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1.
SYF2 is a putative homolog of human p29 in Saccharomyces cerevisiae. It seems to be involved in pre-mRNA splicing and cell cycle progression. Disruption of SYF2 leads to reduced α-tubulin expression and delayed nerve system development in zebrafish. Due to the potential of SYF2 in modulating microtubule dynamics in nervous system, we investigated the spatiotemporal expression of SYF2 in a rat sciatic nerve crush (SNC) model. We found that SNC resulted in a significant upregulation of SYF2 from 3 days to 1 week and subsequently returned to the normal level at 4 weeks. At its peak expression, SYF2 distributed predominantly in Schwann cells. In addition, upregulation of SYF2 was approximately in parallel with Oct-6, and numerous Schwann cells expressing SYF2 were Oct-6 positive. In vitro, we observed enhanced expression of SYF2 during the process of cyclic adenosine monophosphate (cAMP)-induced Schwann cell differentiation. SYF2-specific siRNA-transfected Schwann cells did not show significant morphological change in the process of Schwann cell differentiation. Also, we found shorter and disorganized microtubule structure and a decreased migration in SYF2-specific siRNA-transfected Schwann cells. Together, these findings indicated that the upregulation of SYF2 was associated with Schwann cell differentiation and migration following sciatic nerve crush.  相似文献   

2.
Cigarette smoking (CS) is considered one of the major risk factors to cause neurodegenerative disorders. Nicotine is the main chemical in CS which is responsible for dysfunction of the brain as a neuroteratogen. Also, nicotine dependency is a real mental illness and disease. Recently, chronic nicotine exposure has been shown to cause oxidative/nitrosative stress leading to a deleterious condition to cellular death in different brain regions. However, little is known about the effects of nicotine on mouse neural stem cells (mNSCs). The aim of this study is to investigate the effects of nicotine on mNSCs and elucidate underlying mechanisms involved in expression of a diversity of genes regulated by nicotine. When mNSCs were isolated from the whole brain of embryonic day 16 mice treated with nicotine at vehicle, 100, 400, and 800 μM for 5 d, nicotine significantly decreased the number and size of neurospheres. In immunocytochemistry, nicotine-exposed mNSCs expressing nestin showed the shortened filaments and condensed nuclei. In RT-PCR, messenger RNA (mRNA) levels of proliferating cell nuclear antigen (PCNA) and sirtuin1 (SIRT1) were significantly decreased, while the production of nitric oxide and mRNA levels of cyclooxygenase2 (COX-2), tumor necrosis factor-alpha TNF-α, and histone deacetylase 1 (HDAC1) were increased in a dose-dependent manner. In addition, sodium butyrate and valproic acid, HDAC inhibitors, partially rescue proliferation of mNSCs via inhibition of HDAC1 expression and NO production. Taken together, these data demonstrate that prolonged exposure of nicotine decreased proliferation of mNSCs by increased NO and inflammatory cytokine through increased HDAC1. Furthermore, this study could help in the development of a therapy for nicotine-induced neurodegenerative disorder and drug abuse.  相似文献   

3.
Human phosphatase and tensin homolog (hPTEN) gene was expressed in vascular smooth muscle cells (VSMCs) to study its effect on VSMC proliferation induced in platelet-derived growth factor (PDGF) conditioned medium. After G418 selection, MTT assay was conducted to examine transfected VSMC proliferation induced in human PDGF conditioned medium. We successfully constructed eukaryotic expression vector pcDNA4/myc-His-PTEN and transferred into VSMC cells. We report that in vitro proliferation of VSMC was inhibited in PTEN transfected VSMCs induced in PDGF conditioned medium. RT-PCR and Western blot results indicated significantly high levels of protein kinase B-PKB and nuclear factor kappa B mRNA and protein, respectively, in PDGF group as compared with the control group.  相似文献   

