Objective To investigate the change of splenocyte subsets in Balb/c mice immunized with transgenic alfalfa(Medicago sativa)containing Eg95-EgA31 fusion gene of Echinococcus granulosus(Eg) and challenged with Eg protoscoleces.Methods Leaf protein was extracted from transgenic alfalfa containing Eg95-EgA31 fusion gene by heat-coagulation method,and concentration of 20 g/L was used in the study.Meanwhile,leaf protein extracted from the transgenic alfalfa containing blank vector(pBI121)and the normal alfalfa was served as control.Thirty-two female Balb/c mice were randomly divided into 4 groups,8 mice in each group.Oral group was immunized with the leaf protein containing Eg95-EgA31 fusion antigen intragastrically(100μl per mouse);intranasal group was immunized with the leaf protein containing Eg95-EgA31 fusion antigen intranasally(10 μl per mouse);blank vector group was vaccinated intranasally with 10μl leaf protein with blank vector(pBI121);and normal control group was given 100μl normal leaf protein intragastrically.All mice in the above mentioned groups were immunized every 3 days for 2 months.Then,the mice were challenged intraperitoneally with Eg protoscoleces(50 protoscoleces per mouse)8 weeks after last vaccination and sacrified 24 weeks pest infection to separate the splenocytes.Flow cytometry was used to measure the percentages of CD4+ and CD8+ T ceils subsets.Resuits Compared with the normal control group(0.166±0.018,0.083±0.006,2.019 ±0.369),the percentages of CD4+(0.286±0.009)and CD8+(0.102±0.004)T cell subsets and the ratio of CD4+/CD8+(2.814±0.014)in oral group increased significantly (P<0.01 or<0.05).The percentage of CD4+ subset(0.269±0.016)and the ratio of CD4+/CD8+(2.955±0.986) in intranasal group was significantly higher than that ofthe normal control group(all P<0.01).The percentage of CD4+ subset in oral group was significantly higher than that of the intranasal group(P<0.05).No significant difference was found in the percentages of CD4+ and CD8+ T cell subsets and the ratio of CD4+/CD8+ between the blank vector group(0.169±0.018,0.093±0.019,1.852±0.188)and the normal control group(all P>0.05).Conclusions CD4+ T cell may play an important role in the protection induced by transgenic alfalfa vaccine against the challenge of Eg protoscoleces.Intragastrical immunization may be a good route.

Objective To investigate the dynamic changes of splenocyte cytokines in mice immunized with the transgenic alfalfa containing Eg95-EgA31 fusion gene of Echinococcus granulosus (Eg). Methods Eighty-eight Balb/c mice were divided into 2 groups randomly according to body weights,and immunized orally or intranasally with 100μl or 10μl extracted leaf protein from the transgenic alfalfa(20 g/L) respectively once per 3 days for 2 months. Four mice randomized from each group were killed to get splenocyte on week 0(control),2,4,6,8,10,12,14,16,18 and 20 after the last immunization. The splenocyte were cultured in medium for 48 hours with EgAg or concanavalin A (ConA) stimulation to induce the interleukin (IL)-12,interferon γ(IFN-γ) and IL-10,and cultured for 72 hours with EgAg or lipopolysaccharide (LPS) stimulus to induce the tumor necrosis factor α (TNF-α). Then the supernatant was collected to measure the level of IL-12,IFN-γ,TNF-α and IL-10 by ELISA. Results In the oral immunization group,the level of IL-12,IFN-γ,TNF-α and IL-10 increased significantly from week 4 to week 6,week 2 to week 8,week 2 to week 6 and week 4 to week 12,respectively,reaching the highest level(25.0±5.8)ng/L on week 4,(575.0±28.9)ng/L on week 2,(50.0±11.5)ng/L on week 2 and (42.5±2.9)ng/L on week 8,respectively,as compared with the values on week 0[(11.3±2.5),(125.0±28.9),(11.3±2.5),(12.5±2.9)ng/L,all P < 0.01]; in the intranasal immunization group,it was similar about the values of IL-12,IFN-γ,TNF-α and IL-10 could be seen from week 4 to week 6,week 2 to week 10,week 4 to week 10 and week 6 to week 16,respectively,reaching the highest level(25.0±5.8)ng/L on week 6,(725.0±28.9)ng/L on week 4,(27.5±2.9)ng/L on week 6 and (60.0±11.5)ng/L on week 6,respectively,as compared with the values on week 0[(11.3±2.5),(125.0±28.9),(11.3±2.5),(12.5±2.9)ng/L,all P < 0.01]. The cytokine levels in the groups with EgAg,ConA or LPS stimulus were significantly higher than those in the corresponding splenocyte suspension groups(P < 0.05 or < 0.01),and the cytokine levels in the groups with ConA or LPS stimulus were obviously higher than those in the corresponding groups with EgAg stimulation (P < 0.05 or < 0.01). Conclusion The mixed responses of Th1 and Th2 types can be induced in mice immunized with the transgenic alfalfa in the early period post immunization(2-10 weeks).

