Favorable neuroprotective effect of intra-arterial application of edaravone dexborneol in ischemic stroke rats.

OBJECTIVE: The aim of this study was to investigate the neuroprotective effects of intra-arterial administration of edaravone dexborneol in rats with acute ischemic stroke and determine the optimal dose. MATERIALS AND METHODS: Firstly, 120 male Sprague-Dawley rats (265-300 g) were selected to establish ischemic stroke models and were randomly divided into groups of sham-operation (Sham group), cerebral ischemia-reperfusion (IS group), permanent focal ischemia (PI group) and treatment (2MG group: 2 mg/kg, 4MG group: 4 mg/kg, 6MG group: 6 mg/kg) groups. There are 20 rats in each group, and ten rats in each group were randomly selected for Longa score and 2,3,5-triphenyl tetrazolium chloride staining to observe the changes in neurological function and the proportion of cerebral infarct volume in each group. Secondly, the remaining ten rats in each group were scored for Longa and tested for free radicals (hydroxyl radical; peroxynitrite; nitric oxide) and pro-inflammatory cytokines (interleukin 6; interleukin-1ß; tumor necrosis factor-α). We monitored changes in the indicators in each group of rats. RESULTS: There were no significant differences among the enrolled Sprague-Dawley rats concerning age, sex, and feeding conditions. Edaravone dexborneol could significantly reduce the cerebral levels of hydroxyl radical, interleukin 6, interleukin-1ß, tumor necrosis factor-α, and their behavioral scores of acute ischemic stroke rats after a single dose in the carotid artery. The results suggested that 4 mg/kg might be an appropriate dose. CONCLUSIONS: A single intra-arterial administration of edaravone dexborneol can improve neurobehavioral function and alleviate cerebral injury in acute ischemic stroke rats through anti-inflammatory and free radical scavenging effects.

Effect of the CYP2D6 gene polymorphism on postoperative analgesia of tramadol in Han nationality nephrectomy patients.

OBJECTIVE: Tramadol is a synthetic opioid which has analgesic efficacy in the postoperative pain. It is metabolized by polymorphic enzyme cytochrome P450 (CYP2D6). Patients with different CYP2D6 genotypes would have different responses to tramadol in pain relief. The CYP2D6*10 allele is the most common allele in a Chinese population. The aim of this study was to evaluate whether the different CYP2D6*10 genotypes have an effect on the postoperative tramadol analgesia in the Chinese population after elective nephrectomy. METHODS: One hundred and twenty patients after performed elective nephrectomy were enrolled in this study after being approved by the local Ethics Committee. The patients were given patient-controlled analgesia (PCA) which included 10 mg/ml tramadol after receiving a loading dose of 100 mg tramadol and 1 mg granisetron intravenously. Blood samples were collected after induction of anesthesia. The CYP2D6*10 polymorphism was analyzed by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP). According to the results, the patients were divided into three groups (CYP2D6*1/*1, n = 33; CYP2D6*1/*10, n = 28; CYP2D6*10/*10, n = 50). The total consumption of tramadol, visual analogue scale (VAS) score, and PCA control times among the three genotype groups for 2, 4, 24, 48, and 72 h after operation were compared. RESULTS: Nine out of 120 patients were dropped out of the study; 111 patients completed the study. The frequency of CYP2D6*10 allele was 57.7%. The demographic data among the three groups were comparable. The consumption of tramadol, patient self-control times of pump, and VAS score in CYP2D6*10/*10 group were significantly higher than that in CYP2D6*1/*1 or CYP2D6*1/*10 group at 2 and 4 h (P < 0.05), while it did not differ between CYP2D6*1/*1 and CYP2D6*1/*10 group (P > 0.05). There was no difference in the incidence of nausea and vomiting among the three groups (P > 0.05). No sever apnea was recorded in these groups. CONCLUSIONS: Different CYP2D6*10 genotypes have an influence on the analgesic effect of tramadol in Han nationality patients after elective nephrectomy.

Influence of Dexmedetomidine on Myocardial Injury in Patients with Simultaneous Pancreas-Kidney Transplantation.

