Xue-Lian Zheng

RNA Interference Against Peroxisome Proliferator-Activated Receptor δ Gene Promotes Proliferation of Human Colorectal Cancer Cells

PurposeThis study was designed to investigate the effects of peroxisome proliferator-activated receptor δ (PPAR δ) on the proliferation and apoptosis of human colorectal cancer cells.MethodsFor RNA interfering (RNAi), HCT-116 cells were transfected with short hairpin RNA (shRNA)-expressing plasmids against PPAR δ or negative control vectors, and the stably transfected cells were selected with G418. The efficacy of RNAi was assessed by real-time reverse transcription polymerase chain reaction (RT-PCR) and Western blotting analysis. The proliferation, cell cycle, and apoptosis of HCT-116 cells treated by RNAi, compared with those containing control vectors or untreated, were analyzed respectively by using MTT (methyl thiazolyl tetrazolium), flow cytometry, and TdT (terminal deoxynucleotidyl transferase)-mediated dUTP nick end-labeling (TUNEL) assay.ResultsRNAi targeting PPAR δ resulted in substantial suppression of PPAR δ expression and significantly promoted the proliferation of HCT-116 cells relative to those with control vectors or untreated, obviously decreasing the frequency of G1-phase cells but had no effect on cell apoptosis.ConclusionsPPAR δ may inhibit the proliferation of CRC cells and increase the number of cells in G1 phase, without any function in cell apoptosis.

Microscopic spread of low rectal cancer in regions of the mesorectum: detailed pathological assessment with whole-mount sections

Background and aimsThe aim was to investigate the regional spread of microscopic tumor nodules in the mesorectum of patients with low rectal cancer, and to provide further pathological evidence for optimal procedure selection of radical resection for rectal cancer.Patients and methodsSixty-two patients with low rectal cancer underwent low anterior resection and total mesorectal excision (TME). Surgical specimens were sliced transversely on serial embedded blocks at 2.5-mm intervals, and stained with hematoxylin and eosin. On whole-mount sections the mesorectum was divided into three regions: the outer region of the mesorectum (ORM), the middle region of the mesorectum (MRM), and the inner region of the mesorectum (IRM). Microscopic metastatic foci were investigated for metastatic mesorectal region, frequency, types, involvement of the lymphatic system, and correlation with the primary tumor.ResultsMicroscopic spread of the tumor was observed in 38.7% of patients (24 out of 62) and that in the ORM was observed in 25.8% of the patients (16 out of 62). Microscopic tumor foci spread in the circumferential resection margin (CRM) occurred in 6.5% of the patients (4 out of 62), and distal mesorectum (DMR) involvement was detected in 6.5% (4 out of 62), with the spread extending to within 3xa0cm from the lower margin of the tumor.ConclusionsThe results of the present study support the theory that complete excision of the mesorectum without destruction of the ORM is essential for surgical management of low rectal cancer, and an optimal DMR clearance resection margin should be no less than 4xa0cm. Four patients with microscopic tumor nodule spread in the ORM observed in the study suggested that microscopic metastases exist in pelvic lateral areas and in the mesorectum simultaneously, indicating the significance of preoperative and/or postoperative radiochemotherapy.

Microsatellite Instability Did Not Predict Individual Survival in Sporadic Stage II and III Rectal Cancer Patients

Objectives: Tumors with high-frequency microsatellite instability (MSI-H) have unique biological behavior and the predictive role of microsatellite instability (MSI) status on survival of colorectal cancer is still debated. The prognostic significance of MSI status in sporadic stage II and III rectal cancer patients needs to be more precisely defined. So we investigated the relationship between MSI status and clinicopathological features and prognosis in these patients. Methods: DNAs from fresh-frozen paired samples of tumors and corresponding normal tissue from 128 stage II and III rectal cancer patients were analyzed for MSI by PCR amplification using markers recommended by a National Cancer Institute workshop on MSI. To assess prognostic significance, Cox proportional hazards modeling was used. Results: Twelve (9.3%) tumors in our study were MSI-H, 28 (21.9%) were low-frequency MSI (MSI-L) and 88 (68.8%) were microsatellite stable (MSS). Most of the MSI-H tumors compared with MSI-L and MSS tumors were found in female patients (p = 0.031), had mucinous histology (p = 0.023), high grade of differentiation (p = 0.002) and high level of preoperative serum carcinoembryonic antigen (p = 0.005). Rectal cancer patients with MSI-H did not show a better clinical outcome than those with MSI-L/MSS, neither in all cases (p = 0.986) nor in stage II and stage III disease analyzed separately (p = 0.705 and p = 0.664, respectively). Conclusions: Data provided here demonstrated there was high incidence of MSI-H and MSI was not a prognostic factor in sporadic stage II and III rectal cancers from the Chinese Han population included in this study. Tumor stage is more suitable than MSI status for prediction of individual survival in sporadic stage II and III rectal cancer patients.

