ObjectiveTo research the hepatitis B virus (HBV) replication and immune tolerance status of transgenic mice for elucidating the pathogenesis of hepatitis B and evaluating new drugs against HBV.Methods SPE grade HBsAg negative nontransgenic and transgenic mice with the same genetic background were recruited in this study.HBsAg,HBeAg and HBV DNA were detected by chemiluminescent method.Pre-S1 and HBcAg were detected by enzyme linked immunosorbont assay (ELISA).Liver pathology was examined and HBsAg expressions at different stages were determined by immunohistochemical staining.The lymphocyte proliferation of mice was detected by flow cytometry and interferon (IFN)γ-producing T lymphocytes was determined by enzyme linked immunospot (ELISPOT).The expressions of Toll-like receptor (TLR)2 and TLR9 in splenocyte suspension and splenic dendrite cells (DC) were determined by double-labeling immunofluorescence.The data were analyzed by t test and F test.ResultsHBsAg,preS1,HBeAg,HBcAg were expressed and HBV DNA was replicated in HBV transgenic mice,while anti-HBs,anti-HBc,and anti-HBe were all negative.There were no obvious pathological changes in liver tissues.HBsAg was expressed in cytoplasm and HBcAg in nucleus of hepatocytes.After stimulated with HBsAg,T lymphocyte proliferation capacity of HBV transgenic mice was (697.6±67.3) cpm,which was much lower than that of nontransgenic mice [( 1315.5 ±191.6) cpm].The number of spot forming cells of IFNγ-producing splenocytes from transgenic mice after HBsAg stimulation was 8.25 ± 1.10,which was obviously lower than that of nontransgenic mice (28.50±4.21) (F=155.967,P=0.000).The expressions of CD11c+,TLR2 and TLR9 on DC from both HBV transgenic and nontransgenic mice were not different significantly (all P＞0.05).The HBsAg expressions in liver tissues were observed in 18-day-old fetal mice and 1-day-old newborn mice.ConclusionsThe HBV transgenic mice can express HBV-related antigens,and are immune tolerant to the antigens.The innate and acquired immunity of the HBV transgenic mice are normal,which is similar to chronic asymptomatic HBV carriers of human.Therefore,HBV transgenic mouse is an ideal animal model.

[ ABSTRACT] AIM:To investigate the change of phosphorylation of tau protein and expression of cyclooxygenase 2 ( COX-2) in colon submucosal neurons of enteric nerve system in inflammatory bowel disease ( IBD) rats induced by tri-nitrobenzene sulfonic acid (TNBS).METHODS:Male rats (n=30) were randomly assigned to 3 groups (n=10 each):control group, IBD group and TNBS group.The IBD rats were induced by TNBS+ethanol enema for 14 d.The control and TNBS rats were given an equal volume of saline and TNBS, respectively.The general situation and the histopathologic change of the rat colon were observed.Immunofluorescence was used to check the change of phosphalated tau protein and COX-2 expression in the submucosal neurons of the colon.The expression of COX-2 and phosphorylated tau231 and tau262 in the rat colon submucosal neurons was observed by double immunofluorescence staining.RESULTS:Compared with con-trol group, the number of neurons in the colon of IBD rats decreased obviously and the expression of phospholated tau231 and tau262 was significantly increased.The number of neurons in the colon of TNBS rats showed no significant difference compared with control rats.The rat neurons in control group and TNBS group did not express COX-2.COX-2 expression was observed in the nucleus and cytoplasm of colonic neurons in IBD rats, which showed significantly different from control and TNBS rats.CONCLUSION:The decreased neurons in the enteric nerve system of IBD rats might be associated with the phosphorylation of tau protein and the expression of inducible COX-2.