4.
STAT3 pathway plays an important role in the growth of diffuse large B-cell lymphoma (DLBCL) cells. Here we investigated the antitumor activity of Quercetin, a flavonoid compound, in combination with rituximab in DLBCL cell lines in vitro. We found that Quercetin synergistically enhanced rituximab-induced growth inhibition and apoptosis in DLBCL cell lines. Moreover, we found Quercetin exerted inhibitory activity against STAT3 pathway and downregulated the expression of survival genes. These results suggest that combining the Quercetin with rituximab may present an attractive and potentially effective way for the treatment of DLBCL.  相似文献   

5.
Esophageal squamous cell carcinoma (ESCC) is one of the most malignant tumors. The aim of this study was to investigate the biology characteristics of ESCC by analyzing microRNA and mRNA expression profile. We used BRB-array tools to analyze the deregulated microRNA and mRNA between esophageal squamous cell carcinomas and paired normal adjacent tissues. We used miRTrail and protein–protein interaction methods to explore the related pathways and networks of deregulated microRNA and mRNA. By combining the results of pathways and networks, we found that the deregulated microRNA and their deregulated target mRNA are enriched in the following pathways: DNA replication, cell cycle, ECM-receptor interaction, focal adhesion, mismatch repair, and pathways in cancer. The results showed that many deregulated microRNAs and mRNAs may play a vital role in the pathogenesis of ESCC, and the systems biology approach is very helpful to explore molecular mechanism of ESCC.  相似文献   

6.
The tumor suppressor PTEN is a lipid phosphatase that is found mutated in different types of human cancers. PTEN suppresses cell proliferation by inhibiting the PI3K-Akt signaling pathway at the cell membrane. However, PTEN is also demonstrated to localize in the cell nucleus where it exhibits tumor suppressive activity via a different, unknown mechanism. In this study we report that PTEN also localizes to the nucleolus and that nucleolar PTEN plays an important role in regulating nucleolar homeostasis and maintaining nucleolar morphology. Overexpression of nuclear PTEN in PTEN null cells inhibits Akt phosphorylation and reduces cell size. Knockdown of PTEN in PTEN positive cells leads to nucleolar morphologic changes and an increase in the proportion of cells with a greater number of nucleoli. In addition, knockdown of PTEN in PTEN positive cells increased ribosome biogenesis. These findings expand current understanding of function and relevance of nuclear localized PTEN and provide a foundation for the development of novel therapies targeting PTEN.  相似文献   

7.
Interferon gamma (IFN-γ) is an important immunoregulatory cytokine that has a central role against viral and bacterial infections. In this study, the cDNA encoding 141 amino acids of mature IFN-γ from mice splenocytes was cloned in a prokaryotic expression vector pQE 30. Optimization of expression conditions resulted in high IFN-γ protein. Western blot showed that recombinant IFN-γ was specifically recognized by its counterpart anti-mouse IFN-γ antibodies. In vitro dose-dependent studies, with A549 and HeLa cell lines, showed that cloned IFN-γ was safe and had no effect on cell proliferation. The protein prediction and analysis using SOPMA program, revealed that IFN-γ had 80 α-helices, 8 β-turns jointed by 9 extended strands and 44 random coils. A total of four major clusters were observed with murine IFN-γ sharing 39 % homology with human IFN-γ. Pair-wise alignment studies with human revealed 26 % identity and 43.3 % similarity. The recovery of bioactive proteins from inclusion bodies (IBs) is a complex process and various protocols have been developed. We report here a simple, robust and inexpensive purification approach for obtaining recombinant IFN-γ protein expressed as IBs in E.coli.  相似文献   

