Antigenotoxicity and Cytotoxic Potentials of Cell-Free Supernatants Derived from Saccharomyces cerevisiae var. boulardii on HT-29 Human Colon Cancer Cell Lines.

Microbial-derived postbiotics are of interest recently due to their lower side effects than chemotherapy for cancer treatment and prevention. This study aimed to investigate the potential antigenotoxic and cytotoxic effects of cell-free-supernatant (CFS) postbiotics derived from Saccharomyces boulardii by applying SOS chromotest and MTT assay on HT-29 cell lines. Also, further cellular pathway-related assays such as cell cycle, DAPI, and annexin V-FITC/PI staining were performed. Real-time PCR was utilized to assess the expression levels of some genes involved in apoptosis. Based on the outcomes, the CFSs of S. boulardii showed significant antigenotoxic effects (20-60%, P < 0.05), decreased cell viability (with the significant IC50 values of 33.82, 22.68, and 27.67 µg/mL after 24, 48, and 72 h respectively), suppressed the initial (G0/G1) phase of the cell's division, influenced the nucleus of the treated cells, induced apoptosis, and increased the expression of Caspas3 and PTEN genes after 48 h, while the RelA and Bcl-XL genes indicated diminished expression in treated HT-29 cells. Consequently, CFS postbiotics of S. boulardii exhibited significant antigenotoxic and cytotoxic effects and induced apoptosis responses in HT-29 cancer cells. The results of this investigation lead us to recommend that the CFS postbiotics generated from Saccharomyces cerevisiae var. boulardii be taken into consideration as a potential anticancer agent or in the design of supplementary medications to treat and prevent colon cancers.

An Apoptosis-Related Gene Prognostic Index for Colon Cancer.

Purpose: To construct an apoptosis-related gene prognostic index (ARGPI) for colon cancer, and clarify the molecular and immune characteristics of the risk subgroup as defined by the prognostic index and the benefits of adjuvant chemotherapy. Integrating the prognostic index and clinicopathological risk factors to better evaluate the prognosis of patients with colon cancer. Methods: Based on the colon adenocarcinoma data in the TCGA database, 20 apoptosis-related hub genes were screened by weighted gene co-expression network analysis (WGCNA). Five genes constituting the prognosis model were determined by Cox regression and verified by the Gene Expression Omnibus (GEO) dataset. Then the molecular and immune characteristics of risk subgroups defined by the prognostic index and the benefits of adjuvant chemotherapy were analyzed. Finally, nomograms integrating ARGPI and four clinicopathological risk factors were used to evaluate the prognosis of patients with colon cancer. Results: The ARGPI was constructed based on the FAS, VWA5A, SPTBN2, PCK1, and TIMP1 genes. In the TCGA cohort, patients in the low-risk subgroup had a longer progression-free interval (PFI) than patients in the high-risk subgroup, which coincided with the results of the GEO cohort. The comprehensive results showed that the high-risk score was related to the enrichment of the cell cycle pathway, high mutation rate of TP53 and KRAS, high infiltration of T regulatory cells (Tregs), immunosuppressive state, and less chemotherapeutic benefit. However, low-risk scores are related to drug metabolism-related pathways, low TP53 and KRAS mutation rates, high infiltration of plasma cells, more resting CD4 memory cells and eosinophils, active immune function, and better chemotherapeutic benefits. Receiver operating characteristic curve of two-year progress prediction evaluation showed that the ARGPI had higher prognostic accuracy than TNM staging. Nomograms integrating ARGPI and clinicopathological risk factors can better evaluate the prognosis of patients with colon cancer. Conclusions: The ARGPI is a promising biomarker for determining risk of colon cancer progression, molecular and immune characteristics, and chemotherapeutic benefit. This is a reliable method to predict the prognosis of colon cancer patients. It also can assist doctors in formulating more effective treatment strategies.

Good or not good: Role of miR-18a in cancer biology.

miR-18a is a member of primary transcript called miR-17-92a (C13orf25 or MIR17HG) which also contains five other miRNAs: miR-17, miR-19a, miR-20a, miR-19b and miR-92a. This cluster as a whole shows specific characteristics, where miR-18a seems to be unique. In contrast to the other members, the expression of miR-18a is additionally controlled and probably functions as its own internal controller of the cluster. miR-18a regulates many genes involved in proliferation, cell cycle, apoptosis, response to different kinds of stress, autophagy and differentiation. The disturbances of miR-18a expression are observed in cancer as well as in different diseases or pathological states. The miR-17-92a cluster is commonly described as oncogenic and it is known as 'oncomiR-1', but this statement is a simplification because miR-18a can act both as an oncogene and a suppressor. In this review we summarize the current knowledge about miR-18a focusing on its regulation, role in cancer biology and utility as a potential biomarker.