Objective To investigate variation of lumbar spine stability in children with spastic cerebral palsy (SCP) after two types selective posterior rhizotomy (SPR) at lumbosacral and conical sites.Methods Forty-five cases of SCP have undergone with lumbosacral SPR and 38 with conical SPR. Posteroanterior, lateral, 40°-double-oblique, and dynamic (hyperextension and hyperflexion) position lumbar Xray films were taken for all of them before and three months to seven years ( 19 months in average) after operation to observe postoperative lumbar deformity, lumbosacral angle, lateral Cobb' s angle, arch-vertex distance, lordotic index, Posner's definition, and other stability indicators pre- and post-operation of the two groups. Results ① There was statistically significant difference in lumbosacral angle, lateral Cobb's angle, arch-vertex distance, lordotic index and Posner's definition at the 1 st to 2nd lumbar vertebrae (L1-L2 ), the 4th to 5th lumbar vertebrae ( L4 - L5 ), and the 5th lumbar to the 1st sacral vertebrae ( L5 - S1 ) among those with lumbosacral SPR before and after operation (P ＜0. 05). But, only Posner's definition at the 12th thoracic vertebra to the 2nd lumbar vertebra ( T12 - L2 ) varied significantly ( P ＜ 0. 05 ) among those with conical SPR ② Various lumbar deformity was observed in six cases ( 13% ) with lumbosacral SPR, three of them with instable neurological symptoms; while two cases (5%) did so after conical SPR,one with neurological symptoms, with statistical significance ( P ＜ 0. 05). Conclusions Little variation of lumbar spine stability is found among children with spastic cerebral palsy in mid-short term after SPR, while influence of conical SPR is much less on lumbar stability. Their long-term postoperative influence has to be followed-up further.

Objective To observe the protection effect of thrombopoietin on adriamycin-inducod acute myocardium injury in rats model.Methods 32 Wistar rats were randomized into four groups (n=8):Control group,ADM group,ADM +TPOL group and ADM +TPOH group.All agents were given by intraperitoneal injection.The control group was given normal saline,while the other three groups were given adriamycin at the dosage of 20 mg/kg once.TPO groups were injected TPO at different dosages of 10 μg/kg or 30 μg/kg three times on alternate days.ELISA was used to detect CK-MB and cTnI content of serum in the rats.By HE staining,pathological change was found and grade of tissue morphology was scored.The ultrastructure change of cadiocyte was observed by the electron microscope.Results The energy of CK-MB (14.65±1.91,14.21±1.70) and cTNI (9.66±1.31,10.07±1.20) in TPO groups were obviously lower than that of the ADM group(19.58±3.49,12.50±1.62) (P<0.05),respectively.Pathological score of ADM group was higher than TPO groups (P<0.01).The ultrastructure of myecard tissue in the ADM group was damaged more severely than TPO groups.Above-mentioned indexes were with no significant difference between ADM+TPOL group and ADM+TPOH group.Conclusion TPO can protect the heart which was injured by the ADM.

Objective To inwst the antagonistic effect of thrombopoietin on adriamycin induced myocardium injury in rats and explore the mechanism.Methods 32 Wistar rats were randomized into four groups(n=8):Control group,ADM group,ADM+TPOL group and ADM+TPOH group.All agents were given by intraperitoneal injection.The control group was given saline.While the other three groups were given adriamycin at the dosage of20mg/kg.TPO group were injected TPO at the dosages of 10μg/kg or 30μg/kg three times on alternale days.ELISA was used to detect the concentration of CK-MB and cTnI in the serum of rats.The change of cardiocyte ultrastructure was observed by the electron microscope,and pathological change Was observed by immunohistochemistry staining.The expression level of 8- hydroxy-2'-deoxyguanosin(8-OHdG)produced by DNA oxidative damage in myocard tissue were detected.IPP6.0 software was used to detect IOD and calculate the 8-OHdG index.Results The energy of CK-MB and cTNI of TPO group was obviously lower than that in ADM group(P<0.01).The ultragtrueture of cardiocyte in the ADM group Wag damaged more severely than that in TPO group.Pathological Score,IOD and 8-OHdG index of TPO groups were lower than ADM group(P<0.05).These indexes had no significant statistics difference between ADM+TPOL group and ADM+TPOH group.Conclusions TPO can provide heart protection by antagonizing oxidative damage of myocardial cell induced by edriamycin.

