Inhibition of Ubiquitin-specific Peptidase 8 Suppresses Adrenocorticotropic Hormone Production and Tumorous Corticotroph Cell Growth in AtT20 Cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Two recent whole-exome sequencing researches identifying somatic mutations in the ubiquitin-specific protease 8 (USP8) gene in pituitary corticotroph adenomas provide exciting advances in this field. These mutations drive increased epidermal growth factor receptor (EGFR) signaling and promote adrenocorticotropic hormone (ACTH) production. This study was to investigate whether the inhibition of USP8 activity could be a strategy for the treatment of Cushing's disease (CD).</p><p><b>METHODS</b>The anticancer effect of USP8 inhibitor was determined by testing cell viability, colony formation, apoptosis, and ACTH secretion. The immunoblotting and quantitative reverse transcription polymerase chain reaction were conducted to explore the signaling pathway by USP8 inhibition.</p><p><b>RESULTS</b>Inhibition of USP8-induced degradation of receptor tyrosine kinases including EGFR, EGFR-2 (ERBB2), and Met leading to a suppression of AtT20 cell growth and ACTH secretion. Moreover, treatment with USP8 inhibitor markedly induced AtT20 cells apoptosis.</p><p><b>CONCLUSIONS</b>Inhibition of USP8 activity could be an effective strategy for CD. It might provide a novel pharmacological approach for the treatment of CD.</p>

Effect of hypoxia on the expressions of hypoxia-inducible factor-1α,glucocorticoid receptor and opiomelanocortin genes in AtT-20 cells / 中华神经医学杂志

Objective To investigate the effect of CoCl2-induced hypoxia on the expressions of hypoxia-inducible factor-1α(HIF-lα),glucocorticoid receptor(GR)and opiomelanocortin(POMC)in AtT-20 cells.Methods AtT-20 cells were exposed to different concentrations(25,50,100,and 200 μmol/L)of CoCl2 to induce hypoxia.MTT assay was employed to assess the changes in the cell proliferation after the exposure,and real-time PCR and Western blotting were performed to detect the expressions of HIF-1α,GR and POMC genes at both the mRNA and protein levels.Results Compared with the cells in normoxic conditions,the AtT-20 cells exposed to CoCl2 treatment at 25,50,and 100 μmol/L for 24 and 48 h showed obviously enhanced cell proliferation.CoCl2 significantly increased the mRNA and protein expressions of HIF-1α,GR and POMC genes in a dose-dependent manner(P＜0.05).Conclusion CoCl2-induced hypoxia up-regulates the mRNA and protein expressions of HIF-1α,GR and POMC genes in AtT-20 cells.

Construction and characterization of mutants in the neurofibromatosis type 2 tumor suppressor gene / 中华神经医学杂志

Objective To construct the eukaryotic expression vector of mutants in the neurofibromatosis type 2 (NF2) tumor suppressor gene and observe its expression in rat schwannoma cell line RT4. Methods Site-directed mutagenesis was performed to induce the mutation of the codons for the residue Ile 546 in pEGFP-N1-NF2 into Met to construct the muton of pEGFP-N1-NF2~(△Ⅱe546Met). After lipofectin-mediated transient transformation of RT4 with the plasmids containing the mutation and the one without mutation, respectively, the mRNA and protein expression levels of NF2 were determined using fluorescence imaging and Western blotting. The cell proliferation was determined by the MTT assay. Results DNA sequence analysis confirmed the success of site-directed mutagenesis and Western blotting showed that pEGFP-NF2 protein could be expressed in the RT4 cells. The RT4 cell inhibition rate in the pEGFP-N1-NF2~(△Ⅱe546Met) transfection group was statistically lower than that in the pEGFP-N1-NF2 transfected group (P<0.05).Conclusion The recombinant plasmids pEGFP-N1-NF2~(△Ⅱe546Met) has been successfully constructed with efficient expressions in RT4.

Expression of free fatty acid receptor GPR40 in adult monkey hippocampus after ischemia / 中华神经医学杂志

Objective To investigate the expression of free fatty acid receptor GPR40 and evaluate the possible function of GPR40 in the adult monkey hippocampus after ischemia. Methods According to the post-ischemic adult monkey model of Yamashima, a total of 24 adult monkeys were randomly divided into 4 groups: sham-operated group (control) and post-ischemic day 4, 9, 15 (d4, d9,d15). The expression of free fatty acid receptor GPR40 was detected at the protein level by immunoblotting and immunohistochemistry in the adult monkey hippocampus. Results Immunoblotting analysis showed the expression of GPR40 was decreased in CA1 and increased in DG after ischemia (posfischemic group vs control, P＞0.05). Immunohistochemistry data revealed that the double stained cells of GPR40 and NeuN were also decreased by 50% (d15) in CA1 and had no significant changes in DG after ischemia. Interestingly, the co-labeled cells of GPR40 and GFAP were increased 2.5 folds (d4) in post-ischemic SGZ. Conclusions There is the different expression of GPR40 in adult monkey hippocampal CA1 and DG regions after ischemia. Co-labeled cells of GPR40 and GFAP are increased in post-ischemic SGZ, which indicates that polyunsaturated free fatty acid such as DHA, a ligand of GPR40, may alleviate neuronal injury in post-ischemic hippocampus.