The effect of Candida cell wall beta-glucan on treatment-resistant LL/2 cancer cell line: in vitro evaluation.

Candida albicans (C. albicans) cell wall beta-glucan has been considered as a potential agent in the treatment of cancers due to its anti-tumor properties. Therefore, in the present study, we investigated the anti-cancer effects of Candida cell wall beta-glucan on Lewis lung carcinoma cell line (LL/2) cells. Beta-glucan of C. albicans cell wall was extracted. LL/2 cell line was cultured, then sphere cells and parental cells were exposed to the different concentrations of beta-glucan extracted from C. albicans (10-6000 µg/ml), for 24, 48 and 72 h. Cytotoxicity of beta-glucan was assayed by MTT test, then RNA extracted from cells population (treated and untreated cells), cDNA synthetized and expression level of Sox2, Oct4, C-myc, Nanog genes were also investigated using Real-time methods. At optimal concentrations of 800 and 1000 µg/ml, the extracted beta-glucan showed a significant cytotoxic effect on both parental and sphere cell populations (p < 0.05). Real-time PCR analysis revealed a decreased expression of Oct4 and Sox2 genes in treatment of cells with beta-glucan compared with control group. Since the extracted beta-glucan showed an inhibitory effect on the expression of Oct4 and Sox2 genes involved in LL/2 metastasis, therefore, beta-glucan can be considered as an anti-tumor agent because of its anti-metastatic properties, however, more in vitro and in vivo studies are needed to provide further evidence on this topic in the future.

Effects at molecular level of multi-walled carbon nanotubes (MWCNT) in Chironomus riparius (DIPTERA) aquatic larvae.

Nowadays, due to the physical, chemical, electrical, thermal and mechanical properties of carbon nanotubes (CNT), its have been currently incorporated into biomedical products and they are employed in drug delivery drug administration, biosensor design, microbial treatments, consumer products, and new products containing CNT are expected in the future. CNT are hydrophobic and have a tendency to accumulate in sediments if they are released into aquatic ecosystems. Vertebrate studies have revealed concerns about the toxicity of carbon nanotubes, but there is very limited data on the toxic effects in aquatic invertebrate species. The aim of the present study is to determine the effects of MWCNT in Chironomus riparius at the molecular level, understanding its mode of action and analyzing the suitability of this species to monitor and assess risk of nanomaterials in aquatic ecosystems. To evaluate possible toxic effects caused by carbon nanotube environmental dispersion with regard to aquatic compartment, we study the mRNA levels of several related genes with DNA repairing mechanisms, cell stress response, cell apoptosis and cytoskeleton by Real-Time PCR and proposed a freshwater invertebrate C. riparius, which is a reference organism in aquatic toxicology. The obtained results show a transcriptional alteration of some genes included in this study, indicating that different cell processes are affected and providing one the first evidences in the mechanisms of action of MWCNT in invertebrates. Moreover, this data reinforces the need for further studies to assess the environmental risk of nanomaterial to prevent future damage to aquatic ecosystems.

Influence of pristine and hydrophobic ZnO nanoparticles on cytotoxicity and endoplasmic reticulum (ER) stress-autophagy-apoptosis gene expression in A549-macrophage co-culture.

