Effects of probiotic on intestinal mucosa of patients with ulcerative colitis.

AIM: To investigate the effects of probiotic on intestinal mucosae of patients with ulcerative colitis (UC), and to evaluate the role of probiotic in preventing the relapse of UC. METHODS: Thirty patients received treatment with sulphasalazine (SASP) and glucocorticoid and then were randomly administered bifid triple viable capsule (BIFICO) (1.26 g/d), or an identical placebo (starch) for 8 wk. Fecal samples were collected for stool culture 2 wk before and after the randomized treatments. The patients were evaluated clinically, endoscopically and histologically after 2 mo of treatment or in case of relapse of UC. p65 and IkappaB expressions were determined by Western blot analysis. DNA-binding activity of NF-kappaB in colonic nuclear extracts was detected by electrophoretic mobility shift assay (EMSA). mRNA expressions of cytokines were identified by semi-quantitative assay, reverse transcriptase- polymerase chain reaction (RT-PCR). RESULTS: Three patients (20%) in the BIFICO group had relapses during 2-mo follow-up period, compared with 14 (93.3%) in placebo group (P<0.01). The concentration of fecal lactobacilli, bifidobacteria was significantly increased in BIFICO-treated group only (P<0.01). The expressions of NF-kappaB p65 and DNA binding activity of NF-kappaB were significantly attenuated in the treatment group than that in control (P<0.05). The mRNA expression of anti-inflammatory cytokines was elevated in comparison with the control group. CONCLUSION: The probiotic could impede the activation of NF-kappaB, decrease the expressions of TNF-alpha and IL-1beta and elevate the expression of IL-10. These results suggest that oral administration of this new probiotic preparation is effective in preventing flare-ups of chronic UC. It may become a prophylactic drug to decrease the relapse of UC.

Contribution of eIF-4E inhibition to the expression and activity of heparanase in human colon adenocarcinoma cell line: LS-174T.

AIM: Heparanase degrades heparan sulfate proteoglycans (HSPGs) and is a critical mediator of tumor metastasis and angiogenesis. Recently, it has been cloned as a single gene family and found to be a potential target for antimetastasis drugs. However, the molecular basis for the regulation of heparanase expression is still not quite clear. The aim of this study was to determine whether the expression of eukaryotic initiation factor 4E (eIF-4E) correlated with the heparanase level in tumor cells and to explore the correlation between heparanase expression and metastatic potential of LS-174T cells. METHODS: A 20-mer antisense s-oligodeoxynucleotide (asODN) targeted against the translation start site of eIF-4E mRNA was introduced into LS-174T cells by lipid-mediated DNA-transfection. eIF-4E protein and mRNA levels were detected by Western blot analysis and RT-PCR, respectively. Heparanase activity was defined as the ability to degrade high molecular weight (40-100 kDa) radiolabeled HS (heparan sulfate) substrate into low molecular weight (5-15 kDa) HS fragments that could be differentiated by gel filtration chromatography. The invasive potential of tumor cell in vitro was observed by using a Matrigel invasion assay system. RESULTS: The 20-mer asODN against eIF-4E specifically and significantly inhibited eIF-4E expression at both transcriptional and translational levels. As a result, the expression and activity of heparanase were effectively retarded and the decreased activity of heparanase resulted in the decreased invasive potential of LS-174T. CONCLUSION: eIF-4E is involved in the regulation of heparanase production in colon adenocarcinoma cell line LS-174T, and its critical function makes it a particularly interesting target for heparanase regulation. This targeting strategy in antisense chemistry may have practical applications in experimental or clinical anti-metastatic gene therapy of human colorectal carcinoma.

A new three-layer-funnel-shaped esophagogastric anastomosis for surgical treatment of esophageal carcinoma.

