Inhibition of all-trans retinoic acid on MDM2 gene expression in astrocytoma cell line SHG-44

Objective To investigate the impact of all-trans retinoic acid （ATRA） on MDM2 gene expression in astrocytoma cell line SHG-44, and to provide basic data for further research on the progression mechanism and gene therapy of human astrocytoma. Methods The differential expressions of MDM2 gene and protein in SHG-44 cells were detected by cDNA microarray and Western blot, respectively, before and after treatment of ATRA. The expressions of MDM2 protein in WHO grade Ⅱ and grade Ⅳ astrocytomas were determined by immunohistochemical streptavidin-peroxidase method. Some differentially expressed genes were selected randomly for Northern blot analysis. Results The intensity ratio of ATRA-treated to untreated SHG-44 cell was 0.37 in the cDNA microarray, suggesting that the expression of MDM2 gene was down-regulated in SHG-44 cells after treatment with ATRA. Some genes differentially expressed in the microarray were confirmed by Northern blot. Western blot demonstrated that the optical density ratios of MDM2 to β-actin in ATRA-treated and untreated SHG-44 were 14.02±0.35 and 21.40±0.58 （t = 24.728, P = 0.000）, respectively, suggesting that the expression of MDM2 protein was inhibited in ATRA-treated SHG-44 cells. Moreover, the percentages of MDM2-positive protein were 24.00% （6/25） and 56.52% （13/23） （x^2 = 5.298, P = 0.021） in WHO grade Ⅱ and grade Ⅳ astrocytomas, respectively, suggesting that the expression of MDM2 protein may increase along with the elevation of astrocytoma malignancy. Conclusion ATRA can inhibit MDM2 gene expression in SHG-44 cells, and MDM2 is related to astrocytoma progression.

SMAD3和CD44在甲状腺乳头状癌组织中的表达水平与临床病理特征及预后的关系

目的探讨信号转导分子3(SMAD3)和白细胞分化抗原44(CD44)在甲状腺乳头状癌(PTC)组织中的表达水平与临床病理特征及预后的关系。方法选取2019年6月-2020年6月收治的PTC患者88例为研究对象,同期取距离癌组织3 cm以上癌旁组织作为对照,实时荧光定量PCR(qRT-PCR)检测SMAD3和CD44的mRNA表达水平;免疫组化法检测SMAD3和CD44的阳性表达情况。Spearman相关性分析PTC患者癌组织中SMAD3和CD44的mRNA表达水平的关系。SMAD3和CD44与PTC患者预后的关系采用Kaplan-Meier法进行分析。COX回归分析影响PTC患者预后的危险因素。结果与癌旁组织相比,PTC组SMAD3的表达水平显著降低(t=8.484,P<0.05),CD44表达水平显著升高(t=11.232,P<0.05)。相关性分析显示,PTC患者癌组织中SMAD3和CD44的mRNA表达水平呈负相关(r s=-0.519,P<0.05)。与癌旁组织相比,PTC组SMAD3阳性表达率显著降低(χ^(2)=41.905,P<0.05),CD44阳性表达率显著升高(χ^(2)=62.888,P<0.05)。SMAD3表达与TNM分期(χ^(2)=7.678,P=0.006)、淋巴结转移(χ^(2)=20.398,P<0.05)、侵犯包膜(χ^(2)=9.881,P=0.002)有关。CD44表达与TNM分期(χ^(2)=3.959,P=0.047)、淋巴结转移(χ^(2)=20.702,P<0.05)、侵犯包膜(χ^(2)=10.363,P=0.001)有关。术后随访3年发现,SMAD3阳性表达PTC患者生存率高于阴性表达患者生存率(χ^(2)=4.644,P=0.031)。CD44阳性表达PTC患者生存率显著低于阴性表达患者生存率(χ^(2)=5.331,P=0.021)。多因素COX回归分析显示,淋巴结转移、SMAD3和CD44表达是影响PTC患者预后的危险因素(P<0.05)。结论PTC组织中SMAD3呈异常低表达,CD44呈高表达,其水平与PTC患者临床病理特征及预后有关。

