Objective: To study the preparation technique and release characteristic of 5- fluorouracil-loaded chitosan microspheres for the intranasal administration. Methods:Using the liquid paraffin as the oil phase,and span-80 as the emuifier; 5- fluorouracil-loaded chitosan microspheres were achieved by emulsion-chemical crosslink technique. The orthogonal experimental design was applied to optimize the preparation procedure. Dynamic dialysis method was used to determine the releasing characteristic of microspheres in vitro and it influencing fators.Swelling behavior was expressed by swelling ratio.The degree of mucoadhesion was investigated by determining the mucociliary transport rate(MTR) of the microparticle across a frog palate. Results: Microspheres with a good shape and narrow size distribution were prepared. The average diameter was (43?4) ?m. The drug loading was 38.5%? 1.0%. The entrapment efficiency was 79.0%?1.8%.The drug release profile in vitro could be described by Higuichi eqution Q=0.1035t 1/2 +0.0284 (r=0.9965). Chitosan had good mucoadhesive property and caused a siginificant reduction in MTR(P

0.1%EDTA≈5% HP ? CD≈1%lecithin. Conclusion: The three methods have good correlation.

BACKGROUND:The microRNAs are involved in regulation of stem cel proliferation, differentiation and aging. To study the effect of Let-7c, a member of Let-7, on the neural differentiation of bone marrow mesenchymal stem cels provides new ideas for stem cel therapy. OBJECTIVE: To investigate the role of Let-7c in the neural differentiation of bone marrow mesenchymal stem cels. METHODS: The lentiviral vectors of Let-7c-up and Let-7c-inhibition were constructed and transfected into rat bone marrow mesenchymal stem cels. Optimal multiplicity of infection was screened. The cels were divided into non-transfected group, negative control group (transfected with empty virus), transfected enhancement group (transfected with LV-rno-Let-7c-up), transfected inhibition group (transfected with LV-rno-Let-7c-5p-inhibition). Bone marrow mesenchymal stem cels were treated with fasudil as an inducer for triggering the cels to differentiate into neurons. The fluorescence expressed by transfected cels was observed under inverted fluorescence microscope. The expression of neuron-specific markers, neuron-specific enolase and microtubule-associated protein 2, were measured by immunocytochemical method. The mRNA expression of microtubule-associated protein 2 was detected by RT-PCR. The cel viability was determined by MTT method. RESULTS AND CONCLUSION:Under the inverted fluorescence microscope, the cels were successfuly transfected with LV-rno-Let-7c-up and LV-rno-Let-7c-5p-inhibition. Fasudil induced bone marrow mesenchymal stem cels to differentiate into neurons. The transfection efficiency and expression levels of neuron-specific enolase and microtubule-associated protein 2 in the transfected enhancement group were significantly higher than those in the negative control group (P < 0.05), while in the transfected inhibition group, they were lower than those in the negative control group (P < 0.05). These findings indicate that the differentiation percentage of bone marrow mesenchymal stem cels is increased by fasudil after transfection with LV-rno-Let-7c-up, and Let-7c may promote the differentiation of bone marrow mesenchymal stem cels into neurons.

BACKGROUND: Basic fibroblast growth factor (bFGF) has significant promotion effects on repair in trauma, but local application cannot play a role for a long time.OBJECTIVE: To observe expression of bFGF gene in rabbit bone marrow mesenchymai stem cells (BMSCs) following transfection.DESIGN, TIME AND SETTING: The cytogene in vitro observation was performed at the College of Life Science, Yunnan University from March to August 2009.MATERIALS: One Japan flap-eared rabbit was purchased from the Department of Animal, Kunming Medical College. pCDNA3.1plasmid (Invitrogen, USA) was used in this study.METHODS: Bone marrow was extracted from rabbit anterior superior iliac spine. BMSCs were harvested by the adherent method.Cells were digested and subcultured when 80% confluent. According to GeneBank bFGF cDNA sequence, gene was designed and synthesized. Following electropherosis, the gel was retrieved using xhol I, BamH I enzyme digestion. Restriction enzyme was used to perform enzyme digestion, electropherosis and gene recovery in PcDNA Vector plasmid. bFGF DNA was connected with PcDNA Vector plasmid. PcDNA-bFGF eukaryotic expression vector was constructed. Recombinant was transfected into rabbit BMSCs using liposome infection protocol, and stable transfected line was screened.MAIN OUTCOME MEASURES: BMSC surface antigen expression was measured. Western blot was utilized to determine the expression of target protein.RESULTS: Results of flow cytometry showed that cultured cells were positive for CD90 and CD44, but negative for CD45.Results of immunohistochemistry demonstrated that vessel-derived BMSCs were negative for CD34, but positive for CD44. In cell disruption liquid of bFGF-transfected BMSCs, a significant positive zone of hybridization was visible at M, 23 000. However, no positive band was found in protein from pCDNA3.1(-) blank vector-transfected BMSCs.CONCLUSION: The bFGF gene was successfully transfected into BMSCs, and this target gene can stably express.