Exposure to ZnO nanoparticles (NPs) might modulate endoplasmic reticulum (ER) stress-autophagy gene expression, but the possible influence of hydrophobic surface coating on these responses was less studied. This study used A549-macrophage co-culture as the in vitro model for lung barrier and investigated the toxicity of pristine and hydrophobic ZnO NPs. Pristine and hydrophobic NPs exhibited different Zeta potential and solubility in water, which suggested that hydrophobic surface coating might alter the colloidal aspects of ZnO NPs. However, pristine and hydrophobic ZnO NPs induced cytotoxicity and reduced the release of soluble monocyte chemotactic protein-1 (sMCP-1) in A549-macrophage co-culture to a similar extent. Exposure to pristine ZnO NPs significantly promoted the expression of ER stress-apoptosis genes, namely DDIT3, XBP-1s, CASP9, CASP12 and BAX (p < 0.05), but hydrophobic ZnO NPs only significantly promoted the expression of BAX (p < 0.05). Exposure to pristine ZnO NPs also significantly reduced the expression of autophagic gene BECN1 (p < 0.05) but not ATG7 (p > 0.05), whereas hydrophobic ZnO NPs significantly reduced the expression of ATG7 and BECN1 (p < 0.01). Moreover, the expression of XBP-1s, HSPA5, CASP9, CASP12, BAX and ATG7 in pristine ZnO NP-exposed co-culture was significantly lower than that in hydrophobic ZnO NP-exposed co-culture (p < 0.05). In conclusion, hydrophobic surface coating might influence the colloidal aspects of ZnO NPs and alter ER stress-apoptosis-autophagy gene expression pattern by pristine ZnO NPs in A549-macrophage co-culture.

Tanshinone Ⅱ A Induce Human Small Cell Lung Cancer Cells Apoptosis and Molecular Mechanism / 医学研究杂志

Objective To know the Tanshinone Ⅱ A induce human small cell lung cancer cells (H446 cells) apoptosis,and discuss the possible molecular mechanism.Methods Determined by MTT method of different concentrations of Tanshinone Ⅱ A impact on human small cell lung cancer H446 cells proliferation;Determined by Hoechst 33258 method of different concentrations Tanshinone Ⅱ A influence on human small cell lung cancer H446 cells apoptosis;Determined by qPCR method of different concentrations of Tanshinone Ⅱ A impact on human small cell lung cancer H446 cells apoptosis gene;Determined by Westen blot method of different concentrations of Tanshinone lⅡ A effect on lung cancer H446 cells apoptosis.Results Different drug concentrations of Tanshinone Ⅱ A (0.3,0.6,1.25,2.5,5 and 10μg/ml) inhibit human small cell lung cancer H446 cells proliferation and depend by the concentration and time;With the increase of the concentration of the drug in the formation of human small cell lung cancer H446 cells apoptosis corpuscle number increase,significant difference was found in high concentration group (P ＜ 0.05);High concentration group of Tanshinone Ⅱ A promote apoptosis gene (Bax,caspase-9,caspase-3) expression,and inhibit apoptosis inhibiting genes (Akt,the Bcl-2) expression and there were significant differences (P ＜ 0.05);High concentration group of Tanshinone Ⅱ A promoting apoptosis protein (Bax,caspase-9,caspase-3)expression,and inhibit apoptosis inhibiting protein (Akt,the Bcl-2) expression and there were significant differences (P ＜ 0.05).Conclusion Tanshinone Ⅱ A may be through Akt-Bax/Bcl-2-caspase-9-caspase-3 signaling pathways induced human small cell lung cancer H446 cells apoptosis.

Baculovirus antiapoptotic protein P35 regulated the host apoptosis to enhance virus multiplication.

The baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV) possesses p35 gene, which encodes antiapoptosis protein P35 (or Ac-P35). A previous study showed that deleting p35 promoted premature cytolysis and caused the reduction of virus yield. In this report, we examined the role of P35 in regulating the expression of Ac-IAPs (inhibitor of apoptosis proteins in AcMNPV) and SfP53 (an apoptosis protein in Sf9 cells), and its effect on the production of progeny virus. Results showed that the overexpression of P35 caused a delay in the increase process of SfP53 before 36-h post infection and improved the transcription levels of iaps gene dramatically; it was more favorable to improve the transcription level of iap1 at 24-72 h post infection. Moreover, P35 could promote the production of progeny virus. This is the first study to disclose the relationship among p35, iap1, iap2, Sfp53 and the replication of the virus in the AcMNPV infection process, which provides the basis for improving the insecticidal activity of recombinant AcMNPV in terms of theory and technology.

Interleukin-22 ameliorates acute severe pancreatitis-associated lung injury in mice.