重度子痫前期患者胎盘组织中CD44 mRNA和CD24 mRNA及蛋白表达水平的临床价值研究

目的探讨重度子痫前期(severe preeclampsia,SPE)患者胎盘中细胞表面跨膜糖蛋白分子(cell surface transmembrane glycoprotein molecules,CD)44 mRNA,细胞表面跨膜糖蛋白分子24(CD24)mRNA及蛋白表达水平表达的临床价值研究。方法选取2019年6月~2022年6月在聊城市第二人民医院接受剖宫产分娩的SPE患者,根据发病孕龄的不同,进一步将其分为早发型SPE组(孕龄≤34周,n=45)和晚发型SPE组(孕龄>34周,n=55)。选取同期产检正常者100例为对照组。采用荧光定量PCR和免疫组织化学检测SPE患者胎盘中CD44和CD24表达,Pearson法分析其表达水平差异以及与SPE疾病临床特征的相关性,多因素Logistic回归分析发生SPE的影响因素。结果相较于对照组,SPE胎盘组织中CD44 mRNA(0.55±0.12 vs 1.02±0.33),CD24 mRNA的表达水平(0.68±0.19vs 1.05±0.11)均降低,差异具有统计学意义(t=13.385,16.853,P<0.05)。免疫组织化学染色结果显示,CD44,CD24在SPE组胎盘组织中多呈阴性表达或弱阳性表达,而在对照组中多呈阳性表达,且SPE胎盘组织中CD44,CD24阳性率低于对照组,差异具有统计学意义(χ^(2)=9.696,14.346,P<0.05)。相较于早发型SPE组,晚发型SPE胎盘组织中CD44(0.65±0.17 vs 0.42±0.11),CD24(0.77±0.23 vs 0.58±0.13)mRNA的表达水平均较高,差异具有统计学意义(t=7.830,4.932,P<0.05)。相较于对照组,SPE组BMI,收缩压、舒张压、尿蛋白、Cr,LDH和BUN均显著升高,差异有统计学意义(t=5.360~30.241,均P<0.05);SPE组分娩孕周较早、MPV,ALB较低、新生儿出生身长较短和体质量较轻均低于对照组,差异具有统计学意义(t=3.232~11.109,均P<0.05)。且SPE胎盘组织中CD44与CD24表达呈正相关(r=0.698,P<0.05),SPE胎盘组织中CD44的表达分别与CD24,分娩孕周、MPV和新生儿出生身长呈正相关(r=0.611,0.639,0.612,0.465,均P<0.05);与收缩压、尿蛋白和LDH呈负相关(r=-0.604,-0.569,-0.593,均P<0.05)。CD24的表达分别与分娩孕周、MPV和新生儿出生身长呈正相关(r=0.605,0.584,0.640,均P<0.05);与收缩压、尿蛋白和LDH呈负相关(r=-0.637,-0.593,-0.561,均P<0.05)。经Logistic回归分析结果显示,MPV(95%CI:1.429~4.350),尿蛋白(95%CI:1.529~2.709),LDH(95%CI:1.425~3.932)均是发生SPE的独立危险因素(均P<0.05)。高水平CD44(95%CI:0.561~0.940),CD24(95%CI:0.495~0.814)是发生SPE的独立保护因素(均P<0.05)。结论SPE患者胎盘中CD44,CD24表达水平较低,高水平CD44,CD24均是SPE发生的独立保护因素,可为后续SPE的治疗提供方向。

Ardisia gigantifolia ethanolic extract inhibits cell proliferation and targets cancer stem cells in gastric cancer

Objective:To evaluate the effects of ethanol extract from Ardisia gigantifolia leaves on cell proliferation and cancer stem cell(CSC)number in gastric cancer.Methods:The inhibitory effect of Ardisia gigantifolia extract on the proliferation of MKN45 and MKN74 gastric cancer cells was assessed using 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide assay.Non-adherent culture(3D)model was used to evaluate the effect of the extract on tumorsphere size and number.Moreover,the expression of CD44,ALDH,and p21 was determined by immunofluorescence analysis.Flow cytometric analysis was performed to evaluate cell cycle arrest and the expression of gastric CSC markers CD44 and ALDH.Real-time PCR analysis was also carried out to assess the effect of the extract on the expression of cell cycle-regulated genes.Results:Ardisia gigantifolia extract effectively inhibited cell proliferation with an IC_(50)of 55.7μg/m L in MKN45 cells and 123.6μg/m L in MKN74 cells.The extract also arrested cell cycle in the G_(0)/G_(1)phase as well as significantly reduced the size and number of tumorspheres.The markedly increased expression of p21 was observed at both m RNA and protein levels in the extract-treated adherent cells and tumorspheres.In addition,Ardisia gigantifolia extract significantly reduced the number of CD44-and/or ALDH-expressing gastric CSC.Conclusions:The development of gastric CSC can be inhibited by the ethanol extract of Ardisia gigantifolia.