8.
Cinnamoyl-CoA reductase (CCR, EC 1.2.1.44), which catalyzes the reduction of cinnamoyl-CoA esters to their respective cinnamaldehydes, is considered as a key enzyme in lignin formation. The substrates of CCR, cinnamoyl-CoA esters, are products of 4-Coumarate-CoA ligase (4CL, EC 6.2.1.12), which is an enzyme upstream of CCR. The PtCCR and Pt4CL were isolated from Populus tomentosa and expressed in E. coli. Results showed that 4CL can catalyze the conversion of hydroxycinnamic acids to cinnamoyl-CoA esters, with high efficiency. The purification of esters using SPE cartridges suggested that 40 % methanol with 0.1 M of acetic acid was the optimal elution buffer for cinnamoyl-CoA esters. The optimization of prokaryotic expression demonstrated that the best expression conditions for recombinant PtCCR was 6 h of 0.4 mM IPTG induction at 37 °C. PtCCR enzyme assay illustrated that the recombinant protein can catalyze the reduction of cinnamoyl-CoA esters. Kinetics analysis showed that feruloyl-CoA has higher affinity to PtCCR with faster reaction speed (Vmax), indicating that feruloyl-CoA was the most favorable substrate for PtCCR catalysis. The recombinant protein was expressed in E. coli, purified through affinity column chromatography, and characterized by SDS-PAGE. SPE cartridges were used to purify the ester products of the Pt4CL reaction. HPLC-MS was used to analyze the structure of esters and evaluate their purity or quantity. Furthermore, the enzyme activity of recombinant CCR to feruloyl-CoA at different pHs indicated that compartmentalization may be an important factor in lignin monomer formation.  相似文献   

9.
Suppression of the activity of pro-apoptotic Bcl-2-family proteins frequently confers chemoresistance to many human cancer cells. Using subcellular fractionation, the ER calcium (Ca++) channel inhibitor dantrolene and small interfering RNA (siRNA) against Bax or Bak, we show that the new synthetic bichalcone analog TSWU-CD4 induces apoptosis in human cancer cells by releasing endoplasmic reticulum (ER)-stored Ca++ through ER/mitochondrial oligomerization of Bax/Bak. Blockade of the protein kinase RNA-like ER kinase or the unfolded protein response regulator glucose-regulated protein 78 expression by siRNA not only suppressed oligomeric Bax/Bak-mediated pro-caspase-12 cleavage and apoptosis but also resulted in an inhibition of Bcl-2 downregulation induced by TSWU-CD4. Induction of the ER oligomerization of Bax/Bak and apoptosis by TSWU-CD4 were suppressed by Bcl-2 overexpression. Inhibition of lipid raft-associated phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling by TSWU-CD4 induced ER stress- and oligomeric Bax/Bak-mediated apoptosis, which were substantially reversed by overexpression of the wt PI3K p85α subunit. Taken together, these results suggest that suppression of lipid raft-associated PI3K/Akt signaling is required for the ER stress-mediated apoptotic activity of Bax/Bak, which is responsible for the ability of TSWU-CD4-treated cancer cells to exit the ER-mitochondrial apoptotic cell death pathway.  相似文献   

10.
The need for glioma biomarkers with improved sensitivity and specificity has sparked research into short non-coding RNA known as microRNA (miRNA). Altered miRNA biogenesis and expression in glioma plays a vital role in important signaling pathways associated with a range of tumor characteristics including gliomagenesis, invasion, and malignancy. This review will discuss current research into the role of miRNA in glioma and altered miRNA expression in biofluids as candidate biomarkers with a particular focus on glioblastoma, the most malignant form of glioma. The isolation and characterization of miRNA using cellular and molecular biology techniques from the circulation of glioma patients could potentially be used for improved diagnosis, prognosis, and treatment decisions. We aim to highlight the links between research into miRNA function, their use as biomarkers, and how these biomarkers can be used to predict response to therapy. Furthermore, increased understanding of miRNA in glioma biology through biomarker research has led to the development of miRNA therapeutics which could restore normal miRNA expression and function and improve the prognosis of glioma patients. A panel of important miRNA biomarkers for glioma in various biofluids discovered to date has been summarized here. There is still a need, however, to standardize techniques for biomarker characterization to bring us closer to clinically relevant miRNA-based diagnostic and therapeutic signatures. A clinically validated biomarker panel has potential to improve time to diagnosis, predicting response to treatment and ultimately the prognosis of glioma patients.  相似文献   

