM3 and HepG2 cells,UBE2S knockdown significantly inhibited cell clone formation and tumor sphere formation(P<0.05).Conclusion UBE2S is highly expressed in T cells in HCC microenvironment in close correlation with a poor prognosis.High UBE2S expression promotes the stemness of HCC cells.

Purpose To investigate the clinicopathological features of primary mucoepidermoid carcinoma of the liver(PMCL).Methods The history,clinical data,pathological diagnosis,differential diagnosis,treatment and prognosis of 4 patients with PMCL were retrospectively analyzed.Results All four tumors were located in the left half of the liver and were sol-itary.Microscopic examination showed that tumor cells were composed of three types of cells:epidermal like cells,interme-diate cells,and mucinous cells,with varying proportions.Im-munophenotype:tumor cells showed diffuse strong positivity for CA199 and CK19,while mucous cells showed focal positivity for CEA.CK7 mainly expressed in epidermal like cells and mucous cells,while CK5/6,p40,and p63 expressed in epidermal like cells and intermediate cells.Special staining with AB,PAS,and mucin carmine confirmed the presence of mucous cells,Ki67 proliferation index ranged from 30％to 40％.FISH testing:no MAML2 gene breakage or rearrangement was detected in 4 cases of PMCL,and MAML2 gene amplification occurred in 1 case of tumor.Conclusion The incidence of PMCL is low and present with non-specific clinical manifestations,which often leads to misdiagnosis.The diagnosis should be differentiated from bile duct adenocarcinoma and adenosquamous carcinoma.Moreover,there is currently no established standard treatment for PMCL and has a poor prognosis.

Purpose To investigate the clinicopathological features of primary mucoepidermoid carcinoma of the liver(PMCL).Methods The history,clinical data,pathological diagnosis,differential diagnosis,treatment and prognosis of 4 patients with PMCL were retrospectively analyzed.Results All four tumors were located in the left half of the liver and were sol-itary.Microscopic examination showed that tumor cells were composed of three types of cells:epidermal like cells,interme-diate cells,and mucinous cells,with varying proportions.Im-munophenotype:tumor cells showed diffuse strong positivity for CA199 and CK19,while mucous cells showed focal positivity for CEA.CK7 mainly expressed in epidermal like cells and mucous cells,while CK5/6,p40,and p63 expressed in epidermal like cells and intermediate cells.Special staining with AB,PAS,and mucin carmine confirmed the presence of mucous cells,Ki67 proliferation index ranged from 30％to 40％.FISH testing:no MAML2 gene breakage or rearrangement was detected in 4 cases of PMCL,and MAML2 gene amplification occurred in 1 case of tumor.Conclusion The incidence of PMCL is low and present with non-specific clinical manifestations,which often leads to misdiagnosis.The diagnosis should be differentiated from bile duct adenocarcinoma and adenosquamous carcinoma.Moreover,there is currently no established standard treatment for PMCL and has a poor prognosis.

AIM: To explore whether Agkistrodon Halys venom antitumor component-I (AHVAC-I) affects the migration of gastric cancer cells by human primary gastric cancer-associated fibroblast (GCAFs). METHODS: Tissue block culture and trypsin digestion were used to separate and culture human primary gastric cancer-associated fibroblasts (GCAFs); the GCAFs-CM

