BACKGROUND:In literatures, the recombinant human bone morphogenetic protein-2 (rhBMP-2) loaded microcapsules can be fabricated by double emulsion solvent evaporation method with different organic solvents, such as methylene chloride, ethyl acetate or their mixture. But so far it is not determined yet which one is better. OBJECTIVE: To optimize the preparation method of microcapsules encapsulating rhBMP-2 and to compare the effects of different organic solvents on the microcapsules. METHODS:Polylactic acid-polyethylene glycol-polylactic acid copolymer as microcapsules was used to prepare rhBMP-2 loaded microcapsules with double emulsion solvent evaporation method. Four kinds of organic solvents, methylene chloride (group A), mixture of methylene chloride and ethyl acetate (group B), ethyl acetate (group C) and acetyl acetone (group D) were chosen as oil phases to compare their effects on microcapsule's morphology, diameter, and encapsulation efficiency. Passage 3 bone marrow mesenchymal stem cels from rats were co-cultured with prepared microcapsules for 14 days, and then alkaline phosphatase activity was detected. RESULTS AND CONCLUSION:Compared with the other organic solvents, dichloromethane could cause microcapsules with the smaler and more uniform shape (4-10 microns) and the highest encapsulation efficiency; the microcapsules prepared by mixture of methylene chloride and ethyl acetate had relatively wide size distribution and moderate encapsulation efficiency; the microcapsules prepared by acetylacetone were difficult to form and keep the bioactivity of rhBMP-2. After cultured with rat bone marrow mesenchymal stem cels for 14 days, the alkaline phosphatase activity in groups A, B and C was significantly higher than that in group D and there was no significant difference between group A and group B; the alkaline phosphatase activity in groups A and B was significantly higher than that in group C (P< 0.05). The results show the rhBMP-2-loaded microcapsules prepared by methylene chloride as organic solvent have good shape, high encapsulation efficiency, and good biological activity.

Objective To analyze the relationship between expression of Her-2 protein and the infection of human papillomavirus (HPV) and clinicopathological features in upper gastrointestinal multiple primary cancers, and to explore the relationship between the different histological malignancies in a single-system. Methods 39 patients were primary esophageal squamous cell carcinoma and gastric/cardia adenocarcinoma. By using immunohistochemistry (IHC) methods, the expression of Her-2 protein in 39 cases of multiple primary cancers specimens were examined, and by using insitu hybridization (ISH) technology, the infection of HPV in the same ones were detected. Results The over-expression of Her-2 protein in esophageal squamous cell carcinoma was not significantly relative to the degree of differentiation, the depth of penetration, the lymph node metastasis and the patientsˊage and gender (P> 0.05). The over-expression of Her-2 protein in gastric adenocarcinoma was closely correlated to the invasion depth and lymph node metastasis (P<0.05), no obvious correlation with the factors of patientsˊage and gender and the degree of differentiation (P > 0.05). The infection of HPV in esophageal squamous cell carcinoma and gastric adenocarcinoma was not significantly relative to the degree of differentiation, the depth of penetration,the lymph node metastasis, and the age and gender of the patients (P> 0.05). In the upper gastrointestinal multiple primary cancers, the Her-2 protein over-expression had the consistency (κ= 0.56, P< 0.05). Meanwhile, the HPV-DNA expression also had the consistency (κ=0.80, P<0.05). Conclusion Both Her-2 protein and HPV expression show the consistency in upper gastrointestinal multiple primary cancers,which suggests that Her-2 protein and HPV may be the common oncogenic factors for both esophageal squamous cell carcinoma and gastric adenocarcinoma.

p75 neurotrophin receptor (p75 NTR) is a member of the tumor necrosis factor receptor superfamily, which interacts with tropomyosin receptor kinase (Trk) receptor or binds neurotrophic factors. It mediates a variety of complex signal transduction pathways, induces synaptic growth and affects cell survival. After acute cerebral ischemia, p75 NTR binds effector factors such as pro-nerve growth factor (proNGF) and sortilin, activating downstream apoptotic signal molecules and leading to neuronal death. Therefore, elucidating the pathways and molecular mechanisms of p75 NTR that mediate neuronal apoptosis in acute cerebral ischemia is of great significance for the development of new therapeutic drugs for acute cerebral ischemia.

