High-resolution melting PCR analysis for genotyping the gene polymorphism of TNF-α, TGF-ß1, IL-10, and IFN-γ in lung transplant recipients.

BACKGROUND: High-resolution melting (HRM) analysis is a genotyping method which has the advantages of simple, rapid, low-cost and closed-tube operation. OBJECTIVES: This study evaluated HRM analysis as an option for detecting the single nucleotide polymorphism (SNP) of cytokine, and profiled the distribution of cytokine gene polymorphism in the lung transplant recipients (LTRs). MATERIAL AND METHODS: High-resolution melting-polymerase chain reaction (HRM-PCR) assays for genotyping tumor necrosis factor alpha (TNF-α) (-308 A/G), tumor growth factor beta 1 (TGF-ß1) (+869 T/C), interleukin 10 (IL-10) (-592 C/A, -819 T/C, -1082 G/A), and interferon gamma (IFN-γ) (+874 T/A) SNPs were developed on the LightCycler® 480. The SNPs of the aforementioned cytokine genes in 322 LTRs and 266 normal controls were detected using HRM-PCR approach. To confirm the accuracy of the HRM-PCR assay, we randomly selected 100 samples from the LTRs and detected the aforementioned SNPs with sequence-specific primer-polymerase chain reaction (SSP-PCR) method, using a commercial kit. RESULTS: The data show that the HRM-PCR assay can distinguish all the cytokine SNPs, and the results of HRM-PCR analysis are in complete concordance to the genotyping results obtained using a commercial kit (κ = 1.0). Our data also show that the allele and genotype frequencies of the abovementioned cytokine are not significantly different between the LTRs and the control groups (p > 0.05). In addition, we found the genotypes of TGF-ß1 +869 associated with high expression phenotype were prevalent in the LTRs. On the contrary, for TNF-α -308, IL-10 and IFN-γ, the genotypes associated with low expression phenotype were most common in the LTRs. CONCLUSIONS: In this study, we described a rapid, low-cost and high-throughput HRM-PCR technology for genotyping cytokine SNPs. Our data may be utilized for future studies examining the associations of cytokine gene polymorphisms with the prognosis of the LTRs.

Effects of Dexmedetomidine and ACE Genotype on Cardiovascular Response During the Decannulation Period of General Anesthesia in Patients With Essential Hypertension.

PURPOSE: This study investigated the effects of dexmedetomidine on cardiovascular response during the decannulation period of general anesthesia in patients with different genotypes of angiotensin-converting enzyme (ACE) and essential hypertension. METHODS: The present study enrolled patients with essential hypertension and American Society of Anesthesiologists class II or III who were scheduled to undergo abdominal surgery under general anesthesia. Patients were assigned to 1 of 6 groups according to ACE genotype, as detected by polymerase chain reaction-restriction fragment length polymorphism, as follows: DD; ID; II; and DD, ID, and II each with dexmedetomidine (Dex). Dexmedetomidine was intravenously infused at 0.5 µg/kg/h for 30 min before the end of surgery in groups DD (Dex), ID (Dex), and II(Dex). Anesthesia was induced and maintained by the same anesthetics in all patients. Systolic and diastolic blood pressure, heart rate (HR), ECG, and rate-pressure product were recorded before anesthesia induction; at 30 min before the end of surgery; at the end of surgery; and at 0, 1.5, 5, and 10 min after extubation. FINDINGS: A total of 210 patients were enrolled (n = 35 per genotype). After extubation, systolic and diastolic blood pressure, HR, and RPP were increased markedly from baseline in groups DD, ID, and II; the increases were greater in groups DD and ID than in group II. No significant changes in blood pressure, HR, or RPP were found, and proper sedative was achieved in groups DD (Dex), ID (Dex), and II(Dex). The prevalences of cardiac arrhythmia were higher in groups DD and ID than in groups II, DD (Dex), ID (Dex), and II(Dex). IMPLICATIONS: Patients essential hypertension and the ACE D allele had a strong hemodynamic response to tracheal extubation, on which dexmedetomidine was found to have both a prevention and treatment effect.

