Objective To explore the significance and the ultrastructural changes of platelets in patients with acute cerebral infarction.Methods Changes of platelets in 20 patients with acute cerebral infarction were observed by transmission electron microscope.Results The ultrastructural changes of platelets within 24 hours after the onset were significant,including more pseudopodium,aggregation and fusion;there were remarkable fewer alpha granules and mitochondria,the remain of mitochondria swelled and more disruption of platelet membrance were found.The degree of platelet changes consisted with the severity and sizes of cerebral infarction.The ultrastructural change of platelets was obviously restored after 2 weeks,but no restoration was found in 3 patients with large area infarction.Conclusion Ultrastructural changes of platelets in acute period of cerebral infarction were remarkable,especially a lack of alpha granules,this might be regarded as an objective index in the judgment of severity and prognosis of cerebral infarction.

Objective To explore the changes and significances of platelet granule membrane protein-140 (GMP-140),GMP-140 in plasma,PAdT and PAgT in patients with acute cerebral infarction.Methods The platelet GMP-140 was measured with enzyme-linked immunosorbent assay (ELISA) competitive method and GMP-140 in plasma by ELISA double antibody method in blood collecting in 3 d and 2 weeks after onset in large(n=22),small(n=25) size and lacuna (n=20) cerebral infarction groups.Results The platelet GMP-140,GMP-140 in plasma,PAdT and PAgT in large cerebral infarction group in 3 d after onset were much higher than control group (P<0.001),and the small size and lacuna groups were higher (P<0.05～0.001) than control but lower than large group (P<0.05).There was no differences between the small and lacuna groups (P>0.05).The PAdT and PAgT in 3 groups were higher (P<0.01).The platelet GMP-140 and GMP-140 in plasma in all 3 groups in 2 weeks after onset had been clearly lower but still higher than control group (P<0.05),the PAdT and PAgT were normal or even more lower.Conclusion Platelet activation was significant in different types acute cerebral infarction,the concentration of GMP-140 in plasma can reflect the degree of platelets activities more well and truly than PAdT and PAgT.

Objective To dynamically monitor the effect of Cinepazide maleate (CM) on ischemic region of the brain and to elucidate the neuroprotection of CM on acute cerebral ischemic rat model and related mechanisms.Methods The rat model of acute brain ischemia-reperfusion was established with tighting threads.They were randomly divided into ischemia-reperfusion group and CM treatment group(n=8 in each group).Rats in CM treatment group were given 3 mg / kg CM by caudal intravenous injection immediately after brain ischemia-reperfusion.T2 weighted imaging(T2WI) and proton magnetic resonance spectroscopy (1HMRS) were performed before operation and at 6 hours,24 hours and 3 days after reperfusion.Results At 6 h,24 h and 3 d after operation there was no difference for areas of cerebral ischemia on T2WI between 2 groups.The values of N-acetylaspartate(NAA) (Phosphocreatine(PCr) +Creartine(Cr)) in ischemia-reperfusion group rats were (1.53±0.20),(0.50 ±0.17),(0.44±0.13),(0.40±0.10) before operation and at 6 h,24 h,3 d after operation,respectively.And in CM treatment group the values of NAA/(PCr+Cr) were (1.44±0.22),(0.68±0.13),(0.61±0.10),(0.53±0.09) at the same time points.The values of lactate(Lac)/(PCr+Cr) in ischemia-reperfusion group were (0.03±0.01),(1.10 ±0.28),(1.30± 0.23),(1.23± 0.19) before operation and at 6 h,24 h,3 d after operation,respectively.And in CM treatment group the values of Lac/(PCr+Cr) were (0.02±0.01),(0.85±0.25),(0.99±0.20),(0.90±0.15) at the same time points.The rats in ischemia-reperfusion group had lower value of NAA/(PCr+Cr) (P＜0.01) and higher value of Lac/(PCr+Cr) at all time points after operation than those before operation.Compared with rats in ischemia-reperfusion group,the value of NAA/(PCr+Cr) increased(P＜0.05) and the value of Lac/(PCr+Cr) decreased(P＜0.05) decreased significantly for CM treatment group rats.Conclusion CM treatment can decrease the intracellular accumulation of lactic acid and reduce neuronal necrosis in acute brain ischemia-reperfusion rats.

