RESUMO
Objective:To investigate the differences of integral alpha 3 (ITGA3) mRNA and protein expressions in gliomas of different grades and different cell lines, and gliomas tissues of different clinical and molecular characteristics, and evaluate their prognostic values in brain glioma patients.Methods:(1) ITGA3 mRNA expression data in the brain gliomas and clinical data of these glioma patients were obtained from The Cancer Genome Atlas (TCGA) and Chinese Glioma Genome Atlas (CGGA). The differences of ITGA3 mRNA expressions in glioma patients with different gender, ages, WHO grading, isocitrate dehydrogenase 1 ( IDH) mutation statuses, and 1p/19q co-deletion statuses were compared. Kaplan-Meier method was used to plot and compare the survival curves of patients with ITGA3 mRNA high expression and ITGA3 mRNA low expression. Receiver operating characteristic (ROC) curve was used to analyze the predictive efficiency of ITGA3 mRNA expression in survival rate of gliomas. Univariate and multivariate Cox regression analyses were used to explore the independent influencing factors for prognoses in glioma patients. The independent influencing factors for prognosis were used to construct nomograms and the calibration diagram was used to verify the reliability of nomograms in predicting the prognoses of these patients. (2) Intracellular localization of ITGA3 and ITGA3 protein expression in low- and high-grade gliomas were determined by on-line database of Human Protein Atlas (HPA). (3) Brain glioma cells U87, U118, U251 and human astrocytes SVG were cultured in vitro, and the ITGA3 mRNA and protein expressions in cells were detected by Western blotting and reverse transcription (RT)-PCR, respectively. Results:(1) In TCGA database, the ITGA3 mRNA expressions in gliomas of WHO grading II, III and IV increased successively, with significant differences (P<0.05). In CGGA database, the ITGA3 mRNA expression in glioma of WHO grading IV was statistically higher than that in glioma of WHO grading II and III ( P<0.05). In TCGA and CGGA databases, the ITGA3 mRNA expressions in glioma patients aged ≤40 years and >40 years, patients with IDH wild-type and IDH mutation, and patients with chromosome 1p/19q deletion and chromosome 1p/19q non-deletion were statistically different ( P<0.05). The survival rate of patients with low ITGA3 mRNA expression was significantly higher than that of patients with high ITGA3 mRNA expression, no matter in low-grade glioma, glioblastoma, or entire glioma samples ( P<0.05). ROC curve showed that, in TCGA database, the area under the curve (AUC) of ITGA3 mRNA in predicting 1, 3, and 5 years survival was 0.791, 0.786, and 0.708 in glioma patients; in CGGA database, the AUC of ITGA3 mRNA in predicting 1, 3, and 5 years survival was 0.661, 0.667, and 0.659. Multivariate Cox regression analysis showed that, in TCGA database, age, WHO grading, IDH mutation, chromosome 1p/19q deletion and ITGA3 mRNA expression ( HR=1.018, 95%CI: 1.006-1.030, P=0.0.003) were independent influencing factors for prognoses of glioma patients ( P<0.05); and in CGGA database, WHO grading, IDH mutation, chromosome 1p/19q deletion, and ITGA3 mRNA expression ( HR=1.445, 95%CI: 1.132-1.844, P=0.003) were independent influencing factors for prognoses of glioma patients ( P<0.05). Nomograms showed that age had the greatest influence in survival, followed by ITGA3 mRNA expression. Calibration plots showed that nomogram was reliable in predicting 1-, 3-, and 5-year survival in glioma patients. (2) Immunofluorescence localization showed that ITGA3 protein mainly aggregated in cell membrane and vesicles. Immunohistochemical staining showed that the ITGA3 protein expression in high-grade glioma tissues was obviously higher than that in low-grade glioma tissues. (3) The results of RT-PCR and Western blotting revealed that the ITGA3 mRNA and protein expressions in glioma cell lines U87, U118 and U251 were significantly higher than those in SVG cells ( P<0.05). Conclusion:The ITGA3 mRNA and protein expression levels are correlated with the malignant degrees of glioma; patients with ITGA3 mRNA low expression tend to have a high overall survival; ITGA3 mRNA expression can be used as an index to evaluate the prognoses of glioma patients.