Association of leptin, visfatin, apelin, resistin and adiponectin with clear cell renal cell carcinoma.

Although an association between obesity and the occurrence of renal cell carcinoma (RCC) has been identified, the mechanism by which obesity functions to increase this risk of cancer remains unclear. Leptin, visfatin, apelin, resistin and adiponectin are peptide hormones secreted by adipocytes; it is considered that these may affect RCC development by exerting effects on proliferation, cell growth and inflammation. The aim of the present study was to investigate the association between the aforementioned adipokine genes and clear cell RCC (CC-RCC). The GSE6344 dataset was downloaded from the Gene Expression Omnibus database, and the relative expression levels of the adipokine genes were analyzed. To verify the results of the mRNA microarray, 77 paired samples of CC-RCC and corresponding adjacent normal tissue were allocated into two groups. The extraction of total RNA was conducted, and the mRNA expression of adipokine genes was analyzed using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The data from the GSE6344 dataset indicated that the expression of visfatin and apelin was upregulated (P<0.0001 and P<0.01, respectively), and adiponectin was downregulated (P<0.001) in the CC-RCC tissues compared with the adjacent normal tissues. The data from RT-qPCR demonstrated that visfatin and resistin gene expression was increased (P<0.01 and P<0.05, respectively) in the CC-RCC tissues. Furthermore, the mRNA expression level of leptin and adiponectin in the adjacent normal tissue was higher than those in the cancer tissue (P<0.01). The current study verifies that visfatin and adiponectin are associated with an increased risk of CC-RCC, which presents further insights into the molecular mechanisms of CC-RCC tumorigenesis.

High-resolution Melting PCR Analysis for Genotyping Lys109Arg and Gln223Arg in Patients with Renal Cell Carcinoma.

Although several studies have documented the role of leptin receptor gene polymorphisms in cancers, the association between leptin receptor gene polymorphisms and renal cell carcinoma (RCC) remains unknown. The aim of this study was to develop a high-resolution melting (HRM) approach for genotyping single nucleotide polymorphisms of leptin receptor gene on the LightCycler 480, and to explore the relation between polymorphisms of the leptin receptor gene and RCC. The study population consisted of 83 patients with renal cell carcinoma and 161 healthy control subjects. The Lys109Arg (A/G) and Gln223Arg (A/G) polymorphisms of leptin receptor gene were examined with HRM assay. Direct DNA sequencing and PCR-restriction fragment length polymorphisms were used as a reference method for genotyping Lys109Arg and Gln223Arg, respectively. Three genotypes of Lys109Arg or Gln223Arg were clearly distinguishable from the melting curve shapes with HRM assay. The data also showed the results of the direct DNA sequencing or PCR-restriction fragment length polymorphisms analysis were in complete concordance to genotyping results obtained by HRM (kappa=1.0). In addition, the data showed the G-G haplotype frequency was higher (p<0.05), and that the A-G (p<0.001) and G-A (p<0.01) haplotypes frequencies were lower in the RCC than controls. We developed a rapid, low cost, high-throughput and reliable single-tube technology for genotyping Lys109Arg and Gln223Arg polymorphisms. In addition, our data suggest that Lys109Arg and Gln223Arg gene polymorphisms are associated with RCC in Chinese Han studied population.

The effect of angiotensin-converting enzyme polymorphism on hemodynamic response to endotracheal intubation in hypertensive patients.