DAPT suppresses the proliferation of human glioma cell line SHG-44

Objective:To explore the suppressing effect ofγ-secretase inhibitor DAPT on proliferation of human glioma cell line SHG-44 in vitro and its mechanism.Methods:The SHG-44 cell was treated by DAPT with different concentration.The proliferation of cells was detected by MTT assay;cell cycle and TSC of CD133^+were determined by flow cytometry analysis technique;the key factor in Notch signaling pathway(Notch-1,Delta-1,Hes-1)was measured by reverse transcrip tase-polymerase chain reaction and western blotting.Results:DAPT inhibited the growth and proliferation of SHG-44 cells significantly(P<0.05).And the inhibiting effect on SHG-44 cells produced by DAPT showed a dose-dependent manner.DAPT increased the rate of cells in G_0/G_1 phase of SHG-44 cells,while it decreased the rate of cells in S phase.TSC of CD133^+was significantly reduced after DAPT treated SHC-44 cells.The expression of protein and mRNA of Notch-1,Delta-1 and Hes-1 were gradually downregulated with the increase of DAPT doses.Conclusions:DAPT can downregulate these key factor in Notch signaling pathway,reduce the TSC of CD133+and inhibit the proliferation of SHC-44 cells.

Endothelial adhesion of synchronized gastric tumor cells changes during cell cycle transit and correlates with the expression level of CD44 splice variants

AIM To study adhesion capacity and CD44 expression of human gastric adenocarcinoma MKN45 cells at different stages of a first cell cycle. METHODS MKN45 cells were synchronized by aphidicolin and assayed for adhesion to an endothelial cell （HUVEC） monolayer. Surface expression of CD44 and CD44 splice variants on MKN45 cells was evaluated by flow cytometry. Functional relevance of CD44 adhesion receptors was investigated by blocldng studies using anti CD44 monodonal antibodies or by hyaluronan digestion. RESULTS： Adhesion of MKN45 to HUVEC was increased during G2/M transit, after which adhesion returned to baseline levels with cell cycle completion. In parallel, CD44 splice vadants CD44v4, CD44v5, and CD44v7 were all upregulated on MKN45 during cell cycle progression with a maximum effect in G2/M. The function of CD44 surface receptors was assessed with specific receptor blocking monoclonal antibodies or removal of hyaluronan by digestion with hyaluronidase. Both strategies inhibited tumor cell adhesion to HUVEC by nearly 50%, which indicates that MKN45-HUVEC-interaction is CD44 dependent. CONCLUSION CD44 expression level is linked to the cell cycle in gastrointestinal tumor cells, which in turn leads to cell cycle dependent alterations of their adhesion behaviour to endothelium.

Effect of CD44 Suppression by Antisense Oligonucleotide on Attachment of Human Trabecular Meshwork Cells to HA

The effects of suppression of CD44 by CD44-specific antisense oligonucleotide on attachment of human trabecular meshwork cells to hyaluronic acid (HA) were observed and the possible relationship between CD44 and primary open-angle glaucoma (POAG) investigated. CD44-specific antisense oligonucleotide was delivered with cationic lipid to cultured human trabecular meshwork cells. The expression of CD44 suppressed by CD44-specific antisense oligonucleotide was detected by RT-PCR and Western blotting. The effect of CD44 suppression by specific antisense oligonucleotide on attachment of trabecular meshwork cells to HA was measured by MTT assay. Results showed that expression of CD44 was suppressed by CD44-specific antisense oligonucleotide. Antisense oligonucleotide also suppressed the adhesion of human trabecular meshwork cells to HA in a concentration dependent manner. It was concluded that attachment of human trabecular meshwork cells to HA was decreased when CD44 was suppressed by specific antisense oligonucleotide. CD44 might play a role in pathogenesis of POAG by affecting the adhesion of trabecular meshwork cells to HA.

