ABSTRACT
Objective:To explore the effect of sulforaphane (SFN) preconditioning on the cold myocardial ischemia-reperfusion injury (IRI) in the rats through PI3K/Akt signaling pathway.Methods:Sixty-four health male Sprague-Dawley (SD) rats were randomly divided into cold IRI group,SFN group,LY (LY294002) + cold IRI group,and LY+SFN group (n=16).The allogeneic heterotopic heart transplantation model was established by donor heart into recipient abdomen.The myocardium tissue was taken 24 h after reperfusion for the detection of histological changes using HE staining.The expression levels of Akt,p-Akt,Bax and Bcl-2 proteins were detected by immunohistochemistry and Western boltting methods.Results:The morphological results showed that the myocardium tissue damage was serious in cold IRI group and LY+cold IRI group,it was light in SFN group;the myocardium tissue damage of the rats in SFN+ LY group was ranged between cold IRI group and SFN group.Compared with IRI group,the expression levels of p-Akt protein and Bcl-2 protein in SFN group were increased (P＜0.05),and the expression level of Bax protein was decreased (P＜0.05).After treatment of blockage LY294002,compared with LY-+-cold IRI group,the expression level of p-Akt protein in LY-+-SFN group was not statistically significant (P＞0.05),the expression level of Bcl2 protein was increased (P＜0.05),the expression levels of Bax protein was decreased),and the ratio of Bcl-2/Bax was also increased (P＜0.05).Conclusion:SFN may attenuate cold IRI of heart transplantation through PI3K/Akt signaling pathway in the rats.
ABSTRACT
Objective:To explore the effect of sulforaphane (SFN) preconditioning on the cold myocardial ischemia-reperfusion injury (IRI) in the rats through PI3K/Akt signaling pathway.Methods:Sixty-four health male Sprague-Dawley (SD) rats were randomly divided into cold IRI group,SFN group,LY (LY294002) + cold IRI group,and LY+SFN group (n=16).The allogeneic heterotopic heart transplantation model was established by donor heart into recipient abdomen.The myocardium tissue was taken 24 h after reperfusion for the detection of histological changes using HE staining.The expression levels of Akt,p-Akt,Bax and Bcl-2 proteins were detected by immunohistochemistry and Western boltting methods.Results:The morphological results showed that the myocardium tissue damage was serious in cold IRI group and LY+cold IRI group,it was light in SFN group;the myocardium tissue damage of the rats in SFN+ LY group was ranged between cold IRI group and SFN group.Compared with IRI group,the expression levels of p-Akt protein and Bcl-2 protein in SFN group were increased (P＜0.05),and the expression level of Bax protein was decreased (P＜0.05).After treatment of blockage LY294002,compared with LY-+-cold IRI group,the expression level of p-Akt protein in LY-+-SFN group was not statistically significant (P＞0.05),the expression level of Bcl2 protein was increased (P＜0.05),the expression levels of Bax protein was decreased),and the ratio of Bcl-2/Bax was also increased (P＜0.05).Conclusion:SFN may attenuate cold IRI of heart transplantation through PI3K/Akt signaling pathway in the rats.
ABSTRACT
Objective To investigate the inflammation levels of 2-diabetes patients before and after 3 months of improving glycemic control.Methods A longitudinal study was performed in a subgroup of 48 subjects with T2D and poor glycemic control.Forty-four healthy individuals were taken as control group.The serum concentration of C-reactionprotein (CRP),interleukin-6 (IL-6),interleukin-6 (IL-8),transforming growth factor-β1 (TGF-β1) and transforming growth factor-β1 (MCP1) in all participants were measured simultaneously by multiplexed Luminex assay.Results The serum levels of CRP,MCP-1 of 2-diabetes patients were 3.96 (3.45,5.58) mg/L and (195.0± 129.8) ng/L,significant higher than those in control group (2.25 (1.24,3.22) mg/L,(148.5±85.7) ng/L),and the differences were significant(t=-2.580,P=0.010;t=-2.118,P =0.047).No significant difference was found in the serum levels of IL-6,IL-8,TGF-β lbetween the two groups (P＞0.05).TGF-β1 level in patients with good glycemic control decreased to 26.85 (23.17-31.12) ng/l,significant lower than that before glycemic control (43.5(26.5-62.25) g/L;Z=-2.191,P=0.028),and there were no significant differences among the other 4 kinds of inflammatory factors before and after blood glucose control(CRP:Z =-0.937P =0.372;IL-6:Z =-0.875,P =0.396;IL-8:Z =-1.215,P =0.286;MCP-1:t =-1.846,P=0.065).Conclusion Low grade systemic inflammation status in T2D patients.Improvement of glycemic control reduces TGF-β1 levels and plays a key role in delaying the development of diabetic nephropathy.