Background: Diabetes is one of the most common chronic diseases in the world. End-stage renal disease (ESRD) caused by diabetes is the most serious long-term complication. The main cause of death in patients with simultaneous pancreas-kidney transplantation (SPKT) is cardiovascular disease. Although dexmedetomidine (Dex) has unique advantages in heart protection against ischaemic/reperfusion injury, few clinical studies have been conducted on its cardioprotective effect in SPKT. This study aimed to explore the influence of Dex on myocardial injury in patients undergoing SPKT and to analyze its possible mechanism. Methods: A randomized controlled trial (RCT) was performed from July 1, 2018 to December 1, 2020. Eighty patients, regardless of gender, scheduled for SPKT were randomly allocated into a Dex group (D group) receiving Dex at a rate of 1 µg/kg for 10 minutes before anaesthesia induction and then continuous infusion at 0.5 µg/kg/hour until the end of surgery and control group (C group) receiving equivalent capacity of saline. Serum cardiac troponin I (cTnI), creatine kinase isoenzyme (CK-MB), tumour necrosis factor-α (TNF-α), and interleukin-6 (IL-6) were recorded at 5 minutes after anaesthesia induction (baseline,T0), 5 minutes before renal arteriovenous opening (T1), 30 minutes after renal arteriovenous opening (T2), 30 minutes after pancreatic related arteriovenous opening (T3), immediately after surgery (T4), 4 hours after surgery (T5), and 24 hours after surgery (T6). Adverse cardiovascular events were recorded during the perioperative period. Changes in ECG S-T segments and T waves were monitored at T0-T6. Myocardial infarction and percutaneous coronary intervention were recorded with an average follow-up of one year. Results: Compared with T0, TNF-α and IL-6 concentrations significantly increased at T1-T6 in the C and D groups (P < 0.05). IL-6 concentration increased significantly after renal artery opening and reached the peak after the opening of pancreatic blood vessels. Compared with the C group, TNF-α, and IL-6 concentrations were significantly reduced in group D at T2-T6 (P < 0.05). Compared with T0, cTnI and CK-MB concentrations were significantly increased at T3-T6 in the C and D groups (P < 0.05). cTnI and CK-MB concentrations increased significantly after the opening of renal artery, and reached the peak after the opening of pancreatic blood vessels. Compared with the C group, cTnI and CK-MB concentrations were significantly reduced in the D group at T3-T6 (P < 0.05). There was no significant difference in patient characteristics amongst groups, including the proportion of intraoperative vasoactive drug use and adverse cardiovascular events during the follow-up period. Heart rate, mean blood pressure, central venous pressure, and cardiac output were not remarkably different between the two groups at any time point. Conclusions: Perioperative reperfusion could aggravate myocardial injury in SPKT. Dex may be considered a way to reduce myocardial injury caused by inflammatory action by decreasing the release of inflammatory factors. Trial Registration Number: Chinese Clinical Trial Registry ID: ChiCTR2200060084.

Effects of electroacupuncture with dexmedetomidine on myocardial ischemia/reperfusion injury in rats.

BACKGROUND: To investigate the protective effect of electroacupuncture combined with dexmedetomidine (EA + Dex) on oxidative stress injury in myocardial ischemia/reperfusion (I/R) rats. METHODS: A total of 50 male Sprague-Dawley (SD) rats were randomly divided into 5 groups: sham operation (sham group); I/R group; dexmedetomidine group (Dex group); electroacupuncture group (EA group); and EA + Dex group. The myocardial I/R model was established. The EA group received EA at the Neiguan acupoint [pericardium 6 (PC6)] every day for 1 week before modeling. Rats in the EA + Dex group received EA at PC6 every day for 1 week before modeling, and intraperitoneal injection of Dex was performed 15 minutes before modeling. Dex was injected intraperitoneally in the Dex group 15 minutes before modeling. The rats were sacrificed 1 hour after reperfusion, and myocardial tissue was obtained to measure the myocardial infarction area. The myocardial tissue pathologic changes were shown by hematoxylin and eosin (HE) staining, and the superoxide dismutase (SOD), malondialdehyde (MDA), adenosine triphosphate (ATP), and reactive oxygen species (ROS) content in serum was determined. RESULTS: Compared with the sham group, the myocardial infarction area was significantly increased (P<0.01), SOD and ATP content was significantly decreased (P<0.01), and MDA and ROS content was significantly increased (P<0.01) in the I/R group; this change was significantly reduced in the Dex, EA, and EA + Dex groups (P<0.01). The indicators in the EA + Dex group were better than those in the EA and Dex groups (P<0.05). There was no significant change in the above indices in the Dex group compared with the EA group (P>0.05). CONCLUSIONS: EA + Dex pretreatment improved the damage of myocardial I/R by increasing SOD and ATP content and reducing the generation of MDA and ROS in an oxygen-free radical system.