AIM: To investigate the potential protective effect of exogenous recombinant interleukin-22 (rIL-22) on L-arginine-induced acute severe pancreatitis (SAP)-associated lung injury and the possible signaling pathway involved. METHODS: Balb/c mice were injected intraperitoneally with L-arginine to induce SAP. Recombinant mouse IL-22 was then administered subcutaneously to mice. Serum amylase levels and myeloperoxidase (MPO) activity in the lung tissue were measured after the L-arginine administration. Histopathology of the pancreas and lung was evaluated by hematoxylin and eosin (HE) staining. Expression of B cell lymphoma/leukemia-2 (Bcl-2), Bcl-xL and IL-22RA1 mRNAs in the lung tissue was detected by real-time PCR. Expression and phosphorylation of STAT3 were analyzed by Western blot. RESULTS: Serum amylase levels and MPO activity in the lung tissue in the SAP group were significantly higher than those in the normal control group (P < 0.05). In addition, the animals in the SAP group showed significant pancreatic and lung injuries. The expression of Bcl-2 and Bcl-xL mRNAs in the SAP group was decreased markedly, while the IL-22RA1 mRNA expression was increased significantly relative to the normal control group (P < 0.05). Pretreatment with PBS did not significantly affect the serum amylase levels, MPO activity or expression of Bcl-2, Bcl-xL or IL-22RA1 mRNA (P > 0.05). Moreover, no significant differences in the degrees of pancreatic and lung injuries were observed between the PBS and SAP groups. However, the serum amylase levels and lung tissue MPO activity in the rIL-22 group were significantly lower than those in the SAP group (P < 0.05), and the injuries in the pancreas and lung were also improved. Compared with the PBS group, rIL-22 stimulated the expression of Bcl-2, Bcl-xL and IL-22RA1 mRNAs in the lung (P < 0.05). In addition, the ratio of p-STAT3 to STAT3 protein in the rIL-22 group was significantly higher than that in the PBS group (P < 0.05). CONCLUSION: Exogenous recombinant IL-22 protects mice against L-arginine-induced SAP-associated lung injury by enhancing the expression of anti-apoptosis genes through the STAT3 signaling pathway.

Association of lumbar disc herniation with the interaction between environmental factors and Fas/FasL/Caspase-9 apoptosis-associated gene polymorphisms / 西安交通大学学报(医学版)

Objective To investigate the etiologic roles of apoptosis-associated genes,environmental factors and their interactions in lumbar disc herniation (LDH).Methods A case-control trial was conducted.We recruited 128 outpatients with LDH as case group and 132 normal people matched by age and gender as control group.Peripheral venous blood samples were collected and DNA was extracted from leukocytes.By using a modified Brucker Autoflex MALDI-TOF mass spectrometer,we analyzed 3 genes with 9 polymorphic sites,namely,Fas-1377G/A rs2234767,Fas-670G/A rs1800682,Fas rs2147420,Fas rs2296603,Fas rs7901 656,Fas rs1 57101 9,FasL-844C/T rs7631 10,CASP-9-1263A > G rs4645978,and CASP-9-712C > T rs4645981.The correlations between polymorphism of Fas,FasL and CASP-9 genes and the risk of LDH were evaluated by non-conditional Logistic regression model.Multiple Logistic regression model was performed to assess the interaction between apoptosis-associated genes and environment factors,such as lumbar vertebral loads,bed type,spare-time exercises and spare-time activities. Results There were preferable balances in case and control groups in age and gender without significant differences.However,the two groups differed significantly (P G (rs4645978),and FasL-844C/T TT and CASP-9-1263A>G GG genotypes might be the high risk genotypes of LDH.The gene-environment interaction analysis revealed that super-multiplicative and sub-multiplicative interactions respectively between FasL-844TT genotype and lumbar vertebral loads (3-4 level),and between CASP-9-rs4645978 GG and lumbar vertebral loads (3-4 level).Conclusion FasL,CASP-9 genes and lumbar vertebral loads and their interactions play important roles in the pathogenesis of LDH.It suggests that the risk of LDH may be codetermined by environmental factors and inherited susceptibility genes,and that the mechanisms of interactions vary in different genotypes and the same or different environmental factors.

The association of apoptotic protein expressions sensitive to apoptosis gene, p73 and p53 with the prognosis of cervical carcinoma.