Effect of Inducible Cyclooxygenase Expression on Local Microvessel Blood Flow in Acute Interstitial Pancreatitis

OBJECTIVEnTo investigate the role of inducible cyclooxygenase (COX-2) mRNA expression in local microvessels in rats with acute interstitial pancreatitis (AIP) induced by caerulein injection.nnnMETHODSnThe reverse transcription polymerase chain reaction (RT-PCR) was used to detect COX-2 gene expression in pancreatic tissue. Parameters of acute pancreatitis, such as serum amylase (AMS) and plasma myeloperoxidase (MPO) activities, were assayed using spectrophotometry. Intravital fluorescence microscopy with fluorescein isothiocyanate-labelled erythrocytes was used to study the pancreatic microvessels of rats with AIP and normal control rats.nnnRESULTSnHighly significant increases in COX-2 expression and AMS and MPO activity were seen in rats with AIP compared with controls. After caerulein injection, pancreatic capillary blood flow was decreased (4 hours, p > 0.05; 8 hours, p < 0.001), functional capillary density was reduced (4 hours, p > 0.05; 8 hours, p < 0.001), and there was irregular and intermittent capillary perfusion at 8 hours. There was also a positive correlation between the level of COX-2 expression and MPO activity (plasma, r = 0.5449, p < 0.05; tissue, r = 0.5698, p < 0.05).nnnCONCLUSIONSnThe correlations between increased COX-2 expression and decreased capillary perfusion and blood flow and increased oedema following AIP may show that COX-2 expression can induce neutrophil sequestration to the pancreas, which may be one of the cascading inflammatory factors in the development of AIP.

Regional micrometastasis of low rectal cancer in mesorectum: a study utilizing HE stain on whole-mount section and ISH analyses on tissue microarray.

Aim: To investigate the regional spread of microscopic tumor nodules in the mesorectum of patients with low rectal cancer, and to provide further pathological evidence for optimal procedure selection of radical resection for rectal cancer. Methods: Sixty-two patients with low rectal cancer underwent low anterior resection and total mesorectal excision (TME). Surgical specimens were sliced transversely on serial embedded blocks at 2.5-mm intervals, and stained with hematoxylin and eosin (HE). On whole-mount sections the mesorectum was divided into 3 regions: the outer region of the mesorectum (ORM), the middle region of the mesorectum (MRM), and the inner region of the mesorectum (IRM). Microscopic metastatic foci were investigated for metastatic mesorectal region, frequency, types, involvement of the lymphatic system, and correlation with the primary tumor. Tumor-suspect nodules previously considered disease free by HE stain on whole-mount section were examined by in situ hybridization (ISH) on tissue microarray (TMA) through detecting mRNAs of CEA and CK20 with non radioactive biotin-tagged oligonucleotide probes. Results: Microscopic spread of the tumor was observed in 50.0 percent of patients (31 out of 62, 24 by HE stain on whole-mount section and 7 by ISH on TMA) and that in the ORM was observed in 38.7 percent of the patients (24 out of 62, 16 observed by HE stain on whole-mount section and 8 by ISH on TMA). Microscopic tumor foci spread in the circumferential resection margin (CRM) occurred in 8.1 percent of the patients (5 out of 62, 4 observed by HE stain on whole-mount section and one by ISH on TMA), and distal mesorectum (DMR) involvement was detected in 6.5 percent (4 out of 62, all observed by HE stain on whole-mount section), with the spread extending to within 3 cm from the lower margin of the tumor. Most (26 of 31) of the patients with microscopic spread in mesorectum had TNM Stage III diseases. Conclusions: The results of the present study support the theory that complete excision of the mesorectum without destruction of the ORM is essential for surgical management of low rectal cancer, and an optimal DMR clearance resection margin of no less than 4 cm was referenced. Five patients with microscopic tumor nodule spread in the CRM observed in the study suggested that microscopic metastases exist in pelvic lateral areas and in the mesorectum simultaneously, indicating the significance of preoperative and/or postoperative radiochemotherapy.

An improved primary culture system of pancreatic duct epithelial cells from Wistar rats

Pancreatic duct epithelial cells (PDEC) are involved in most common pancreatic diseases. Primary cultivation of PDEC is a prerequisite for in vitro studies, in which in vivo situations can be simulated and molecular mechanisms investigated better than in cultured cell lines. However, some problems still exist regarding rat PDEC primary cultivation. In this study, an improved primary culture system of rat PDEC is presented. Some modifications, especially regarding specimen chosen, digestive control and epithelium purification, were made to simplify the procedure, increase cell yield, and improve epithelium purification. Cultures were identified as PDEC by morphological characteristics, reverse transcription-polymerase chain reaction and immunocytochemistry staining with cytokeratin 19. In addition, growth characteristics of rat PDEC are described in detail. This improved technique, which is more efficient and cost-effective, will be useful for in vitro pancreatic studies.