Activation of caspase-12 at early stage contributes to cardiomyocyte apoptosis in trauma-induced secondary cardiac injury / 生理学报

Trauma-induced secondary cardiac injury (TISCI) is associated with increased adverse cardiac events and death. We have previously reported that TISCI results in myocardial apoptosis and secondary cardiac dysfunction. However, the underlying mechanism is unclear. To identify the time course of trauma-induced cardiomyocyte apoptosis and possible apoptotic pathway, traumatic rat models were built with Noble-Collip drum. Meanwhile, normal rat cardiomyocytes were cultured with traumatic plasma (TP) for 48 h. Cardiomyocyte apoptosis, cardiac function and the apoptosis related enzymes, including caspase-3, -8, -9, and -12, were determined. The results showed that there was no direct injury of rat hearts immediately after trauma. However, compared with hearts from the sham rats, hearts isolated from traumatic rats exhibited reduced +dP/dTand -dP/dT24 h after trauma. In traumatic rats, myocardial apoptotic index and caspase-3 activity obviously increased 6 h after trauma, and achieved the maximal value 12 h after trauma. The activity and expression of caspase-12, an endoplasmic reticulum (ER) stress-specific caspase, elevated markedly 3 h after trauma and reached its peak 6 h after trauma. Otherwise, caspase-8 (extrinsic apoptotic pathway) and caspase-9 (intrinsic apoptotic pathway) in the myocardial tissue of traumatic rats were activated 24 h after trauma. Meanwhile, incubation of normal rat cardiomyocytes with TP increased caspase-12 activity at 6 h, caspase-3 activity at 12 h, caspase-8 and -9 activities at 24 h, respectively. TP-induced cardiomyocyte apoptosis was virtually abolished by Z-ATAD-FMK (a caspase-12 specific inhibitor). In addition, there was a significant negative correlation between myocardial caspase-12 activity and trauma-induced secondary cardiac dysfunction. Our present study demonstrated that caspase-12 is firstly activated and plays an important role in TISCI rats. Inhibition of caspase-12 mediated apoptosis may be a novel strategy in ameliorating posttraumatic cardiomyocyte apoptosis and secondary cardiac injury.

Effect of amygdala H2S system on depression-like behavior in posttraumatic stress disorder rats / 中国病理生理杂志

AIM:To explore the depression-like behavior and cystathionine β-synthase (CBS)/hydrogen sulfide (H2S) levels of amygdala in posttraumatic stress disorder (PTSD) rats and to study the effect of exogenous H2S on PTSD rats.METHODS:Single prolonged stress paradigm was adopted to prepare PTSD animal model.Forced swimming test and sucrose preference test were used to evaluate the depression-like behavior.The content of CBS/H2S in amygdala tissue was measured by Western blot and methylene blue method.In vivo extracellular single unit recordings was used to examine the frequency of spontaneous discharges of amygdala neurons.RESULTS:The immobility time in forced swimming test of PTSD group increased and sucrose preference in sucrose preference test of PTSD group decreased compare with normal group (P<0.01).CBS/H2S level in amygdala tissue of PTSD group decreased compared with normal group (P<0.01).The immobility time of the rats in forced swimming test of NaHS+PTSD group decreased and the sucrose prefe-rence in sucrose preference test of NaHS+PTSD group increased compare with PTSD group (P<0.01).L-cysteine increased the frequency of spontaneous discharges of amygdala neurons (P<0.01).CONCLUSION:Depression-like behavior is aggravated in PTSD model rats owing to the inhibition of CBS/H2S content in amygdala tissue.The mechanism of behavior-improving effect of H2S on PTSD model rats is possibly related to excitating amygdala neurons and increasing the frequency of spontaneous discharges.

Construction and verification of SIRT1 and its mutant T200I, E420K eukaryotic expression vector / 国际生物医学工程杂志

Objective To clone silent information regulator 1 (SIRT1) gene full-length cDNA,construct recombinant eukaryotic expression vector containing SIRT1 gene and its mutant T200I,E420K,so as to lay the foundation for further research of SIRT1 gene function.Methods RT-PCR amplified SIRT1 gene full-length cDNA.PCR products were cloned into the eukaryotic expression vector pcDNA3.1 (+) through double digestion and pcDNA3.1(+)-SIRT1 recombinant plasmid was obtained.Meanwhile,site-directed mutagenesis was applied to build its mutant pcDNA3.1 (+)-T200I and pcDNA3.1 (+)-E420K expression vector.Recombinant plasmid was identified by enzyme digestion and DNA sequencing and the recombinant eukaryotic expression vector of success was screened out.Results SIRT1 gene full-length cDNA was successfully cloned,and pcDNA3.1 (+)-SIRT1 eukaryotic expression vector and its mutant were also successfully constructed.Positive recombinant plasmid sequencing was compared after enzyme digestion,and it was completely consistent with the expected sequence.Transfected 293T cell line was established and HIS tagged SIRT1 protein was expressed.Conclusions We successfully constructed pcDNA3.1 (+)-SIRT1 and its mutant pcDNA3.1 (+)-T200I,pcDNA3.1 (+)-E420K eukaryotic expression vector,which may provide genetic material for biological function study of SIRT1 gene and its mutant T200I,E420K.