ABSTRACT
<p><b>BACKGROUND AND OBJECTIVE</b>The p53 as a transcription factor in cell stress was activated to regulate cell cycle and programmed cell death to inhibit tumor growth. Usually, p53 is kept in non-activated state through various mechanisms, including the action of p53 C-terminal negative regulatory sequences. The purpose of the study is to prepare the two types p53 recombinant adenoviruses that carry full-length p53 as well as deletion of negative regulatory sequences at p53 C-terminus and to detect exogenous GFP expression in human lung cancer cell infected-virus by FCM scatter plot.</p><p><b>METHODS</b>Using pAdEasy-Track vector system the p53 recombinant plasmids was constructed and the homologous recombinants in E. coli was produced. The three kinds of recombinant adenovirus in L293 cells was generated, sequencing proved. Exogenous GFP expression in human lung cancer 801D cells infected-virus was detected by FCM scatter plot.</p><p><b>RESULTS</b>p53 recombinant adenoviruses named Ad-p53(wtp), Ad-p53(del) and Ad-(empty carrier) were produced. Results of sequences indicate that the Ad-p53(del) was deletion of 111 bases before stop codon TGA and of 3 untranslated region at p53, the Ad-p53(wtp) no loss of any p53 base, the Ad-(empty carrier) no p53 sequence. FCM scatter plot indicate the percentage of 801D cells expressed GFP with three kinds of viral infection was almost same and was increased with the virus density. 801D contains ratio of cells with different fluorescence intensity.</p><p><b>CONCLUSION</b>The preparation of recombinant adenovirus, Ad-p53(del), pA-p53(wtp) and Ad-(empty carrier). The cells expressed-GFP can be quantitatively detected by FCM scatter plot. It was provide that the reliability of the virus system and accurate method for selecting viruses density to infecting cells.</p>
Subject(s)
Animals , Humans , Mice , Adenoviridae , Genetics , Flow Cytometry , Methods , Genes, p53 , Green Fluorescent Proteins , Genetics , Recombination, GeneticABSTRACT
Objective To explore the relation and measures prevention between aspirin and relapsing haemorrhage after operation in cerebral haemorrhage patients. Method It' s a prospective control study. A total of 725 patients with hypertensive basal ganglia cerebral haemorrhage admitted to department of neurosurgery from January 2001 to May 2007 were enrolled. They were diagnosed according to the diagnostic criteria set by the fourth national cerebrovascular disease conference in 1995. Haematoma volume was ＞ 50 mL. All patients were treated with craniotomy. And those with respiration and circulation failure, neurologic function deficit before the onset of the disease,major organ dysfunction, haemorrhagic disease and bleeding tendency or applied medicines affecting coagulation function excepted aspirin were excluded. The patients without use of aspirin before the onset of the disease were operated as the control group(group A), and there were 389 patients in group A.The patients with use of aspirin before the onset of the disease were randomly assigned to group B and C group,and there were 168 patients in group B or group C.The patients in group C received the frozen apheresis platelets. We counted different haematoma volume of relapsing haemorrhage after operation,death rate,ADL scores grades by 6 months follow-up survey in three groups. Quantitative data were expressed as mean ± standard deviation (-x ± s). The data were analyzed by using Chi-square test and Student's t test and rank sum test with SPSS 13.0 statistical package. A P value less than 0.05 indicated statisticals significance. Results Haematoma volume of relapsing haemorrhage was (40.59 + 20. 061 )mL, (53.21 ± 21.260) mL, (40.68 ± 19.517) mL in groups A, B, C,respectively. There was significant difference between group A and group B ( P ＜ 0.01 ), between group B and group C ( P ＜ 0.05), but there was no significant difference between group A and group C(P ＞ 0.05). ADL scores grades at 6-month follow-up was (67.04 ± 26. 176), (54.47 ± 29.403 ), (68.21 ± 25.254) in groups A, B, C, respectively. There was more significant difference between group A and group B, in ADL scores grades and the death rate between group B and group C (P ＜ 0.01), but there was no significant difference between group A and group C (P ＞ 0.05). Conclusions Aspirin can increase the occurrence rate of haemorrhage after operation, disablement and death in cerebral haemorrhage patients, but frozen apheresis platelets can reduce the occurrence rate.