Modified Iron Deposition in Nigrosomes by Pharmacotherapy for the Management of Parkinson's Disease.

Background: Increased iron deposition in nigrosome as assessed by susceptibility-weighted imaging (SWI) is involved in the pathogenesis of Parkinson's disease (PD). This study investigated the effects of antiparkinson drugs on iron deposition in the nigrosome of PD patients. Methods: Based on the retrospective analysis of clinical data, alterations in iron deposition in the substantia nigra were investigated in 51 PD patients across different types of therapies and in nine Parkinson-plus syndrome patients. The Movement Disorder Society revision of the Unified Parkinson's Disease Rating Scale (MDS-UPDRS) Part Ⅲ/Ⅳ (UPDRS Ⅲ/Ⅳ) was utilized to evaluate motor function and complications. SWI (slice = 0.6 mm) was used to detect iron deposition in the nigrosome and substantia nigra. Nigrosome loss was scored on a 1-point nigrosome visibility scale. Visual assessment of dorsolateral nigral hyperintensity (DNH) was separately performed for each side of the nigrosome with SWI. Results: Increased UPDRS Ⅲ scores were correlated with low nigrosome scores based on correlation analysis at a disease duration of 6-12 months (r = -0.8420). The loss of the nigrosome on SWI was clearly inhibited in PD patients with a 3-5-year duration of administration of antiparkinson medications compared with no treatment. Decreased UPDRS Ⅲ scores and increased nigrosome scores were observed in the regular treatment of PD patients with a 6-7-year disease duration. For patients with Parkinson-plus syndromes, such as multiple system atrophy, iron accumulation was apparent in the corpus striatum and substantia nigra compared with that for patients with progressive supranuclear palsy. Conclusions: Early and regular treatment with antiparkinson drugs not only alleviates the chance of PD disability but also prevents the loss of DNH, namely, iron accumulation in the nigrosome.

8-Nitro-quinoline.

The molecule of the title compound, C(9)H(6)N(2)O(2), is almost planar, with a dihedral angle of 3.0 (9)° between the pyridine and benzene rings.

Correlation between cognitive impairment during the acute phase of first cerebral infarction and development of long-term pseudobulbar affect.

PURPOSE: The relationship between cognitive impairment during the acute phase of first cerebral infarction and the development of long-term pseudobulbar affect (PBA) has not been elucidated. Therefore, in this study, we aimed to determine if cognitive impairment during the acute phase of cerebral infarction will increase the risk of long-term post-infarction PBA. PATIENTS AND METHODS: This was a nested case-control study using a prospective approach. A consecutive multicenter matched 1:1 case-control study of cognitive impairment cases following acute cerebral infarction (N=26) with 26 sex-, education years-, and age-matched controls. Univariate and multivariate conditional logistic regression analyses were performed to study the clinical features and changes in cognitive domain as well as the risk factors for PBA. RESULTS: Long-term PBA was independently predicted by low Montreal cognitive assessment (MoCA) scores at baseline. Multivariable regression models showed that post-infarction low MoCA scores remained independent predictors of long-term PBA (odds ratio [OR]=0.72; 95% confidence interval [CI]=0.54-0.95; P=0.018). Among all cognitive disorders, digit span test (DST) scores (OR=0.39; 95% CI=0.16-0.91, P=0.030), StroopC time (OR=1.15; 95% CI=1.01-1.31; P=0.037), and clock-drawing task (CDT) scores (OR=0.62; 95% CI=0.42-0.90; P=0.013) were found to be the independent risk factors for PBA. CONCLUSION: Cognitive impairment during the acute phase of cerebral infarction increased the risk of cerebral infarction-induced long-term PBA. Development of PBA was closely associated with executive function, attention, and visuospatial disorder.

