Oxymatrine could promote mesenchymal stem cell therapy in hepatic fibrosis rats: an experimental research / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To investigate whether oxymatrine (OM) could promote mesenchymal stem cell (MSC) therapy in CCl4-induced hepatic fibrosis (HF) in rats and to initially explore its mechanisms.</p><p><b>METHODS</b>Totally 50 male SD rats were randomly divided into five groups,i.e., the normal control group, the model group, the MSC therapy group, the OM therapy group, and the MSC combined OM therapy group, 10 in each group. Except the normal control group, the HF model was duplicated by CCl4 induction. After successful modeling, rats in the MSC therapy group received 5 x10(6) MSCs by intravenous injection via caudal vein, once a week. Rats in the OM therapy group received 50 mg/kg OM by intramuscular injection, three times a week. Rats in MSC combined OM therapy group received 5 x 10(6) MSCs by intravenous injection via caudal vein, once a week and 50 mg/kg OM by intramuscular injection three times a week. Equal volume of normal saline was given to those in the normal control group and the model group. All medication lasted for 8 weeks. Serum levels of ALT and AST were detected 8 weeks later. The hepatic histopathological injury and extracellular matrix deposit were assessed using HE and Masson staining. Expressions of serum interleukin-4 (IL-4) and interleukin-10 (IL-10) were detected using enzyme linked immunosorbent assay (ELISA).</p><p><b>RESULTS</b>(1) Compared with the normal control group, serum levels of ALT and AST significantly increased in the model group (P < 0.05). Compared with the model group, serum levels of ALT and AST significantly decreased in the OM therapy group, the MSC therapy group, and the MSC combined OM therapy group at the end of 8 weeks of treatment (P < 0.05). But serum levels of ALT and AST were significantly lower in the MSC combined OM therapy group than in the OM therapy group and the MSC therapy group (P < 0.05). (2) Compared with the model group, the hepatic injury was significantly lessened and the area of extracellular matrix deposit was significantly reduced in the OM therapy group, the MSC therapy group, and the MSC combined OM therapy group (P < 0.05). Besides, they wer more significant in the MSC combined OM therapy group (P < 0.05). (3) Compared with the model group, the serum IL-4 level was significantly higher in the MSC therapy group and the MSC combined MO group (P < 0.05). It was higher in the MSC combined MO group (P < 0.05). Although the serum IL-4 level also increased in the OM therapy group, but with no statistical difference (P > 0.05). (4) The serum IL-10 level significantly increased in the OM therapy group, the MSC therapy group, and the MSC combined OM therapy group (P < 0.05), and it was the highest in the MSC combined OM therapy group among the three groups (P < 0.05). (5) Two-photon fluorescence imaging showed no signals of MSCs in liver with or without OM injection.</p><p><b>CONCLUSION</b>OM could promote mesenchymal stem cell therapy in hepatic fibrosis rats, which might be involved in increasing serum levels of IL-4 and IL-10.</p>

Differential proteomic analysis of rat hepatic stellate cells treated by oxymatrine liposomes using two-dimensional electrophoresis / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To analyze differentially expressed proteins of hepatic stellate cells (HSCs) treated with oxymatrine (OMT) liposomes, thus further exploring the molecular mechanism of OMT liposomes for treating liver fibrosis.</p><p><b>METHODS</b>A rat model of CCl4 induced chronic liver fibrosis was established. HSCs were perfusion isolated from modeled SD rats and cultured in vitro . Passage 2 HSCs were divided into the model group (Group A), the OMT-liposome-treated group (Group B), and the liposome-treated control group (Group C). HSCs from normal rats were taken as the normal control group (Group D). The total proteins of HSCs cells were extracted from Group B and D after 7 days of treatment, and separated with isoelectrofocusing two-dimensional electrophoresis (2-DE). A 2-DE system was established to analyze the differences in the protein profile between Group B and Group C. Tow protein dots with most obvious difference were selected to determine the structures and functions of different proteins using peptide mass fingerprinting (PMF).</p><p><b>RESULTS</b>(1) The total number bf proteins decreased after treated with OMT liposomes, with 864 spots before treatment and 756 spots after treatment, and the matching rate was 63%. (2) According to 2-DE results, 10 differential protein spots were found by image analysis of magnifying images in local regions. (3) Two most differently expressed proteins were identified to be ATM (46. 236 kD) and Miz1 (54. 051 kD) by PMF and SWISS-PROT protein database retrieval.</p><p><b>CONCLUSION</b>Action of OMT liposomes on HSCs of rats with chronic liver fibrosis caused different protein expressions, which might be involved in the signaling pathways of inducing the apoptosis of HSCs.</p>