AIM: To reduce the incidence of postoperative anastomotic leak, stenosis, gastroesophageal reflux (GER) for patients with esophageal carcinoma, and to evaluate the conventional method of esophagectomy and esophagogastroplasty modified by a new three-layer-funnel-shaped (TLF) esophagogastric anastomotic suturing technique. METHODS: From January 1997 to October 1999, patients with clinical stage I and II (IIa and IIb) esophageal carcinoma, which met the enrollment criteria, were surgically treated by the new method (Group A) and by conventional operation (Group B). All the patients were followed at least for 6 months. Postoperative outcomes and complications were recorded and compared with the conventional method in the same hospitals and with that reported previously by McLarty et al in 1997 (Group C). RESULTS: 58 cases with stage I and II (IIa and IIb) esophageal carcinoma, including 38 males and 20 females aged from 34 to 78 (mean age: 57), were surgically treated by the TLF anastomosis and 64 by conventional method in our hospitals from January 1997 to October 1999. The quality of swallowing was improved significantly (Wilcoxon W=2 142, P=0.0 001) 2 to 3 months after the new operation in Group A. Only one patient had a blind anastomatic fistula diagnosed by barium swallow test 2 months but healed up 3 weeks later. Postoperative complications occurred in 25 (43 %) patients, anastomotic stenosis in 8 (14 %), and GER in 13 (22 %). The incidences of postoperative anastomotic leak, stenosis and GER were significantly decreased by the TLF anastomosis method compared with that of conventional methods (chi(2)=6.566, P=0.038; chi(2)=10.214, P=0.006; chi(2)=21.265, P=0.000). CONCLUSION: The new three-layer-funnel-shaped esophagogastric anastomosis (TLFEGA) has more advantages to reduce postoperative complications of anastomotic leak, stricture and GER.

A novel method for preparation of tissue microarray.

AIM: To improve the technique of tissue microarray (tissue chip). METHODS: A new tissue microarraying method was invented with a common microscope installed with a special holing needle, a sampling needle, and a special box fixing paraffin blocks on the microscope slide carrier. With the movement of microscope tube and objective stage on vertical and cross dimensions respectively, the holing procedure on the recipient paraffin blocks and sampling procedure of core tissue biopsies taken from the donor blocks were performed with the refitted microscope on the same platform. The precise observation and localization of representative regions in the donor blocks were also performed with the microscope equipped with a stereoscope. RESULTS: Highly-qualified tissue chips of colorectal tumors were produced by a new method, which simplified the conventional microarraying procedure, and was more convenient and accurate than that employing the existing tissue microarraying instruments. CONCLUSION: Using the refitted common microscope to produce tissue microarray is a simple, reliable, cost-effective and well-applicable technique.

Effect of high temperature on gastrin, somatostatin and motilin production in ulcerous gastric antral mucosa of rats.

OBJECTIVE: To investigate the changes of gastrin, somatostatin and motilin production in the gastric antral mucosa of rats with experimental gastric ulcer. METHODS: Rat models of gastric ulcer model were induced successfully by injection of acetic acid into the gastric antral wall of 2 groups of Wistar rats (7 in each group) that were subjected to environment of either high or normal temperature. Another 2 groups of rats (n=7) receiving normal saline injection in the same manner, along with still another 2 groups (n=7) without any treatment, all of which were kept under conditions with different temperatures accordingly, constituted the control groups. The levels of gastrin, somatostatin and motilin in the gastric antral mucosa of the rats were measured with radioimmunoassay. RESULTS: In rats with gastric ulcer, the levels of gastrin and motilin in the antral mucosa increased, but in a lesser scale in rats with ulcer kept in high temperature than in normal temperature group, while that of somatostatin was reduced. The level of somatostatin declined less in the high temperature group with ulcer than in the normal temperature group with ulcer. CONCLUSION: High temperature can affect gastrin, somatostatin and motilin production in the gastric antral mucosa of rats with gastric ulcer.

[In vitro regulation of HLA and B7 expression in multidrug-resistant breast cancer cells].

OBJECTIVE: To study the modulation of the expressions of human leucocyte antigen (HLA) and co-stimulatory molecules B7 (CD80, CD86) in multidrug-resistant (MDR) breast cancer cells. METHODS: The MDR phenotype MCF-7/ADR cells were treated with recombinant human interferon (IFN)-alpha2b (rhIFN-alpha2b) at different doses for varied length of time, and the expressions of HLA and B7 were determined by flow cytometry (FCM) and compared with those of non-treated cells. Statistical analysis was performed using SPSS10.0 software. RESULTS: At the doses smaller than 1x10(3) IU/ml, rhIFN-alpha2b did not exhibit significant inhibition on MCF-7/ADR cell growth, but HLA and B7 expressions were enhanced in a dose- and time-dependent fashion and significant up-regulation occurred 24 h after 400 IU/ml rhIFN-alpha2b treatment. Treatment with rhIFN-alpha2b at the doses of 100, 400, 800 IU/ml respectively for 24 h produced significant increases in the positive cell ratios for HLA class I expression (P<0.05), HLA-DR expression (P<0.005) and B7-2 expression (P<0.05), while changes in B7-1 expression was not obvious (P>0.05). The mean relative linear fluorescence intensity (RLFI) of HLA class I was also significantly increased (P<0.001) but alterations in the mean RLFI of HLA-DR and B7 were not significant. CONCLUSIONS: rhIFN-alpha2b within the dose range from 400 to 800 IU/ml can effectively enhance the expression of HLA and B7 molecules, suggesting that it may have the potential to reverse immune tolerance of breast cancer cells. Cytokine treatment may be effective in reversing immune tolerance caused by down-expression of HLA and B7.

