Differential gene expression profiles of peripheral blood mononuclear cells in childhood asthma.

OBJECTIVE: Asthma is a common childhood disease with strong genetic components. This study compared whole-genome expression differences between asthmatic young children and healthy controls to identify gene signatures of childhood asthma. METHODS: Total RNA extracted from peripheral blood mononuclear cells (PBMC) was subjected to microarray analysis. QRT-PCR was performed to verify the microarray results. Classification and functional characterization of differential genes were illustrated by hierarchical clustering and gene ontology analysis. Multiple logistic regression (MLR) analysis, receiver operating characteristic (ROC) curve analysis, and discriminate power were used to scan asthma-specific diagnostic markers. RESULTS: For fold-change>2 and p < 0.05, there were 758 named differential genes. The results of QRT-PCR confirmed successfully the array data. Hierarchical clustering divided 29 highly possible genes into seven categories and the genes in the same cluster were likely to possess similar expression patterns or functions. Gene ontology analysis presented that differential genes primarily enriched in immune response, response to stress or stimulus, and regulation of apoptosis in biological process. MLR and ROC curve analysis revealed that the combination of ADAM33, Smad7, and LIGHT possessed excellent discriminating power. CONCLUSIONS: The combination of ADAM33, Smad7, and LIGHT would be a reliable and useful childhood asthma model for prediction and diagnosis.

Gene therapy in pancreatic cancer.

Pancreatic cancer (PC) is a highly lethal disease and notoriously difficult to treat. Only a small proportion of PC patients are eligible for surgical resection, whilst conventional chemoradiotherapy only has a modest effect with substantial toxicity. Gene therapy has become a new widely investigated therapeutic approach for PC. This article reviews the basic rationale, gene delivery methods, therapeutic targets and developments of laboratory research and clinical trials in gene therapy of PC by searching the literature published in English using the PubMed database and analyzing clinical trials registered on the Gene Therapy Clinical Trials Worldwide website (http://www. wiley.co.uk/genmed/ clinical). Viral vectors are main gene delivery tools in gene therapy of cancer, and especially, oncolytic virus shows brighter prospect due to its tumor-targeting property. Efficient therapeutic targets for gene therapy include tumor suppressor gene p53, mutant oncogene K-ras, anti-angiogenesis gene VEGFR, suicide gene HSK-TK, cytosine deaminase and cytochrome p450, multiple cytokine genes and so on. Combining different targets or combination strategies with traditional chemoradiotherapy may be a more effective approach to improve the efficacy of cancer gene therapy. Cancer gene therapy is not yet applied in clinical practice, but basic and clinical studies have demonstrated its safety and clinical benefits. Gene therapy will be a new and promising field for the treatment of PC.

Effects of azathioprine and infliximab on mesenchymal stem cells derived from the bone marrow of rats in vitro.

Mesenchymal stem cell (MSC) transplantation has been demonstrated to be promising in the treatment of inflammatory bowel disease (IBD). Azathioprine (AZA) is widely used in IBD patients. Infliximab, as a representative of biological therapy for IBD, is important in the treatment regimen. In the present study we investigated the effects of AZA and infliximab on the cell proliferation, cell cycle and apoptosis of the MSCs derived from the bone marrow of Sprague­Dawley (SD) rats in vitro in order to provide preliminary data for optimizing the treatment of IBD. MSCs derived from the bone marrow of rats were either cultured in various concentrations of AZA­ or infliximab­supplemented medium for 24, 48 and 72 h, respectively. The growth curves of MSCs were obtained. The apoptosis and the cell cycle of the MSCs were analyzed by flow cytometry. AZA decreased the proliferation of MSCs by 66% and increased apoptosis at 0.20 mg/ml for 72 h (P<0.05). The percentage of necrotic cells increased markedly in MSCs treated with 0.30 mg/ml AZA for 72 h (P<0.05). As the exposure time increased, the percentage of MSCs in phase G0­G1 increased and that in phase S decreased in AZA groups exceeding 0.20 mg/ml (P<0.05). However, infliximab had a minimal impact on the cell proliferation, apoptosis and cell cycle of the MSCs. AZA was able to inhibit cell proliferation and induce apoptosis of the MSCs in vitro. Infliximab did not affect the cell proliferation, apoptosis and cell cycle of the MSCs derived from rats.