In vivo transplantation of bone marrow mesenchymal stem cells accelerates repair of injured gastric mucosa in rats / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Adult stem cells provide a promising alternative for the treatment of injured tissues. We aimed to investigate the effect of in vivo transplantation of bone marrow mesenchymal stem cells (BMMSCs) on injured gastric mucosa in rats.</p><p><b>METHODS</b>The gastric ulcer in rats was induced by indomethacin. BMMSCs from male rats, labeled with the fluorescent cell linker 5,6-carboxyfluorescein diacetate succinimidyl ester (CFDA SE), were transplanted into the female rats via tail vein injection. The healing process of gastric ulcers was monitored by HE staining. The protein levels of vascular endothelial growth factor (VEGF) and the epidermal growth factor receptor (EGFR) in the injured gastric mucosa were determined by immunohistochemistry.</p><p><b>RESULTS</b>At 48 and 72 hours after BMMSCs transplantation, the CFDA SE labeled cells were found scattered in the injured gastric mucosa, but not in the gastric mucosa of control rats. At 72 hours after BMMSCs transplantation, the mean ulcer index was 12.67 ± 2.16 in the BMMSCs transplanted group and 17.33 ± 1.97 in vehicle-treated controls (P < 0.01). Both VEGF and EGFR protein expression levels were significantly higher in the gastric section from the rats that received BMMSCs transplantation as compared to rats without BMMSCs transplantation.</p><p><b>CONCLUSION</b>Autologous BMMSCs transplantation can accelerate gastric ulcer healing in injured gastric mucosa in a rodent model.</p>

Expression of S100A6 in primary and metastatic human gastric cancer / 中华肿瘤杂志

<p><b>OBJECTIVE</b>Some members of the S100 gene family have been suggested to be associated with cancer development and metastasis. Our previous cDNA micro-array studies have showed S100A6 expression is elevated in gastric cancer compared with that in paired normal mucosa. To validate our previous results and further investigate the possible role of S100A6 gene in gastric cancer, we carried out this detailed S100A6 expression analysis in more matched gastric cancer samples.</p><p><b>METHODS</b>S100A6 expression was detected in 20 paired fresh surgical samples of gastric tumor tissue and matched non-cancerous mucosa by QRT-PCR. A gastric cancer tissue microarray (TMA) containing 1020 duplicate matched normal mucosa, gastric cancer tissue and metastatic lymph node tissue cores from 208 gastric cancer patients was constructed. S100A6 expression was detected by immunohistochemistry and the correlation between S100A6 expression with clinicopathological factors and survival was analyzed.</p><p><b>RESULTS</b>As quantitated by QRT-PCR, S100A6 transcript level was elevated in 73.7% of the primary cancer lesions with an average 2.25-fold up-regulation than that in matched non-neoplastic mucosa. As displayed by immunohistochemistry, the positive rate of S100A6 in non-neoplastic mucosa, tumor lesions and metastatic lymph nodes was 34.3%, 84.1% and 90.9%, respectively. S100A6 expression level in cancer and metastatic lymph node was significantly higher than their matched non-neoplastic mucosa (P < 0.05). 65.5% of patients showed an increased S100A6 expression in cancer tissue compared with that in matched normal mucosa. S100A6 overexpression was associated with larger tumor size and deeper invasion (P = 0.022 and P = 0.009). No evidence was found for an association between S100A6 expression level and other variables, including tumor grade, nodal metastases, and TNM stage. There was no association between S100A6 expression level and survival. But compared with paired non-neoplastic mucosa, an increased S100A6 expression in tumor lesion predicated a decreasing suvival if compared with a decreased S100A6 expression, though the difference was statistically not significant.</p><p><b>CONCLUSION</b>Elevated expression of S100A6 gene may be an early event in the development and progression of gastric cancer. Further study of this gene may be helpful for understanding the nature of gastric carcinoma.</p>

Gene expression profile of human normal gastrointestinal tract tissues: bioinformatic study / 中国应用生理学杂志