OBJECTIVE: To evaluate the expressions of several apoptotic pathway proteins in relation to clinical parameters and survival in patients with cervical carcinoma. METHODS: A total of 20 patients with clinically advanced staged carcinoma of cervix (International Federation of Gynecology and Obstetrics [FIGO] stage IIB-IVA) aged from 40 to 75 years were included in this study. The expression profile of anti-apoptotic protein (sensitive to apoptosis gene [SAG]), mitochondrial apoptotic proteins (B-cell lymphoma-extra-large [Bcl-xL] and Bcl-2 homologous antagonist/killer [Bak]), and tumor suppressor proteins (p73 and p53) were examined by real-time polymerase chain reaction experiments along with their relation to clinical parameters and survival analyses during follow-up for 5 to 8 years. RESULTS: No significant difference was found in the expressions of SAG, Bcl-xL, Bak, p73 and p53 proteins with respect to stage and grade of tumor. A significant positive correlation was noted between SAG and Bcl-xL genes (r=0.752, P<0.001) and between SAG and Bak genes (r=0.589, P=0.006). Among genes determined to be significantly associated with overall survival in the univariate analysis (P=0.026 for SAG, P=0.002 for Bcl-xL, and P=0.027 for p53), only p53 was identified as the significant predictor in the multivariate analysis (hazard ratio: 8.53, 95% confidence interval: 1.34-54.2, P=0.023). CONCLUSION: In conclusion, our findings demonstrated a reverse correlation of SAG, Bcl-xL, and p53 expressions with overall survival of patients. No association of apoptotic pathway proteins with clinicopathological characteristics of cervical carcinoma patients was noted. Low SAG, Bcl-xL, and p53 expression levels revealed to be useful as prognostic predictors in patients with cervical carcinoma.

Chromosomal microarray analysis of consecutive individuals with autism spectrum disorders or learning disability presenting for genetic services.

Chromosomal microarray analysis is now commonly used in clinical practice to identify copy number variants (CNVs) in the human genome. We report our experience with the use of the 105 K and 180K oligonucleotide microarrays in 215 consecutive patients referred with either autism or autism spectrum disorders (ASD) or developmental delay/learning disability for genetic services at the University of Kansas Medical Center during the past 4 years (2009-2012). Of the 215 patients [140 males and 75 females (male/female ratio=1.87); 65 with ASD and 150 with learning disability], abnormal microarray results were seen in 45 individuals (21%) with a total of 49 CNVs. Of these findings, 32 represented a known diagnostic CNV contributing to the clinical presentation and 17 represented non-diagnostic CNVs (variants of unknown significance). Thirteen patients with ASD had a total of 14 CNVs, 6 CNVs recognized as diagnostic and 8 as non-diagnostic. The most common chromosome involved in the ASD group was chromosome 15. For those with a learning disability, 32 patients had a total of 35 CNVs. Twenty-six of the 35 CNVs were classified as a known diagnostic CNV, usually a deletion (n=20). Nine CNVs were classified as an unknown non-diagnostic CNV, usually a duplication (n=8). For the learning disability subgroup, chromosomes 2 and 22 were most involved. Thirteen out of 65 patients (20%) with ASD had a CNV compared with 32 out of 150 patients (21%) with a learning disability. The frequency of chromosomal microarray abnormalities compared by subject group or gender was not statistically different. A higher percentage of individuals with a learning disability had clinical findings of seizures, dysmorphic features and microcephaly, but not statistically significant. While both groups contained more males than females, a significantly higher percentage of males were present in the ASD group.

Induction effect of benzene on apoptosis of mouse bone marrow cells through mitochondrial-dependent apoptosis pathway and its mechanism / 吉林大学学报(医学版)