PEI-PEG as a siRNA Genetic Vector Demonstrating Interference in the Expression of CD44v6 Protein in Gastric Cancer Cells

OBJECTIVE To investigate the effect of polyethylene imine glycol （PEI-PEG）/siRNA nanocomposites in the in vitro transfection of human gastric cancer SGC7901 cell lines and the down-regulation of gene expression of the adherence factor CD44v6. METHODS PEI-PEG/siRNA nanoparticles, in different N/P ratios, were synthesized and transfected into gastric cancer cells. Lipo2000/siRNA was used in the control group. The transfection efficiencies were observed under fluorescence microscope. The cytotoxicity of the nanoparticles was measured using the MTT assay （mononuclear cell direct cytotoxicity assay）, and the down-regulation effect of siRNA on CD44v6 gene was evaluated by Western blot. Based on the different N/P ratios, PEI-PEG/siRNA composites were synthesized and transfected into gastric cancer cells. Lipo2000/siRNA was used in the controls. The transfection efficiency was observed under fluorescence microscope. The cytotoxicity of the nanoparticles was measured using the MTT assay and the down-regulation effect of siRNA on CD44v6 gene was evaluated by Western blot. RESULTS After transfection, the transfection efficiency of the PEI-PEG/siRNA nanocomposites increased incrementally in N/P ratio value. The transfection efficiency improved with an increase in N/P ratio. When the N/P value was 15, fluorescence became more intense in the PEI-PEG/siRNA group than in the Lipo2000/siRNA group. At the same time, cell viability was （80.4 ± 5.6）% in the MTT reduction assay, which was similar to that in the Lipo2000/siRNA group. The results of Western blot analysis showed that the expression level of CD44v6 protein decreased to （59.7 ± 3.0）% after siRNA-CD44v6 was inhibited. CONCLUSION PEI-PEG could effectively form the nanocomposite in combination with siRNA, be transfected into the SGC7901 gastric cancer cell lines and inhibit CD44v6 protein expression. Moreover, as a genetic carrier, PEI-PEG copolymer has greater advantages, including high transfection e. ciency, less cytotoxicity and an easily alterable vector structure.

Effects of all-trans retinoic acid on metabolic gene expression in the glioma cell line SHG-44

BACKGROUND： Genetic abnormalities and changes in gene expression have been shown in various grades of glioma. However, the relationship between gene expression patterns and pathways related to malignant transformation of glioma remains poorly understood. OBJECTIVE： To screen differentially expressed genes between normal and all-trans retinoic acid-treated glioma cell line SHG-44 cells with a complementary DNA （cDNA） microarray. DESIGN, TIME AND SETTING： The genomics, in vitro study was performed at the Laboratory of Neurobiology, Third Military Medical University of Chinese PLA, China from January to October 2007. MATERIALS： The glioma cell line SHG-44 was provided by the Third Military Medical University of Chinese PLA. AII-trans retinoic acid was purchased from Sigma, USA. cDNA microarray was purchased from City University of Hong Kong. METHODS： The glioma cell line SHG-44 was treated with 10 μmol/L all-trans retinoic acid for 3 days Differentiation-related genes were determined using cDNA microarray. MAIN OUTCOME MEASURES： Gene expression patterns were compared between normal and all-trans retinoic acid-treated SHG-44 cells. Differentially expressed genes were randomly selected and determined by Northern blot analysis. RESULTS： Northern blot analysis revealed downregulated RPL 13 gene expression and upregulated SOD2 gene expression, which was identical to cDNA microarray results. Five differentially expressed genes （TPI1, BPGM, ALDOA, LDHA, and RRM1） were shown to be involved in cell metabolism, in six metabolic pathways. Four differentially expressed genes （TPI1, BPGM, ALDOA, and LDHA） were associated with carbohydrate metabolism, such as fructose metabolism, pyruvic acid metabolism, pentose phosphate pathway, glycolysis, and gluconeogenesis. One differentially expressed gene （RRM1） was correlated with purine and pyrimidine metabolism. CONCLUSION： Five metabolic genes （TPI1, BPGM, ALDOA, LDHA, and RRM1）, which participate in cell carbohydrate and nucleotide metabolism, were shown to closely correlate with glioma development.