<p><b>AIM</b>To identify up-regulated genes specific to human normal gastrointestinal tissues.</p><p><b>METHODS</b>Study was made on human normal tissue gene expression database open to the public. Tissue-specific genes were identified using one-tailed student T test. Online software including Ingenuity and KEGG were applied for physiological function analyses. Unsupervised two-way hierarchical clustering method was used to analyze the expression profile of stomach-specific genes in gastric cancer gene expression datasets.</p><p><b>RESULTS</b>The analyses identified 196 stomach-specific genes, 203 ileum-specific genes and 224 colon-specific genes, respectively. The gene expression profiles reflect major organ-specific physiological functions on the molecular level. Some putative oncogenes and tumor suppressor genes were found in the tissue-specific gene list. Hierarchical clustering analysis revealed that the stomach-specific genes were up-regulated in normal stomach tissues but down-regulated in stomach cancer tissues. The normal tissues clustered together, so did the cancer tissues. At the meantime, clustering could also distinguish the moderate and severe differentiated stomach cancer.</p><p><b>CONCLUSION</b>Human normal stomach, ileum and colon possess tissue-specific up-regulated genes, which are closely associated with physiological functions.</p>

Construction of a gastric cancer related gene GCRG213-specific siRNA expression vector and its effect on gastric cancer cells in vitro / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate the effect of gene GCRG213 siRNA transfection into gastric cancer cell line MKN45 cells.</p><p><b>METHODS</b>Two pairs of DNA sequences containing small hairpin structure to GCRG213 were designed and synthesized. The complement form was obtained by annealing and inserted into RNAi expression vector IMG-800. They are IMG-800-1 and IMG-800-2 correspondingly. The recombinant plasmid IMG-800-1, IMG-800-2 and the vector IMG-800 were separately transfected into MKN45 cells conducted by lipofectamine 2000. After G418 selecting, the cells were transfected steadily. Expression of GCRG213 was detected by semi-quantitative RT-PCR and Western Blot. The growth graph of six steady transfected cell cultures was protracted by cell counting. FACS was used to detect the cell cycle, and Annexin V FITC/PI double labeling were used to detect the effects on cell apoptosis in the above-mentioned cells. The clone formation rate in plate and in nude mice was tested to investigate the tumorigenic characteristics of the six steadily transfected cells in vitro and vivo.</p><p><b>RESULTS</b>Through sequencing, two pairs of DNA sequences containing small hairpin structure to GCRG213 were proved to be successfully cloned into siRNA expression vector IMG-800, correspondingly called IMG-800-1 and IMG-800-2. The recombinant plasmid IMG-800-1, IMG-800-2 and vector IMG-800 were transfected separately into MKN45 cells conducted by lipofectamine 2000. After G418 selecting, the cells were transfected steadily. Transfecting the siRNA vector (IMG-800-1, IMG-800-2 ) into the MKN45 cells significantly decreased the expression of GCRG213, at both mRNA and protein levels. The growth graph showed that the growth of IMG-800-1 and IMG-800-2 transfected cells were slower than that of vector transfected cells. The proportion of cells in G2/M and/or S phase decreased in the cells transfected with IMG-800-1 and IMG-800-2 and cell apoptosis increased. The average clone formation rate in vitro decreased in the cells transfected with IMG-800-1 and IMG-800-2, compared with those transfected with vector. In vivo, the time of tumor formation of IMG-800-1 and IMG - 800-2 transducted cells in nude mice was prolonged and the tumor size was smaller.</p><p><b>CONCLUSION</b>GCRG213 SiRNA transfection may induce inhibition of growth and proliferation of tumor cells, promote cell apoptosis, and inhibit the tumorigenicity in vitro and vivo.</p>