Objective To establish mouse poisoning model by inhaling benzene, and to investigate the induction effect of benzene on the apoptosis of mouse bone marrow cells and its mechanism, and to provide an experimental basis for study on bone marrow toxicity mechanism.Methods 24 male mice were randomly divided into four groups (n=6).The mice in one group were exposed to ambient air (control group)and the mice in the other three groups were exposed to different doses (400,800,1 600 mg·m-3 )of benzene (low,middle and high doses of benzene groups)for 1 5 d in the respective inhalation chambers. At the end of the experiment, the mice were killed. The bone marrow of the mice was obtained. The pathological changes of the bone marrow cells of the mice in various groups were observed under light microscope with HE staining.The apoptotic rates and mitochondrial membrane potential (MMP ) of the mice in various groups were detected by flow cytometry, and the expressions of mitochondrial-deperdent apoptosis related gene proteins were determined with immunohistochemistry method. Results The number of distal and central cells in different doses of benzene groups were significantly reduced,and accompanied by blood sinus expansion in high dose of benzene group.The apoptotic rates of the cells in middle and high doses of benzene groups were obviously higher than that in control group (Ρ<0.01),and there were also significant differences between high dose group and low,middle doses of benzene groups (Ρ<0.05).The MMP was significantly decreased with the increasing of benzene doses, and there were significant differences between middle,high doses of benzene groups and control group (Ρ<0.05).The number of Bax,CytC positive cells in different doses of benzene groups and the number of Caspase-9,Caspase-3 positive cells in middle and high doses of benzene groups were significantly increased compared with control group(Ρ<0.05);the number of Bcl-2 positive cells in different doses of benzene groups was decreased(Ρ<0.05),and number of Bcl-2 positive cells in middle and high doses of benzene groups was decreased compared with low dose of benzene group (P<0.05). Conclusion Benzene with certain dose can induce the apoptosis of mouse bone marrow cells, and promote the expressions of mitochondrial apoptosis related gene proteins. Benzene-induced apoptosis through mitochondrial-dependent apoptosis pathway may be an important mechanism of bone marrow toxicity induced by benzene.

Analysis of effects of allicin prodrug on proliferation of esophageal cancer cell line Eca9706 and the expression of apoptosis gene / 中国生化药物杂志

Objective To explore and analyze the effects of allicin prodrug on proliferation of esophageal cancer cell line Eca9706 and the expression of apoptosis gene.Methods Different concentrations of allicin prodrugs were divided into a total of four groups:A1 group(10μg/mL),A2 group(20 μg/mL),A3 group (40 μg/mL),A4 group (normal saline,0 μg/mL),and respectively applicated in esophageal carcinoma cell line Eca9706.After culturing for 1 days,2 days,3 days,cell proliferation was detected by MTT and expression of p53 at the mRNA level was detected by RT-PCR.Results The optimum concentration of proliferation inhibition of allicin prodrug on esophageal cancer cell line Eca9706 was 20μg/mL and the best inhibition time was 2 days;the gene level of esophageal cancer cell line Eca9706 was inhibited with a dose-dependent.Conclusion Allicin prodrugs could effectively inhibit the proliferation of esophageal cancer cell line Eca9706,and control the proliferation of esophageal cancer cells by regulating the expression of apoptosis associated genes,so as to eliminate tumor cells.

Effect of arctigenin on apoptosis of mouse erythroleukemia FBL-3 cells and its mechanism / 中华实用儿科临床杂志

Objective To investigate the effect of arctigenin on the apoptosis of FBL-3 cells and its mechanisms.Methods The mouse erythroleukemia FBL-3 cells were taken as subjects.The untreated FBL-3 parental cells were taken as the control group,and the FBL-3 cells treated with 20 mg/L arctigenin for 24 h were taken as the experimental group.The effects of arctigenin on the apoptosis of the mouse erythroleukemia FBL-3 cells were determined by agarose gel electrophoresis and flow cytometry assay.The changes of apoptosis-related genes were analyzed by reverse transcriptase-polymerase chain reaction(RT-PCR).Results The agarose gel electrophoresis results revealed that the " DNA ladder" was displayed in the experimental group compared to the control group.The flow cytometry data demonstrated that the apoptosis number of FBL-3 cells in the experimental group was markedly increased compared to that in the control group (t =60.681,P =0.000).The RT-PCR assay results suggested that the expressions of Bcl-2 and IAP-1 gene were down-regulated in the experimental group compared to the control group (t =14.732,29.702,all P =0.000),while the expressions of Bax and Smac gene were up-regulated(t =6.721,8.499;P =0.003,0.001),but the expression of Bcl-XL did not change(t =0.209,P =0.844).Conclusions Arctigenin can induce the apoptosis of the mouse erythrolenkemia FBL-3 cells.Apoptosis of the mouse erythrolenkemia FBL-3 cells induced by arctigenin may be correlated to the down-regulation of Bcl-2 and IAP-1 and up-regulation of Bax as well as Smac.

Prognostic role of sensitive-to-apoptosis gene expression in rectal cancer.

