CD64 expression is increased in patients with severe acute pancreatitis: clinical significance.

BACKGROUND/AIMS: Upregulated CD64 expression on neutrophils is the most useful marker for acute bacterial infections and systemic inflammation. However, it is unknown whether CD64 is involved in the pathogenesis of acute pancreatitis (AP). This study was designed to determine whether CD64 is implicated in severe acute pancreatitis (SAP), and thus, is a suitable marker for SAP. METHODS: SAP was induced in rats with an intraperitoneal injection of L-arginine. CD64 expression in the rat pancreas was determined by quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry. Additionally, the CD64 mRNA expression in peripheral blood leukocytes from 21 patients with mild acute pancreatitis (MAP) and 10 patients with SAP was investigated at the time of admission and during remission by qRT-PCR. RESULTS: CD64 mRNA and protein expression in the pancreas was significantly higher in rats with SAP, compared to the controls. The CD64 expression was higher in the patients with SAP than in the patients with MAP. During remission, CD64 mRNA decreased in both the MAP and SAP patients. The area under the curve of CD64 expression for the detection of SAP was superior to both the Ranson and the Acute Physiology and Chronic Health Evaluation II scores. CONCLUSIONS: The CD64 level was significantly increased in correlation with the disease severity in SAP and may act as a useful marker for predicting the development of SAP.

Estradiol regulates miR-135b and mismatch repair gene expressions via estrogen receptor-ß in colorectal cells.

Estrogen has anti-colorectal cancer effects which are thought to be mediated by mismatch repair gene (MMR) activity. Estrogen receptor (ER) expression is associated with microRNA (miRNA) expression in ER-positive tumors. However, studies of direct link between estrogen (especially estradiol E2), miRNA expression, and MMR in colorectal cancer (CRC) have not been done. In this study, we first evaluated the effects of estradiol (E2) and its antagonist ICI182,780 on the expression of miRNAs (miR-31, miR-155 and miR-135b) using COLO205, SW480 and MCF-7 cell lines, followed by examining the association of tissue miRNA expression and serum E2 levels using samples collected from 18 colorectal cancer patients. E2 inhibited the expressions of miRNAs in COLO205 cells, which could be reversed by E2 antagonist ICI 182.780. The expression of miR-135b was inversely correlated with serum E2 level and ER-ß mRNA expression in CRC patients' cancer tissues. There were significant correlations between serum E2 level and expression of ER-ß, miR-135b, and MMR in colon cancer tissue. This study suggests that the effects of estrogen on MMR function may be related to regulating miRNA expression via ER-ß, which may be the basis for the anti-cancer effect in colorectal cells.

Establishment of a human hepatoma multidrug resistant cell line in vitro.

AIM: To establish a multidrug-resistant hepatoma cell line (SK-Hep-1), and to investigate its biological characteristics. METHODS: A highly invasive SK-Hep-1 cell line of human hepatocellular carcinoma, also known as malignant hepatoma was incubated with a high concentration of cisplatin (CDDP) to establish a CDDP-resistant cell subline (SK-Hep-1/CDDP). The 50% inhibitory dose (IC(50)) values and the resistance indexes [(IC(50) SK-Hep-1/CDDP)/(IC(50) SK-Hep-1)] for other chemotherapeutic agents and the growth curve of cells were all evaluated using cell counting kit-8 assays. The distribution of the cell cycles were detected by flow cytometry. Expression of acquired multidrug resistance P-glycoprotein (MDR1, ABCB1) and multidrug resistance-associated protein 1 (MRP1, ABCC1) was compared with that in parent cells by Western blotting and immunofluorescence combined with laser scanning confocal microscopy. RESULTS: The SK-Hep-1/CDDP cells (IC(50) = 70.61 +/- 1.06 microg/mL) was 13.76 times more resistant to CDDP than the SK-Hep-1 cells (IC(50) = 5.13 +/- 0.09 microg/mL), and CDDP-resistant cells also demonstrated cross-resistance to many anti-tumor agents such as doxorubicin, 5-fluorouracil and vincristine. Similar morphologies were determined in both SK-Hep-1 and SK-Hep-1/CDDP groups. The cell cycle distribution of the SK-Hep-1/CDDP cell line exhibited a significantly increased percentage of cells in S (42.2% +/- 2.65% vs 27.91% +/- 2.16%, P < 0.01) and G2/M (20.67% +/- 5.69% vs 12.14% +/- 3.36%, P < 0.01) phases in comparison with SK-Hep-1 cells, while the percentage of cells in the G0/G1 phase decreased (37.5% +/- 5.05% vs 59.83% +/- 3.28%, P < 0.01). The levels of MDR1 and MRP1 were overexpressed in the SK-Hep-1/CDDP cells exhibiting the MDR phenotype. CONCLUSION: Multiple drug resistance of multiple drugs in the human hepatoma cell line SK-Hep-1/CDDP was closely related to the overexpression of MDR1 and MRP1.