Objective To investigate the protectve effects and underlying mechanisms of deferroxamine on glutamate-induced injury in cultured hippocampal neurons.Methods Primarily cultured hippocampal neurons from fetal rat were used in a model of glutamate induced neurotoxicity.There were two experimental groups.Neurons were pretreated with deferroxamine before glutamate in the deferroxamine group, and neurons were treated with glutamate only in the control group.The morphological change was examined under microscope.Hoechst 33342 DNA staining method was used to study the ratio of condensed nuclei.The levels of lactate dehydrogenase (LDH), malonaldehyde (MDA) and hydroxyl radical were determined using biochemistry.The change in calcium signal was detected using microfluorescent technique.Results The neurons pretreated by deferroxamine had intact morphology with the ratio of condensed nuclei at 14% ± 6% compared to 58% ± 6% (t= 8.98, P ＜0.01 ) in the control group.LDH level was (36.42 ± 8.99) U/L in the deferroxamine group and was (68.06 ± 11.26) U/L in the control group ( t =3.25,P＜0.05).The respective levels of hydroxyl radical were (34.21 ±4.23) U/L and (47.06 ±8.79) U/L (t = 3.11, P ＜0.05 ).The respective levels of MDA were (12.26 ± 2.78 ) nmol/mg and (28.86±5.19) nmol/mg(t =4.88,P＜0.01).Conclusion Deferroxamine can protect neurons from glutamate induced damage.The mechanisms include an inhibition of Ca2+ overload and reduction in the levels of MDA and hydroxyl radicals.

Objective To develop a method for dynamically observing the biological efficacy of botulinum toxin A (BTX-A) and to investigate the dose-effect relationship between BTX-A dosage and muscle strength.MethodsFifty-four male Sprague Dawley rats were randomly divided into 9 groups.Groups 1-7 were injected intramuscularly with 0.1 ml BTX-A (0.01 U to 4.0 U) into the gastrocnemius on the right side.Rats in group 8 were injected intramuscularly with an equal volume of saline solution as the control group,and group 9 was used to determine the location of injection.Gastrocnemius muscle strength was evaluated using a self-made evaluation system before and after the toxin injection and on the 3rd,7th,14th,21st,30th,45th,60th and 75th day following.ResultsMuscle strength reached its lowest level on days 3 to 7,with a significant difference in the decline of muscle strength between the test groups and the control group up to day 60.With the lower BTX-A doses (0.01 U,0.1 U,0.5 U,1.0 U),muscle strength had decreased significantly on the 21st day,but recovered to its initial levels in all groups at the same time.There was no significant difference among the 1.0 U,1.5 U,2.0 U and 4.0 U groups.ConclusionsStandardized gastrocnemius injection combined with neuromuscular functional evaluation can establish a model of BTX-A dosage and muscle paralysis which can be used to assess the evolution of the biological efficacy of BTX-A.