AIM: To investigate the association between prognosis of rectal cancer treated with chemoradiotherapy (CRT) and expression of sensitive-to-apoptosis (SAG), B-cell lymphoma-extra large (Bcl-X(L)) and Bcl-2 homologous antagonist/killer (Bak). METHODS: Real-time quantitative polymerase chain reaction was used to determine the expression of proteins of interest, namely SAG, Bcl-X(L), Bak and ß-actin, in rectal carcinoma patients who had a follow-up period of 3 years after CRT. Biopsy specimens were excised from the rectal tumor preceding CRT. RESULTS: SAG, Bcl-X(L) and Bak proteins showed significant correlations with each other. In multivariate analysis, patients with high vs low SAG expression showed a statistically significant difference in 2-year survival rates: 56% vs 73%, respectively (P = 0.056). On the other hand, there were no significant correlations between the expression levels of all three genes and metastatic rates or tumor responses to CRT. Mean overall survival in the patients with elevated SAG expression was 27.1 mo ± 3.9 mo [95% confidence interval (CI): 19.3-34.9], and in patients with reduced expression, it was 32.1 mo ± 2.5 mo (95% CI: 27.3-36.9). The corresponding values for Bcl-X(L) were 28.0 mo ± 4.1 mo (95% CI: 19.9-36.1) and 31.7 mo ± 2.9 mo (95% CI: 26.0-37.5), and those for Bak were 29.8 mo ± 3.7 mo (95% CI: 22.5-37.2) and 30.6 mo ± 2.4 mo (95% CI: 25.5-35.0), respectively. CONCLUSION: Two-year survival rates significantly correlated with low SAG expression, and SAG may be a candidate gene for good prognosis, independent of therapeutic response of different individuals.

Survivin isoforms and clinicopathological characteristics in colorectal adenocarcinomas using real-time qPCR.

AIM: To investigate three isoforms of survivin in colorectal adenocarcinomas. METHODS: We used the LightCycler Technology (Roche), along with a common forward primer and reverse primers specific for the splice variants and two common hybridization probes labeled with fluorescein and LightCycler-Red fluorophore (LC-Red 640). Real time quantitative polymerase chain reaction (PCR) was performed on cDNAs from 52 tumor specimens from colorectal cancer patients and 10 unrelated normal colorectal tissues. In the patients group, carcinoembryonic antigen (CEA) and CA19-9 tumor markers were also measured immunochemically. RESULTS: Wild type survivin mRNA isoform was expressed in 48% of the 52 tumor samples, survivin-2b in 38% and survivin-ΔΕx3 in 29%, while no expression was found in normal tissues. The mRNA expression of wild type survivin presented a significant correlation with the expression of the ratio of survivin-2b, survivin-ΔΕx3, survivin-2b/wild type survivin and survivin-ΔΕx3/wild type survivin (P < 0.001). The mRNA expression of wild-survivin and survivin-ΔΕx3 was related with tumor size and invasion (P = 0.006 and P < 0.005, respectively). A significant difference was found between survivin-2b and morphologic cancer type. Also, the ratio of survivin-ΔEx3/wild-survivin was significantly associated with prognosis. No association was observed between the three isoforms and grade, metastasis, Dukes stage and gender. The three isoforms were not correlated with CEA and CA19-9. CONCLUSION: Survivin isoforms may play a role in cell apoptosis and their quantification could provide information about clinical management of patients suffering from colorectal cancer.

The effect of neural stem cell transplantation on the hindlimb motor function and caspase-3 expression of motor cortex in spinal cord transected rats / 中华行为医学与脑科学杂志

Objective To observe the effect of neural stem cell (NSC)transplantation on the hindlimb motor function and caspase-3 expression of motor cortex (MC) in spinal cord transected (SCT) rats. Methods S prague-Dawley (SD) rats were randomly divided into sham operated group, operation group( T9 transection), and subacute NSC transplantation group. The MC of each group( n= 8 )was harvested 7 days post operation (dpo), then western blot was employed to detect the level of caspase-3( β-actin was used as control ). The left 5 animals of each group were subjected to BBB score evaluation at 16th week,then the animals were sacrificed and the MC was harvested and performed immunostain by using caspase-3 rabbit antibody. Results Following NSC transplanta tion, BBB scores( ( 7.58 ± 0. 99 ) ) increased significantly than seen in the SCT animals( ( 5.16 ± 1.19) ). The expressional level of caspase-3 at 7day post operation was( (0.89 + 0.12)) in MC of SCT rats,while it decreased significantly to( (0.76 + 0.11 ) ) in NSC transplantation rats(P＜0.05). The immunoreactive stain of caspase-3 was seen in the cytoplasm of pyramidal neuron in the cortex. Conclusion NSC engraft can downregulate the expression of caspase-3, corresponding to a significant improvement in hindlimb motor function. These findings indicate that NSC transplantation probably regulate the expression of apoptosis genes in MC to promote neurological function recovery in rats subjected SCT.