Endotracheal intubation elicits a hemodynamic response associated with increased heart rate and blood pressure that is influenced by the angiotensin-converting enzyme (ACE) insertion (I)/deletion (D) genetic polymorphism which may be of importance also for the pressure response to anesthesia. A total of 337 patients underwent abdominal surgery in general anesthesia and 16% were D/D-homozygotes, 45% were I/D heterozygotes and 39% of the patients were I/I homozygotes. Before surgery most patients were in treatment for arterial hypertension. Systolic and diastolic pressure, heart rate and concentrations of catecholamines in blood were determined before and after induction of anesthesia and for up to 10 minutes following endotracheal intubation. Anesthesia decreased blood pressure and for patients presenting ID and DD, blood pressure and heart rate reached similar levels but compared to II-homozygotes, D-carriers demonstrated significantly higher levels for systolic pressure and heart rate before and after intubation (p < 0.05). The blood levels of catercholamines were similar in the three genotype groups. The incidence of ECG-determined myocardial ischemia was higher in D-allele carriers compared to I-allele homozygotes (DD 22%, ID 25% vs. II 5%). In response to anesthesia and intubation and independent of sympathetic nervous activity, D-allele carriers for ACE polymorphism increased blood pressure response and higher risk for development of cardiovascular complications compared to patients homozygous for the I-allele.

Association of HLA and cytokine gene polymorphisms with idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a rare, progressive, and lethal interstitial lung disease with unknown etiology. Divergent observations have suggested that genetic factors contribute to IPF susceptibility. This study investigated the relationship between human leukocyte antigen (HLA), cytokine gene polymorphisms, and IPF in a Chinese Han population. The gene polymorphisms of HLA-A, -B, -DRB1, tumor necrosis factor alpha [TNF-α (-308 A/G)], transforming growth factor beta [TGF-ß1 (+869 T/C)], interleukin 10 [IL-10 (-592 C/A, -819 T/C, and -1082 G/A)], and interferon gamma [IFN-γ (+874 T/A)] were detected in 102 individuals with IPF and 266 unrelated normal controls using PCR with sequence-specific primers and a high-resolution melt (HRM) approach. The data showed that there was no difference in HLA allele frequencies between the IPF and control groups. However, the data showed the frequency of HLA-A*02-DRB1*04 haplotype in the IPF group was significantly higher than that in the control group [odds ratio (OR) = 4.69, 95% confidence interval (CI) = 1.82-12.08, p < 0.001]. In addition, no differences in the allele and genotype distributions of the cytokines were found between the IPF and control groups (p > 0.01). Our findings suggest that there is an association between specific HLA haplotype and IPF genetic susceptibility and that the genetic variability of some cytokines may not be involved in the pathogenesis of IPF.

Glucocorticoid Receptor ß Acts as a Co-activator of T-Cell Factor 4 and Enhances Glioma Cell Proliferation.

We previously reported that glucocorticoid receptor ß (GRß) regulates injury-mediated astrocyte activation and contributes to glioma pathogenesis via modulation of ß-catenin/T-cell factor/lymphoid enhancer factor (TCF/LEF) transcriptional activity. The aim of this study was to characterize the mechanism behind cross-talk between GRß and ß-catenin/TCF in the progression of glioma. Here, we reported that GRß knockdown reduced U118 and Shg44 glioma cell proliferation in vitro and in vivo. Mechanistically, we found that GRß knockdown decreased TCF/LEF transcriptional activity without affecting ß-catenin/TCF complex. Both GRα and GRß directly interact with TCF-4, while only GRß is required for sustaining TCF/LEF activity under hormone-free condition. GRß bound to the N-terminus domain of TCF-4 its influence on Wnt signaling required both ligand- and DNA-binding domains (LBD and DBD, respectively). GRß and TCF-4 interaction is enough to maintain the TCF/LEF activity at a high level in the absence of ß-catenin stabilization. Taken together, these results suggest a novel cross-talk between GRß and TCF-4 which regulates Wnt signaling and the proliferation in gliomas.

Association of TNF-α, TGF-ß1, IL-10, IL-6, and IFN-γ gene polymorphism with acute rejection and infection in lung transplant recipients.