RESUMO
Objective To detect the down-regulation ofmiRNA-99b expression in cell invasion and its mechanism in human glioma cell line U251.Methods Glioma cell line U251 were routinely cultured in vitro.(1) U251 cells were divided into blank control group,negative control group and miRNA-99b inhibitor group;cells in the latter two groups were transfected with negative control sequences and miRNA-99b inhibitors,respectively;and cells in the blank control group did not give any treatment;mRNA expressions of miRNA-99b and mammalian target of rapamycin (mTOR) in U251 cells were measured by reverse transcription (RT)-PCR;the changes of mTOR,eIF4E-binding protein 1 (4E-BP1) andphosphorylated (p)-4E-BPl protein expressions in U251 cells were detected by Westem blotting;cell invasion was evaluated by Transwell assay.(2) U251 cells were divided into negative control group Ⅰ and mTOR siRNA group,and cells in the two groups were transfected with negative control sequences and mTOR siRNA,respectively;the miRNA-99b and mTOR mRNA expressions in U251 cells were measured by RT-PCR;the mTOR and p-4E-BP1 protein expressions in U251 cells were measured by Western blotting.(3) U251 cells were divided into miRNA-99b inhibitor+negative control group and miRNA-99b inhibitor+mTOR siRNA group,and cells in the two groups were transfected with miRNA-99b inhibitor+negative control sequences and miRNA-99b inhibitor+mTOR siRNA,respectively;the p-4E-BP1 protein expression in U251 cells was measured by Western blotting;cell invasion was evaluated by Transwell assay.Results (1) As compared with those in the blank control group and negative control group,the miRNA-99b rnRNA expression was significantly decreased,the mTOR mRNA and protein expressions and p-4E-BP1 protein expression were significantly increased,and the number of transmembrane cells was significantly larger in U251 cells of miRNA-99b inhibitor group (P＜0.05);there were no significant differences in 4E-BP1 protein expression among the three groups (P＞0.05).(2) As compared with those in the negative control group Ⅰ,the mTOR mRNA and protein expressions and p-4E-BP1 protein expression were significantly decreased in U251 cells of mTOR siRNA group (P＜0.05);there was no significant difference in miRNA-99b mRNA expression between the two groups (P＞0.05).(3) As compared with those in the miRNA-99b inhibitor+negative control group,the p-4E-BP1 protein expression and number of transmembrane cells were significantly decreased/smaller in U251 cells ofmiRNA-99b inhibitor+mTOR siRNA group (P＜0.05).Conclusions Down-regulation ofmiRNA-99b expression promotes glioma cell invasion,and its mechanism is related to the regulation of mTOR/4E-BP1 signaling pathway.
RESUMO
Objective To study the effect of soluble amyloid precursor protein α (sAPPα) on nerve cell apoptosis and neurological function in subarachnoid hemorrhage (SAH) rats.Methods Sixty male SD rats were randomly divided into control group (n=20),SAH+saline group (n=20) and SAH+sAPPα group (n=20).A SAH model was established by injecting autologous blood into cistern magna in rats.After a SAH model was established for SAH + saline group and SAH + sAPPα group by injecting saline and sAPPα respectively into the cistern magna of rats,the apoptotic cells were detected by immunofluorescene with TUNEL staining and the neurological function was scored in 10 rats from each group on day 3 after injection of sAPPα and saline.Results The number of apoptotic cells in brain tissue was significantly greater in SAH+saline group than in control group (P＜0.05) and was significantly smaller in SAH+sAPPα group than in SAH+ saline group (P＜0.05).The neurological function score was 26.7±0.5,13.9±0.7 and 23.0±0.8 respectively in control group,SAH + saline group and SAH + sAPPα group.Conclusion sAPPα alleviates secondary damage of neurological function by inhibiting the apoptosis of nerve cells in rats after SAH and can thus improve their neurological function.
RESUMO
Objective To identify the reasons and treatment strategies of epidural fluid collection (EFC) secondary to cranioplasty in patients with traumatic brain injury after decompressive craniectomy.Methods From June 2013 to July 2017,a retrospective analysis was performed on clinical data of 150 patients with traumatic brain injury after decompressive craniectomy in our hospital.A total of 47 patients experienced EFC following cranioplasty and 103 not.Risk factors of EFC after cranioplasty were analyzed by multiple factor Logistic regression.Results For the 47 EFC patients,32 patients had no obvious clinical symptoms and EFC was absorbed gradually through conservative therapy;15 patients had clinical symptoms,such as mental deterioration,headache,or limb weakness.EFC disappeared through vacuation in 4 patients and subcutaneous drainage in 11.The proportions of patients with skull defect＞80 cm2,dural defect and dural calcification in patients with EFC were significantly higher as compared with those without EFC (P＜0.05).Multiple factor Logistic regression analysis showed that skull defect＞80 cm2 and dural mater calcification were independent risk factors for EFC after cranioplasty.Conclusions Patients with large skull defect＞80 cm2 and dural calcification are prone to have EFC after cranioplasty.Careful evaluation of imaging data,good surgical skills and strengthening postoperative management can reduce incidence of EFC after cranioplasty.