CD44蛋白在胃癌中的表达情况及临床意义分析

目的:分析胃癌中白细胞分化抗原44(CD44)蛋白的表达情况与临床意义。方法:回顾性分析2019年1月—2022年1月于苏州市立医院接受手术治疗的胃癌患者139例的临床资料,并采用免疫组织化学法检测患者胃癌组织病理标本中CD44蛋白表达情况。分析CD44蛋白表达情况与临床病理特征及预后的关系。结果:胃癌组织中CD44蛋白表达率高于正常胃黏膜组织,差异有统计学意义(P<0.001)。阳性病例中,低分化胃癌患者CD44蛋白免疫组化评分高于高/中分化胃癌患者,肿瘤最大径≥4 cm胃癌患者CD44蛋白免疫组化评分高于肿瘤最大径<4 cm胃癌患者,差异有统计学意义(P<0.001);CD44蛋白阳性与CD44蛋白阴性胃癌患者性别、年龄、Lauren分型比较,差异无统计学意义(P>0.05);CD44蛋白阳性胃癌患者肿块最大径≥4 cm、低分化、血管侵犯阳性、淋巴结转移阳性、TNM分期为Ⅲ或Ⅳ期占比高于CD44蛋白阴性者,差异有统计学意义(P<0.05);CD44阳性表达者生存期短于阴性表达者,差异有统计学意义(P=0.003)。结论:不同分化程度、肿瘤最大径的胃癌患者CD44蛋白表达存在明显差异,CD44蛋白阳性与CD44蛋白阴性胃癌患者肿块最大径、分化程度、血管侵犯情况、淋巴结转移情况、TNM分期存在明显差异,可将CD44蛋白作为胃癌浸润、转移及判读预后的生物学标记物。

The expression and clinical significance of β-catenin and colorectal cancer stem cells marker EpCAM^(high)/CD44^+ in colorectal cancer

Objective： The aim of the study was to explore the role of Wnt βcatenin signalling pathway in the maintenance, invasion and metastasis of colorectal cancer stem cells. Methods： Double immunohistochemical staining was used to detect the expression of EpCAMhigh/CD44~ which is regarded as the marker of colorectal cancer stem cells in 80 cases of colorectal cancer and their corresponding liver metastases. The SP method of immunohistochemistry was used to detect the expression of the key protein βcatenin in the Wnt pathway in these tissue. The expression and correlation of ^-catenin and EpCAMh^gh/ CD44＋ in colorectal cancer were analyzed and their role on the biological behavior of colorectal cancer was explored. Results： The abnormal expression of βcatenin was significantly higher in colorectal cancer than in the paraneoplastic normal intes- tinal mucosa [55% （44/80） vs 10% （2/20）, P 〈 0.05]. The positive expression of EpCAMhigh/CD44＋ was significantly higher in colorectal cancer than in the paraneoplastic normal intestinal mucosa [66.25% （53/80） vs 0% （0/20）, P 〈 0.05]. In the 80 cases of colorectal cancer, the abnormal expression of ^-catenin has no correlation with gender （P = 0.079）, age （P = 0.416） and the magnitude （P = 0.816） of the tumor （P 〉 0.05）, but it was significantly correlated with degree of differentiation （P = 0.001）, depth of invasion （P = 0.001）, clinical stage （P = 0.000） and metastasis （P = 0.000）. In the colorectal cancer, the expression of EpCAMhi^h/CD44~ cells has no correlation with gender （P = 0.934） and the magnitude （P = 0.160） of the tumor （P 〉 0.05）, but was significantly correlated with age （P = 0.021）, degree of differentiation （P = 0.013）, depth of invasion （P = 0.000）, clinical stage （P = 0.000） and metastasis （P = 0.000）. In the corresponding liver metastases, we could also detecte EpCAMhih/CD44＋ cells. In cases with abnormal expression of βcatenin, the positive expression rate of EpCAMhigh/CD44＋ was significantly higher than those with normal expression of β-catenin （84.1% vs 44.4%）, and the difference was statistically significant （P 〈 0.05）. Conclusion： The abnormal activation of Wnt β-catenin signalling pathway may prompt the abnormal proliferation of the colorectal cancer stem cells, which leads to the recurrence and metastasis of the cancer.

