Objective To study low frequency ultrasound combined with microbubbles to promote the transfection of P53 gene in human prostate cancer cells mediated by liposome.Methods Ultrasound equipment was used with a frequency of 21 kHz and intensity was 46 mW/cm2 and the working time was controlled at 20% (i.e.,2 s “on”time and 8 s “off”time)lasting 5 minutes.The human prostate cancer cell line PC-3 suspension was prepared,the cell concentration was adjusted to 1 × 10 5 cell/ ml,and cells were divided into 8 groups:control group,single microbubbles group,single ultrasound group,ultrasound combined with microbubbles group,single liposome group,liposome combined with microbubbles group, liposome combined with ultrasound group,liposome combined with ultrasound and microbubbles group. Each microbubbles group was added SonoVue 200 μl and the wild type P53 plasmid,plasmid∶liposome is 1 ∶2.At 24 hours after irradiation,Western-blot and RT-PCR were used to detect the gene transfection efficiency,CCK-8 was used to detect cell proliferation,then cell survival rate was calculated,cell apoptosis was detected by flow cytometry.Results After transfection,compared with single liposome group and control group,liposomes combined with ultrasound and microbubbles group can significantly improve expression of the human wild type p53 gene and protein (P <0.001).After transfection,the apoptosis rate of human prostate cancer PC-3 cells in liposomes combined with ultrasound and microbubbles group was significantly higher than that of the liposome group and control group (P <0.001).And after transfection, cell survival rate of liposome combined with ultrasound and microbubble group decreased significantly than those of single liposome group and control group (P <0.001).Conclusions Low frequency and low energy ultrasound combined with microbubbles can promote the transfection of human wild-type P53 gene mediated by liposome.

Objective To investigate the effect of repetitive transcranial stimulation (rTMS) on learning and memory and on the neuron and synapse ultrastructures of the CA3 region of the hippocampus.Methods Forty Sprague-Dawley (SD) rats were randomly divided into 4 groups (n =l0 in each group):a normal control group,a depression group,an rTMS group and a sham group.Unpredictable mild stress was used to establish depression models in the rats of the latter3 groups.The sucrose water consumption test and open-field test were used to evaluate any depressive behavior of each group.The rTMS group rats were given 15 Hz rTMS for 21 days while the sham group received sham stimulation.The orientational navigation and spatial probe tests were performed on each group using a Morris water maze to evaluate their learning and memory abilities.In addition,changes in the ultrastructure of the CA3 region of the hippocampus were detected using transmission electron microscopy.Results The modelling induced significant differences in the sucrose water consumption test results and in horizontal and vertical behavior in the open-field tests.Escape latency and spatial probe time were significantly different between the rTMS group and the sham and depression groups.There was no significant difference in the behavioral indexes between the depression group and the sham group.Electron microscopy showed pathological changes in the ultrastructures of the neurons and synapses in the CA3 region of the hippocampus among the depression group,while in the rTMS group those ultrastructures tended to be basically normal.Conclusion rTMS can improve learning and memory during depression,at least in rats.A possible mechanism is that rTMS can induce changes in the ultrastructures of neurons and synapses in the CA3 region of the hippocampus.

According to relevant national laws and regulations, practitioner training was included into laboratory animal science teaching reform.By adjusting the training content and teaching method and use of animal models of typical human diseases, the transformation of training mode was realized and improved.By the assessment of basic theory in combination with practical operation, the thinking ability and hands-on skill of the practitioners are much improved. Through classroom instruction, experimental teaching, quality assessment and tracking survey, the evaluating process of the training quality of training teaching is performed.Therefore, the teaching reform of the laboratory animal science based on the training of practitioners is established.