Purpose To discuss the clinical pathological features, diagnosis and differential diagnosis, treatment and prognosis of pri-mary breast diffuse large B-cell lymphoma ( DLBCL) . Methods 7 cases of primary breast DLBCL were collected, their clinical path-ological characteristics and immunophenotypes were also observed, and the treatment and prognostic factors were discussed. Results All of the 7 patients were women, aged 28~75 years, with the median 51 years. 5 cases involved left breast, 2 cases were located in the right breast. Painless mammary mass and ipsilateral axillary lymphadenopathy were the commonest clinical manifestations. On mi-croscopic observation, tumor cells were large to medium-sized which characterized as diffuse infiltration between the lobules of mamma-ry gland, around the duct, interstitial and fat tissue, some were single file cord pattern. 1 case was ALK-1 positive DLBCL, 6 cases were DLBCL of the non special type. The immunophenotype showed 5 cases were of non-GCB, 2 cases were of GCB type. Ki-67 index were from 60% to 95%. According to Ann Arbor staging, 2 cases were stage I EA, 5 cases were stage II EA. IPI score:4 cases with 0 score, 3 cases with 1 score. Of 7 patients, one case was modified radical mastectomy, 4 cases were lumpectomy, 2 cases were diag-nosed by core needle biopsy. 2 patients died without chemotherapy, 5 patients were chemotherapy or radiotherapy for 4 to 6 cycles ( CHOP/R-CHOP) , 4 patients received complete response, one died. Patients were followed up from 1 to 114 months, the median fol-low-up time was 18 months. The 1 year OS rate was 57. 1%, 5 years OS rate was 14. 3%. Conclusion The diagnosis of primary breast DLBCL is confirmed by pathological biopsy and immunohistochemical markers. The immunophenotype was mainly non-GCB type. Comprehensive treatment including chemotherapy and radiotherapy is appropriate. The prognosis should be comprehensively eval-uated by multiple factors.

Objective Previous study found TGF-βpathway might be the molecular pathway influencing the prognosis of colo-rectal cancer, while it was uncertain whether Chinese population is associated with the disease.The article was to evaluate the genetic factors associated with prognosis in colorectal cancer. Methods 52 cases patients with colorectal cancer were followed-up for 36 months in our hospitals from January 2013 to August 2014.Their DNAs were extracted and stored and gene typing were carried out in 5 candidate genes to detect the association between SNPs and the prognosis in colorectal cancer. Results The results showed that within the TGF-βsignaling pathway, after adjusting for Bonferroni multiple testing, allele A of SNP rs10749971 located in gene POU2AF1 was associated with the recurrence of patients with stage III disease under additive and recessive genetic models ( HR =1.968, P=0.004;HR=2.174, P=0.010).Allele C of SNP rs961253 in the gene BMP2 could increase the recurrence risk (HR=1.992, P=0.005) and the death risk (HR=3.161, P=0.007) of patients with stage III disease under recessive genetic models.Allele A of SNP rs4464148 in SMAD7 gene could significantly decrease the death risk of patients with stage II and III colorectal cancer under dominant genetic model (HR=0.382, P=0.017;HR=0.230, P=0.006).In addition, accumulated effects of several adverse genes showed gene high risk group could increase the risk of death for patients with stage III colorectal cancer significantly ( HR=15.512, P=0.036;95%CI:1.611-149.360). Conclusion In different genetic models, SNP locus mutation within gene POU2AF1, BMP2 and SMAD7 on TGF-βpathway was associated with the prognosis of patients with colorectal cancer.With the increase of the number of unfavorable genes, the death risk increases accordingly.

Purpose To investigate the clinicopathlogic characteristic, diagnosis and histogenesis of pancreatic solid-pseudopapillary tumors ( SPPT) . Methods Combined with relevant literature, the clinical history, histopathological features and immunohistochemi-cal characteristics were analyzed in 11 cases of SPPT. Results There were 10 female patients and only 1 male in total 11 cases, aged from1 7 to 60 years (mean 33). The sizes of tumors were from 3. 2 to 10. 0 cm. Histologically, they were composed of papillary and microcysticsolid structures. Pseudopapillary with a fibrovascular core was remarkable. Immunohistochemically, the tumors expressed EMA (1/11), vimentin (10/11), NSE (11/11), Syn (7/11), CgA (1/11), CD56 (11/11), CD10 (11/11), PR (9/11), CD99 (9/11),α-AT (11/11),β-catenin (11/11), E-cadherin (11/11), Cyclin D1 (11/11), c-Myc (11/11). 6 patients were followed up for a period of 20 to 112 months, and they were all alive and had no recurrence and metastasis. Conclusions SPPT is a tumor with low malignancy of the pancreas that most frequently affect young females. SPPT may be derived from multipotent stem cells and closely related withβ-catenin signaling pathway. Pathological morphology and immunohistochemistry are very important to the diag-nosis and differential diagnosis of SPPT.