RESUMO
Objective To investigate whether the monitoring of cerebral blood flow can be used to evaluate the successful preparation of rat models with focal cerebral ischemia.Methods With the line plug inserted into the left internal cerebral artery of 30 SPF Wistar Han rats at (16.0±0.5) mm,(18.0±0.5) mm and (20.0±0.5) mm respectively,three kinds of focal cerebral ischemia models (10 each) were prepared.All the rats were divided into incomplete occlusion group,complete occlusion group and deep occlusion group according to whether the blood clots were found in skull base and the infarction was appeared in the middle cerebral artery territory after the whole brain was stained with 2,3,5-triphenyl tetrazolium chloride.The cerebral blood flow in the middle cerebral artery territory of each animal was monitored by laser Doppler at the time of before and after blocking with the line plug inserted and reperfusion with the line plug pulled out and recorded for statistical analysis.The cerebral blood flow was expressed as the relative flow unit (perfusion unit,PU);The changes of the cerebral blood flow after occlusion and reperfusion were expressed as a percentage of that blood flow and those before occlusion.Results During the process of modeling,1 rats died,and the other in the incomplete occlusion group (n=9),in the complete occlusion group (n=15),and in the deep occlusion group (n=5).The depths of inserting of 8 rats in the incomplete occlusion group were about (16±0.5) mm,and the blood flow from the anterior cerebral artery to middle cerebral artery could not be prevented completely,the Longa score was evaluated 6 h after ischemia and ranged from 0 to 1,there were no blood clots in skull base and no infarction after TTC staining.The depths of inserting of 9 rats in the complete occlusion group were about (18±0.5) mm,and the anterior cerebral artery blood flow was completely blocked,the neurological dysfunction was significant after 6 h ischemia and the Longa score was ranged from 2 to 3,the blood clots in skull base were absent and the infarctions in middle cerebral artery territory were obvious after TTC staining.The depth of inserting of 5 rats in the deep occlusion group were about (20±0.5) mm,and the anterior cerebral artery blood flow was also completely blocked,the neurological dysfunction was serious after 6 h ischemia and the Longa score was ranged from 3 to 4,there existed blood clots in skull base and obvious infarctions in middle cerebral artery territory after TTC staining.The cerebral blood flows after the line plug inserted compared with those before occlusion in the incomplete occlusion group,complete occlusion group and deep occlusion group were all decreased (94±17 vs.256±36,43±9 vs.286±44,44±6 vs.294±46,respectively,all P0.05);The percentages of cerebral blood flow after occlusion and before were (36.93±0.06)%,(15.09±0.02)%,(15.52±0.04)% respectively,and the difference between groups was statistically significant (F=39.14,P<0.01).The cerebral blood flow after reperfusion were 213±31,147±17,96±14,respectively,and the difference was also statistically significant (F=50.05,P<0.01),and the cerebral blood flow in deep occlusion group was less than that in complete occlusion group(P<0.05);The percentages of cerebral blood flow after reperfusion and before occlusion were (83.10±0.02)%,(51.83±0.05)%,(33.49±0.09)% respectively,and the difference was also statistically significant (F=93.23,P<0.01).Conclusion The cerebral blood flow monitored by laser Doppler can be used as a real-time,convenient,micro invasive,objective and reliable standard to evaluate the successful preparation of rat MCAO models with line plug method.