[Diagnosis of hepatocellular carcinoma with bone metastasis using fine needle aspiration biopsy: report of 2 cases].

The clinical presentations and imaging characteristics of hepatocellular carcinoma (HCC) are usually various and complex. This article reported 2 cases of HCC with the initial symptoms being bone metastasis that did not exhibit typical clinical manifestations or distinct feature in liver imaging, without elevated serum alpha-fetoprotein. The diagnoses were certified by fine needle aspiration biopsy, which is recommended for differential diagnosis in cases of complex HCC with bone metastasis, to avoid liver biopsy and surgical bone biopsy.

[Construction and application of colorectal polyp database].

OBJECTIVE: To study the construction and application of computerized database of colorectal polyp in the clinical management and research of this disease. METHOD: A colorectal polyp database and its management system was constructed on the basis of Microsoft Access 2000. Clinical, endoscopic and pathological data, which went through standardized and elemental processing, of 2 627 cases (4 850 records) of colorectal polyp collected from 1990 to 2000 in Nanfang Hospital was entered into this database. RESULTS: Using this new database, the information on the population and age distribution, location and clinical features of colorectal polyps were obtained. Comparative study of the clinical and pathological findings in the cases, evaluation of the therapeutic effects, statistical review of the identification of the polyp and its canceration in the previous years as well as the analysis of other relevant factors were successfully accomplished, which greatly facilitated the follow-up study of some chosen cases that may be of clinical significance. CONCLUSIONS: Applications of modern informatics and computer technology greatly facilitates case management and clinical research of colorectal polyps, and standardized and elemental processing of the clinical data offers a new possibility for easy case information management.

[Construction and identification of the cDNA phage expression library for human colorectal cancer antigens].

OBJECTIVE: To construct a cDNA phage expression library for human colorectal carcinoma antigens. METHODS: After the total RNA was extracted from human colorectal cancer tissues, the single-strand and double-strand cDNA were synthesized through reverse transcriptase PCR and long-distance PCR, with the cDNA fragments smaller than 500 bp removed and the remaining cDNA combined with the right and left arms of dephosphorylated lambdaTriplEx2 phage vector. The recombinant phage were then packaged in vitro by MaxPlax Packaging extract, and a small portion of the packaged phage was used to infect E.coli XL1-Blue. Titer measurement was performed so as to determine the capacity of the library. SfiI restriction endonucleases was used to cut the recombined phage DNA in order to identify the size of inserted cDNA. RESULTS: The constructed cDNA phage expression library for human colorectal cancer antigens consisted of 2.39 x 10(6) pfu/ml bacteriophages with a recombination rate of 97.5% and the length of the inserted cDNA fragment ranged from 600 to 4,000 bp with an average of 1,400 bp. CONCLUSION: The cDNA phage expression library of human colorectal cancer antigens is successfully constructed to meet the currently recognized standards, and can be well applicable in screening cDNA-cloned genes of human colorectal cancer-associated antigens by immunoscreening.

[A novel method for fabrication of tissue microarray].

BACKGROUND & OBJECTIVE: Tissue chip (tissue microarray, TMA) is one of the most important biochip techniques, just following the gene chip and the protein chip, which is one of the most important functional genomics and proteomics research methods in the post-genomic era. However, the present TMA technology has certain shortcomings, such as lack of advanced instruments, tedious procedure, and low sampling accuracy, etc. This new method was designed to improve the technology of TMA. METHODS: A common microscope was installed with a special holing needle, a sampling needle, and a proper box to fix paraffin blocks on the microscope carrier. With the precise mechanical control of microscope, the holing procedure on the recipient paraffin blocks and sampling procedure of core tissue biopsies were performed with the re-equipped microscope. The precise observation and localization of sampling regions were also performed using the same microscope equipped with a stereoscope. RESULTS: The new method simplified TMA procedure, and the whole process of holing, locating, and sampling was performed with the same instrument on the same platform. The single-use holing and sampling needles were first applied to maintain higher accuracy and to avoid the tissue remains and contamination among different samples. And high-qualified tissue chips of colorectal tumors were produced successfully by the new method. CONCLUSION: Using the re-equipped common microscope to fabricate tissue microarrays is a simple, reliable, cost-effective,and well-applicable technique.