Objective To explore the effects of hydrogen sulfide (H2S) on neuron specific-enolase (NSE),neurotrophic protein S100B and neurons apoptosis in hippocampus CA1 region in the early stage of cardiopulmonary resuscitation (CPR) in rabbits. Methods Twenty-five Japanese white rabbits were randomly divided into three groups:sham group (S group),cardiac arrest group (CA group) and H2S treatment group (H2S group). Rabbits were anaesthetized with 5％ halothane,trachea was exposed and intuhated,right femoral vein was cannulated for medical agent administration,and right carotid artery was cannulated for monitoring of blood pressure and blood samples taken. Cardiac arrest was produced by suffocation with clamping the endotracheal tube and turning off mechanical ventilation.Mter 8 min of the endotracheal tube clamping, rabbits received CPR. After the restoration of spontaneous circulation (ROSC),rabbits in groups CA and H2S inhaled 30％ O2 or 30％ O2 containing 80 × 10-6 H2S,respectively.Blood samples were taken before,and 30 min and 60 min after ROSC for detection of the concentrations of NSE and S100B in the plasma. As 60 min after ROSC,rabbits were decapitated after perfusion with 500 ml phosphate-buffered saline and followed by 4％ paraformaldehyde 500 ml through aortic artery,and then the hippocampus was removed rapidly and fixed in 4％ paraformaldehyde for the histological examination.All values were expressed in mean ± standard deviation ((x) ± s).Comparisons were carried out among different groups with SNK-q test of one-way analysis of variance ( One-Way ANOVA plus SNK).Results In comparison with group S,the concentrations of NSE and S100B were significantly increased 30 min and 60 min after ROSC (P ＜ O.05),the viable neurons were decreased and cleaved caspase-3 positive neurons in hippocampus CA1 region increased 60 min after ROSC in groups CA and H2S (P ＜0.05).In comparison with group CA,the concentration of S1OOB decreased 60 min after ROSC (P ＜ 0.05) ; the viable neurons were increased while cleaved caspase-3 positive neurons in hippocampus CA1 region decreased 60 min after ROSC in group H2S ( P ＜ 0.05 ). Conclusions H2S can inhibit the neurons apoptosis in hippocampus CA1 region,increase the viable neurons,decrease the concentration of S100B in the plasma,and then attenuate the cerebral injury after cardiopulmonary resuscitation in rabbits.

Objective To investigate the application of fusion imaging in the initial prostate biopsy.Methods Retrospective analysis was made on 40 patients who underwent initial ultrasound-guided prostate biopsy in Shanghai Jiao Tong University Affiliated 6th People's Hospital from August 2014 to May 2015.All patients underwent preoperative magnetic resonance imaging (MRI) scans one week prior to surgery and the results showed that all patients had suspicious positive prostate cancer lesions,while there were no positive findings in the same area in sonography.All patients with prostate cancer have been confirmed by pathologic examination.Image fusion technology was used to guide the biopsy of lesions which were suspected as prostate cancer by MRI,and then the prostate underwent systematic biopsy by 10 needles under the guide of ultrasound.R× C Chi-square test was used to compare the positive ratio among imaging fusion biopsy,systematic biopsy and combined method in the diagnosis of prostate cancer.Fourfold table Chi-square test was used to compare to undetected rate between fusion imaging biopsy and systematic biopsy in the diagnosis of prostate cancer.The differences of detection rate in the Gleason score more than 7 points of prostate cancer between fusion imaging biopsy and systematic biopsy was compared by Fisher exact test.Results In this group of 40 patients with prostate biopsy,14 cases (35.0％) were diagnosed prostate cancer by systematic biopsy,19 cases (47.5％) were diagnosed prostate cancer by image fusion biopsy,and 22 cases (55.0％) were diagnosed prostate cancer by systematic biopsy combined with image fusion.The difference of diagnostic positive rate among combined method of biopsy,image fusion biopsy and systematic biopsy had not statistically significant (P ＞ 0.05).Three cases (13.6％,3/22) of prostate cancer patients were missed in image fusion method group,and 8 cases (36.4％,8/22) of prostate cancer patients have not been diagnosed by systematic biopsy,which indicated the umdetected rate of systematic biopsy is higher than that of image fusion biopsy in prostate cancer diagnosis,and there was a statistical difference between the two groups (x2=8.338,P=0.005).Among 19 cases of prostate cancer patients who were diagnosed by image fusion biopsy method,Gleason score were greater than 7 points in 15 cases (78.9％,15/22).Among 14 cases of prostate cancer patients who were diagnosed by systematic biopsy method,Gleason score were greater than 7 points in 6 cases (42.9％,6/14).The positive rate of Gleason score ≥ 7 points in fusion imaging biopsy was higher than that of systematic biopsy,which had a statistical difference (Fisher exact probability method,P=0.039).Conclusion Image fusion method can be used to reduce the tndetected rate of prostate cancer and improve the detection rate of the high-grade prostate cancer.