BACKGROUND:Immune rejections after organ transplantation or serious adverse reactions due to immunosuppressive drugs show a lack of effective treatments and poor therapeutic outcomes. Therefore, we try to find an effective immune suprresion method in combination of the latest immunomodulatory achievements. OBJECTIVE:To construct a lentiviral vector overexpressing indoleamine 2,3-dioxygenase (IDO). METHODS:(1) The IDO gene that was successful y contructed was inserted into lentiviral packaging plasmids GV308 to construct GV308-IDO lentivirus packaging plasmids. (2) The 293T cel s with 80%confluence were co-cultured with 5'LTR and 3'LTR, basic elements of lentiviral packaging auxiliary components, including Psi, cPPT, 3FLAG, TetR, IRES, WRPE, TetIIP, Ubiquitin Promoter, SV40 origin and HIV. RESULTS AND CONCLUSION:Western blot assay showed that in 10 g/L agarose gel electrophoresis, there was a target fragment at Mr 48 000. This value was consistent with the size of IDO protein. RT-PCR results showed visible IDO expression in 293T cel s. These findings suggest that IDO fusion gene has been successful y reorganized in the lentiviral packaging plasmids.

OBJECTIVE:To discuss the feasibility of the development of pharmaceutical care by clinical pharmacist based on micro letter public platform. METHODS:The content and method of pharmaceutical care based on micro letter public platform were introduced by registering micro letter public platform public number,building reasonable medication guidance team,releasing reasonable medication information regularly. RESULTS:40 pieces of reasonable medication information had been pushed by micro letter public platform of our department during Aug.-Nov. 2014,and 350 users added our public platform. The pushed information was mainly about reasonable drug use(57.5%),read by 18 585 persons,and read by 465 person per time in average. Reposted or collected 2 190 times in total,reposted or collected 55 times in average,each piece of information was read by 4 219 person per time at the most. CONCLUSIONS:The pharmaceutical care by pharmacists depending on micro letter public platform contribute to put pharmacist team together,build professional image and provide pharmaceutical care for the public in the information age.

Objective To investigate the relationship between the plasma NT-pro BNP level and the hear function in children pa-tients with heart failure.Methods 30 children patients with heart failure and 30 healthy children were selected as the patients group and the control group respectively.The plasma NT-pro BNP level was detected in the patients group before and after treatment and the control group.The changes of heart functional indexes and the clinical symptoms in the patients group were detected,the detec-tion results and the plasma NT-pro BNP levels were performed the correlation retrospective analysis.Results The plasma NT-pro BNP level before treatment in the patients group was significantly higher than that in the control group,there was the statistically significant difference between them(P <0.05 ).After treatment,the clinical symptoms in the patients group were obviously im-proved,the plasma NT-pro BNP level was significantly decreased and LVEF by ultrasound was also increased,the differences were statistically significant(P <0.05).The plasma NT-pro BNP level after treatment in the patients group was obviously higher than that in the control group with statistical difference between them(P <0.05).The plasma NT-pro BNP level before and after treat-ment in the patients group demonstrated a negative correlation with LVEF(r=-1.32,r=-1.78).Conclusion Detecting the plas-ma NT-pro BNP level in children patients with heart failure has the important clinical guidance significance for the diagnosis,disease condition evaluation,prognostic evaluation and cardiac function grading in the children patients with heart failure.