11.
Temozolomide (TMZ) is an alkylating agent that is widely used in chemotherapy for cancer. A key mechanism of resistance to TMZ is the overexpression of O6-methylguanine-DNA methyltransferase (MGMT). MGMT specifically repairs the DNA O6-methylation damage induced by TMZ and irreversibly inactivates TMZ. Regulation of MGMT expression and research regarding the mechanism of TMZ resistance will help rationalize the clinical use of TMZ. In this review, we provide an overview of recent advances in the field, with particular emphasis on MGMT structure, function, expression regulation, and the association between MGMT and resistance to TMZ.  相似文献   

12.
Receptors for advanced glycation end-products (RAGE) are members of the immunoglobulin superfamily of cell-surface receptors implicated in mechanisms of pulmonary inflammation. In the current study, we test the hypothesis that RAGE mediates inflammation in primary alveolar macrophages (AMs) exposed to diesel particulate matter (DPM). Quantitative RT-PCR and immunoblotting revealed that RAGE was up-regulated in Raw264.7 cells, an immortalized murine macrophage cell line and primary AMs exposed to DPM for 2 h. Because DPM increased RAGE expression, we exposed Raw264.7 cells and primary AMs isolated from RAGE null and wild-type (WT) mice to DPM prior to the assessment of inflammatory signaling intermediates. DPM led to the activation of Rat sarcoma GTPase (Ras), p38 MAPK and NF-κB in WT AMs and, when compared to WT AMs, these intermediates were diminished in DPM-exposed AMs isolated from RAGE null mice. Furthermore, cytokines implicated in inflammation, including IL-4, IL-12, IL-13 and TNFα, were all significantly decreased in DPM-exposed RAGE null AMs compared to similarly exposed WT AMs. These results demonstrate that diesel-induced inflammatory responses by primary AMs are mediated, at least in part, via RAGE signaling mechanisms. Further work may show that RAGE signaling in both alveolar epithelial cells and resident macrophages is a potential target in the treatment of inflammatory lung diseases exacerbated by environmental pollution.  相似文献   

13.
Recently, it was found that α-Calcitonin gene-related peptide (CGRP) was associated with breast cancer metastases, but the role of CGRP in interaction between breast cancer and osteoblast during bone metastases is not clear. Here, we investigated the effect of CGRP on osteoblast in co-culture system with breast cancer. Using a breast cancer–osteoblast co-culture system, we chose MDA-MB-231 for breast cancer and human cell line MG-63 for osteoblast. CGRP was added to this co-culture system. The expression levels of the Runx2, RANK1, and osteoprotegerin (OPG) were analyzed using real-time PCR and western blot. CGRP receptors were investigated by immunofluorescence. We found that breast cancer cells cause osteolysis lesions by upregulating Runx2 expression, decreasing OPG expression, and increasing RANKL expression in osteoblasts. Our data prove that CGRP can regulate osteoclast coupling genes in osteoblast by increasing OPG, and decreasing RANKL and Runx2 expressions in a time-dependent manner; and inhibit those osteolytic factors induced by interaction between breast cancer cells and osteoblast. This inhibition could be abolished by the CGRP antagonist, CGRP8–37. In conclusion, calcitonin receptor-like receptor is the key player for CGRP’s effect in this co-culture system.  相似文献   

14.
This study aimed to evaluate the immunolocalization and messenger RNA (mRNA) expression for transforming growth factor-beta (TGF-β) and its receptors (TGF-βRI and RII), as well as mRNA expression for P450 aromatase and FSH receptor in caprine preantral follicles. The effects of TGF-β, FSH alone, or in association on the in vitro follicular development were also assessed. Immunohistochemical analyses showed the expression of TGF-β and its receptors in oocytes of all follicle stages and granulosa cells of primary and secondary follicles. mRNA for TGF-β receptors and for FSH receptor (FSHR) was present in preantral follicles as well as in oocytes and granulosa cells of antral follicles. Isolated secondary follicles were cultured in α-minimum essential medium (MEM) alone or supplemented with either FSH (100 ng/ml), TGF-β (10 ng/ml), or TGF-β + FSH for 18 d. TGF-β increased significantly oocyte diameter when compared to FSH alone and control. After 18 d of culture, all groups showed a significant reduction in P450 aromatase and FSHR mRNA levels in comparison to fresh control. In contrast, treatment with FSH significantly increased the mRNA expression for TGF-β in comparison to fresh control and other treatments. In conclusion, the findings showed that TGF-β and its receptors are present in caprine ovarian follicles. Furthermore, they showed a positive effect on oocyte growth in vitro.  相似文献   