ABSTRACT
<p><b>BACKGROUND</b>To isolate and clone the cisplatin genes in 801-D cell line, a kind of lung cancer cell line, with the emphasis of the objective genes regulated by wild type p53 (wtp53).</p><p><b>METHODS</b>Total RNA was extracted from transfected 801-D-wtp53, 801-D-vector cells which were both treated by cisplatin and 801-D-wtp53 cells. Using mRNA differential display, the DNA bands on gel were displayed by silver stain method. The DNA bands obtained from differential display were recovered and reamplified by PCR. The isolated genes were further proved by reverse Northern dot blot and were cloned to pGEMT easy vector.</p><p><b>RESULTS</b>Six positive genes were identified and cloned. Out of them, 2 related fragments were found to have an open reading frame. One was partly homologous to ribonucleoside-diphosphate reductase A, and the other was no homologous to the known genes.</p><p><b>CONCLUSIONS</b>There are obvious differences in gene expression in 801-D-wtp53 after induced by cisplatin than two other controls. It is possible for p53 to regulate the sensitization of lung cancer cells to cisplatin through its downstream target genes.</p>
ABSTRACT
<p><b>BACKGROUND</b>To study the effects of extraneous p53 antisense RNA on malignant growth and sensitivity to cisplatin of human lung cancer cell line.</p><p><b>METHODS</b>801D cell line with p53 deletion and mutation at 248 codon was selected as a parent cell line. An 1.8 kb human p53 full length cDNA was inserted into a mammalian expression vector PEGFP to construct a p53 antisense RNA recombined plasmid PEGFP-p53(AS) and GFP gene at plasmid was a report gene to monitor extraneous gene expression. The extraneous gene was detected by PCR. The p53 mutation protein was examined by immunohitochemical stain of p53 monoclonal antibody. The inhibition growth efficacy of extraneous p53 in vitro was determined by clonogenic survival assay. Sensitivity of cells to cisplatin was examined with MTT assay. FCM analysis was performed to measure the effect of p53 antisense RNA on cell cycle.</p><p><b>RESULTS</b>Two cell lines, PEGFP-p53(AS)-801D and PEGFP-801D, were established after transfection of 801-D cells by lipofection and selection. Presence of extraneous p53 gene in PEGFP-p53(AS)-801D was proved by PCR and expression of extraneous p53 was estimated when green fluorescence in those cells was found out under the fluorescent microscopy. Mutated p53 protein in parent cell line 801D was positive and in PEGFP-p53(AS)-801D was negative with immunochemical stain. The inhibition rate of colony formation was 61% for PEGFP-p53(AS)-801D (P < 0.001). The sensitivity of PEGFP-p53(AS)-801D cells to cisplatin was increased. FCM analysis showed that the cell line was arrested at G1 phase.</p><p><b>CONCLUSIONS</b>p53 mutation at 248 code plays an important role on malignant growth and resistance to cisplatin of human lung cancer cell line 801D. Malignant growth of cells with p53 deletion and mutation at 248 codon can be inhibited by extraneous p53 antisense RNA, and simultaneously the sensitivity to cisplatin is also increased.</p>
ABSTRACT
<p><b>BACKGROUND</b>To construct and express a phage display library of anti human lung cancer monoclonal antibody 5F-11.</p><p><b>METHODS</b>Immunoglobulin variable regions (VH,VL) were amplified from 5F-11 hybridrom by RT-PCR. ScFv genes consisting of VH DNA and VL DNA joined together by a linker DNA were cloned into a phage vector pCANTAB5E. After 4 rounds of screening with lung adenocarcinoma cell line A2 as antigen, an enriched secondary phage display library was obtained.</p><p><b>RESULTS</b>A recombinant phage display library with total of 8×10⁷ pfu/ml was established. Randomized clones from unselected library digested with BstNⅠ showed different patterns, however, those from selected library showed that phages with special pattern were enriched. Twenty-three out of 30 clones were found to respond strongly to A2 cell lines.</p><p><b>CONCLUSIONS</b>The ScFv of anti-lung adenocarcinoma monoclonal antibody 5F-11 can be successfully produced, which may be useful to widen the application of the antibody.</p>