The association between IFN-γ and IL-4 genetic polymorphisms and childhood susceptibility to bronchial asthma.

The present study aims to investigate the association between the genetic polymorphisms of interferon (IFN)-γ and interleukin (IL)-4 with childhood susceptibility to asthma and the levels of IFN-γ, IL-4, and immunoglobulin (Ig) E among asthmatic children. A total of 100 asthmatic children and 122 control children were enrolled in the present study. The genotypes of the IFN-γ gene at the -179G/T locus and the IL-4 gene at the -33C/T and -589C/T loci were detected using polymerase chain reaction with restriction fragment length polymorphism. The IFN-γ gene at the +874A/T locus and the IFN-γ CA repeats were tested using allele-specific and capillary electrophoresis, respectively, whereas the IFN-γ, IL-4, and total IgE levels were measured using enzyme-linked immunosorbent assays. The 100 asthmatic children and the 122 control children were all GG homozygous in the -179 locus of the IFN-γ gene, which shows that the IFN-γ gene is not mutated at the -179 locus. No significant differences were found in terms of genotypic and allelic frequency distribution in the IFN-γ gene or the CA repeat at the +874A/T locus between the asthmatic children and the control (P>0.05). An association was found between the polymorphism of the IFN-γ gene at +874A/T and IFN-γ levels. IFN-γ expression was lower among patients with the AA genotype than those with the AT genotype (P<0.05); the genotypic and allelic frequency distributions of the IL-4 gene at -33C/T and -589C/T were significantly different between the asthmatic children and the control (P<0.05). The levels of IL-4 and IgE among children with TT genotype at the -33 and -589 loci were higher than those with the CT genotype, but only the polymorphism at -33C/T was associated with IL-4 levels (P<0.05). The polymorphisms of the IFN-γ gene at +874A/T or the CA repeats are not correlated with susceptibility to asthma. Thus, the polymorphism at +874A/T is correlated with IFN-γ level. The TT genotypes of the IL-4 gene at the -33 and -589 loci are associated with asthma susceptibility in children, and polymorphism at the -33 locus may be associated with IL-4 level.

Expression of IFN-γ and IL-4 in the esophageal mucosa of patients with reflux esophagitis and Barrett's esophagus and their relationship with endoscopic and histologic grading.

INTRODUCTION: This study investigated the expression of interferon-γ (IFN-γ) and interleukin-4 (IL-4) in the esophageal biopsies from patients with reflux esophagitis (RE) and Barrett's esophagus (BE) and their relationships with endoscopic grading and histologic grading. METHODS: Ninety individuals were recruited (30 RE, 30 BE, and 30 normal control) and underwent upper gastrointestinal endoscopy with esophageal biopsy. Immunohistochemistry was used to semi-quantitatively detect the expression of IFN-γ and IL-4 in the specimens. RESULTS: IFN-γ was overexpressed in BE and RE compared with the control (P < 0.01), whereas IL-4 was up-regulated in BE compared with RE and control (P < 0.01). A positive correlation between the level of IFN-γ and the endoscopic grading (r = 0.509, P < 0.01) and histologic grading of RE (r = 0.493, P < 0.01) was observed. CONCLUSIONS: Th1 and Th2 immune responses play different roles in the development of RE and BE. Th1 immune response is the major pathway for RE development, whereas both Th1 and Th2 immune responses are implicated in BE pathogenesis. The expression of IFN-γ is associated with the endoscopic and histologic grading of RE.

Knockdown of hTERT by SiRNA suppresses growth of Capan-2 human pancreatic cancer cell via the inhibition of expressions of Bcl-2 and COX-2.