Infection and rejection are common complications faced by lung transplant recipients (LTRs) and have become major impediments to long-term survival. Cytokines may play an important role in the development of these complications. In this study, we explored the correlation between TNF-α (-308 A/G), TGF-ß1 (+869 T/C, +915 G/C), IL-10 (-592 C/A, -819 T/C, -1082 G/A), IL-6 (-174 G/C), and IFN-γ (+874 T/A) gene polymorphisms and the incidence of acute rejection and infection. Transplant outcomes were reviewed in a retrospective cohort of 113 LTRs from a single center between December 2004 and November 2012. Cytokine polymorphisms were measured using sequence-specific primer-based PCR. HLA typing was performed for the donors and recipients. We found that the LTRs with the IL-10 -819 CC and -592 CC genotypes had a significantly decreased risk of infection (p = 0.017, OR = 0.177, 95% CI = 0.04-0.85). However, we found no significant association between cytokine polymorphisms and acute rejection. Furthermore, the data revealed that the occurrence of acute rejection was strongly associated with infection episodes (χ(2)  = 8.5256, p < 0.01). These results suggest that LTRs possessing the IL-10 -819 CC and -592 CC genotype may be protected from the occurrence of infection. Our results demonstrated that infection is an important cause of acute rejection for LTRs.

Association of leptin receptor Lys109Arg and Gln223Arg polymorphisms with increased risk of clear cell renal cell carcinoma.

BACKGROUND: Although roles of genetic polymorphisms of leptin receptor (LEPR) gene in several cancers have been documented, the association between polymorphisms of LEPR and clear cell renal cell carcinoma (CC-RCC) remains unknown. The aim of this study was to explore any relation. MATERIALS AND METHODS: The study population consisted of 77 patients with CC-RCC and 161 healthy control subjects. Polymorphism analyses of Lys109Arg and Gln223Arg were performed by direct DNA sequencing and PCR-restriction fragment length polymorphism approaches respectively. RESULTS: Comparisons of allelic and genotypic frequencies in Lys109Arg and Gln223Arg showed no significant difference between the cases and controls. However, when evaluating the combined genotype of Lys109Arg and Gln223Arg, risk with GG/GG was increased (OR=1.85, 95%CI=1.04-3.30) and with GA/GG or GG/GA was decreased (OR=0.07, 95%CI=0.01-0.54; OR and 95%CI of the latter could not be calculated for a value of zero) . Furthermore, the G-G haplotype frequency of Lys109Arg and Gln223Arg in the cases was higher (OR=1.68; 95%CI=1.02-2.76). In contrast, the A-G and G-A haplotype frequencies in the cases were lower than those in the controls (OR=0.06; 95%CI=0.01 to 0.47; OR and 95%CI of the latter could not be calculated for a value of zero). In addition, the Lys109Arg A allele was in LD with the Gln223Arg A allele (d'=0.9399) in the CC-RCC subjects, but not in the controls. CONCLUSIONS: Our data suggest that the GG/GG combined genotype and G-G haplotype of Lys109Arg and Gln223Arg can act as evaluating factors for CC-RCC risk.

[Association between suppressors of cytokine signaling mRNA expression and Th1/Th2 balance in children with asthma].

OBJECTIVE: Suppressors of cytokine signaling (SOCS) have been shown to play an important role in regulating cytokines, such as intracellular interferon (IFN) and interleukin (IL), in the Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway. At present, the association between SOCS and asthma is still under study. The aim of this study is to explore the relationship of SOCS-1 and SOCS-3 mRNA expression in peripheral blood mononuclear cells (PBMCs) with the intracellular IFN-'/IL-4 ratio in CD4+ T cells and specific IgE (sIgE) level in children with asthma. METHODS: BMCs were collected from 44 children with allergic asthma (4-14 years) and 30 healthy children. The intracellular IFN-'/IL-4 ratio in CD4+ T cells was measured by flow cytometry. Total RNAs were extracted from the PBMCs and SOCS-1 and SOCS-3 mRNA expression was measured by SYBR Green I quantitative RT-PCR. RESULTS: Compared with the healthy children, children with allergic asthma showed a lower level of intracellular IFN-' in peripheral blood [(15.7±2.0)% vs (19.1±2.7)%] and IFN-'/IL-4 ratio (3.4±1.5 vs 4.8±2.9) and higher SOCS-1 mRNA expression (-Ct, 11.1±1.9 vs 12.6±2.8). There was a negative relationship between SOCS-1 mRNA expression and the percentage of IFN-'-producing CD4+ T cells in peripheral blood in both asthmatic and healthy children (P<0.05). No correlation was found between SOCS-1 and SOCS-3 expression and sIgE level. CONCLUSIONS: Children with allergic asthma have elevated levels of SOCS-1 mRNA in PBMCs, which is associated with Th2-skewed immune response.

