Objective To establish an acurate and convenient method to distinguish human platelet antigen(HPA) SNPs based on Target Enriched Multiplex-PCR(TEM-PCR),fluorescent probe melting curve analysis and blood direct PCR.Methods Design TEM-PCR primers and probes of HPA1-17,Cab alleles,amplify target sequences of all 18 alleles by blood direct PCR and distinguish different SNPs by melting curve of probes.Results The TEM-PCR could amplify all target sequences of 18 alleles and the melting curve analysis could distinguish those SNPs,the accuracy was equal to PCR-SSP method and the process was more convenient without blood genomic DNA extraction and subsequent gel electrophoresis thus decrease the cross-contamination risk.Conclusion Successfully established a HPA1-17,Cab alleles distinguishing method based on TEM-PCR,blood direct PCR and fluorescent probe melting curve analysis technique.

Objective To investigate the treatment of septic shock(SS) from upper urinary obstruction(UUO). Methods Continuous veno-venous haemofiltration(CVVH) combined with surgical method was applied to 42 SS patients from UUO. Their general conditions, liver and kidney functions, APACHE Ⅱ, therapeutic intervention scorisystem(TISS), multiple organ dysfunction syndrome (MODS), complication rate and main outcomes were analysed comparing with traditional therapies groups(n=30). Results Compared with those traditional therapies, the APACHEⅡ , MODS and TISS score decreased (P＜0.05). Thirty-seven out of 42 patients survived and the survival rate was 88.0% during ICU. Conclusion CVVH combined with surgical methods may effectively decrease the incidence of complications and mortality of SS from UUO, and the mechanism may be related to the remove of mediators of inflammation.

Objective The aim of this study was to evaluate whether the reoperation is the risk factor for mortality after Sun's procedure(Total aortic arch replacement + frozen elephant trunk) for Stanford type A aortic dissection involving aortic arch.Methods Between February 2009 to February 2012,data from 383 patients who underwent Sun's procedure for Stanford type A aortic dissection involving aortic arch were collected retrospectively.35 patients had history of cardiac surgery.Of these patients,16 patients had underwent Bentall procedure,7 patients ascending aortic replacement,4 patients Wheat surgery,4 patients aortic valve replacement,2 patents Bentall combined with mitral valve replacement or plasty,1 patient bivalve replacement,1 patient atrial septal defect repair,1 patient coronary artery surgery.All the risk factors related to mortality were analyzed by univariate statistical analysis.Significant univariate variables were entered into multiple logistic analysis.Results Total 31 patients died in the hospital and the mortality was 8.07％.Of the 35 patients with history of cardiac surgery,3 patients died and the mortality was 8.33％.Univariate analysis showed that symptom onset before surgery less than 1 week,preoperative limb ischemia,combining with coronary artery surgery and cardiopulmonary bypass time longer than 300 minutes in the operative were risk factors for mortality.After these factors were entered into multiple logistic regression analysis,the result showed that symptom onset before surgery less than 1 week (P =0.038,OR =2.43),cardiopulmonary bypass time longer than 300 minutes(PP ＜0.001,OR =12.05) were final independent risk factors for mortality.Reoperation was not the independent risk factor for mortality.The intensive care unit and mechanical ventilation length of reoperation group was (2.09 ± 1.89) days and(30.09 ±33.42) hours respectively,while that of primary group was(2.71 ±3.01) days (P =0.25) and(33.86 ±40.98) hours(P =0.61) respectively.The incidence of postoperative bleeding of reoperation group was 3.03％,while that of primary group was 1.88％ (P =0.50).Conclusion Reoperation was not the independent risk factor for mortality after Sun's procedure for Stanford type A aortic dissection involving aortic arch and the morbidity was also not higher than primary surgery.For these patients,sun' s procedure should be advocated.

Objective To evaluate the risk factors of Sun' s procedure for acute Stanford A type aortic dissection involving aortic arch.Methods Between February 2009 to February 2012,data from 233 patients who underwent Sun' s procedure for acute Stanford type A aortic dissection involving aortic arch were collected retrospectively.All the risk factors related to mortality were analyzed by univariate statistical analysis.Significant univariate variables were entered into multiple logistic analysis.Results Total 23 patients died in the hospital and the mortality was 9.87 ％.Univariate analysis showed that age,limb ischemia,and cardiopulmonary bypass longer than 268 minutes were risk factors for mortality.After these factors were entered into multiple logistic regression analysis,multiple logistic regression showed that age(P =0.017,OR =1.062),cardiopulmonary by pass time longer than 268 minutes(P =0.001,OR =6.150) were final independent risk factors for mortality.ConclusionAge and cardiopulmonary bypass time longer than 268 minutes were final independent risk factors for mortality.Longer cardiopulmonary bypass time should be avoided.

