miR-423-5p enhances chemoresistance of glioblastomas cells to temozolomide by regulating PDCD5 / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To investigate the miR-423-5p expression in brain glioma tissues and cell lines, and its promotive effect on temozolomide (TMZ) chemoresistance by targeting PDCD5 (programmed cell death protein 5). Methods: Tumor tissues and matched peritumoral tissues were collected from 20 brain glioma patients who were surgically treated in the Department of Neurosurgery, Affiliated Hospital of Beihua University between January 2017 and December 2018. Glioblastoma cell lines (U251, U87, SHG-44) and human normal glial cell line HMC-3 were also used in the study. The relative expression of miR-423-5p and PDCD5 in brain glioma and peritumoral tissues and cell lines was detected by qPCR. The synthesized miR-423-5p mimics and miR-NC were respectively transfected into U251 and U87 cells; meanwhile, TMZ at different concentrations (50, 100, 150 and 200 μmol/L) were also used to treat the cells. Then, the chemoresistance of cells to TMZ were determined. MTT assay and colony formation assay were used to examine the proliferation of U251 and U87 cells, andWestern blotting was used to detect the expression of c-caspase 3, Bcl-2 and PDCD5 proteins in U251 and U87 cells. The targeting relationship between PDCD5 and miR-423-5p was validated through Dual luciferase reporter gene assay. Results: miR-423-5p was highly expressed in glioma tissues and glioma cell lines (all P<0.01). As compared with the miR-NC group, the proliferation and TMZ-chemoresistance of U251 and U87 cells in miR-423-5p mimics group significantly increased (all P<0.01). Dual luciferase reporter gene assay validated that miR-423-5p could bind with PDCD5 3' UTR to suppress the expression of PDCD5. Conclusion: High expression of miR-423-5p enhances the chemoresistance of glioma cells to TMZ, and miR-423-5p may serve as a potential therapeutic target in the treatment of brain glioma.

Over-expression of programmed cell death 5 gene enhances chemosensitivity of brain glioma cells to temozolomide / 中国肿瘤生物治疗杂志

@# Objective: To explore the role of tumor suppressor gene programmed cell death 5 gene (PDCD5) in the growth and temozolomide (TMZ) sensitivity of brain glioma cells. Methods:Atotal of 116 patients with cerebral glioma admitted to the Department of Neurosurgery, First Clinical Hospital of Jilin University from January 2009 to December 2014 were enrolled in this study. QPCR, WB and immunohistochemistry method were used to detect the mRNAand protein expressions of PDCD5 in glioma cell lines (U87, U251), U87 cell line with stable PDCD5 expression (U87-PDCD5), glioma cells with si-PDCD5 transfection and primary cerebral glioma tissues, respectively. MTT assay was used to detect the effect of over-expression or knockdown of PDCD5 on the growth and TMZ-sensitivity of glioma cells. The subcutaneous tumor-bearing model of glioma cell line U87 was established in nude mice, and then the experimental mice were randomly divided into control group, TMZ group, PDCD5 group and TMZ+exogenous PDCD5 recombinant expression vector group.After 20 days, the animals were sacrificed by cervical dislocation and the tumor tissue was excised to measure the tumor volume and weigh. The expression of PDCD5 in tumor tissues was detected by qPCR and WB methods, and the effects of PDCD5 combined with TMZ on the growth of gliomas were also analyzed. Results: The relative mRNA and protein expressions of PDCD5 in U87 cells were significantly lower than those in U251 cells (both P<0.05), and the mRNA and protein expressions of PDCD5 in high level glioma tissues were significantly lower than those in low level tissues (all P<0.05). The sensitivity of U87-PDCD5 cells and U251 cells to TMZ was higher than that of U87 cells (all P<0.05). The sensitivity of cells to TMZ in U87-PDCD5-siRNA group and U251siRNA group was significantly lower than that of the control group (both P<0.05). The tumor volume and weigh to fnudemicexenografts were compared,and the results showed control group>TMZ group>PDCD 5group>combined group(allP<0.05);however, the mRNA and protein expressions of PDCD5 in the transplanted tumor tissues of each group showed the opposite trend (all P<0.05). Conclusion: PDCD5 over-expression can enhance the chemosensitivity of braingliomato the chemotherapy drug TMZ, while silencing of PDCD5 expression exertsthe opposite effect.The combination of PDCD5 and TMZ can better inhibit the growth of xenografts in nude mice.