RESUMO
Objective To detect the expression of micro RNA (miR)-433 and histone deacetylase 6 (HDAC6) in glioma tissues and investigate the effect of miR-433 on cell proliferation and invasion of human glioma cell line U251. Methods (1) Forty-two glioma samples, collected from patients accepted surgical resection and conformed by pathology in our hospital from January 2010 to December 2014, and 13 healthy brain tissues, collected from patients accepted surgery for craniocerebral trauma at the same time period, were used in our study; reverse transcription (RT)-PCR was used to detect the mRNA expression levels of miRNA-433 and HDAC6 in the glioma samples and brain tissues. (2) Human glioma cell line was routinely cultured and divided into blank control group, nonsense sequence control group and miRNA-433 mimics group;cells in the later two groups were transfected with nonsense sequences or miRNA-433 mimics, and cells in the blank control group did not give any treatment;the mRNA expression levels of miRNA-433, P21 and HDA C6 in these 3 groups were detected by RT-PCR;the cellular viability was measured by CCK-8 assay;flow cytometry was used to monitor the changes of cell cycle and apoptosis; cell invasion was evaluated by Transwell assay; HDAC6 protein expression was detected by Western blotting. (3) Wide-type (WT)HDAC63'-UTR and mutant type (MUT)HDAC63'-UTR luciferase report vectors were established; miR-433 mimics+WT HDAC63′-UTR and nonsense sequences+WT HDAC63'-UTR were transfected into the U251 cells, and dual-luciferase experiment was used to detect the fluorescence intensity of the cells; miR-433 mimics+MUT HDAC63'-UTR and nonsense sequences+MUT HDAC63'-UTR were transfected into the U251 cells, and dual-luciferase experiment was used to detect the fluorescence intensity of the cells. (4) U251 cells were divided into nonsense sequence control group, HDAC6 expression plasmids group and HDAC6 siRNA group, and nonsense sequences, HDAC6 expression plasmids or HDAC6 siRNA were transfected respectively; RT-PCR was used to detect the P21 and HDAC6 mRNA expressions and miRNA-433 expression; U251 cells were divided into miR-433 mimics group and miR-433 mimics+HDAC6 expression plasmids group, and miR-433 mimics or miR-433 mimics+HDAC6 expression plasmids were transfected, respectively, and one-5 d after that, CCK-8 was used to detect the cellular viability. Results (1) The miRNA-433 expressions gradually increased and HDA C6 mRNA expressions gradually decreased in the high-grade gliomas, low-grade gliomas and normal brain tissues, and significant differences were noted among each two groups (P<0.05);the miRNA-433 expression was negatively correlated with HDA C6 mRNA expression in the glioma tissues (r=0.829, P=0.000). (2) As compared with blank control group and nonsense sequence control group, miRNA-433 mimics group had significantly higher miRNA-433 and P21 mRNA expressions, cell percentage at G0/G1 stage, and apoptotic rate (P<0.05), and had statistically lower HDAC6 mRNA expression, cellular viability on 2-5 d of culture, number of transmembrane cells and HDAC6 protein expression (P<0.05). (3) The luciferase activity in cells from miR-433 mimics+WT HDAC63'-UTR group was significantly lower as compared with that in the nonsense sequences+WT HDAC63'-UTR group (P<0.05);the luciferase activity in cells from miR-433 mimics+MUT HDAC63'-UTR group and nonsense sequences+MUT HDAC63'-UTR group showed no significant differences (P>0.05). (4) The HDA C6 mRNA expressions were gradually increased, and P21 mRNA expressions were gradually decreased in the HDAC6 siRNA group, nonsense sequence control group, and HDAC6 expression plasmids group, with significant differences (P<0.05);on 2-5 d of culture, the cellular viability in the miR-433 mimics+HDAC6 expression plasmids group was significantly higher than that in the miR-433 mimics group (P<0.05). Conclusions The miRNA-433 expression level is low in human glioma tissues;miRNA-433 over-expression may inhibit the cell activity and promote cell apoptosis of glioma cell line U251 in vitro via inhibiting the HDAC6 expression.
RESUMO
Objective:To observe the effect of Xingnaojing injection in the treatment of severe traumatic brain injury .Methods:From August 2013 to December 2014 , 110 patients with severe traumatic brain injury were randomly divided into the control group and the research group with 55 ones in each .The control group was treated with the conventional treatment , including removal of intracrani-al hematoma, bone disc decompression according to surgical indications , anti-infection, dehydration intracranial pressure reduction and nutritional support treatment .The research group was treated with Xingnaojing injection additionally .After the one-week, two-week and 4-week treatment , the GCS score of the two groups were compared .The rate of lung infection and stress ulcer in 4 weeks were com-pared.The clinical outcome of the two groups after the 4-week treatment was also evaluated .Results:The GCS score of the research group was higher than that of the control group after the 2-and 4-week treatment (P<0.05).There was no significant difference in the incidence of stress ulcer between the two groups , while the incidence of pulmonary infection in the research group was lower than that in the control group (P<0.05).The total effective rate of clinical prognosis in the research group (90.9%) was significantly higher than that in the control group (72.7%, P<0.05).Conclusion:Xingnaojing injection combined with the conventional treatment for the patients with severe traumatic brain injury can decrease the clinical complications and improve the clinical symptoms of patients , which is worthy of clinical promotion .