Objective To harvest pancreatic tissues from rats , prepare decellularized bio-derived pancreatic scaffolds ( DBPS) , and to examine the integrity and biocompatibility of the scaffolds .Methods Normal pancreases were harvested from healthy adult SD rats .DBPS was prepared by perfusing SDS and Triton X-100 through bile duct and the portal vein, respectively.After decellularization, normal pancreatic tissue and DBPS were compared via HE staining , and transmission electron microscopy ( TEM ) . Abdominal wall and subcutaneous implantations were used to compare biocompatibility , and the remain quantity of residual protein and growth factors were determined via enzyme linked immunosorbent assay(ELISA).MTT assay was used to test the scaffolds’ cytotoxicity.The scaffolds were co-cultured with endotheliocyte .Results HE staining and TEM study indicated no residual cells in the DBPS as well as preservation of the complete extracellular matrix .The remain quantity of residual protein and growth factors in ECM was high .The abdominal wall and subcutaneous implantation revealed that DBPS triggered a lower immune response as compared to the control group.MTT assay showed little cytotoxicity .Endotheliocyte assembled and growed with the scaffolds together .Conclusion DBPS are completely decellularized , and exhibit a higher level of biocompatibility in vivo.Using the way of vessels can make the integrity of extracellular matrix to be fully preserves and contain more growth factors .So using vessels way is better than bile duct .

Objective:To analyze the application value of the mechanical thrombectomy system in the treatment of acute limb ischemia.Methods:The clinical data of 50 patients with lower limb ischemia who were treated with the Rotarex mechanical thrombectomy system from Jun 2017 to Sep 2019 were retrospectively analyzed.Results:In 4 cases of popliteal artery rupture occurred during the operation. The success rate of the operation was 92%. Catheter-directed thrombolysis was used in 7 cases, percutaneous transluminal angioplasty was used in 4 cases and percutaneous transluminal angioplasty combined with stent implantation was used in 39 cases. The ankle-brachial index of these 50 patients before and after operation was 0.18±0.24 and 0.64±0.28 respectively ( t=12.87, P<0.001). Treatment was successful in 43 cases. Follow-up ranged from 1 to 24 months, 5 cases were amputated, 2 cases had no improvement of toe ulcer gangrene, 9 cases had thrombus recurrence, and no complications such as bleeding were observed. The primary patency rates at 3, 6 and 12 months were 92%, 84% and 74%, respectively. Conclusion:The mechanical thrombectomy system is safe and effective in the treatment of acute lower limb ischemia with ideal short-term patency.

Objective To explore the curative effect of the microsurgical excision and spinous process and vertebral plate complex orthotopic replantation to spinal canal plasty for the treatment of spinal canal schwannoma.Methods The clinical data of 18 patients with spinal schwannomas,admitted to our hospital from September 2011 to June 2014,were analyzed retrospectively.All these 18 patients were treated by microsurgical excision and spinous process and vertebral plate complex orthotopic replantation to spinal canal plasty,and were followed up for 3-18 months (mean:10 months).After the surgery,vertebral canal MRI was adopted to evaluate the tumor,and CT three-dimensional reconstruction or X-ray of the spine was used for assessing the vertebral canal formation,and Frankel grading standard was employed to evaluate the recovery of spinal cord function.Results All tumors in 18 patients were excised with the help of a microscope,and no endorachis and nerve roots were damaged during the surgery.After the surgery,the patients showed significantly improved symptoms and signs without the leakage of cerebrospinal fluid,infection of incisional wound,tumor recurrence,spinal stenosis and spinal instability.At 3 months after the surgery,Frankel grading standard was adopted to assess the recovery of the spinal cord function,and the results showed grade D in 4 patients and grade E in 14 patients.Conclusion Microsurgical excision and spinous process and vertebral plate complex orthotopic replantation to spinal canal plasty is an effective way in treating spinal canal schwannoma as it can completely expose the tumor,maximize the excision extension and decrease the spinal cord injury;meanwhile,the spinal canal is formed well and the spine is stable after the surgery.