[Effect of glucosylceramide synthase on P-gp expression by ERK signal transduction pathway in leukemia multi-drug resistance cell line].

OBJECTIVE: To investigate the effect of glucosylceramide synthase (GCS) on P-glycoprotein (P-gp) expression via extracellular signal-regulated kinase (ERK) pathways in leukemia K562/A02 cell line. METHODS: K562/A02 multidrug resistance cells were treated with GCS siRNA and U0126, respectively. Expression of multidrug resistance protein 1 (MDR1) mRNA was analyzed with qRT-PCR. Phosphorylated ERK1/2, total ERK1/2 protein and P-gp in different groups were measured with Western blotting. RESULTS: After treated with U0126, P-ERK1/2 was decreased along with the increased U0126 concentration. P-ERK1/2 and P-gp were apparently down-regulated by U0126 at the concentrations of 20 µmol/L, 40 µmol/L and 60 µmol/L. After being transfected with GCS siRNA, GCS mRNA was inhibited by 70.50% (58.00%-76.00%) in K562/A02 cells. Compared with the negative control, both P-ERK1/2 and P-gp were inhibited significantly after RNAi for 72 hours (P<0.01 and P<0.05, respectively. CONCLUSION: GCS may affect the expression of P-gp by ERK signal transduction pathway in leukemia cells.

Serum angiotensin I-converting enzyme levels and the therapeutic effects of octreotide in esophageal variceal hemorrhage.

INTRODUCTION: The aim of this study is to investigate the relationship between serum angiotensin I-converting enzyme (ACE) levels and therapeutic effects of octreotide in the treatment of esophageal variceal hemorrhage (EVH) as a result of liver cirrhosis. METHODS: Serum ACE levels were measured by ultraviolet light colorimetric analysis in 80 cases of liver cirrhosis with EVH before and after the treatment of various doses of octreotide (25 and 50 µg/hr treatment in 40 cases, respectively), which were compared with 20 healthy controls. RESULTS: Between the octreotide treatment groups, there were no significant differences in the Child-Pugh score, the endoscopic severity of esophageal varices and ACE levels before octreotide treatment. Pretreatment levels of serum ACE were markedly higher in patients with EVH compared with healthy controls (P < 0.001). Serum ACE levels were significantly higher before octreotide treatment than 72 hours after treatment in patients with EVH. Serum ACE after octreotide treatment declined more evidently in the 50 µg/hr than in the 25 µg/hr treatment group. The hemostatic rate within 6 hours after octreotide treatment was significantly higher in the 50 µg/hr than in the 25 µg/hr treatment group. The rebleeding rate within 72 hours after octreotide treatment was markedly lower in the 50 µg/hr than in the 25 µg/hr treatment group. CONCLUSIONS: Octreotide treatment in patients with EVH can result in decreased serum ACE levels, which correlated with the dose of octreotide. The decline in serum ACE levels may be involved in the mechanisms by which octreotide lowers portal vein pressure in EVH treatment.

The effect of glucosylceramide synthase on P-glycoprotein function in K562/AO2 leukemia drug-resistance cell line.