OBJECTIVE: To investigate the effects of human telomerase reverse transcriptase (hTERT) on the growth of Capan-2 human pancreatic cancer cell and apoptosis. METHODS: Cell proliferation, apoptosis and cell cycle were analyzed by cell counting and flow cytometry. mRNA and protein expressions of hTERT, Bcl-2 and cyclooxygenase (COX)-2 were assessed by real time PCR and Western blot. RESULTS: Cell growth was significantly inhibited by 26.39 percent 24 h after hTERT-small interference RNA (siRNA) transfection (50 nmol/L) with a 100 percent silencing efficiency (P < 0.05). The inhibition rates of cell proliferation were 46.77 percent, 70.61 percent, 84.71 percent and 85.99 percent at 2, 3, 5 and 7 days after transfection, respectively (P < 0.001). Early and late apoptotic cells increased significantly (especially 24 h after transfection) (P < 0.001). The cell cycle was suppressed significantly as manifested by the increase of cells in the G0/G1 phase and the decrease of cells in the S phase and G2/M phase (P < 0.01). The expressions of Bcl-2 mRNA and COX-2 mRNA were inhibited significantly 48 h after transfection: the inhibition rates were 86.86 percent and 100 percent, respectively (P < 0.001). Levels of Bcl-2 protein were downregulated by 58.54 percent and 63.44 percent and the levels of COX-2 protein were downregulated by 50.06 percent and 82.77 percent at 48 h and 72 h after transfection, respectively. CONCLUSION: Knockdown of hTERT by siRNA can inhibit the growth of Capan-2 cell. The inhibitory effect is associated with the downregulation of Bcl-2 and COX-2.

[Toxicity of cationic liposome Lipofectamine 2000 in human pancreatic cancer Capan-2 cells].

OBJECTIVE: To investigate the toxicity of cationic liposome Lipofectamine 2000 (Lipo) in human pancreatic cancer Capan-2 cells. METHODS: Capan-2 cells were cultured in the presence of Lipo at toxic concentrations, and the cell growth, apoptosis and cell cycle changes were evaluated by cell counting and flow cytometry. RESULTS: The concentrations of both Lipo and siRNA affected the transfection efficiency. In a transfection volume of 2 ml, the presence of 5 microl Lipo resulted in slowed growth of Capan-2 cells, which was especially obvious after 3 days (P<0.001). Prolonged culture of the transfected cells caused significant increases in early apoptotic cells (P<0.05) and in the damaged or necrotic cells (P<0.001), and resulted in reduced viable cells (P<0.01); these changes became obvious after a 48-hour culture, which also increased the ratio of G(0)/G(1) phase cells (P<0.05) and decreased those of G(2)/M phase cells (P<0.01), S phase cells (P<0.01), and the late apoptotic cells (P<0.05). CONCLUSION: Toxic concentrations of Lipo can affect the growth, apoptosis and cell cycles of Capan-2 cells in vitro, and this urges careful concentration selection when using Lipo for gene transfer into different cells.

[A randomized and case-control clinical study on trimebutine maleate in treating functional dyspepsia coexisting with diarrhea-dominant irritable bowel syndrome].

OBJECTIVE: To observe the efficacy and adverse drug reaction of trimebutine maleate in treating patients with functional dyspepsia (FD) coexisting with diarrhea dominant irritable bowel syndrome (IBS-D). METHODS: 129 patients were enrolled in this randomized, case-control and prospective study and divided into 3 groups. Group A was treated with trimebutine maleate and bacillus licheniformis, Group B with trimebutine maleate and Group C with bacillus licheniformis. The symptoms of the patients were described with grading score and efficacy of treatment assessed according to the changes of grading score of symptoms. RESULTS: There was a significant decrease in the scores of postprandial fullness (4.55 +/- 0.85, 1.26 +/- 0.52; 4.36 +/- 0.66, 1.48 +/- 0.61), early satiation (4.05 +/- 0.96, 1.01 +/- 0.51; 3.89 +/- 0.81, 1.25 +/- 0.76), abdominal pain (9.26 +/- 0.68, 0.68 +/- 0.43; 9.57 +/- 1.60, 0.76 +/- 0.54) and total symptom score (20.00 +/- 1.25, 3.06 +/- 0.91; 19.05 +/- 2.28, 3.89 +/- 2.12) before and after treatment in Group A and B (P < 0.05), but there was no such significance in Group C (P > 0.05). There was a significant decrease in diarrhea score before and after treatment in the 3 groups (A: 4.78 +/- 0.76, 0.65 +/- 0.53; B: 4.13 +/- 0.65, 1.25 +/- 0.62; C: 4.65 +/- 0.88, 1.45 +/- 0.70) (P < 0.05). After treatment for 4 weeks, there was significant difference in the scores of postprandial fullness, early satiation, abdominal pain and total symptom score as well as the effective rate of every symptom and total effective rate between Group A or B and Group C (P < 0.05). The ratio of cost and effect was 4.07, 1.19 and 6.65 in Group A, B and C respectively, the Group B being the best. The rate of adverse drug reaction was 22.9% and 23.7% in Group A and B, and the main adverse drug reactions were mild thirst and constipation. CONCLUSIONS: In treating patients with functional dyspepsia coexisting with diarrhea dominant irritable bowel syndrome, trimebutine maleate has the advantage of high efficacy, low cost and few adverse reactions.