Expression and location of IL-17A, E, F and their receptors in colorectal adenocarcinoma: Comparison with benign intestinal disease.

The research aimed to investigate secretion, expression and location of IL-17 relative ligands, IL-17 relative receptors, infiltrating inflammatory cells and parenchymal structural cells in colorectal cancer (CRC) compared with ulcerative colitis (UC) and benign hyperplastic polyp. 29 human intestinal tissues with CRC, 17 with UC and 7 with polyp were stained using immunohistochemistry to evaluate immunoreactivity for IL-17 family relative ligands including IL-17A, E, F and their respective relative receptors such as IL-17RA, IL-17RB and IL-17RC. At the same time the infiltration of inflammatory cells including lymphocytes, phagocytes, mast cells and neutrophils and parenchymal structural cell changes involving vascular endothelial cells and CD90+ fibroblast cells were also evaluated using the same methods The immunoreactivity or positive inflammatory cells of all the sections were analyzed using professional image analysis software to determine statistical significance. The immunoreactivity for IL-17A, IL-17RA, IL-17E, IL-17RB and IL-17F showed significant decrease in CRC tissue when compared to UC (p = 0.00001. respectively). The reduction of above IL-17 relative ligands and receptors was accompanied by an obvious decrease in the number of infiltrating neutrophils and mast cells in CRC (p = 0.00001 and p = 0.007, respectively) but accompanied by a marked increase of CD31+ blood vessels (p = 0.001). The immunoreactivity of IL-17A, IL-17RA, IL-17E, IL-17RB and IL-17F and the numbers of infiltrating neutrophils and mast cells showed significant decrease in CRC tissues when compared to those in polyp (p < 0.05). In contrast, the immunoreactivity of IL-17RC and the numbers of CD3+ 1ymphocytes were elevated in CRC when compared with those in polyp (p = 0.0001, p = 0.007, respectively). In CRC tissues, positive correlations between IL-17A, IL-17RA with CD68+ macrophages were observed respectively (r = 0.621, p = 0.0001; r = 0.75, p = 0.0001). IL-17 cytokine family including ligands and their corresponding receptors were secreted and expressed by infiltrating inflammatory cells. Not only infiltrating lymphocytes but also increased blood endothelial cells were relative significantly to genesis and progression of CRC.

Immune analysis of expression of IL-17 relative ligands and their receptors in bladder cancer: comparison with polyp and cystitis.

BACKGROUND: Bladder cancer, cystitis and bladder polyp are the most common urinary system diseases all over the world. Our former research results show that IL-17A and IL-17 F contribute to the pathogenesis of benign prostatic hyperplasia (BPH) and prostate cancer (Pca) while IL-17E interacting with IL-17RB might have an anti-tumor effect. RESULTS: Using imunohistochemistry, we systemically compared immunoreactivity of ligands (IL-17A, E and F) and receptors (IL-17RA, IL-17RB and IL-17RC) of IL-17 family, infiltration of inflammatory cells and changes of structural cells (fibroblast cells, smooth muscle and vascular endothelial cells) in sections of bladder tissues from subjects with bladder cancer, cystitis and bladder polyp. Compared with subjects with cystitis, immunoreactivity for IL-17A, IL-17 F and IL-17RC was significantly elevated in the group of bladder cancer (p < 0.01), while immunoreactivity of IL-17E, IL-17RA and IL-17RB, and the infiltrating neutrophils were decreased (p < 0.05). The numbers of infiltrating lymphocytes and phagocytes and CD31+ blood vessels and immunoreactivity of CD90+ fibroblasts were also elevated in patients with bladder cancer compared with those of cystitis. The patterns of IL-17 ligands and receptors, and inflammatory cells and structural cells varied in cystitis, bladder polyp and bladder cancer. In bladder cancer, immunoreactivity of IL-17E and IL-17 F was positively correlated with smooth muscles and lymphocytes, respectively. In addition, immunoreactivity of IL-17A and IL-17E was positively correlated with their receptors IL-17RA and IL-17RB respectively. CONCLUSIONS: The data suggest that changed patterns of expression of the IL-17 cytokine family ligands and receptors might be associated with infiltration of inflammatory cells and structural cells (CD90+ fibroblasts and CD31+ blood vessels), which might also contribute to occurrence and development in bladder cancer.

[Knockdown of STAT3 reduces the level of survivin and promotes the apoptosis of U87MG glioma cells].