Platelet membrane receptor P2Y12 H1/H2 polymorphism is highly associated with cerebral infarction: a case-control study.

OBJECTIVES: This study aimed to determine the relationship between the polymorphisms of the H1/H2 gene of platelet membrane receptor P2Y12 and cerebral infarction (CI) in a Han population in North Shandong Province, People's Republic of China. PATIENTS AND METHODS: A case-control study, which involved 168 nonstoke subjects (contrast group) and 152 CI patients (CI group), was conducted. The state of subjects in the CI group was validated by computed tomography or MRI. The clinical data were categorized into two groups. The data included age, gender, smoking, drinking, shrinkage pressure, diastolic blood pressure, blood glucose, cholesterol, triglyceride, low-density lipoprotein, high-density lipoprotein, serum uric acid, fibrinogen and homocysteine. The polymorphisms were genotyped with PCR and restriction fragment length polymorphism analysis. The distribution characteristics of nonstoke subjects and CI patients and the relationship between the polymorphisms of the H1/H2 gene of platelet membrane receptor P2Y12 and ischemic stroke were analyzed. RESULTS: No significant difference was found between the contrast group and CI group (P>0.05) in terms of age, gender composition, smoking, alcohol consumption, blood glucose, cholesterol, triglyceride, low-density protein, high-density lipoprotein cholesterol, uric acid and homocysteine. In contrast, significant differences were found between these two groups (P<0.01) in terms of SBP, DBP and plasma fibrinogen level. The genotyping revealed 112 carriers of the wild-type H1/H1 genotype and 40 carriers of the mutational H2 allele of P2Y12 H1/H2 in the CI group and 140 carriers of the wild-type H1/H1 genotype and 28 carriers of the mutational H2 allele of P2Y12 H1/H2 in the contrast group. Furthermore, the H1/H2 and H2/H2 gene frequencies (26.3%) were significantly higher in the CI group (χ2=4.440, P<0.05) than those in the contrast group (16.7%). Moreover, the frequencies of the H2 allele in the CI and contrast groups were 14.5% and 8.6%, respectively, and the difference was statistically significant (χ2=5.392, P<0.05). Multiple logistic regression analysis results revealed that factors associated with CI include systolic blood pressure and plasma fibrinogen level, which carry the -893T gene. After adjusting for potential confounding factors, the H2 allele carriers had a 1.928-fold increased risk for CI (OR=1.928, 95% confidence interval: 1.137-3.188; P=0.038) when compared with noncarriers. CONCLUSION: The present study found that hypertension and elevated plasma fibrinogen levels are significant risk factors for ischemic stroke and confirmed that the H1/H2 and H2/H2 genes of platelet membrane glycoprotein receptor P2Y12 are risk factors of ischemic stroke.

Activation of bone morphogenetic protein-6 gene transcription in MCF-7 cells by estrogen.

BACKGROUND: Bone morphogenetic protein-6 (BMP-6) is closely correlated with tumor differentiation and skeletal metastasis. Estrogen is considered as a stimulant for the initiation and promotion of breast cancer. Previous studies demonstrated that 17beta-estadiol (E2) can selectively increase the expression of BMP-6. This experiment is designed to detect the molecular mechanism of estrogen activating BMP-6 gene transcription in human estrogen receptor positive (ER+) breast cancer cell line MCF-7. METHODS: After the treatment of MCF-7 cells with E2 at different concentrations (10(-11) mol/L, 10(-9) mol/L, 10(-7) mol/L), the BMP-6 expression level was examined through real-time polymerase chain reaction. Through restriction enzyme digestion, human BMP-6 1.2 kb long promoter, BMP-6 0.7 kb long promoter was cloned into pGL-3 basic vector; after the treatment with 10(-7) mol/L E2, luciferase activities of the two promoters were detected. Site-directed mutagenesis was performed to obtain the mutant forms of estrogen response element half-site (1/2 ERE) element and Sp1 sites in the BMP-6 promoter, the activities of these mutant form promoters were detected following the methods mentioned above. Chromatin immunoprecipitation (ChIP) assay was also used to confirm the binding of estrogen receptor alpha (ERalpha) on BMP-6 promoter in the presence of E2. RESULTS: E2 dose dependently increased BMP-6 mRNA expression in human ER+ breast cancer cell line MCF-7. At a dose of 10(-7) mol/L E2, human BMP-6 1.2 kb promoter activity was increased by 90% compared with the control group treated with ethanol (P < 0.05). Both the 1/2 ERE response element mutant form and the Sp1 site mutant form of the BMP-6 promoter abolished the activation of the BMP-6 promoter's response to E2. Through ChIP assay, the binding of ERalpha on 1/2 ERE response element in BMP-6 promoter was further validated. CONCLUSION: Estrogen induces BMP-6 expression in human ER+ breast cancer cell line MCF-7 through its receptor ERalpha binding on 1/2 ERE element in the BMP-6 promoter.