Objective EuroSCORE is a widely used objective risk scoring model.The aim of this study was to evaluate the validation of EuroSCORE Ⅱ in predicting mid-term survival after total aortic arch replacement with stented elephant trunk implantation(sun' s procedure) for Stanford Type A aortic dissection.Methods Total 90 patients entered the study randomly.All the patients underwent aortic surgery using total aortic arch replacement with stented elephant trunk implantation (Sun's procedure).The patients were divided into three groups based on the predicting mortality by EuroSCORE Ⅱ.Ggroup 1:0 ＜:P ≤ 5 ％,Group 2:5 ％ ＜ P ≤ 10％,Group 3:P ＞ 10％.Kaplan-Meier method was used to evaluate the long term survival of three groups.Receiver operating characteristic curve was used to test discrimination of the EuroSCORE.Calibration was assessed with a Hosmer-Lemeshow goodness-offit statistic.Results 87 patients were followed umil October,2013.The mean follow-up time was(33.32 ± 11.11) months.Total 5 patients died during the follow-up time.Three patients died in group 1,2 patients died in group 2 and 1 in group 3.There was no statistical difference for the mid-term survival rate between 3 groups (P =0.054).Conclusion Although EuroSCORE Ⅱ is the newest risk model for cardiac surgery,it is not accurate when it is applied for predicting mid-term survival after aortic surgery.A new risk evaluating system specially designed for aortic surgery should be developed in the future.

Background Retinal microvascular pericytes (RMPs) have been played increasing attention as an emerging key in pathogenesis of various retinal angiogenic diseases including diabetic retinopathy,and RMPs are thought to be a potential target for treatment.Yet the study has been hindered by the difficulty of obtaining source of tissue and isolating pure population.Objective This study was to establish a simple method of isolation,purification and cultivation of primary RMPs for rat.Methods Eyeballs were extracted from clean male Sprague Dawley rats and immersed by 75％ alcohol for 1 minute.The retinas were isolated and mechanical morcel.Trypsin (2.5 g/L) was firstly used and followed by type Ⅰ collagenase (2 g/L) for the digestion of the retina for 15 minutes,respectively.Retinal microvascular fragments were screened by 100 μm and 55 μm filter screen.DMEM containing 20％ fetal bovine serum was added for the cultivation and passaged of the cells.The cells were purified by exchanging medium and partial enzymatic digestion.The morphology and growth status were monitored under the phase contrast microscope,and α-smooth muscle actin (α-SMA),platelet-derived growth factor receptor-β (PDGFR-β),yon Willebrand factor (vWF),glial fibrillary acidic protein (GFAP) antibodies were used for the identification of RMPs.Results RMPs migrated out of fragments after 24-48 hours of plating.On day 7,RMPs appeared in primary cultures as loose colonies.The cells reached confluence to about 80％-90％ on day 14-16.The subcultures grew faster than the primary and reached confluence on day 12-14.The culture showed typical morphology of pericyte with large irregular triangular cell body and multiple long processes,and they could be repeatedly passaged 9 times without obvious loss of characteristic phenotype.Fluorescence assay exhibited that 96％ of the cells showed positive immunofluorescence for α-SMA and PDGFR-β,confirming the purity of RMPs in culture.However,only a few of them were positive for GFAP and the cells response for vWF was absent.Conclusions High purity of rat RMPs can be obtained easily by our method without high cost-consuming.Hcrc wc cstablished a simple mcthod for the primary culture of rat RMPs.

Objective To explore the diagnosis, treatment strategies and clinical outcomes of spontaneous subarachnoid hemorrhage in elderly patients. Methods A total of 68 patients aged over 60 years presenting with spontaneous subarachnoid hemorrhage underwent cerebral angiography. And 72 intracranial aneurysms were detected in 60 patients, among whom 47 patients with 59 aneurysms underwent endovascular coil embolization, 6 aneurysms in 6 patients were clipped in microneurosurgery operation, and 7 patients chose conservative treatment. Results A total of 57 aneurysms (96.6%) were embolized successfully, among which 40 aneurysms (70.2%) reached dense occlusion and 17 aneurysms (29.8%) reached incomplete occlusion. Among 47 patients undergoing endovascular treatment, 39 patients (83.0%) were evaluated as good, 7 patients (14.9%) were moderately to seriously disabled according to Glasgow Outcome Scale when they were discharged, and one patient died. Conclusions Active treatment such as endovascular coil embolization can acquire good outcome in elderly patients with spontaneous subarachnoid hemorrhage, especially in elderly patients detected with aneurysms.