Radiosensitization effect of recombinant adenoviral-mediated PUMA gene on pancreatic carcinoma cells / 中华放射医学与防护杂志

Objective To study the effect of PUMA gene mediated by recombinant adenovirus vector combined with radiation on the pancreatic carcinoma. Methods The PANC-1 cells were infected with Ad-PUMA (MOI = 10, 50 and 100, respectively) for 48 h. The expression of PUMA mRNA and protein was detected by RT-PCR and Western blot, respectively. PANC-1 cells were divided into 4 groups: control group, transfection group, irradiation group and combined treatment group. The cell growth inhibition rate and apoptotic rate of PANC-1 cells were assessed by MTT assay and flow cytometry. Human pancreatic carcinomas were transplanted subcutaneously in nude mice, which were randomized into 4 groups: control group, transfection group, irradiation group and combined treatment group. Tumor growth rate and apoptotie index at different time points were recorded in 35 days. Results The expression of PUMA mRNA and protein was increased with the increase of MOI of Ad-PUMA, which was does-dependant (MOI = 10, mRNA = 0.46 ± 0.02, protein = 0. 75 ±0.09;MOI=50, mRNA= 1.12±0.09, protein = 1.01 ±0.18;MOI= 100, mRNA= 1.50±0.08, protein=1.80 ± 0.15 ;P < 0.05). The proliferation of PANC-1 cells was suppressed significantly when transfected by Ad-PUMA in a dose-dependent manner(r = -0.986 55), which was more significant combined with radiation (r = -0.971 26, P < 0.05). Meanwhile, the apoptotie rate was increased in the same manner [for pre- and post-irradiation,which was (45.4 ± 5.26) % and (73.2 ± 6.62) %, respectively, P < 0.05]. From 7 to 35 d after PUMA gene transfection and radiotherapy, the tumor growth was significantly slower than those of irradiation group, transfection group and control group [35 d after therapy, the volume of tumor was (19.82 ± 6.45)mm3 ,(39.5 ± 9.23)mm3 , (33.6 ±3 10.3)mm3 and (52.0 ± 11.43)mm3 , respectively, P < 0.05]. And the apoptotic index was increased in the same manner (AI = 0.43 ± 0.05, 0.29 ± 0.10, 0.24 ± 0.05 and 0.00 ± 0.00, respectively, P < 0.05). Conclusions Recombinant adenoviral-mediated PUMA gene combined with irradiation could increase the cell-killing effect on pancreatic carcinoma. It is better than that of either one kind of therapy.

Effect of Estrogen on Expression of Osteoblast Apoptosis Related Genes Induced with Serum Hungry / 中国康复理论与实践

@#ObjectiveTo explore the mechanism of estrogen inhibiting osteoblast apoptosis induced with serum hungry.MethodsOsteoblasts of the second or third generation from newly born SD rats calvaria were divided randomly into three groups: control group,serum hungry group,serum hungry with estrogen group.Cells of each group were incubated for 1,2,3,5,7 or 14 d,and then were stained immunohistochemically.The rates of positive cells of each group were analyzed.ResultsThere was a little positive expression of Bax,Bcl2 and Fas in control group.The expression of Bax and Fas were significantly increased(P<0.05)in serum hungry group,peak time was 14 d,but the expression of Bcl-2 were not affected.Compared with that of serum hungry group,the expression of Bax and Fas significantly decreased(P<0.05) in serum hungry and estrogen group,peak time was still 14 d,while that of Bcl-2 increased(P<0.05).ConclusionSerum hungry can increase the expression of Bax and Fas in osteoblast,that can be inhibited by estrogen.Estrogen can also increase the expression of Bcl-2 in osteoblast.All of these may play a role in inhibiting osteoblast apoptosis induced with serum hungry.