Objective To investigate the effect of signal transducer and activator of transcription (STAT3)-specific siRNA (siSTAT3) on the apoptosis of U87MG human glioma cells and its mechanism. Methods U87MG glioma cells were cultured in vitro. The effect of siSTAT3 on U87MG human glioma cells was evaluated by MTT assay to observe cell proliferation and by flow cytometry and acridine orange staining to observe cell apoptosis. Reverse transcription PCR and Western blotting were used to detect the expression of survivin. Results Compared with mock and si-scrambled groups, siSTAT3 expression plasmid inhibited the proliferation of U87MG cells obviously. Flow cytometry and acridine orange staining showed that siSTAT3 plasmid significantly promoted glioma cell apoptosis. In addition, siSTAT3 plasmid significantly inhibited the mRNA and protein levels of survivin. Conclusion Knockdown of STAT3 in U87MG cells can downregulate the level of survivin and remarkably promote the apoptosis of glioma cells.

High expression of VEGF and PI3K in glioma stem cells provides new criteria for the grading of gliomas.

Glioma is a type of tumor derived from glial cells, which is associated with a high level of incidence and mortality. At present, the generation of a fast and efficient method to evaluate the malignancy grade of glioma is required. Cancer stem cells (CSCs) are currently attracting attention in oncological studies; therefore, the present study aimed to investigate novel biomarkers of glioma CSCs, in order to provide new criteria for the grading of glioma. The mRNA expression levels of CD133, (sex determining region Y)-box 2, nestin, vascular endothelial growth factor (VEGF) and phosphoinositide-3-kinase (PI3K) were detected in 15 human samples of high-malignancy glioma and 12 human samples of low-malignancy glioma in vitro. The mRNA expression levels of VEGF and PI3K were higher in the high-malignancy group, as compared with in the low-malignancy group. In conclusion, the mRNA expression levels of VEGF and PI3K in glioma CSCs may be considered a novel criteria for the grading of glioma.

Expression of IL-17A, E, and F and their receptors in human prostatic cancer: Comparison with benign prostatic hyperplasia.

BACKGROUND: Benign prostatic hyperplasia (BPH) and prostate cancer (PCa) are the most common urological diseases in elderly men. Although studies suggest the cytokine family might be associated with BPH and PCa, there has been no systematic comparisons of expression of IL-17A, E, F and their receptors, infiltration of inflammatory cells, and changes in structural cells in PCa and BPH. METHODS: Immunohistochemistry was employed to evaluate immunoreactivity for IL-17A, E, F and their receptors IL-17RA, IL-17BR, and IL-17CR, infiltration of inflammatory cells, and changes in structural cells including endothelial cells, fibroblasts, and smooth muscle cells in prostate tissues from subjects with PCa or BPH as well as controls. RESULTS: Immunostaining showed that expression of immunoreactivity for IL-17A, IL-17RA, IL-17E, and IL-17F was significantly elevated in prostatic tissue from BPH and PCa compared with that in controls, which was accompanied by increased numbers of infiltrating inflammatory cells and CD31(+) blood vessels. Compared with BPH, PCa was characterized by reduced immunoreactivity for IL-17BR and reduced numbers of CD68(+) macrophages, fibroblasts, and smooth muscle cells, although there was a trend for these changes to correlate with disease severity in both PCa and BPH. CONCLUSION: Our data are compatible with hypothesis that IL-17A acting through IL-17RA, but not IL-17CR contribute to the pathogenesis of BPH and PCa. In contrast, IL-17E interacting with the IL-17BR might have an anti-tumor effect.

Survivin small interfering RNA suppresses glioblastoma growth by inducing cellular apoptosis.

A survivin small interfering RNA sequence specific for a human and mouse homogenous sequence was constructed. Survivin small interfering RNA could significantly inhibit glioma cell proliferation and induce apoptosis when it was transfected into either a human glioma cell line U251 or rat glioma C6 cells in vitro. In addition, treatment of rat orthotopic glioma models with survivin small interfering demonstrated the inhibition of glioma growth in vivo. Our experimental findings suggest that the use of RNA interference techniques to target the survivin sequence may be useful in the treatment of glioma.

Survivin-specific small interfering RNAs enhance sensitivity of glioma U-87MG cells to paclitaxel by promoting apoptosis.

A survivin siRNA expression vector was transfected into glioma U-87MG cells and these cells were then treated with paclitaxel. The results showed that survivin-specific siRNA combined with paclitaxel treatment synergistically inhibited glioma U-87MG cell proliferation and promoted apoptosis. This treatment also inhibited the expression of the cell cycle regulatory proteins, survivin, cyclinD1, c-Myc and CDK4 and enhanced the sensitivity of U-87MG cells to paclitaxel.