Objective To understand the prevalence of autism spectrum disorders (ASD) in children aged 0-6 years old and influencing factors in Hainan province.Methods A total of 37 862 children aged 0-6 years were selected from 18 counties in Hainan province for a screening by using questionnaire of "warning signs in child development",then field diagnosis was made,and general descriptive statistic analysis was conducted.The prevalence of ASD and related factors were analyzed with x2 test and unconditional logistic regression model.Results Among 37 862 children aged 0-6 years,235 were diagnosed with ASD,the prevalence of ASD was 0.62％ (0.99％ in boys,0.17％ in girls),the differences was significant (x2=101.91,P=0.000).The prevalence of ASD increased with age (x2=288.62,P=0.000).The prevalence of ASD was significantly higher in urban area than in other areas (x2=114.77,P=0.000).Factors such as full term pregnancy or not,neonatal asphyxia,father' s characteristics,father' s habit of chewing areca or smoking,mother' s general mood,and mother' s induced abortion history were the influencing factors for ASD.Conclusion The prevalence of ASD in children aged 0-6 years was high in Hainan and was influenced by genetic factors,pregnancy and delivery process,parents unhealthy habit before and during pregnancy and other factors.

The disorder of autophagy lysosomal pathway (ALP) is an important pathogenesis of neuronal damage after cerebral ischemia, and the restoration of ALP may alleviate neuronal damage after cerebral ischemia. As the main transcription factor regulating ALP, transcription factor EB (TFEB) can directly regulate autophagosome generation, autophagosome-lysosome fusion, and autophagic flux by regulating the expression of autophagic genes and lysosomal genes. Therefore, regulating TFEB can alleviate ALP dysfunction and thereby reduce cerebral ischemic damage. This article reviews the structure, biological function of TFEB and its role in regulating ALP to alleviate neuronal damage after cerebral ischemia.

Objective To explore the appropriate target ranges of blood glucose in intensive insulin therapy (ⅡT) for acute hyperglycemia following traumatic brain injury (TBI).Methods A randomized,open-label and controlled clinical trial was performed on 208 patients,admitted to our hospitals from Junuary 2014 to Sepember 2016.They were divided into ⅡT group (n=156),who were subdivided into slight (10.1-13.0 mmol/L),moderate (7.1-10.0 mmol/L),and strict (4.4-7.0 mmol/L) control blood glucose groups (n=52),and non-ⅡT group (n=52).Survival analysis 6 months after treatment was performed by Kaplan-Meier method.Modified Rankin scale (mRS) scores and Barthel index (BI),Glasgow Outcome scale (GOS) scores,concentrations of lactic acid in cerebrospinal fluid (CSF) and glycosylated hemoglobin,Glasgow coma scale (GCS) scores,Acute Physiology and Chronic Health Evaluation (APACHE Ⅱ) scores,Length of staying in intensive care unit (ICU) and incidence of adverse events were compared between the patients from different groups at different treatment times.Results Blood glucose level within 7 d of admission in patients ofⅡT group was in target ranges.The survival rate of patients from slight and moderate control blood glucose groups was significantly higher than that in the non-ⅡT group and strict control blood glucose group 6 months after treatment (x2=4.237,P=0.040;x2=5.621,P=0.018).As compared with those in the non-ⅡT group and strict control blood glucose group,the mRS scores 3 months after treatment were significantly decreased,and GOS scores and BI one,3 and 6 months after treatment were significantly increased in patients from slight and moderate control blood glucose groups (P＜0.05).As compared with that in the non-ⅡT group,and slight and moderate control blood glucose groups,the glycosylated hemoglobin level 7 d after treatment was significantly decreased in strict control blood glucose group (P＜0.05).As compared with those in the non-ⅡT group and strict control blood glucose group,the concentration of lactic acid in CSF 7 d after treatment,APACHE Ⅱ scores 7 and 14 d after treatment,length of staying in ICU and incidence of adverse events were significantly decreased in patients from slight and moderate control blood glucose groups (P＜0.05).The mean value of blood glucose in slight and moderate control blood glucose groups was (8.40±0.39) mmol/L.Conclusion Proper ⅡT improves the outcomes of TBI patients and (8.40±0.39) mmol/L are established as the target ranges in ⅡT for TBI.