Objective To investigate the variance and significance of Th1/Th2 type cytokines in plasma of ovarian cancer patients. Methods The levels of IL-2, IFN-?, TNF-?, IL-4, IL-6, IL-10 in 34 plasma samples of ovarian cancer patients and 14 normal women were detected by cytometric bead array. Results The levels of IL-2, IFN-?, TNF-? in ovarian cancer patients were obviously lower than those in normal womens but the levels of IL-4, IL-6, IL-10 were obviously higher. Furthermore, the variance changed along with clinic stage. Conclusion It is suggested that the imbalance of Th1/Th2 type cytokines in ovarian cancer patients may provide clinical index for the evaluation of progression and prognosis.

BACKGROUND:MicroRNA plays an important role in the process of growth and aging of living body. To know the role of let-7d in inducing bone marrow mesenchymal stem celldifferentiation into neurons can promote the stem celltransplantation. OBJECTIVE:To investigate the role of let-7d in inducing bone marrow mesenchymal stem celldifferentiation into neurons. METHODS:(1) The lentiviral vector of let-7d was constructed and transfected into rat bone marrow mesenchymal stem cells. The cells were divided into non-transfected group, negative control group (transfected with FU-RNAi-NC-LV), transfected enhancement group (transfected with let-7d-LV), transfected inhibition group ( transfected with let-7d-inhibition-LV). (2) Rat bone marrow mesenchymal stem cells were treated with fasudil as an inducer for triggering the cells to differentiate into neurons. The expression of neuron-specific markers, neuron-specific enolase and microtubule-associated protein 2, were measured by immunocytochemical method. The mRNA expression of microtubule-associated protein 2 was detected by RT-PCR. The viability of bone marrow mesenchymal stem cells was determined by MTT method. RESULTS AND CONCLUSION:Under inverted fluorescence microscope, the cells were successful y transfected with let-7d. Fasudil induced bone marrow mesenchymal stem cells to differentiate into neurons. The transfection efficiency and expression levels of neuron-specific enolase and microtubule-associated protein 2 in transfected enhancement group were higher than those in the negative control group (P<0.05);while in the inhibition group, they were lower than those in the negative control group (P<0.05). These findings indicate that let-7d can promote the differentiation of bone marrow mesenchymal stem cells into neurons induced by fasudil, and by control ing the expression of let-7d we can influence the differentiation efficiency from bone marrow mesenchymal stem cells to neurons.