Propofol prevents neuronal mtDNA deletion and cerebral damage due to ischemia/reperfusion injury in rats.

Propofol is a commonly used intravenous anesthetic that has been demonstrated to be neuroprotective against cerebral ischemia-reperfusion (I/R) injury. It remains unclear whether this protective effect has any relationship with the prevention of neuronal mitochondrial deoxyribonucleic acid (mtDNA) deletion. In this study, 81 Wistar rats were randomly divided into three groups (n = 27 each): sham (S group), ischemia/reperfusion (I/R group), or propofol (P group). Cerebral ischemia was induced by clamping the bilateral common carotid arteries for 10 min. A polymerase chain reaction (PCR) was conducted to determine mtDNA deletion. The mitochondrial membrane potential (MMP) changes were detected via microplate reader. The neuronal ultrastructure was visualized via electron microscope. MMP significantly decreased after I/R (P<0.05 compared with the S group). Severe damage to the ultrastructure of neuronal mitochondria was observed in cerebral I/R injury. When propofol (1.0mg/kg/min) was administered intravenously for 1h prior to the induction of I/R, the neuronal structure and MMP were well preserved, and mtDNA deletion was reduced after ischemia/reperfusion injury compared with the I/R group (P<0.05). These data suggested that propofol prevented mtDNA deletion and preserved a normal structure and MMP, which are important for normal mitochondrial function and increase neuronal resistance to I/R injury.

Effects of propofol and sevoflurane on aquaporin-4 and aquaporin-9 expression in patients performed gliomas resection.

Post-operative cerebral edema is a threat for patients performed gliomas resection. Some studies have shown that general anesthesia drugs, such as, propofol had neuroprotective effect. Aquaporin-4 (AQP4) and Aquaporin-9 (AQP9) play an important role in maintaining brain water homeostasis under various conditions. The aim of this study was to compare the effect of propofol or sevoflurane on expression of AQP4 and AQP9 in patients performed gliomas resection. 30 patients performed gliomas resection were included in this study. The patients were randomly divided into two groups: propofol group and sevoflurane group. Fresh human gliomas specimens were obtained and hematoxylin eosin (HE) staining, immunohistochemical staining and Western blot analysis were used for observation of the expression of AQP4 and AQP9. The immunohistochemical staining of the sections showed that the percentage of AQP4 positive cells in the propofol group (14.3±4.61%) was significantly lower than that in sevoflurane group (37.3±10.01%) (n=15, P<0.05). There was no significant difference in the percentage of AQP9 positive cells in propofol group and sevoflurane group (25.8±2.67 versus 28.1±7.81%, n=15, P>0.05). Western blot analysis confirmed the immunohistochemistry results. AQP4 protein level in propofol group was significantly lower than that in sevoflurane group (1.4±0.13 versus 1.7±0.12, P<0.05). Western blot analysis did not show any difference of expression of AQP9 protein between the propofol group and sevoflurane group (2.0±0.13 versus 2.1±0.13, P>0.05, n=6). AQP4 expression was lower in patients of propofol group than that in sevoflurane group. Our results suggested that propofol could inhibit the expression of AQP4.

Effects of APP 5-mer peptide analogue P165 on the synaptic proteins and insulin signal transduction proteins.