Establishment of a cisplatin-induced multidrug resistance cell line SK-Hep1/DDP.

BACKGROUND AND OBJECTIVE: Multidrug resistance (MDR) is a major obstacle in the chemotherapy of cancer patients. The aim of this study was to establish a mutidrug-resistant cell line SK-Hep1/DDP and explore its molecular mechanism of the MDR. METHODS: SK-Hep1/DDP cell line was induced by pulse treatment using a high concentration of cisplatin (DDP) in vitro. The chemoresistance indexes of cells were evaluated by CCK-8 assays. The protein of MDR1 (ABCB1), MRP1 (ABCC1), MRP2 (ABCC2) and Bax were detected by Western blotting, and the effect of MDR1 inhibitor cyclosporine A (CsA) on expression of MDR1 proteins in SK-Hep1 and SK-Hep1/DDP cell lines. Flow cytometry was performed to determine the distribution of the cell cycle and cell apoptosis ratio. RESULTS: The SK-Hep1/DDP cells were 13.76 times more resistant to DDP in comparison with SK-Hep1 cells, and SK-Hep1/DDP cells also exhibited cross-resistance to many other chemotherapeutic agents (adramycin and 5-fuorouracil). MDR1, MRP1, and MRP2 protein expressions were significantly higher in the SK-Hep1/DDP than in the SK-Hep1 (P < 0. 01), but Bax was lower in the SK-Hep1/DDP than in the SK-Hep1(P < 0. 01). There was no obvious influence between SK-Hep1 and SK-Hep1/DDP cells in the expression of MDR1 by MDR1 inhibtor CsA (P > 0. 05). The percentages of cells in G(2)/M and S phase were significantly increased in SK-Hep1/DDP in comparison with those in SK-Hep1 [(20.67 +/- 5.69)% vs. (12.14 +/- 3.36)%; (42.20 +/- 2.65)% vs. (27.91 +/- 2.16)%; P < 0. 01]. After the cells were exposed to 10 µg/mL DDP for 24 h, the cell apoptosis rate of SK-Hep1/DDP was decreased in comparison with SK-Hep1, but it was increased in those with pretreatment of MDR1 inhibitor CsA as compared with those without pretreatment. CONCLUSIONS: A reliable multi-drug resistant human hepatoma cell line SK-Hep1/DDP is successfully established. The MDR mechanisms of this cell lines are closely related to the over-expression of MDR1 MRP1 and MRP2, lower expression of Bax and the attenuated cell apoptosis induced by chemotherapeutic agents.

Mitochondrial microsatellite instability in gastric cancer and its precancerous lesions.