Objective:To explore the application value of automated machine learning (autoML) model in predicting the risk of preeclampsia in the first trimester.Methods:From January 2017 to October 2020, 2 180 singleton pregnant women who were registered in Jinan Second Maternal and Child Health Hospital and underwent pregnancy examination at 12 weeks of gestation were selected. The pregnant women were divided into preeclampsia group (103 cases) and control group (2 077 cases) according to the occurrence of preeclampsia. The differences in clinical data and hematological indexes in the two groups were compared, and the correlation between each index and the risk of preeclampsia was analyzed too. All the pregnant women were randomly divided into training set and test set according to the ratio of 7∶3, and the autogluon autoML algorithm was used to build a variety of machine learning models, and training and cross-validation were performed in the training set to compare the accuracy of the different models. The importance of each index in the autoML model was analyzed, and the autoML model and the logistic regression model were used to predict the risk of preeclampsia in pregnant women in the test set respectively, and the receiver operating characteristic (ROC) curve was used to evaluate the prediction performance of the autoML and the logistic regression model.Results:The age, pre-pregnancy body mass index, body mass index at 12 weeks of gestation, waist circumference at 12 weeks of gestation, proportion of drinking history, high-sensitivity C-reactive protein (hs-CRP), triglyceride, low-density lipoprotein cholesterol (LDL-C), aspartate aminotransferase (AST), platelet distribution width (PDW), mean platelet volume, thyroid stimulating hormone (TSH) and β-human chorionic gonadotropin were all significantly higher than those in the control group (all P<0.05), and the free tri-iodothyronine (free T3), free thyroxine (free T4), placental growth factor (PIGF), soluble fms-like tyrosine kinase-1 (sFlt-1) and pregnancy-associated plasma protein-A (PAPP-A) were all significantly lower than those in the control group (all P<0.05). Correlation analysis showed that the correlation between pre-pregnancy body mass index, body mass index at 12 weeks gestation, waist circumference at 12 weeks gestation, hs-CRP, triacylglycerol, AST, TSH, free T3, free T4, β-HCG, PIGF, sFlt-1, PAPP-A and preeclampsia risk were obviously higher; but the correlation between each index were lower. A total of 18 models in 8 categories were constructed with the autoML model algorithm, and the neural network _L2 based on FastAI had the highest accuracy in the training set (0.963) and the validation set (0.971). The TSH, LDL-C, PDW, waist circumference at 12 weeks of gestation, sFlt-1, AST were more important in the model, while the free T4, total cholesterol, pregnancy times, drinking history, parity and family history of hypertension were less important indicators. The area under the ROC curve of the autoML model for predicting the risk of preeclampsia in the first trimester was significantly higher than that of the logistic regression model (0.984 vs 0.765, P=0.002), while there was no statistical difference in the prediction accuracy of the two prediction models in the training set ( P>0.05). The prediction accuracy and sensitivity of the autoML model in the test set were both significantly higher than those of the logistic regression model (99.54% vs 98.32%, 93.75% vs 75.00%, both P<0.05). Conclusions:Factors such as TSH, LDL-C, PDW, waist circumference, sFlt-1 and AST in the first trimester of pregnancy have a certain correlation with the risk of preeclampsia. The autoML model based on the indicators of the first trimester has a high predictive value for the risk of preeclampsia.

Proline-rich transmembrane protein 2 (PRRT2) is the leading cause of paroxysmal kinesigenic dyskinesia (PKD), benign familial infantile epilepsy (BFIE), and infantile convulsions with choreoathetosis (ICCA). Reduced penetrance of PRRT2 has been observed in previous studies, whereas the exact penetrance has not been evaluated well. The objective of this study was to estimate the penetrance of PRRT2 and determine its influencing factors. We screened 222 PKD index patients and their available relatives, identified 39 families with pathogenic or likely pathogenic (P/LP) PRRT2 variants via Sanger sequencing, and obtained 184 PKD/BFIE/ICCA families with P/LP PRRT2 variants from the literature. Penetrance was estimated as the proportion of affected variant carriers. PRRT2 penetrance estimate was 77.6% (95% confidence interval (CI) 74.5%-80.7%) in relatives and 74.5% (95% CI 70.2%-78.8%) in obligate carriers. In addition, we first observed that penetrance was higher in truncated than in non-truncated variants (75.8% versus 50.0%, P = 0.01), higher in Asian than in Caucasian carriers (81.5% versus 68.5%, P = 0.004), and exhibited no difference in gender or parental transmission. Our results are meaningful for genetic counseling, implying that approximately three-quarters of PRRT2 variant carriers will develop PRRT2-related disorders, with patients from Asia or carrying truncated variants at a higher risk.
Dystonia ; Epilepsy, Benign Neonatal/genetics* ; Humans ; Membrane Proteins/genetics* ; Mutation ; Nerve Tissue Proteins/genetics* ; Pedigree ; Penetrance ; Seizures/genetics*