ABSTRACT
<p><b>BACKGROUND</b>To study the inhibition effects of both extraneous right sense and antisense p53 on malignant phenotype of human lung cancer cell line.</p><p><b>METHODS</b>The named 801D cell line with p53 deletion and mutation at 248 code was selected as a model in vitro. The recombined plasmid pEGFP-p53(RS) and pEGFP-p53(AS) were constructed. The extraneous gene was detected by PCR. The p53 mutation protein was examined by immunohistochemical stain of p53 antibody. The inhibition effect of extraneous p53 on tumor growth in vitro were determined by clonogenic survival assay. FCM analysis was carried out in cells. The inhibition effect on malignant growth of extraneous p53 in vivo was observed by heteroplastic transplant on nude mouse.</p><p><b>RESULTS</b>The transfected cell lines, pEGFP-p53(AS)-801D, pEGFP-p53(RS)-801D and pEGFP-801D were established. Presence of extraneous p53 and neo genes in pEGFP-p53(AS)-801D and pEGFP-p53(RS)-801D was proved by PCR and green fluorescence was found out in those cells under the microscope. Mutant protein in pEGFP-p53(AS)-801D was negative by immunohistochemical stain. The malignant growth of these transfected cell lines was inhibited comparing with parents in vivo and in vitro. Inhibition rate of colony formation was 62.0% for pEGFP-p53(AS)-801D and 80.8% for pEGFP-p53(RS)-801D. The tumorigenicity in nude mice was suppressed. Inhibition effects of extraneous right sense p53 on malignant growth of 801D was more distinct. FCM analysis showed that pEGFP-p53(AS)-801D cells were arrested at G1 phase.</p><p><b>CONCLUSIONS</b>The transfected cell lines with extraneous right sense and antisense p53 appear that malignant growth can be inhibited in vivo and in vitro.</p>
ABSTRACT
Objective To improve the diagnosis and treatment of renal inflammatory pseudotumor (RIP). Methods 10 cases of RIP treated from 1970 to 1999 were reviewed and the diagnosis and treatment were discussed. Results The main clinical manifestation of RIP were fever,lumbago and hematuria.6 of 10 underwent nephrectomy because of the suspicion of kidney cancer whereas the other 4 were cured by antibiotics without recurrence on following up for 1～5 years. Conclusions RIP is rare,the diagnosis being based on clinical symptoms together with dynamic B ultrasound and CT scan.Needle biopsy is indicated to establish the diagnosis if necessary.Antibiotics is usually effective.
ABSTRACT
The aim of this study was to investigate the expression of basic fibroblast growth factor (b FGF) and its relationship with tumor microvasogenesis and biological behavior of human renal carcinoma. Gene expression of b FGF was assayed in 20 cases of renal carcinoma tissue using RNA dot blot hybridization. Meanwhile, the protein expression of b FGF and microvasular density (MVD) was assayed in 35 cases of renal carcinoma tissue with immunohistochemical assay. It was found that the expression of b FGF was significantly higher in renal cancer tissue than in normal renal tissue or normal tissue adjacent to the tumor, There was significant correlation between b FGF expression in cancer tissue and clinical stages as well as clinical grades.Similar results were also observed between MVD and b FGF expression in renal cancer tissue. It seemed that b FGF played an important role in angiogenesis in renal carcinoma. Taken together, the results suggest that MVD and the expression of b FGF in renal cancer tissue might be related to the prognosis of renal carcinoma patients.