Double electrodes simultaneous stimulation and implantation technique in deep brain stimulation / 中华创伤杂志(英文版)

Posttraumatic tremor is often one of the causes of disability in head injury patients. Usually, pharmacotherapy for this type of tremor is not effective. Since early 1970s, surgical ablation of the ventral thalamus has been used to treat various types of tremor. Nowadays, deep brain stimulation (DBS) confirms its efficacy in alleviating different forms of tremor, including posttraumatic tremor. Such therapy has been reported achieving around 80% success rate in the treatment of posttraumatic tremor. These successful results suggest that the application of DBS therapy can be considered as one of the alternative treatments for minimizing the tremor occurring from different pathologies.

Loss of heterozygosity on chromosome 22 in sporadic schwannoma and its relation to the proliferation of tumor cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Schwannoma is the tumor arising mainly from the cranial and spinal nerves. Bilateral vestibular schwannoma is the hallmark of neurofibromatosis type 2 (NF2). The NF2 gene has been cloned with comprehensive analysis of its mutations in schwannoma. However, most studies focused on vestibular schwannoma. There are differences in proliferation of tumor cell and ultrastructure between vestibular and spinal schwannomas. It is unknown whether genetic alterations in vestibular schwannoma are different from those in non-vestibular schwannoma. We analyzed the loss of heterozygosity (LOH) on chromosome 22 in patients with sporadic schwannoma including vestibular and spinal schwannomas and correlated this genetic alteration with tumor proliferation.</p><p><b>METHODS</b>In 54 unrelated patients without clinical NF1 or NF2, 36 patients had sporadic vestibular schwannoma, and 18 dorsal spinal root schwannoma. Four highly polymorphic linkage to NF2 gene microsatellite DNA markers (D22S264, D22S268, D22S280, CRYB2) were used to analyze LOH. The proliferative index was evaluated by Ki-67 and proliferative cell nuclear antigen (PCNA) immunostaining. Student's t test was used to analyze the difference of the proliferative index between schwannoma with LOH and that without LOH. The difference of the frequency of LOH in vestibular and spinal schwannomas was investigated by the chi-square test.</p><p><b>RESULTS</b>Twenty-three schwannomas (42.6%, 23/54) showed allele loss. The frequency of LOH in vestibular schwannoma was significantly higher than that in spinal schwannoma (chi2 = 5.14, P < 0.05). The proliferative index of schwannoma with LOH was significantly higher than that without LOH (tki-67 = 2.97, P = 0.0045; tPCNA = 2.93, P = 0.0051).</p><p><b>CONCLUSIONS</b>LOH on chromosome 22 is a frequent event in the tumorigenesis of sporadic schwannoma. And, there is a correlation between LOH on chromosome 22 and proliferative activity in schwannoma. The frequency of LOH in vestibular schwannoma is significantly different from that in spinal schwannoma.</p>

Immunohistochemical study in dural arteriovenous fistula and possible role of ephrin-B2 for development of dural arteriovenous fistula / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Although there were several clinical and experimental studies discussing the pathogenesis of dural arteriovenous fistula (DAVF), the pathological process leading to intracranial DAVF so far remains unknown. In this study, we investigated the expression of vascular growth factors in order to elucidate the possible role of these factors for the development of DAVF and to study the biological activity of this uncommon lesion.</p><p><b>METHODS</b>We examined the histological features, proliferative and angiogenic capacities of the tissue specimens obtained from 6 patients who underwent surgery at our institution. Immunohistochemical staining for vascular endothelial growth factor (VEGF), its receptors Flk-1 and Flt-1, ephrin-B2, MIB-1 and proliferating cell nuclear antigen (PCNA) was performed using standard immunohistochemical techniques.</p><p><b>RESULTS</b>A positive immunostaining was found for all antibodies studied except MIB-1, whereas nuclear endothelial expression of PCNA was observed in only 3/6 cases. VEGF stained positive in all of the available specimens (6/6). Flk-1 showed a positive immunoreaction in only 2/6 cases and Flt-1 in 4/6 cases. Ephrin-B2 was expressed in the majority (5/6) of the cases.</p><p><b>CONCLUSIONS</b>These results support the hypothesis that DAVFs might be acquired dynamic vascular malformations with low biological activity. Vascular growth factors like VEGF and ephrin-B2 might play a pivotal role in the formation of DAVF.</p>

Relationship of survival rate, hematoma thickness and midline shift in patients with acute subdural hematomas / 中华外科杂志

<p><b>OBJECTIVE</b>To estimate outcomes of patients with acute subdural hematomas by analysing the hematoma thickness, midline shift and the differences between them.</p><p><b>METHODS</b>Ninety-five patients with acute subdural hematoma were retrospectively studied by calculating hematoma thickness, midline shift and their difference with a statistical analysis of Kaplan-Meier, Wilcoxon-Mann-Whitney U test.</p><p><b>RESULTS</b>The hematoma thickness ranged from 5.0 to 40.0 mm and midline shift was from 0 to 35.0 mm. Among these patients, 51% died and 49% survived after surgery. 18 patients (19%) showed good or satisfactory results. Kaplan-Meier analysis proved that the survival for patients with hematoma thickness approximately equal to l7 mm and a midline shift 15 mm or whose midline shift exceeded hematoma thickness by 2.2 mm, the survival rate was 50%. Glasgow outcome scale scores were correlated significantly with these parameters.</p><p><b>CONCLUSION</b>The hematoma thickness, midline shift and their difference provided a database from which criteria could be derived, that is crucial for prognosis estimation.</p>