[Expressions of survivin and GRIM-19 in prostate cancer].

OBJECTIVE: To investigate the expressions of survivin and GRIM-19 in prostatic cancer tissue and their clinical implications. METHODS: We detected the expressions of survivin and GRIM-19 in the tissues of normal prostate (NP), benign prostate hyperplasia (BPH) and prostate cancer (PCa) using immunohistochemical staining, RT-PCR and Western blot, and processed the data by SPSS12. RESULTS: The positive rates of survivin expression were 6.25% , 18.18% and 90.62% in NP, BPH and PCa (P < 0.01), while those of GRIM-19 were 87.50%, 81.82% and 9.37% , respectively (P < 0.01). Semiquantitative RT-PCR and immunohistochemical staining showed that both survivin mRNA and survivin expressions were highly positive in PCa but negative in NP and BPH. Western blot exhibited that the survivin protein was expressed strongly in PCa but weakly in NP and BPH, while the GRIM-19 protein was expressed just contrariwise (P < 0.01). CONCLUSION: The expressions of survivin and GRIM-19 may be closely correlated with the pathogenesis of prostate cancer.

Automatic identification of the reference system based on the fourth ventricular landmarks in T1-weighted MR images.

RATIONALE AND OBJECTIVES: The reference system based on the fourth ventricular landmarks (including the fastigial point and ventricular floor plane) is used in medical image analysis of the brain stem. The objective of this study was to develop a rapid, robust, and accurate method for the automatic identification of this reference system on T1-weighted magnetic resonance images. MATERIALS AND METHODS: The fully automated method developed in this study consisted of four stages: preprocessing of the data set, expectation-maximization algorithm-based extraction of the fourth ventricle in the region of interest, a coarse-to-fine strategy for identifying the fastigial point, and localization of the base point. The method was evaluated on 27 Brain Web data sets qualitatively and 18 Internet Brain Segmentation Repository data sets and 30 clinical scans quantitatively. RESULTS: The results of qualitative evaluation indicated that the method was robust to rotation, landmark variation, noise, and inhomogeneity. The results of quantitative evaluation indicated that the method was able to identify the reference system with an accuracy of 0.7 +/- 0.2 mm for the fastigial point and 1.1 +/- 0.3 mm for the base point. It took <6 seconds for the method to identify the related landmarks on a personal computer with an Intel Core 2 6300 processor and 2 GB of random-access memory. CONCLUSION: The proposed method for the automatic identification of the reference system based on the fourth ventricular landmarks was shown to be rapid, robust, and accurate. The method has potentially utility in image registration and computer-aided surgery.

Stereotactic localization and visualization of the subthalamic nucleus.

BACKGROUND: The subthalamic nucleus (STN) is widely recognized as one of the most important and commonly targeted nuclei in stereotactic and functional neurosurgery. The success of STN surgery depends on accuracy in target determination. Construction of a digitalized atlas of STN based on stereotactic MRI will play an instrumental role in the accuracy of anatomical localization. The aim of this study was to investigate the three-dimensional (3D) target location of STN in stereotactic space and construct a digitalized atlas of STN to accomplish the visualization of the STN on stereotactic MRI, thus providing clinical guidance on the precise anatomical localization of STN. METHODS: One hundred and twenty healthy people volunteered to be scanned by 1.5 Tesla MRI scanning with 1-mm-thick slice in the standard stereotactic space between 2005 and 2006. One adult male was selected for 3D reconstruction of STN. The process of 3D reconstruction included identification, manual segmentation, extraction, conservation and reconstruction. RESULTS: There was a significant correlation between the coordinates and age (P < 0.05). The volume of left STN was significantly larger than the right STN, and there was a significant negative correlation between volume and age (P < 0.05). The surface of the STN nucleus after 3D reconstruction appeared smooth, natural and realistic. The morphological feature of STN on the individual brain could be visualized directly in 3D. The 3D reconstructed STN could be rotated, zoomed and displayed at any direction in the stereotactic space. The anteroposterior diameter of the STN nucleus was longer than the vertical and transverse diameters in 3D space. The 3D reconstruction of STN manifested typical structure of the "dual lens". CONCLUSIONS: The visualization of individual brain atlas based on stereotactic MRI is feasible. However, software for automated segmentation, extraction and registration of MR images need to be further developed.