Gene expression profile of human adenocarcinoma by cDNA microarray and clustering / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the gene expression profile of human gastric adenocarcinoma by means of cDNA microarray and to analyze its biological significance.</p><p><b>METHODS</b>Paired tumor and non-tumor specimens from 18 cases of advanced gastric adenocarcinoma were studied. Total RNA was isolated and labeled by reverse transcription reaction with cy5 and cy3 for cDNA probe. cDNA microarrays containing 148 genes were hybridized with labeled cDNA probe. Data from cDNA microarray experiments were analyzed by average-linkage hierarchical clustering and significance analysis of microarrays (SAM).</p><p><b>RESULTS</b>Eighteen tumor and non-tumor specimens were clearly divided by clustering analysis. Three features of gene expression profile were found in gastric adenocarcinoma and non-tumor tissues. The profile of differential gene expression in tumor and non-tumor tissues was mainly shown in feature B and feature C. In gastric adenocarcinoma tissues, the expression of genes in feature B was lower and that in feature C was higher. The profile of differential gene expression among gastric adenocarcinoma tissues was found in feature A. In feature A, the profile of similar gene expression was found in paired tumor and non-tumor tissues from 13 patients. SAM analysis showed that 19 genes in feature B and 12 genes in feature C were of significant difference between tumor and non-tumor specimens. The expression levels of genes related to cell cycle, growth factor, cell adhesion, and matrix remodeling were higher or lower in gastric adenocarcinoma tissues.</p><p><b>CONCLUSION</b>Data from cDNA microarray experiments can clearly distinguish gastric adenocarcinoma from non-tumor tissues. The profiles show that gene expression in gastric adenocarcinomas is both homogeneous and heterogeneous. The homogeneous gene expression profile is found in both tumor and non-tumor tissues from 13 patients, suggesting that some gene aberrance is an early event of carcinogenesis of gastric adenocarcinoma. This study provides not only a new molecular basis for understanding biological properties of gastric adenocarcinoma, but also useful resources for future development of diagnostic and prognostic markers for gastric adenocarcinoma.</p>

Clinical analysis of 38 elderly patients with early double primary cancers / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the clinical features and proper treatment of 38 elderly patients with early double primary cancers.</p><p><b>METHODS</b>Thirty-eight elderly patients with early double primary cancers treated from January 1980 to March 2003 were retrospectively reviewed for involved organs, treatment and prognosis.</p><p><b>RESULTS</b>Digestive tract was the most frequently involved, followed by urogenital system and lung. Long-term results of endoscopic mucosal resection (EMR), operation and radiotherapy were superior to other methods. The prognosis of gastrointestinal carcinoma was better than that of prostate carcinoma and hematopoietic system. The operation rate decreased with increasing age. The 5-year survival rates of EMR, operation and radiotherapy were 85.7%, 71.1% and 75.0%, respectively. The medium survival time was 120 months in first cancer and 39 months in the second primary cancer. The 5-year survival rates of the first cancer and second primary cancer were 88.6% and 53.8%.</p><p><b>CONCLUSION</b>Yearly follow-up for elderly patients with endoscopy, beta ultrasonic scan and X-ray contribute to finding of early double primary cancers. Operation is the best treatment of early double primary cancers. Endoscopic mucosal resection is especially suitable for old patients with digestive tract and bladder cancer.</p>

Influence of dendritic cell infiltration on prognosis and biologic characteristics of progressing gastric cancer / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the relation between dendritic cell (DC) infiltration and clinicopathologic parameters, biologic characteristics and prognosis of progressing gastric cancer.</p><p><b>METHODS</b>The development of apoptotic cell death (apoptotic index, AI) in 61 progressing gastric carcinoma tissues was analyzed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate biotin nick end labeling (TUNEL) method. The PCNA labeling index (PCNA-LI), density of dendritic cells in the tumor were detected by immunohistochemical method by the LSAB kit using antibody against S-100 protein and PC-10.</p><p><b>RESULTS</b>DC infiltration was negatively correlated with lymph node metastasis, clinical stage and PCNA-LI, but positively with AI. The DCs in gastric cancer groups with and without lymph node metastasis were (5.63 +/- 4.37)/HPF and (8.51 +/- 5.57)/HPF with difference significant (P < 0.05). The DC infiltration in I, II, III stage lesions were (11.23 +/- 6.05)/HPF, (6.28 +/- 4.37)/HPF and (5.53 +/- 5.19)/HPF also with differences significant (P < 0.01). The PCNA-LI was significantly higher in the low DC group (57.10% +/- 14.18%) than that of high DC group (48.15% +/- 10.59%, P < 0.01). AI findings were 3.77% +/- 1.26% and 2.95% +/- 1.07% in the high and low DC groups (P < 0.01). A positive correlation was observed between DC infiltration and AI (r = 0.39, P < 0.01) whereas a negative correlation between DC infiltration and PCNA-LI (r = -0.47, P < 0.01). The prognosis of high DC infiltration patients was significantly better than those with low ones.</p><p><b>CONCLUSION</b>The infiltrating dendritic cells in and around tumor, representing the local immune status of the host, may play an important role in immunological defense mechanism of host versus tumor. Dendritic cells may inhibit the proliferation and induce the apoptosis of the tumor cells, thus affecting the clinical features and improve the prognosis of gastric carcinoma.</p>