Objective:To prepare the resiquimod-loaded lipid microbubbles R848-MBs, evaluate their enhanced ultrasound imaging and high intensity focused ultrasound (HIFU) ablation effects, and explore their ability to improve tumor immune microenvironment synergize with HIFU.Methods:R848-MBs were prepared by the thin film hydration-mechanical shock method; The basic characteristics and safety of R848-MBs were detected, the HIFU controlled-release characteristics were verified in vitro and the drug metabolism and biological distribution were investigated in vivo. The ability of enhancing ultrasound imaging was observed in vitro and in vivo. To investigate the enhanced HIFU ablation effect of R848-MBs, six EMT6 tumor-bearing mice were randomly divided into HIFU group and R848-MBs+ HIFU group, three mice in each group, the changes in contrast average sound intensity before and after ablation in mouse tumor areas and the change of ultrasound image gray value in tumor area were evaluated, the tumor were resected to observe the coagulative necresis by TTC staining and HE staining. Forty-five tumor-bearing mice were randomly divided into control group, Free R848 group, HIFU group, Blank-MBs+ HIFU group and R848-MBs+ HIFU group, nine mice in each group. On the third day after treatment, 3 mice in each group were randomly selected and killed, to evaluate the ability of R848-MBs to improve tumor immune microenvironment synergize with HIFU. The expression level of CRT on the surface of tumor cells were detected by immunofluorescence staining, the proportion of mature DC in lymph nodes, spleen, and CD8 + T cells in spleen were detected by flow cytometry. The treatment effectiveness of each group( n=6) were evaluated by measuring tumor volume, observing and drawing survival curves. Results:The R848-MBs lipid microbubbles with good safety were successfully prepared, with a concentration of 2.58×10 9/ml, as spherical bubbles under optical microscope and laser confocal microscopy, in a particle size of (1.72±0.11)μm, at a surface potential of (-10.16±0.73)mV. The cumulative drug release was up to 83.44% after HIFU (90 W, 3 s) in vitro. The concentration of R848 in plasma decreased rapidly, and the drug concentration in tumor tissue of the R848-MBs+ HIFU group was higher than that of the R848 group 24 hours after treatment ( P<0.01). The ultrasound imaging of R848-MBs was significantly enhanced in contrast mode in vitro and in vivo; R848-MBs can significantly enhance the HIFU ablation effect, the contrast average sound intensity change in the tumor area before and after ablation in the R848MBs+ HIFU group was greater than that in the R848 group ( P<0.05), and the immediate ultrasound grayscale value change in the HIFU+ R848-MBs group was 46.34±3.21, which was significantly greater than that in the HIFU group (10.67±1.53), with statistical significance ( P<0.000 1). Coagulation necrosis was observed in tumor HE staining and TTC staining. The results of treatment efficacy in vivo showed that R848-MBs+ HIFU group had the strongest therapeutic effect, and R848-MBs combined with HIFU treatment could significantly prolong the survival period of mice compared with intravenous injection of free R848 ( P<0.01). Immunofluorescence staining and flow cytometry results showed an increase in the expression level of CRT on the surface of tumor tissue in the R848-MBs combined with HIFU group, and the percentage of mature DC in tumor draining lymph nodes (58.53±1.04)% were significantly higher than those in the HIFU group (37.56±2.13)% ( P<0.001), and the percentage of mature DC in the spleen (70.65±1.91)% were significantly higher than those in the HIFU group (36.46±3.89)% ( P<0.001), the percentage of CD8 + T cells in the spleen (27.46±3.04)% was significantly higher than that in the HIFU group (18.69±0.29)% ( P<0.01). Conclusions:The HIFU controlled-release lipid microbubbles R848-MBs can not only enhance the efficiency of HIFU ablation, but also improve the tumor immune microenvironment.