Objective To explore the effects of repetitive transcranial magnetic stimulation on behaviors and hippocampus neuronal apoptosis in rats with chronic stress-induced depression.Methods 40 male SD rats of SPF grade were randomly divided into normol control group (n =8) and model preparation group (n =30) after screening.Rats in model preparation group were singly housed and given chronic unpredictable mild stress(CUMS) to build depression model.Excluding unsuccessful modeling rats,the model preparation group was divided into three groups:model group(n=8,without any treatment),rTMS group (n=8,with the intervention of 10 Hz rTMS) and shame group (n=8,simulation of rTMS environment without rTMS stimulus).The changes of behaviors in each group were detected by weight measurement,sucrose consumption test and open-field test.The changes of morphology of hippocampal neurons were detected by Nissl's staining.The changes of Bax in hippocampal neuron were detected by Immunohistochemical staining.Results (1) Behavioral results showed stress for 21 d could make rat behavior scores decrease significantly(all P＜0.05),and rTMS intervention could significantly improve their behavior scores (all P＜0.01).Compared with model group,the weight reduction rate (0.32±0.05)％,the score of sucrose consumption test(7.03 ± 1.02) and the score of open field test(8212.41 ± 1416.15,8.75 ± 1.58) in rTMS group was higher(P ＜0.01).(2) Nissl staining showed stress for 21 d could make the number of hippocampal CA3 neurons was reduced,cell morphology was poor,and the number of Nissl bodies was reduced.rTMS intervention could increase the number of hippocampal CA3 neurons,cell morphology was integral and the number of Nissl bodies was increased.(3) Immunohistochemistry results showed stress for 21 d could cause the number of Bax cell were significantly increased(P＜0.01),and rTMS intervention can make the number of Bax cell were significantly lower(P＜0.01).Conclusion rTMS intervention improves the depressive behavior in chronic stress depression model rats and inhibits the apoptosis,which might work through inhibition of neuron apoptosis and decline of Bax expression in hippoeampal neurons.

To observe the effects of Feiyanning Decoction, a compound traditional Chinese herbal medicine, in inhibiting the growth of transplanted A549 tumors in nude mice and the expressions of p-Akt and HIF-1alpha in A549 tumors.

Objective To study whether ultrasound combined with microbubbles induces suppress invasion in androgen-independent prostate cancer cells PC-3 and to identify the probable mechanism.Methods Ultrasound with a frequency of 21 kHz and intensity of 46 mW/cm2 in continuous wave mode was used.Ultrasound combined with microbubbles (200 μl SonoVue) was used to treat androgen-independent human prostate cancer PC-3 cells for 30 seconds.PC-3 cells were divided into three groups:control group,ultrasound group and ultrasound combined with microbubbles group.Twelve hours after treatment,cell growth curve we drawn,and transwell chamber model was used to do cell invasion experiments in vitro.Real-time PCR was used to measure the expression matrix metalloproteinase-2 and matrix metalloproteinase-9 messenger ribonucleic acid.Results Twelve hours after treatment,cell growth was not significant difference among three groups (P ＞ 0.05).Twelve hours after treatment,ultrasound combined with microbubbles could significantly inhibit the invasion of human prostate cancer cells (P ＜0.05).Treatment with ultrasound combined with microbubbles down-regulated the expression of matrix metalloproteinase-2 and matrix metalloproteinase-9 messenger ribonucleic acid.Conclusions Ultrasound combined with microbubbles inhibited the invasion in human androgen-independent prostate cancer cells through down-regulation of matrix metalloproteinase-2 and matrix metalloproteinase-9.