15.
16.
Hyperpolarization-activated cyclic-nucleotide-gated cation nonselective (HCN) channels are involved in the pathology of nervous system diseases. HCN channels and γ-aminobutyric acid (GABA) receptors can mutually co-regulate the function of neurons in many brain areas. However, little is known about the co-regulation of HCN channels and GABA receptors in the chronic ischemic rats with possible features of vascular dementia. Protein kinase A (PKA) and TPR containing Rab8b interacting protein (TRIP8b) can modulate GABAB receptors cell surface stability and HCN channel trafficking, respectively, and adaptor-associated kinase 1 (AAK1) inhibits the function of the major TRIP8b-interacting protein adaptor protein 2 (AP2) via phosphorylating the AP2 μ2 subunit. Until now, the role of these regulatory factors in chronic cerebral hypoperfusion is unclear. In the present study, we evaluated whether and how HCN channels and GABAB receptors were pathologically altered and investigated neuroprotective effects of GABAB receptors activation and cross-talk networks between GABAB receptors and HCN channels in the hippocampal CA1 area in chronic cerebral hypoperfusion rat model. We found that cerebral hypoperfusion for 5 weeks by permanent occlusion of bilateral common carotid arteries (two-vessel occlusion, 2VO) induced marked spatial and nonspatial learning and memory deficits, significant neuronal loss and decrease in dendritic spine density, impairment of long-term potentiation (LTP) at the Schaffer collateral-CA1 synapses, and reduction of surface expression of GABAB R1, GABAB R2, and HCN1, but increase in HCN2 surface expression. Meanwhile, the protein expression of TRIP8b (1a-4), TRIP8b (1b-2), and AAK1 was significantly decreased. Baclofen, a GABAB receptor agonist, markedly improved the memory impairment and alleviated neuronal damage. Besides, baclofen attenuated the decrease of surface expression of GABAB R1, GABAB R2, and HCN1, but downregulated HCN2 surface expression. Furthermore, baclofen could restore expression of AAK1 protein and significantly increase p-PKA, TRIP8b (1a-4), TRIP8b (1b-2), and p-AP2 μ2 expression. Those findings suggested that, under chronic cerebral hypoperfusion, activation of PKA could attenuate baclofen-induced decrease in surface expression of GABAB R1 and GABAB R2, and activation of GABAB receptors not only increased the expression of TRIP8b (1a-4) and TRIP8b (1b-2) but also regulated the function of TRIP8b via AAK1 and p-AP2 μ2, which restored the balance of HCN1/HCN2 surface expression in rat hippocampal CA1 area, and thus ameliorated cognitive impairment.  相似文献   

17.
Decades of research have been devoted to defining the role of GABAergic transmission in nociceptive processing. Much of this work was performed using rigid, orthosteric GABA analogs created by Povl Krogsgaard-Larsen and his associates. A relationship between GABA and pain is suggested by the anatomical distribution of GABA receptors and the ability of some GABA agonists to alter nociceptive responsiveness. Outlined in this report are data supporting this proposition, with particular emphasis on the anatomical localization and function of GABA-containing neurons and the molecular and pharmacological properties of GABAA and GABAB receptor subtypes. Reference is made to changes in overall GABAergic tone, GABA receptor expression and activity as a function of the duration and intensity of a painful stimulus or exposure to GABAergic agents. Evidence is presented that the plasticity of this receptor system may be responsible for the variability in the antinociceptive effectiveness of compounds that influence GABA transmission. These findings demonstrate that at least some types of persistent pain are associated with a regionally selective decline in GABAergic tone, highlighting the need for agents that enhance GABA activity in the affected regions without compromising GABA function over the long-term. As subtype selective positive allosteric modulators may accomplish these goals, such compounds might represent a new class of analgesic drugs.  相似文献   