Previous work from our laboratory demonstrated that glucosylceramide synthase (GCS) and multidrug resistance 1 gene (MDR1) are co-overexpressed in drug-resistant leukemia cells. We hypothesized that GCS and MDR1 may interact. In this study, we used RNA interference (RNAi) to silence the GCS or MDR1 gene in K562/AO2 drug-resistant cells. The sensitivity of cells to different treatments with doxorubicin was evaluated. We used Taqman probe fluorescence real-time quantitative PCR, and detected expression of GCS and MDR1 mRNAs in different interfering groups. Intracellular mean fluorescence intensity (MFI), which represents rhodamine123 (rh123) retention, was determined by flow cytometry (FCM). An MTT cytotoxicity assay showed that the 50% inhibition concentration (IC50) of doxorubicin of K562/AO2 cells (138.25 ± 3.75 µg/ml) was significantly higher than that of K562 drug-sensitive cells (2.125 ± 0.125 µg/ml), and that IC50 was evidently lower in K562/AO2 cells, whether it was transfected with a small interfering RNA (siRNA) targeting GCS (GCSsiRNA) or one targeting MDR1 (MDR1siRNA). Compared with untreated K562/AO2 cells, the inhibition rates of GCS mRNA in the cells transfected with GCSsiRNA for 9 and 36 h were 56.67 ± 9.29% (p < 0.05) and 74 ± 6.38% (p < 0.05), respectively. Interestingly, the expression of MDR1 mRNA was also inhibited to 51.7 ± 4.5% (p < 0.05) 36 h after transfection with GCSsiRNA, but there was no significant difference in MDR1 expression at 9 h post-transfection in cells treated with GCSsiRNA and a negative control. It is well known that rh123 retention in cells results from an efflux function of P-glycoprotein (P-gp). In K562 cells, rh123 retention was much higher than in K562/AO2 cells (p < 0.01). We also noted that rh123 retention in the K562/AO2 cells transfected with GCSsiRNA for 48 h was significantly higher than in the negative control group. In conclusion, we show in the present study that inhibition of the GCS gene affects the expression of MDR1 mRNA and P-gp function.

Down-regulation of glutamine synthetase enhances migration of rat astrocytes after in vitro injury.

Astrocytes undergo reactive transformation in response to physical injury (reactive gliosis) that may impede neural repair. Glutamine synthetase (GS) is highly expressed by astrocytes, and serves a neuroprotective function by converting cytotoxic glutamate and ammonia into glutamine. Glutamine synthetase was down-regulated in reactive astrocytes at the site of mechanical spinal cord injury (SCI) and in cultured astrocytes at the margins of a scratch wound, suggesting that GS may modulate reactive transformation and glial scar development. We evaluated this potential function of GS using siRNA-mediated GS knock-down. Suppression of astrocytic GS by GS siRNA increased cell migration into the scratch wound zone and decreased substrate adhesion as indicated by the number of focal adhesions expressing the adaptor protein paxillin. Migration was enhanced by glutamine and suppressed by glutamate, in contrast to the result expected if enhanced migration was due solely to changes in glutamine and glutamate concomitant with reduced GS activity. The membrane type 1-matrix metalloproteinase (MT1-MMP) was up-regulated in GS siRNA-treated astrocytes, while a broad-spectrum MMP antagonist inhibited migration in both wild type and GS knock-down astrocytes. In addition, GS siRNA inhibited expression of integrin ß1, while antibody-mediated inhibition of integrin ß1 impaired direction-specific protrusion and motility. Thus, GS may modulate motility and substrate adhesion through transmembrane integrin ß1 signaling to the cytoskeleton and by MMT-mediated proteolysis of the extracellular matrix.

[Study on the correlation of GCS and MDR1 genes in inducing multidrug resistance in human K562/A02 cell line].