Objective To explore the effect of different temperature of the forced- air warming system on the prevention of hypothermia during laparotomy of infants. Methods A total of 60 infants undergoing laparotomy under general anesthesia were recruited and divided into three groups by random digits table method with 20 cases each according to admitting time; when used the force- air warming system intraoperatively, the three groups were respectively setting on 45℃(automatic adjustment for 43 ℃ after 45 minutes), 43 ℃ and 38 ℃.The core temperature were respectively recorded before anesthesia and 15, 30, 45, 60 minutes after anesthesia (every 30 minutes after 1 hour).The hypothermia incidence and anesthesia recovery conditions were recorded simultaneously. Results There was no significant difference on the core temperature among three groups before anesthesia (P > 0.05). 30 minutes after the anesthetic, the core temperature of 45 ℃ group was (36.31±0.20) ℃,43 ℃ group was (36.32±0.24) ℃ and 38 ℃ group was (36.08±0.21) ℃.The differences among three groups was statistically significant (F=8.12, P < 0.01), but there was no significant difference between 45 ℃ group and 43 ℃ group (P > 0.05). 60 minutes after the anesthetic, the core temperature of 45 ℃ group was (36.39±0.26) ℃,43 ℃ group was (36.19±0.22) ℃ and 38 ℃ group was (35.92±0.15) ℃. The differences among three groups was statistically significant(F=25.19, P<0.01).The hypothermia incidence of 45℃group, 43℃ group,38℃ group was 10.0%(2/20), 25.0%(5/20), 50.0%(10/20)respectively and the differences among three groups was statistically significant( χ2=8.04, P<0.05). The time to complete consciousness of 45 ℃ group was (15.40±5.09) minutes,43 ℃ group was (19.80±4.10) minutes and 38 ℃ group was (22.00±4.36) minutes. The differences among three groups was statistically significant (F=10.96, P<0.01). The time to tracheal extubation of 45 ℃ group was (18.10±5.97) minutes, 43 ℃ group was (21.85±4.02) minutes and 38 ℃ group was (24.90±5.54) minutes.The differences among three groups was statistically significant (F=9.83, P<0.01). Conclusions The forced-air warming system can increase the infants′peripheral tissue heat content and reduce the heat losing.So that it will help decrease the intraoperative hypothermia incidence and shorten the anesthesia recovery period.Meanwhile the higher temperature of the forced-air warming system is setted ,the better effect it is.

Objective To knockout Rag2 and IL2rg genes and construct severe combined immunodeficiency mice based on CRISPR/Cas9 technology. Method Design and synthesis of 25 bp sgRNA were made according to the Rag2 and IL2rg sequences in Genbank. After annealing, sgRNA was cloned into pX330 vector. Recombination plasmid Rag2?sgRNA, IL2rg?sgRN and Cas9 were then transcribed into RNA, these RNA were microinjected into zygotes and the zygotes were transplanted into recipient ICR mice. F0 founders were born and mutated F0 founders mated with wild type mice to obtain F1 generation heterozygous mice. Mutated F1 mice were crossed and got F2 generation homozygous mice. Genotype and phenotype of the knockout mice were identified by sequencing, flow cytometry and xenograft model. Results Rag2?sgRNA and IL2rg?sgRNA recombination plasmids were constructed and transcribed into RNA. After microinjection and mat? ing, F0 founders were born and F2 homozygous mice were obtained. The results of sequencing showed that there were two types of genotype in IL2rg gene, 10 bp or 11 bp deletion;however, there was only one genotype in Rag2 gene, which was 8 bp deletion. Compared with wild?type BALB/c mice, the number of CD3 +, B220 + and NKp46 + cells in peripheral blood of the knockout mice was reduced significantly. After inoculation of human breast cancer cell line SKBR?2HL cells, tumor size in the xenograft mouse model was increased gradually along with time extension. Conclusion CRISPR/Cas9 is an efficient way to mutate Rag2 and IL2rg gene in mice in vivo, leading to aberrant T cells, B cells and NK cells.