Diabetic encephalopathy (DE) is one of risk factors for Alzheimer's disease (AD). Our previous findings indicated that DE animals had impairment of learning and memory and degeneration of hippocampal neurons, which could be improved by neurotrophic peptide. APP 17-mer peptide is a synthesized peptide sequenced from soluble amyloid precursor protein. APP 17-mer peptide has neural protective effect, but is susceptible to enzyme degradation. Soluble APP 5-mer peptide is the active form of APP 17-mer peptide, and composed of arginine, glutamic acid, arginine, methionine and serine. P165, an APP 5-mer peptide analog reconstructed by our lab, is resistant to enzyme degradation, and can be orally used to protect neurons. In the present study, high glucose and Aß25-35 were used to cause injury to human neuroblastoma cell line SH-SY5Y in vitro, and streptozotocin was used to induce diabetes in mice in vivo. The changes in synaptic proteins and proteins of insulin signal transduction which closely correlate with learning and memory were detected in these cells and the brain of mice. Results showed that P165 could up-regulate the expression of α-synuclein and insulin receptor (IR), down-regulate the expression of insulin receptor substrate-1 (IRS-1), PSD-95, Shank1 and MAPK expression. All these findings suggest that nicorandil might be a potential drug used for the treatment of AD.

Increased argonaute 2 expression in gliomas and its association with tumor progression and poor prognosis.

BACKGROUND: Previous studies have showed that argonaute 2 is a potential factor related to genesis of several cancers, however, there have been no reports concerning gliomas. METHODS: Paraffin specimens of 129 brain glioma cases were collected from a hospital affiliated to Binzhou Medical University from January 2008 to July 2013. We examined both argonaute 2 mRNA and protein expression by real-time quantitative PCR (qRT-PCR), Western blot analysis, and immunohistochemistry (IHC). The survival curves of the patients were determined using the Kaplan-Meier method and Cox regression, and the log-rank test was used for statistical evaluations. RESULTS: Both argonaute 2 mRNA and protein were upregulated in high-grade when compared to low-grade tumor tissues. Multivariate analysis revealed that argonaute 2 protein expression was independently associated with the overall survival (HR=4.587, 95% CI: 3.001-6.993; P=0.002), and that argonaute 2 protein expression and WHO grading were independent prognostic factors for progression-free survival (HR=4.792, 95% CI: 3.993-5.672; P<0.001, and HR=2.109, 95% CI: 1.278-8.229; P=0.039, respectively). Kaplan-Meier analysis with the log-rank test indicated that high argonaute 2 protein expression had a significant impact on overall survival (P=0.0169) and progression-free survival (P=0.0324). CONCLUSIONS: The present study showed that argonaute 2 expression is up-regulated in gliomas. Argonaute 2 might also serve as a novel prognostic marker.

[Effectiveness comparison of triple endobuttons and clavicular hook plate for treatment of fresh acromioclavicular joint dislocation].

OBJECTIVE: To compare the effectiveness of triple Endobuttons and clavicular hook plate fixation in the treatment of fresh acromioclavicular joint dislocation (Rockwood type III-V). METHODS: Between February 2008 and October 2010, 40 patients with fresh acromioclavicular joint dislocation (Rockwood type III-V) were treated with triple Endobuttons in 18 cases (trial group) and with clavicular hook plate in 22 cases (control group). There was no significant difference in gender, age, disease duration, joint dislocation classification, preoperative visual analogue scale (VAS) score and Constant-Murley score between 2 groups (P > 0.05). RESULTS: After operation, wound healed by first intention with no early complication of infection or neurovascular injury. The patients were followed up 12-20 months (mean, 15.8 months) in trial group and 13-24 months (mean, 17.2 months) in control group. Significant differences were found in the VAS score and Constant-Murley score at the last follow-up between 2 groups (P < 0.05). X-ray films showed no loosening of internal fixators or hook displacement occurred. No re-dislocation was observed. CONCLUSION: Triple Endobuttons has less shoulder pain and better shoulder motion than clavicular hook plate, so it is an effective method of treating Rockwood type III-V fresh acromioclavicular joint dislocation.

Propofol improved neurobehavioral outcome of cerebral ischemia-reperfusion rats by regulating Bcl-2 and Bax expression.