OBJECTIVE: To investigate the correlation of glucosylceramide synthase (GCS) gene and multidrug resistance 1 (MDR1) gene in inducing multidrug resistance in human multidrug-resistant K562/A02 cell line, and search for a novel strategy for reversing multidrug resistance of leukemia cells. METHODS: The expression levels of GCS and MDR1 mRNA in K562 and K562/A02 cells were assayed by RT-PCR. siRNAs targeting the GCS and MDR1 gene were transfected into K562/A02 cells with liposome, respectively. The differential expression of GCS and MDR1 mRNAs, as well as their correlation, were detected by RT-PCR and real time quantitative-PCR(QPCR). RESULTS: The expression level of GCS and MDR1 mRNA was dramatically lower in drug-sensitive K562 cells compared with the K562/A02 cells. The GCS mRNA was inhibited by 73%(59%-82%) and MDR1 mRNA expression was down regulated by 67% (38%-82%) in K562/A02 cells after being transfected with GCS siRNA. The expression level of MDR1 mRNA was inhibited by 81%(63%-91%) and GCS mRNA expression had no apparent change in K562/A02 cells treated with MDR1 small interference RNA(siRNA). CONCLUSION: Positive correlation was detected between the expression of GCS and MDR1 mRNA in K562/A02 cells and MDR1 mRNA expression was down regulated after silencing the GCS gene expression.

GCS induces multidrug resistance by regulating apoptosis-related genes in K562/AO2 cell line.

We have previously shown that the expression of glucosylceramide synthase (GCS) gene in drug-resistant K562/AO2 human leukemia cell was higher than that in drug-sensitive K562 cell, and the sensitivity to adriamycin of K562/AO2 cell was enhanced by inhibiting GCS. It is concluded that the overexpression of GCS gene is one of the reasons which lead to multidrug resistance (MDR) of leukemia cell. Meanwhile, we also found that higher expression of Bcl-2 gene and protein were exhibited in K562/AO2 cell compared with K562 cell. Basing on this, we hypothesized that the high expression of GCS gene which results in MDR of leukemia cell is correlated with Bcl-2 signal transduction. In order to validate the hypothesis, the inhibition of GCS gene in K562/AO2 cell was observed by using chemical suppressor PPMP and siRNA targeted at GCS, and applying RT-PCR and flow cytometry, the expression levels of apoptosis-related gene Bcl-2 and Bax were analyzed before and after inhibiting GCS gene in K562/AO2 cell. The results demonstrated that the gene and protein of Bcl-2 in K562/AO2 cell were both down-regulated significantly after GCS gene being inhibited; however, the Bax mRNA expression had no apparent change in different groups. This suggested that GCS gene may contributed to MDR of human leukemia cell K562/AO2 by Bcl-2 signal transduction.

The expression and clinical significance of TGF-beta1 and MMP2 in human renal clear cell carcinoma.

The expression and clinical significance of transforming growth factor beta1 (TGF-beta1) and matrix metalloproteinase-2 (MMP2) in human renal clear cell carcinoma (RCCC) were investigated. The intensity of TGF-beta1 and MMP2 expression in RCCC kidneys was significantly higher than that in normal kidneys. Expression of TGF-beta1 and MMP2 in RCCC tissues was positively correlated with pathological grade and clinical stage. There was also a significant correlation between TGF-beta1 and Msshese analyses indicate that upregulation of TGF-beta1 and MMP2 expression may occur during the progression of RCCC. Thus, TGF-beta1 and MMP2 may be useful molecular markers for evaluating prognosis in RCCC patients.

[Expression of the CDH13 gene and BCR/ABL fusion gene in chronic myeloid leukemia patients and their relationship].

OBJECTIVE: To investigate the expression of the CDH13 gene and BCR/ABL fusion gene in chronic myeloid leukemia(CML) patients at different stages and explore their relationship. METHODS: RNA was isolated from peripheral blood in 30 healthy adults, 22 primary CML patients and 25 blastic crisis of CML patients. We examined the expression of the CDH13 gene and BCR/ABL fusion gene using EVA Green real-time reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: The expression of the BCR/ABL fusion gene in the patients with blastic crisis CML was 4.72 folds as that of the patients with primary CML. The expression of the CDH13 gene in the primary and blastic crisis CML patients was significantly reduced to 36% and 25% of that in healthy controls, respectively. Moreover, negative correlation was found between the expressions of these two genes. CONCLUSION: The study showed that the reduction of the CDH13 expression at different clinical stage of CML may account for the defective cell adhesion in CML, and the expression of the CDH13 gene was probably down-regulated by the BCR/ABL fusion gene.