立体定向大鼠脑内SHG-44人脑胶质瘤模型的建立

目的 :探讨建立大鼠 SHG- 4 4脑胶质瘤模型的方法。方法 :选用雄性 Wistar大鼠 ,接种前连续 3d用地塞米松 1 mg/ 1 0 0 g体重灌胃 ;在立体定向条件下选取大鼠脑右侧尾状核区为靶点 ,接种 2× 1 0 5个处于对数生长期的 SHG- 4 4细胞 ,接种后观察大鼠生长状态 ,分别于第 1、 2周进行核磁共振 (MRI)检查。在实验第 2周时解剖标本 ,行组织病理和 GFAP免疫组化检查。结果 :接种 1周后核磁共振检查 ,脑内形成实体瘤 ;HE染色组织病理证实是胶质瘤 ,GFAP免疫组化阳性 ;成瘤率约 6 0 %。结论 :用预先免疫抑制的方法 。

人脑胶质瘤细胞SHG-44照射后COX-2表达与放射敏感性的关系

目的:探讨人脑胶质瘤细胞SHG-44照射存活后代细胞对放射敏感性的变化,以及环氧合酶-2(COX-2)的表达情况,为临床使用COX-2抑制剂提供理论依据。方法:采用克隆形成率研究SHG-44及SHG-44照射存活后代细胞的放射敏感性;并分别用RT-PCR方法及免疫组化的方法检测COX-2基因mRNA和蛋白的表达情况。结果:与SHG-44细胞相比,SHG-44照射后代细胞对放射的敏感性下降,COX-2 mRNA及蛋白表达均升高。COX-2表达水平与SHG-44照射后代细胞放射敏感性呈负相关。结论:辐射诱导了SHG-44细胞COX-2表达的升高,使SHG-44照射后代细胞对放射敏感性下降,COX-2表达的升高可能是导致其辐射耐受的原因之一。

吴茱萸碱对胶质瘤SHG-44细胞凋亡的促进作用及其机制

目的研究吴茱萸碱对人胶质瘤SHG-44细胞凋亡的促进用及其机制。方法将体外培养的胶质瘤SHG-44细胞分为对照组、吴茱萸碱组(根据吴茱萸碱浓度不同分为3个亚组),CCK-8法观察细胞活性,流式细胞仪测定细胞凋亡率,Hoechst 33258核染色法观察细胞核凋亡,透射电镜观察细胞形态学的变化,Western blot法检测胶质瘤SHG-44细胞内Cleaved Caspase-3及Cleaved Caspase-9的蛋白表达。结果吴茱萸碱对胶质瘤SHG-44细胞增殖有抑制作用。流式细胞仪及Hoechst 33258核染色法的检测结果表明,随着吴茱萸碱作用浓度的递增,胶质瘤SHG-44细胞凋亡率呈剂量依赖性递增;吴茱萸碱作用后可使Cleaved Caspase-3及Cleaved Caspase-9蛋白的表达升高。结论吴茱萸碱通过改变Cleaved Caspase-3及Cleaved Caspase-9蛋白的表达,抑制胶质瘤SHG-44细胞增殖并促进其凋亡。

九节龙皂苷诱导胶质瘤SHG-44细胞凋亡及其机制研究

目的研究九节龙皂苷对胶质瘤SHG-44细胞潜在的治疗作用及其机制。方法用四甲基偶氨唑蓝(MTT)法检测不同剂量九节龙皂苷于不同时间(6、12、24、72h)对人胶质瘤SHG-44细胞活性的影响和细胞流式术检测SGH-44细胞调亡情况;Western-blot检测caspase-3和Bcl-2在SHG-44细胞中的表达情况。结果流式细胞仪检测显示,随着九节龙皂苷浓度的增大和时间延长,SHG-44细胞的凋亡率明显上升,Western-blot结果提示九节龙皂苷下调了凋亡抑制蛋白Bcl-2的表达并激活了凋亡蛋白caspase-3,九节龙皂苷明显抑制SHG-44细胞的生长与增殖。结论九节龙皂苷引起胶质瘤细胞大量凋亡,具有显著的抗肿瘤作用,通过调控caspase-3和Bcl-2诱导胶质瘤SHG-44细胞凋亡。