Growth Differentiation Factor 5 (GDF5) Core Promoter Polymorphism Is Not Associated with Susceptibility to Osteoarthritis of the Knee in the Korean Population

BACKGROUND: Osteoarthritis (OA) is a common disease characterized by degenerating joint cartilage in the knee, hip, and hand. A functional single nucleotide polymorphism (SNP) +104T/C; rs143383 in the 5' untranslated region of the growth differentiation factor 5 (GDF5) gene was recently associated with susceptibility to OA in the Japanese and Chinese populations. METHODS: To investigate whether this association is present in the Korean population, the frequency of the polymorphism was investigated in 276 patients with knee OA and 298 healthy subjects as controls. Polymorphism analysis was performed by amplifying the core promoter region of the GDF5 gene and digesting it with the BsiEI restriction enzyme. RESULTS: The frequency of the TT, CT, and CC genotypes was 54.3% (150/276), 41.7% (115/276), and 4.0% (11/276), respectively, in patients with OA, and 53.4% (159/298), 37.9% (113/298), and 8.7% (26/298), respectively, in healthy controls. No significant differences in genotypic or allelic frequencies of the +104T/C SNP of the GDF5 gene were observed between patients with OA and controls. Also, no significant differences in allelic and genotypic frequencies were found when the individuals were stratified by age and gender. CONCLUSIONS: The results suggest that the +104T/C; rs143383 GDF5 core promoter polymorphism is not a risk factor for OA in the Korean population.

Genetic and Expression Analysis of the SIRT1 Gene in Gastric Cancers

PURPOSE: Silent mating-type information regulation 2 homologue 1 (SIRT1) is a nicotinamide adenine dinucleotide-dependent deacetylase. SIRT1 plays an important role in the regulation of cell death/survival and stress response in mammals. The aim of this study was to investigate whether the SIRT1 gene is involved in the development or progression of gastric cancers. MATERIALS AND METHODS: SIRT1 and p53 genes in 86 gastric cancers were examined for genetic alterations by PCR-single strand conformation polymorphism sequencing, as well as SIRT1 protein expression in 170 gastric cancers by immunohistochemistry. RESULTS: In the genetic analysis, we found SIRT1 and p53 mutations in two and 12 cases, respectively. Two missense mutations, c.599 C>T (T200I) and c.1258 G>A (E420K), were detected in the SIRT1 gene coding region. The SIRT1 and p53 mutation were found in mutually exclusive gastric cancers. The immunohistochemistry revealed that SIRT1 overexpression was found in 95 (55.9%) of 170 gastric cancers. Altered SIRT1 expression was not statistically associated with clinicopathological parameters, including tumor differentiation, location, lymph node metastasis, or p53 expression. Two cases with an SIRT1 mutation showed increased SIRT1 expression. CONCLUSIONS: These results suggest that genetic alterations and overexpression of the SIRT1 gene may contribute to gastric cancer development.

An Association of C/T Polymorphism in the TFF1 Gene and the Susceptibility to Gastric Cancer

PURPOSE: This study investigated whether a single nucleotide polymorphism (SNP) located at position -2 in the Kozak sequence of the TFF1 gene is associated with H. pylori infection and the development of gastric cancer in Koreans. MATERIALS AND METHODS: We enrolled 167 patients with gastric cancer from January 2000 to December 2003 and also 299 healthy controls during the same period. The genotype of the TFF1 SNP was analyzed by polymerase chain reaction-restriction fragment length polymorphism and single strand conformation polymorphism. We also examined the H. pylori infection by Giemsa staining. RESULTS: No significant difference in the allele or the TFF1 SNP genotype frequency was observed between the patients with gastric cancer and the control subjects (P=0.595 and P=0.715, respectively). When stratified by the histological subtype of gastric cancer and the age of the patients, the risk was not statistically significant between the two study groups (P=0.088 and P=0.551, respectively). H. pylori infection was detected in 39 cases and it was not associated with the TFF1 genotype. CONCLUSION: These findings suggest that this TFF1 gene polymorphism is not associated with H. pylori infection and gastric cancer in Koreans and so it doesn't contribute to the susceptibility to gastric cancer in Koreans.

The clinical study on conservative treatment for the patients with tube pregnancy under laparoscopy / 中国微创外科杂志

Ojective To investigate therapeutic results of conservative operation with laparoscopy for all kinds of patients with tube pregnancy. Methods 62 patients with tube pregnancy were performed the conservative surgery, fenestration on the position of tube pregnancy or take out embryos from the tube crevice combined with injection of MTX under laparoscopy. Results all of 61 patients' tube were retained successfully. no complications occurred, urine HCG recovered to normal levels two weeks after operation. Only one patient continue to pregnancy and completely recovered after intramuscular injection MTX. 18 patients were carried out HSG 1～4 months after operations. 15 patients' tube among them were unobstructed. Opening rate was 88.3%. Within postopeative four years, 12 patients were repregnant. Pregnancy rate was 66.7%. Conclusions Tube pregnancy conservative treatments under laparoscopy not only is safe and minimal invasion, but also had less complications, therefore it is a more suitable method for retaining the fertile females' tubes.