Objective To investigate the clinical features and prognosis of hepatic angiomyolipoma(HAML). Methods Clinical data of 13 patients with operatively confirmed HAML treated in our hospital were retrospectively analyzed. Results HAML was usually asymptomatic and the imaging features varied. Only 1 case was correctly diagnosed before operation. HMB45 positive staining was the pathologic characteristic of HAML. Conclusion HAML can be easily misdiagnosed, should be properly treated by surgery and has a good prognosis.

Objective To evaluate the impact of cerebrospinal fluid drain(CSFD) on the incidence of acute spinal cord injury(SCI) following extensive TAAA repair.Methods From February 2009 to July 2016,153 patients underwent extensive TAAA repairs with a consistent strategy of normal thermia,non-circulatory bypass,sequential aortic cross clamping,aortic-lilac bypass,and intercostal artery reconstruction.The repairs were performed with preoperative CSFD (n =78) or without CSFD (n =75).In the former group,CSFD was inserted after the patient has been anaesthetized and continued for 72 hours after surgery.The target CSF pressure was 10 mmHg or less.Results The mean age of patients was (38 ± 10) years and 108 (70.6％) were male.There were 87 (53.8％) patients with previous aortic surgeries and 33 (22％) with Marfan syndrome.The two groups had similar risk factors for paraplegia.Aortic clamp time,operation time and number of reattached intercostal arteries were similar in both groups.In-hospital mortality rates were 1.3％ (one patient) and 6.7％ (five patients) for CSFD and the group without CSFD,respectively (P =0.086).Ten patients (13.3 ％) in the group without CSFD had paraplegia develop.In contrast,only two patients in the CSFD group(2.6％) had postoperative paraplegia(P =0.013).Stepwise logistic regression analysis identified CSFD had spinal cord protection,P =0.026;OR =0.171;95％ CI:0.036-0.809).No patients occurred CSF catheter related complications.Conclusion This randomized clinical trial showed that preoperative CSFD placement could be an effective strategy in preventing SCI following extensive aortic aneurysm repair.Care should be taken to prevent complications related to overdrainage.

BACKGROUND:Oligodendrocytes are mostly differentiated from oligodendrocyte precursor cel s. A suitable medium and cel seeding density have a significant impact on the process of the isolation of oligodendrocyte precursor cel s to obtain oligodendrocytes. OBJECTIVE:To explore the optimization of oligodendrocyte culture conditions. METHODS:Oligodendrocyte precursor cel s isolated from the newborn rats 48 hours after birth were cultured in DMEM/high glucose medium or DMEM/F12 medium using seeding densities of 2×104 cel s/cm2, 4×104 cel s/cm2, 8×104 cel s/cm2, 16×104 cel s/cm2, 32×104 cel s/cm2, and 64×104 cel s/cm2, respectively. Oligodendrocyte precursor cel s were induced to differentiate into oligodendrocytes at 72 hours after cel adhesion. Morphology of differentiated oligodendrocyte precursor cel s were observed under a light microscope, and the differentiation results were identified by immunofluorescence staining after 7-day induced differentiation. RESULTS AND CONCLUSION:Morphology of oligodendrocyte precursor cel s were recognized when cultured in DMEM/high glucose medium or DMEM/F12 medium using seeding densities of 2×104 cel s/cm2, 4×104 cel s/cm2, and 8×104 cel s/cm2, respectively. Immunofluorescence staining showed that myelin basic protein-positive cel s were found after 7-day induced differentiation, and the positive cel number were 16.40±3.30, 49.95±2.33, and 76.95±4.86 in DMEM/F12 medium, and 12.65±2.53, 32.10±1.17, and 54.05±1.56 in DMEM/high glucose medium (P<0.05). These findings indicate that DMEM/F12 medium is more suitable for culturing oligodendrocyte precursor cel s compared with DMEM/high glucose medium to some extent. The number of differentiated oligodendrocytes was gradual y increased with the enhanced seeding density of oligodendrocyte precursor cel s, and the seeding densities from 4×104 to 8×104 cel s/cm2 were appropriate for the observation of cel morphology.