Objective To investigate the impact of polyclonal neural cell adhesion molecule antibody (P-NCAM-Ab) on the potency of botulinum toxin A (BTX-A).Methods Ninety male Sprague-Dawley rats were randomly divided into 3 equal groups:a normal control group,a BTX-A group and a P-NCAM-Ab group.The rats in the normal control group were injected with 100 μl of saline solution in their right gastrocnemius,while those in the BTX-A and P-NCAM-Ab groups were injected with 100 μl of BTX-A (0.5 U).In addition,the rats in the P-NCAM-Ab group were also injected with 100 μl of P-NCAM-Ab (the dosage was 20 U) at the same site on the 3rd day after the BTX-A injection.The rats' gastrocnemius muscle strength was evaluated with a self-made system for evaluating neuromuscular function before and after the toxin injection,on the 3rd day,as well as 1,2,4,6,8,10 and 12 weeks after the BTX-A injection.Any wet weight changes in the muscles were observed,and immunochemistry methods were employed to observe any structural changes in the motor endplates and nerve fibers at the different time points.Results After the saline injection,the average gastrocnemius muscle strength of the control group increased with time,while strength in the BTX-A and P-NCAM-Ab groups demonstrated a decrease in strength followed by a gradual increase.The average gastrocnemius muscle strength of the rats in the BTX-A and P-NCAM-Ab groups was significantly lower than that of the control group at all time points.Compared with the BTX-A group,the muscle strength of the P-NCAM-Ab group rats decreased further.Strength recovery in the BTX-A and P-NCAM-Ab groups was significantly slower than in the control group.The wet weight percentage in the BTX-A and P-NCAM-Ab groups at first decreased and then recovered with time.After the BTX-A injection,the average wet weight percentage of the P-NCAM-Ab group rats was significantly lower than that of the BTX-A group after 3 days,and 1,2 and 4 weeks.Karnovsky-Roots AchE staining showed that the motor endplates' color in the BTX-A and P-NCAM-Ab groups deepened gradually,though the color of the P-NCAM-Ab group was lighter than that of the BTX-A group at each time point.The mean optical density of the motor endplates' positive reaction area increased with time in both groups,but the P-NCAM-Ab group was lower than that of the BTX-A group at 1,2,4,8 and 12 weeks.Counting the nerve fibers dyed by gold chloride showed similar trends with both experimental groups significantly different from the control group.Conclusion P-NCAM-Ab can increase the potency of BTX-A and prolong its action.

Objective To investigate the effect of insulin-like growth factor 1 antibody (IGF-1Ab) on motor endplate function after injection ofbotulinum toxin A (Btx-A).Methods The total 90 male SD rats were randomly divided into 4 groups:control group,Btx-A group and 2 ug and 20 ug IGF-1Ab groups.In Btx-A and IGF-1Ab groups,a volume of 0.5 U (0.1 mL) Btx-A was intramuscularly injected into a site in the fight gastrocnemius muscle;on day 3,equal volumes (0.1 mL) of IGF-1Ab with a dosage of 2 ug and 20 ug were injected to the 2 ug and 20 ug IGF-1Ab groups respectively at the same site.The gastrocnemius muscle strength and the mean optical density (MOI) value of the positive reaction zone of acetylcholine esterase staining were evaluated at different time points.Results The gastrocnemius muscle strength increased from 12.34±0.16 g before injection to 7.70±0.90 g after injection in the Btx-A group;the gastrocnemius muscle strength decreased in other groups after injection of Btx-A;on day 14,28,42,56 and 70,the muscle strength oflGF-1Ab groups was significantly lower than that of Btx-A group (P＜0.05),and on day 42-70,the value of muscle strength of 20 ug IGF-1Ab group was signficantly lower than that of 2 μg IGF-1Ab group (P＜0.05).The MOI values of the positive reaction zone changed with the same trend.Conclusion IGF-1Ab can suppress the restore of motor endplate function after injection of Btx-A.

Objective To evaluate the differentiation of mouse tracheal epithelial cells ( MTEC) at an air-liquid interface and to investigate the influences of influenza virus on the cystic fibrosis transmem-brane conductance regulator ( CFTR) in primary cultured MTEC for further elucidating the possible mecha-nism of imbalanced fluid and salt transportation in respiratory system caused by influenza virus infection . Methods The morphology of primarily cultured MTEC was observed under inverted microscope .Trans epi-thelial electrical resistance ( TEER) was measured by a resistance meter to evaluate the integrity of cultured MTEC.An Ussing chamber apparatus was used to record the short-circuit current of primary cultured MTEC . Results The primarily cultured MTEC clustered together and had a tight pavement-like appearance under light microscope .The TEER was greater than 1000 Ωafter 6 days of culture .Influenza virus could reduce the short-circuit current of CFTR to (52.77±10.30)％in intact cell membrane and to (41.50±1.09)％in monolayer MTEC after increasing the permeability of basement membrane .It had been proved that CFTR was essential to maintaining the balance of fluid and salt transportation in respiratory system.Conclusion Mouse MTEC are efficiently cultured at a air-liquid interface and the primarily cultured cells are highly simi-lar to those in a normal physiologic state .Influenza virus may block the secretion of anions through inhibiting the function of CFTR , which may induce the development of chronic obstructive pulmonary disease and the incidence of asthma .