18.
This study was aimed to elucidate the roles of inhibition of related JAK/STAT pathways in regulating cytotoxicity induced by cisplatin in non-small-cell lung cancer (NSCLC) cell. We treated five non-small-cell lung cancer cell lines with cisplatin alone or with cisplatin and Jak2 inhibitor (ruxolitinib) and assessed cell viability, expression of Jak2 and STAT3 and cell apoptosis. We also investigated the effect of combination treatment inhibited tumor xenograft growth in two human NSCLC xenograft models bearing the cisplatin resistant (H1299) and sensitive (A549) cells. Different cell lines with different genetic background showed half-maximal inhibitory concentrations (IC50) of cisplatin from 4.66 to 68.28 µmol/L. They could be divided into cisplatin intrinsic resistant and cisplatin sensitive cell lines. In cisplatin-resistant cells with higher Jak2 and STAT3 expression, cisplatin and ruxolitinib combination dramatically suppressed the cell growth, down-regulated the expression of phosphorylated STAT3 and induced cleaved caspase-3 expression. Moreover combination with cisplatin and ruxolitinib also significantly inhibited the growth of resistant cell H1299, A549/DDP and H2347 in soft agar model. Finally, combination group significant inhibited the tumor growth and induced the caspase-3 expression compared with either single agent alone (P < 0.05) on the resistant cell xenografts model. The present study indicates that further study is warranted to determine the effectiveness of combination treatment with cisplatin and Jak2/stat3 pathway inhibitor for platinum-resistant NSCLC.  相似文献   

19.
Lung cancer is a worldwide leading cause of cancer-related death. The aim of this study was to identify target genes and specific biomarkers for identification and treatment of different types of lung cancer with DNA microarray. Gene expression profile GSE6044 and miRNA microarray profile GSE17681 were downloaded from Gene Expression Omnibus database. The differentially expressed genes (DEGs) and miRNAs were screened with multtest package in R language. Then, functional enrichment analysis of identified DEGs was performed. Furthermore, the verified target genes based on screened miRNAs were selected from miRTarBase and miRecords databases. Then miRNA-target gene regulation network was constructed. APOE, CDC6 and ATP2B1were involved in most of the functions obtained for adenocarcinomas, small cell lung cancer and squamous cell carcinomas, respectively. The target DEGs of differentially expressed hsa-miR-29a included FGG in adenocarcinoma, RAN and COL4A1 in small cell lung cancer, GLUL in squamous cell carcinoma. The target DEGs of has-miR-7 were SNCA and SLC7A5 in adenocarcinoma and small cell lung cancer, respectively. ICAM1 and KIT were the target DEGs of hsa-miR-222 in adenocarcinoma and squamous cell carcinoma. The miRNAs and their differentially expressed target genes have the potential to be used in clinic for diagnosis and treatment of different kinds of lung cancer in the future.  相似文献   

20.
Polo-like kinase 4 (PLK4) is a centrosomal protein that is involved in the regulation of centrosome duplication. This study aimed to determine whether the genetic abnormality of PLK4 is involved in human gastric cancer. First, we examined the status of PLK4 mRNA expression in 7 gastric cancer cell lines and 48 primary gastric cancers using an RT-PCR analysis. The upregulation of PLK4 mRNA expression was detected in 57.1 % (4/7) of the gastric cancer cell lines, and a novel PLK4 variant with exon 4, but without exon 5, was identified. In the primary gastric cancers, the upregulation of PLK4 mRNA expression in the cancerous cells was detected in 50.0 % (24/48) of the cases, and this upregulation was statistically significant (P value = 0.0139). Next, we established AGS gastric cancer cells capable of inducibly expressing PLK4 using the piggyBac transposon vector system and showed that PLK4 overexpression induced centrosome amplification and chromosome instability using immunofluorescence and FISH analyses, respectively. Furthermore, PLK4 overexpression suppressed primary cilia formation. Our current findings suggested that PLK4 is upregulated in a subset of primary gastric cancers and that PLK4 overexpression induces centrosome amplification and chromosome instability and causes the suppression of primary cilia formation.  相似文献   

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