Effects of sulfasalazine on biopsy mucosal pathologies and histological grading of patients with active ulcerative colitis.

AIM: To investigate the mechanisms of sulfasalazine (SASP) in the treatment of ulcerative colitis (UC). METHODS: Changes of pathological signs and histological grading of 106 patients with active UC were observed before and after the treatment with SASP, 1 g, thrice daily for 6 wk. RESULTS: The effect of SASP on the vasculitis in lamina propria was 48.2% and 17.4% in the mild active UC (P<0.001) and 68% and 26.7% in the moderate active UC (P<0.001) before and after treatment. Fibroid necrosis of vessel wall was found in one case of mild UC and two cases of moderate UC before treatment and was not found after treatment. No thrombosis was found in mild UC before and after treatment, while thrombosis was found in one case of moderate UC before treatment. The effect on mucosal glandular abnormality was 30.4% and 13.0% in mild UC (P<0.05), and 42% and 40% in moderate UC (P>0.05) before and after treatment. The rate of eosinophil infiltration was 98.2% and 80.4% in mild UC (P<0.01), and 100% and 91.1% in moderate UC (P<0.05) before and after treatment. The effect on crypt abscess was 21.4% and 4.4% in mild UC (P<0.05), and 48% and 13.3% in moderate UC (P<0.001) before and after treatment. The effect on mucosal pathohistological grading was 2.00+/-0.84 and 0.91+/-0.46 in mild UC (P<0.001), and 2.49+/-0.84 and 1.31+/-0.75 in moderate UC (P<0.001) before and after treatment. CONCLUSION: SASP can improve small vessel lesions and crypt abscesses and reduce neutrophilic and eosinophilic leukocyte infiltration in inflammatory mucosa of UC.

Characteristics and therapeutic efficacy of sulfasalazine in patients with mildly and moderately active ulcerative colitis.

AIM: To investigate the characteristics and short-term efficacy of sulfasalazine (SASP) in patients with mildly and moderately active ulcerative colitis (UC). METHODS: Two hundred and twenty-eight patients with mildly and moderately active UC were recruited, 106 patients in 1993-1995, and 122 patients in 2000-2002, they were assigned as the 1990s group (n = 106) and the 2000s group (n = 122), prospectively. The general characteristics, clinical manifestations, colonoscopic and histological data were compared between the two groups. The short-term efficacy and safety of SASP 3 g per d were evaluated. RESULTS: Between 2000s and 1990s groups, the gender ratio of men to women was 1:1.18 and 1:1.04, 57.4% and 50.9% of the patients were between 30 and 49 years old. The gender ratio and age of UC patients were not significantly different. The total course of 50.0% and 37.1% of UC patients was less than 1 year (P<0.05), 10.6% and 31.2% of the cases had a duration of more than 5 years (P<0.05) in 2000s and 1990s groups, respectively. The most common clinical type was first episode in 2000s group and chronic relapse in 1990s group. The patients showed a higher frequency of abdominal pain and tenderness in 1990s group than in 2000s group. Erosions were found in 84.4% and 67.9% of patients in 2000s and 1990s groups (P<0.05). Rough and granular mucosa (67.9% vs 43.4%, P<0.05) and polyps (47.2% vs 32.8%, P<0.05) were identified in 1990s group more than in 2000s group. There were no significant differences in clinical, colonoscopic and histological classifications. After SASP (1 g thrice per d) treatment for 6 wk, the clinical, colonoscopic and histological remission rates were 71.8%, 21.8% and 16.4%, respectively. In 79 patients with clinical remission, 58.2% and 67.1% remained grade 1 in colonoscopic and histological findings, respectively. The overall effects in first episode type (complete remission in 10, 18.9%, partial remission in 28, 52.8%, and improvement in 9, 17.0%) were better than in chronic relapse type (complete remission in 3, 7.5%; partial remission in 16, 40.0%; and improvement in 15, 37.5%) and chronic persistent type (complete remission in 1, 5.9%; partial remission in 6, 35.3%; and improvement in 6, 35.3%) respectively (P<0.05). In 110 patients treated with SASP, 18 patients (16.4%) had adverse reactions. Except for two cases of urticaria and one case of WBC decrease, none of the patients had to stop the treatment because of severe adverse reactions. CONCLUSION: Patients with mildly and moderately active UC in 2000s group had a shorter disease course, milder clinical manifestations, more first episode type and higher frequency of acute mucosal lesions in colonoscopy than in 1990s group. The patients in 1990s group had higher proportion of chronic relapse type and chronic mucosal change in colonoscopy than in 2000s group. The short-term efficacy of SASP could be mainly remission of clinical manifestations. But more than half of the patients still had light inflammation in colonoscopy and histology. The overall effects of SASP in first episode type were better than those in other types. SASP was a safe and effective drug to treat mildly and moderately active UC.