Animals ; Brain Injuries ; drug therapy ; Gynostemma ; Learning ; drug effects ; Memory ; drug effects ; Phytotherapy ; Saponins ; pharmacology ; therapeutic use

Objective To compare the effects of different resuscitation pressure on lung function of rats with uncontrolled hemorrhagic shock.Methods Uncontrolled hemorrhagic shock model of rats was built.Sixty Sprague-Dawley rats were randomly sent into six groups: NC group (the control group),NF group (shock but non-resuscitation),NS40,NS60 group (limited fluid resuscitation),NS80,NS100 group (large-volume fluid resuscitation).When the mean arterial pressure (MAP) reached between 35 mm Hg to 40 mm Hg (1 mm Hg=0.133kPa) resuscitation was begun,normal saline infusion was used to maintain the following desired endpoints.After the fluid resuscitation for one hour,rats of each fluid resuscitation group were given haemostasis and all-out fluid resuscitation,including blood infusion,to maintain the rats' MAP at 90 mm Hg.Results Findings showed that the blood loss in limited fluid resuscitation groups was significantly lower than that in large-volume fluid resuscitation groups at the situation of uncontrolled hemorrhagic shock,damage to lung tissue pathology and acidosis were significantly lower than those in large-volume fluid resuscitation groups.Nine rats in NS60 group survived 72 hours.Three rats in NS40 group survived 72 hours.No rat in NF group as well as in both NS80 and NS100 groups survived 72 hours.Conclusions Uncontrolled hem-orrhagic shock in trauma and different pressure of fluid resuscitation can cause lung injury to varying de-grees.Limited fluid resuscitation provide significant protective effects against such injuries.

Adult ; Aged ; Benzamides ; therapeutic use ; Case-Control Studies ; Female ; Humans ; Imatinib Mesylate ; Immunoglobulins ; metabolism ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; drug therapy ; immunology ; Male ; Middle Aged ; Piperazines ; therapeutic use ; Pyrimidines ; therapeutic use ; T-Lymphocyte Subsets

Adolescent ; Dermatitis Herpetiformis ; complications ; Esophageal Diseases ; etiology ; Humans ; Male ; Mucous Membrane ; pathology

Objective To cultivate and identify the transgenic affalfa containing Echinococcus granulosus Eg95 gene. Methods The alfalfa plants were transformed by co-cultivating alfalfa cotyledons via recombinant Agrobacterium tumefaciens LBA4404 harboring pBI-Eg95. The transgenic alfalfa explants were selected by kanamyein after calli formation, shoots and roots regeneration in the selective medium, the seedlings of transgenic plants were obtained which were finally transplanted into pots containing nutrient soil. After 2-3 months growth, the complete transgenic alfalfa plants containing Echinococcus granulosus Eg95 gene were obtained. To identify the transgenic alfalfa plants, the total DNA, RNA and leaf protein were extracted from fresh leaf tissue of the transgenic alfalfa plants and confirmed by PCR, RT-PCR, SDS-PAGE and Western blot assay. Results A specific band around 471 bp was amplified by PCR with total DNA, and the same band was obtained by RT-PCR with total RNA, which confirmed that the Eg95 gene was stably integrated into the transformed alfalfa genome. SDS-PAGE analysis showed that the relative molecular mass(Mr) of the expressed protein was about 16.5×103, consistent with the Eg95 protein, and the level of Eg95 expression was up to 0.06% of total soluble leaf protein by Bio-Rad Quantity one assay. Western blot verified the expressed protein was reactive with the sera of mice infected with Echinococcus granulosus. Conclusion The transgenic alfalfa plants containing Echinococcus granulosus Eg95 gene are successfully cultivated.