AIM: To evaluate the role of mitochondrial microsatellite instability (mtMSI) in gastric carcinogenesis. METHODS: MtMSI was measured with PCR-single strand conformation polymorphism (PCR-SSCP) in 68 cases of advanced gastric cancer, 40 cases of chronic gastritis, 30 cases of intestinal metaplasia and 20 cases of dysplasia. RESULTS: MtMSI was observed in 12.5% (5 of 40) of chronic gastritis, 20.0% (6 of 30) of intestinal metaplasia, 25.0% (5 of 20) of dysplasia and 38.2% (26 of 68) of gastric cancer. These findings showed a sequential accumulation of mtMSI in the histological progression from chronic gastritis to gastric cancer. An association of mtMSI with intestinal histological type and distal location was found (P=0.001 and P=0.002), whereas no significant correlation was found between mtMSI and age at diagnosis, sex, tumor size, depth of invasion, lymph node spread and clinical stages (P>0.05). CONCLUSION: MtMSI may play an early and important role in the gastric carcinogenesis pathway, especially in the intestinal type and distal gastric cancer.

[The specific killing of human melanoma cells by replication selective adenovirus].

OBJECTIVE: To construct replication selective adenovirus AdhepE1 targeting human melanoma and observe its specific killing of human melanoma cells in vitro. METHODS: Adenovirus E1 region, the murine tyrosinase promoter and enhancer DNA sequences were acquired respectively by PCR cloning. The shuttle plasmid of replication-selective adenovirus targeting human melanoma was constructed by DNA recombination. Replication-selective adenovirus AdhepE1 was generated by homologous recombination. The human melanoma cell line SK-Mel-1 and hepatocellular carcinoma cell line HepG2 were attacked separately by lower dose of AdhepE1. Change of cell morphology was observed and the surviving cells were calculated. The expression of E1A was assayed by RT-PCR to verify the specific-replication of AdhepE1. RESULTS: Replication selective adenovirus AdhepE1 targeting human melanoma was acquired by PCR. Human melanoma cell line SK-Mel-1 was sensitive to oncolytic killing of AdhepE1 whereas HepG2 was little responsive. The results of RT-PCR suggested that AdhepE1 replicated specifically in human melanoma cells. CONCLUSION: AdhepE1 can selectively kill human melanoma cells.

[Relationship between mitochondrial DNA instability and interleukin-8 activity in gastric mucosa].

OBJECTIVE: To evaluate the relationship between mitochondrial DNA instability (mtMSI) and interleukin-8 (IL-8) activity in gastric mucosa of various lesions. METHODS: IL-8 level in gastric mucosa was assayed using ELISA method. The mtMSI was detected by PCR-SSCP techniques. RESULTS: mtMSI was observed in 11 out of 30 (36.7%) gastric cancers, 2 of 15 (13.3%) intestinal metaplasia, 2 of 10 dysplasia and 1 of 10 chronic atrophic gastritis. IL-8 level in mtMSI+ group [(76.8 +/- 3.8) pg/mg] was significantly higher than that in mtMSI- group [(48.3 +/- 3.6) pg/mg, P < 0.05]. CONCLUSION: mtMSI closely correlates with IL-8 level in gastric mucosa and is involved in gastric carcinogenesis.

Mutation analysis of APC gene in gastric cancer with microsatellite instability.

AIM: To evaluate the role of APC mutation in gastric carcinogenesis and to correlate APC mutation with microsatellite instability (MSI) in gastric carcinomas. METHODS: APC mutation was measured with multiplex PCR, denaturing gradient gel electrophoresis and DNA sequencing; and MSI was analyzed by PCR-based methods. RESULTS: Sixty-eight cases of sporadic gastric carcinoma were studied for APC mutation at exon 15 and MSI. APC mutations were detected in 15(22.1 %) gastric cancers. Frequence of APC mutation (33.3 %) in intestinal type of gastric cancer was significantly higher than that in diffuse type (13.1 %, P<0.05). On the contrary, no association was observed between APC mutation and tumor size, differentiation, depth of invasion, metastasis or clinical stages. Using five microsatellite markers, MSI in at least one locus was detected in 17 of 68 (25 %) of the tumors analyzed. APC mutations were all detected in MSI-L (only one locus, n=9) or MSS(tumor lacking MSI or stable, n=51), but no mutation was found in MSI-H (> or =2 loci, n=8). CONCLUSION: APC mutation is involved in carcinogenesis of intestinal type of gastric cancer and is independent of MSI phenotype but related to the LOH pathway in gastric cancer.