Propofol is an intravenous anesthetic with neuroprotective effects against cerebral ischemia-reperfusion (I/R) injury. Few studies regarding the neuroprotective and neurobehavioral effects of propofol have been conducted, and the underlying mechanisms are still unclear. Because I/R may result in neuronal apoptosis, the apoptosis regulatory genes B-cell leukemia-2 (Bcl-2) and Bcl-2-associated X protein (Bax) may be involved in the neuroprotective process. In this study, 120 Wistar rats were randomly divided into three groups (sham, I/R-induced, and propofol-treated). Cerebral ischemia was induced by clamping the bilateral common carotid arteries for 10min. Propofol (1.0mg/kg/min) was administered intravenously for 1h before the induction of ischemia. Neuronal damage was evaluated by neurobehavioral scores and histological examination of the brain sections at the level of the dorsal hippocampus at 6h, 24h, 48h, 72h, 4days, 5days, 6days, and 7days after I/R. The apoptotic rate of hippocampal neurons was detected by flow cytometry. The expression of Bcl-2 and Bax was evaluated using immunohistochemical and Western blot methods. The results of this study showed that neurobehavioral scores were higher in propofol-treated rats compared with I/R-induced rats with no propofol treatment. Moreover, the hippocampal expression of Bcl-2 was significantly higher, while the expression of Bax was significantly lower in propofol-treated rats compared with I/R-induced rats at 24h after ischemia. Hence, this study suggests that the neuroprotective effects of propofol against neuronal apoptosis may be a consequence of the regulation of Bcl-2 and Bax.

[Expression of recombinant human BMP6 in CHO cells by fused to the signal peptide and propeptide of another homologue protein].

BMP6 is a member of TGF-beta superfamily, represent more effective osteogenic activity. Two recombinant plasmids were constructed to expression rhBMP6 in mammalian cells, one contained the cDNA encoding the signal peptide, propeptide and mature peptide of human BMP6, wich was named pcDNA-BMP6, the other contained the recombinant DNA encoding the signal peptide, propeptide of human BMP2 and the mature peptide of BMP6, which was named pcDNA-BMP2/6. Transient expression in Cos7 cells demonstrated that the pcDNA-BMP2/6 produced more rhBMP6 than pcDNA-BMP6. For stable expression, the CHO-dhfr- cells were transfected with pcDNA-BMP2/6 and pSV2-dhfr, then screened by G418 and treated with MTX for targeting gene amplification. The partially purified rhBMP6 by heparin affinity chromatography was shown to possess bone induction activity tested by the induction of alkaline phosphatase activity in C2C12 cells.

[Expression, characterization and biological activity analysis of recombinant human bone morphogenetic protein 2 in CHO cells].

Bone morphogenetic protein 2(BMP-2) is a member of the of BMPs family, its osteoinductive capacity has already been demonstrated. We tried to express hBMP-2 in CHO cell. In this study, we inserted hBMP-2 cDNA into vector pCDNA3.1(+) to construct hBMP-2 eukaryotic expression vector pCDNA3.1(+)-hBMP-2. Recombinant Chinese hamster ovary (rCHO) cell line expressing high-level recombinant human bone morphogenetic protein 2(rhBMP-2) was constructed by co-transfecting the expression vectors pCDNA3.1(+)-hBMP-2 and plasmid pSV2-dhfr into dihydrofolate reductase (dhfr)-deficient CHO cells and the subsequent gene amplification in medium containing stepwise increments in methotrexate level such as 0.1 and 1 micromol/L. Western blot analyses showed a specific band of about 18 kD in reduced sample lane and a specific band of about 32 kD in non-reduced sample lane, this indicated that rCHO cells secret rhBMP-2 as a homodimeric glycoprotein form. Finally, we obtained a single clone cell strain expressing a high level (7.83 microg/24 h/10(6) cells) of rhBMP-2 tested by ELISA. Biological activity of rhBMP-2 was tested by the induction of alkaline phosphatase(ALP) activity in C2C12 cells. We treated C2C12 with different concentration of rhBMP-2 condition medium(CM) for 5d. The results showed that the rhBMP-2 could significantly increase the ALP activity of C2C12.