Clinical analysis of primary small intestinal disease: A report of 309 cases.

AIM: To evaluate the major clinical symptom, etiology, and diagnostic method in patients with primary small intestinal disease in order to improve the diagnosis. METHODS: A total of 309 cases with primary small intestinal disease were reviewed, and the major clinical symptoms, etiology, and diagnostic methods were analyzed. RESULTS: The major clinical symptoms included abdominal pain (71%), abdominal mass (14%), vomiting (10%), melaena (10%), and fever (9%). The most common disease were malignant tumor (40%). diverticulum (32%) and benign tumor (10%). Duodenal disease was involved in 36% of the patients with primary small intestinal diseases. The diagnostic rate for primary small intestinal diseases by double-contrast enteroclysis was 85.6%. CONCLUSION: Abdominal pain is the most common clinical symptom in patients with primary small intestinal disease. Malignant tumors are the most common diseases. Duodenum was the most common part involved in small intestine. Double-contrast enteroclysis was still the simplest and the most available examination method in diagnosis of primary small intestinal disease. However, more practical diagnostic method should be explored to improve the diagnostic accuracy.

Clinical features and risk factors of patients with fatty liver in Guangzhou area.

AIM: There is still no accepted conclusion regarding the clinical features and related risk factors of patients with fatty liver. The large-scale clinical studies have not carried out yet in Guangzhou area. The aim of the present study was to investigate the clinical features and related risk factors of patients with fatty liver in Guangzhou area. METHODS: A total of 413 cases with fatty liver were enrolled in the study from January 1998 to May 2002. Retrospective case-control study was used to evaluate the clinical features and related risk factors of fatty liver with logistic regression. RESULTS: Obesity (OR: 21.204), alcohol abuse (OR: 18.601), type 2 diabetes mellitus (OR: 4.461), serum triglyceride (TG) (OR: 3.916), serum low-density lipoprotein cholesterol (LDL-C) (OR: 1.840) and fasting plasma glucose (FPG) (OR: 1.535) were positively correlated to the formation of the fatty liver. The levels of serum alanine aminotransferase (ALT) and gamma-glutamyltransferase (GGT) increased mildly in the patients with fatty liver and were often less than 2-fold of the normal limit. The higher abnormalities of aspartate aminotransferase (AST) levels (42.9%) with AST/ALT more than 2(17.9%) were found in patients with alcoholic fatty liver (AFL) than those with nonalcoholic fatty liver (NAFL) (16.9% and 5.0% respectively). The elevation of serum TG, cholesterol (CHOL), LDL-C was more common in patients with NAFL than with AFL. CONCLUSION: Obesity, alcohol abuse, type 2 diabetes mellitus and hyperlipidemia may be independent risk factors of fatty liver. The mildly abnormal hepatic functions can be found in patients with fatty liver. More obvious damages of liver function with AST/ALT usually more than 2 were noted in patients with AFL.