Objective: To analyze Internet behaviors of children and adolescents and their influencing factors, and to provide scientific basis for health education guidance and prevention measures. Methods: A stratified cluster sampling method was used. From January to June 2019, 30 955 primary school students in 16 provinces (municipalities), from eastern, central, and western economic zones of China were selected to conduct a questionnaire survey, regarding general demographics, diet and exercise, tobacco, alcohol and sleep, Internet and electronics usage, etc. Results: Among them, 43.1% of students had never been online with girls (44.7%) higher than boys (41.8%)(χ2=21.04, P<0.01); 1.3% of students reported online time for more than 4 hours a day, boys were significantly higher than girls(χ2=15.87, P<0.01); Internet addiction was detected in 2.4% of elementary school students, with boys (3.0%) higher than girls(1.7%)(χ2=331.77, P<0.01), and the trend increases with grade. With sixth grade (3.3%) were significantly higher than children from the first grade (0.8%) and other grades(χ2=163.96, P<0.01). Children’s tendency to Internet addiction was 11.9%, of which boys were 13.0% and girls were 10.6%, boys were significantly higher than girls(χ2=117.66, P<0.01). Pearson correlation analysis revealed that, after controlling gender age, boarding school, family relationship, single parents and regrouped families, smoking, drinking sugary drinks, eating fried foods, and left-behind were positively related to Intenet addiction(r=0.06, 0.07, 0.27, 0.44, 0.07, 0.11, 0.09, P<0.01), while medium and high intensity sports, effective physical education, health education in school, AIDS education and parental restrictions on TV and computer games, were negatively associated with internet addiction(r=-0.74, -0.65, -0.04, -0.05, -0.63, P<0.01). Conclusion The internet addiction of Chinese primary school students is common and increases by year. High-intensity, high-density sports, effective physical education, and health education could help prevent Internet addiction among children.

Objective To evaluate the safety and efficacy of botulinum toxin type A for injection in the treatment of post-stroke upper limb spasticity (dosage was 200 U,or 240 U if combined with thumb spasticity).Methods The study was a multi-center,stratified block randomized,double-blind,placebocontrolled trial.All the qualificd subjects were from 15 clinical centers from September 2014 to February 2016.They were randomized (2∶1) to injections of botulinum toxin type A made in China (200-240 U;n =118) or placebo (n =60) in pivotal phase after informed consent signed.The study was divided into two stages.The pivotal trial phase included a one-week screening,12-week double-blind treatment,followed by an expanded phase which included six-week open-label treatment.The tone of the wrist,finger,thumb flexors was assessed at baseline and at weeks 0,1,4,6,8,12,16 and 18 using Modified Ashworth Scale (MAS),disability in activities of daily living was rated using the Disability Assessment Scale and impaction on pain,muscle tone and deformity was assessed using the Global Assessment Scale.The primary endpoint was the score difference between botulinum toxin type A and placebo groups in the tone of the wrist flexor using MAS at six weeks compared to baseline.Results Muscle tone MAS score in the wrist flexor of botulinum toxin type A and placebo groups at six weeks changed-1.00 (-2.00,-1.00) and 0.00 (-0.50,0.00) respectively from baseline.Botulinum toxin type A was significantly superior to placebo for the primary endpoint (Z =6.618,P ＜ 0.01).The safety measurement showed 10 subjects who received botulinum toxin type A had 13 adverse reactions,with an incidence of 8.47％ (10/118),and three subjects who received placebo had three adverse reactions,with an incidence of 5.00％ (3/60) during the pivotal trial phase.All adverse reactions were mild to moderate,none serious.There was no significant difference in adverse reactions incidence between the botulinum toxin type A and the placebo groups.During the expanded phase three subjects had four adverse reactions and the incidence was 1.95％.All adverse reactions were mild,none serious.Conclusion Botulinum toxin type A was found to be safe and efficacious for the treatment of post-stroke upper limb spasticity.Clinical Trial Registration:China Drug Trials,CTR20131191