PTEN基因诱导人脑胶质瘤SHG-44细胞凋亡及bcl-2表达下调

目的 探讨 PTEN基因对人脑胶质瘤 SHG- 4 4细胞凋亡及凋亡相关基因 bcl- 2表达的影响 ,阐明 PTEN基因抑制肿瘤细胞增殖的机制 .方法 PTEN基因体外转染 SHG- 4 4细胞 ,筛选阳性细胞克隆 ,以原位杂交、免疫组化方法检测PTEN基因的表达情况 ;采用透射电镜、流式细胞仪和核DNA琼脂糖凝胶电泳检测细胞的凋亡情况 ;以免疫荧光法检测 bcl- 2基因的表达 .结果 PTEN基因转染的 SHG- 4 4细胞有 PTEN蛋白的表达 .透射电镜下可见转染 PTEN基因后细胞核染色质浓缩边集、胞质浓缩、核碎裂及凋亡小体形成等典型的凋亡表现 ;流式细胞仪显示细胞周期从 G1 期到 S期发生抑制 ,并且在 G1 期峰前出现一明显的凋亡峰 (15 .9% ) ;细胞核 DNA琼脂糖凝胶电泳呈现凋亡细胞特有的梯状条带 ;bcl-2的表达也显著下调 .结论 PTEN基因可诱导 SHG- 4 4细胞凋亡 ,并下调 bcl- 2蛋白的表达 ,这可能是

不同浓度五味子乙素对脑胶质瘤SHG-44细胞增殖的影响

脑肿瘤发病率目前在全球范围内呈上升趋势,脑肿瘤中以胶质瘤最为常见,恶性胶质瘤是肿瘤患者的第4位死亡原因,具有发病率、复发率、死亡率高和治愈率低＂三高一低＂的特点。研究表明,五味子具有保肝、抗肿瘤、抗氧化与抗衰老等作用,但目前对于五味子的研究都主要集中在保肝作用方面,

人脑胶质瘤细胞株SHG-44放射敏感性的测定方法探讨

目的探讨MTT法、细胞计数Kit8(CCK8)法在进行人脑胶质瘤细胞株SHG44细胞放射敏感性测定时能否替代集落形成试验法。方法运用3种方法分别在0、1、2、3、4、6、8、10Gy8个不同剂量点照射后检测其存活分数,采用统计学检查分析3种方法所得存活分数的相关性。结果在一定照射剂量内(照射剂量≤3Gy时),MTT法、CCK8法与克隆形成法相关性较好,而超出该剂量范围时的相关性差。经直线回归分析,3种方法均不存在高度线性相关。CCK8和MTT法相比无更好的相关性。结论集落形成法仍然是测定放射敏感性的“金标准”,MTT法等其他方法可以提供参考,但无法替代集落形成法。

芳香化酶在胶质母细胞瘤细胞系SHG-44细胞中的表达及调控

目的 研究芳香化酶细胞色素P45 0 (AROM)及雌激素受体 (ER α)在胶质母细胞瘤细胞系SHG 4 4细胞中的基因表达。 方法 细胞培养、免疫细胞化学染色、原位杂交染色及RT PCR技术。 结果 在SHG 4 4细胞中分别检测到AROM及ER α的表达 ,进一步发现SHG 4 4细胞中AROM的表达是由多个组织特异性启动子驱动基因的转录。 结论 可能为中枢神经系统肿瘤发生的激素调节提供新的资料。

人恶性胶质瘤细胞系CHG-5和SHG-44细胞骨架的比较研究

目的 比较人恶性胶质瘤细胞系CHG 5和SHG 44的细胞骨架系统成分 ,以进一步了解胶质瘤细胞骨架特征与其分化程度的关系。方法 利用免疫荧光细胞化学染色结合激光共聚焦显微镜观察两种细胞系微管、微丝、中间丝的含量及分布 ,并比较几种常用固定剂和缓冲液对各骨架成分检测的影响。结果 除微管和微丝均被染色外 ,所有细胞均表达Vimentin(波形蛋白 ) ,而只有部分细胞呈胶质纤维酸性蛋白 (GFAP)阳性。微管及中间丝在两种细胞胞质中的分布特征基本相似 ,但两种细胞微丝的定位、分布及荧光强度有一定差异。Vimentin型中间丝在SHG 44细胞中表达较强 ,而GFAP在CHG 5细胞中表达较强。丙酮、75 %酒精和4%多聚甲醛对微管的固定效果较好 ;用醛类固定后中间丝 (尤其是Vimentin)染色极弱。结论 CHG 5分化程度较SHG 44高 。