RESUMO
Objective To explore the effect of sodium valproate (SV) on reducing high-sensitivity C-reactive protein (hsCRP) and attenuating cerebrovascular spasm damage in rats after subarachnoid hemorrhage (SAH).Methods Seventy-two male rats,weighting 300-400 g,were randomized to following experimental groups:sham-operated group,SAH group,SAH+saline treatment group,and SAH+SV treatment group (n=18).The SAH models in the later three groups were induced by injection of autologous blood into the cistern magna.Saline or SV (2 mg/100 g) was given to the rats in the SAH+saline treatment group and SAH+SV treatment group every day via intraperitoneal injection.Serum hsCRP level was measured on 1,3,5 and 10 day.Neurological deficit scale scores were assessed on 3 and 5 day.Results On 1,3,5 and 10 day,HsCRP level in the sham-operated group was (0.09± 0.02) mg/L,(0.09±0.02) mg/L,(0.09±0.02) mg/L and (0.09±0.01) mg/L;that in SAH group was (0.29± 0.01) mg/L,(0.32±0.02) mg/L,(0.35±0.02) mg/L and (0.32±0.02) mg/L;that in SAH+saline treatment group was (0.28±0.02) mg/L,(0.31 ±0.02) mg/L,(0.34±0.02) mg/L and (0.31 ±0.02) mg/L;that in SAH+SV treatment group was (0.15 ±0.02) mg/L,(0.21 ±0.02) mg/L,(0.24±0.02) mg/L and (0.15 ±0.03) mg/L;HsCRP level in the SAH group and SAH+saline treatment group was significantly higher than that in the sham-operated group (P＜0.05);HsCRP level in the SAH+SV treatment group was significantly increased as compared with that in the sham-operated group,but significantly decreased as compared with that in the SAH+saline treatment group and SAH group (P＜0.05).The neurobehavior scale scores on 3 and 5 day in SAH+SV treatment group (23.0±0.8 and 21.8±1.4) were significantly increased as compared with those in the SAH group (14.1±0.8 and 11.9±0.9) and SAH+saline treatment group (13.9± 0.7 and 11.1±1.4,P＜0.05);those in the SAH+SV treatment group (23.0±0.8 and 21.8±1.4) were significantly decreased as compared with that in the sham-operated group,but significantly increased as compared with that in the SAH+saline treatment group and SAH group (P＜0.05).Conclusion SV decreases the inflammatory injury by reducing the hsCRP level and improve the neurological outcome in SAH rat models.
RESUMO
Objective To discuss the feasibility of neuroendoscopic third ventriculostomy for chronic posttraumatic hydrocephalus (PTH).Methods Nineteen cases of chronic PTH treated with neuroendoscopic third ventriculostomy between October 2010 and October 2014 were analyzed retrospectively.There were 13 males and 6 females, aged 11-57 years (mean, 36.3 years).Trauma resulted from traffic accidents in 14 cases, falls in 4 cases and blunt object hitting in 1 case.Of the 19 cases analyzed, 5 had Glasgow Coma Scale (GCS) score of 13-15, 5 had score of 9-12 and 9 had score of 5-8 at admission.Results of operation were assessed with the Canada multicenter evaluation criteria.Prognosis was analyzed with the Glasgow Outcome Scale (GOS).Results All cases were followed up for mean 13.6 months (range, 6-26 months).Improvement of symptoms was achieved in 17 cases, but was not seen in 2 cases.Of the 2 cases, one required ventriculoperitoneal shunt two weeks after ineffective ventriculostomy, and one required second ventriculostomy one month after the presence of stoma blockage.No serious complications occurred.At follow-up, 9 cases had GOS score of 5, 8 cases had score of 4 and 2 cases had score of 3.Conclusions Neuroendoscopic third ventriculostomy is in line with the physical characteristics in cerebrospinal fluid circulation, which implies no shunt implantation, less operative trauma and less complications.The procedure is an effective approach for chronic PTH.
RESUMO
Objective To explore the surgical treatment of posttraumatic epilepsy in functional cerebral area Methods After preoperative evaluation,nineteen patients with intractable posttraumatic epilepsy of functional cerebral area underwent multiple subpial transaction under the electrocorticogram monitoring during the operation, combining with anterior corpus callosotomy, anterior temporal lobectomy and selective amygdalo-hippocampectomy. Results The follow-up results showed seizures of these patients improved considerably. According to Engel' s grading,among 19 cases,10 cases were grade Ⅰ ,7 cases were grade Ⅱ ,2 cases were grade Ⅲ ,and no cases were grade Ⅳ. Conclusions The satisfactory clinical outcome of the surgical treatment of intractable posttraumatic epilepsy in functional cerebral area could be obtained with the help of careful preoperative evaluation and multiple subpial transaction under the electrocorticogram monitoring.