Objective To study the protective immune responses induced by recombinant Bifidobacterium (Bb)-TSO45W-4B-TSOL18 vaccine of Taenia solium (T.solium) in domestic pigs challenged with T.solium eggs.Methods Twenty healthy 40 days old domestic pigs were divided into five groups by random number table according to body weight (15 kg):rBb-TSO45W-4B-TSOL18 vaccine group,rBb-TSO45W-4B vaccine group,rBb-TSOL18 vaccine group,blank vector control group and MRS control group.The content of vaccine in each vaccine group was 1 × 1011 CFU.A total of two immunization times was conducted,once every two weeks.Pigs were challenged with T.solium eggs 4 weeks after the last immunization and killed 3 months after infection.The cysticercus was counted and the reduction of the cysticercus was calculated.Blood was collected to separate sera and prepare peripheral blood lymphocytes (PBMC).The levels of IgG,IgG1 and IgG2a in sera were determined by enzyme linked immunosorbent assay (ELISA).The level of PBMC proliferation was tested using methyltetrazolium (MTT) assay.The levels of interleukin (IL)-2,interferon (IFN)-γ,IL-4 and IL-10 in PBMC culture supernatant were detected using ELISA.Results The reduction of cysticercus was 83.09％,71.36％ and 74.85％ in rBb-TSO45W-4B-TSOL18,rBb-TSO45W-4B and rBb-TSOL18 vaccine groups,respectively.The differences of IgG,IgG1,IgG2a levels in sera between groups were statistically significant (F =132.348,106.336,596.091,all P ＜0.05).The levels of IgG and IgG2a in rBb-TSO45W-4B-TSOL18,rBb-TSO45W-4B and rBb-TSOL18 vaccine groups [(366.81 ± 3.84),(334.94 ± 11.65),(333.52 ± 11.09),(87.74 ± 0.95),(84.48 ± 0.80),(84.30 ± 1.09)mg/L] were higher than those of the MRS control group [(245.94 ± 8.81),(62.61 ± 0.84)mg/L,all P ＜0.05].The levels of IgG1 in rBb-TSO45W-4B-TSOL18,rBb-TSO45W-4B and rBb-TSOL18 vaccine groups [(26.55 ± 1.06),(33.24 ± 1.92),(32.60 ± 1.94)mg/L] were lower than those of the MRS control group [(42.78 ± 0.87)mg/L,all P ＜0.05].The differences of IL-2,IFN-γ,IL-4 and IL-10 levels in PBMC original culture supernatant between groups were statistically significant (F =139.522,1 053.102,769.097,962.298,all P ＜0.05).The levels of IL-2 and IFN-γ in rBb-TSO45W-4B-TSOL18,rBb-TSO45W-4B and rBb-TSOL18 vaccine groups [(212.24 ± 3.12),(205.91 ± 3.18),(205.85 ± 4.35),(28.42 ± 0.28),(25.56 ± 0.28),(25.71 ± 0.35)ng/L] were higher than those of the MRS control group [(174.19 ± 2.14),(17.69 ± 0.28)ng/L,all P ＜0.05],while the levels of IL-4 and IL-10 [(40.45 ± 0.36),(41.38 ± 0.70),(41.52 ± 0.19),(71.45 ± 0.83),(73.38 ± 0.70),(74.77 ± 0.41)rig/L] were lower than those of the MRS control group [(52.57 ± 0.29),(94.82 ± 0.45)ng/L,all P ＜0.05].The differences of PBMC proliferation levels between groups were statistically significant (F =56.318,P ＜0.05).The PBMC proliferation levels in rBb-TSO45W-4B-TSOL18,rBb-TSO45W-4B and rBb-TSOL18 vaccine groups (0.543 ± 0.074,0.481 ± 0.028,0.530 ± 0.053) were higher than those of the MRS control group (0.242 ± 0.053,all P ＜0.05).Conclusions Recombinant Bb-TSO45W-4B-TSOL18 vaccine of T.solium could induce certain protection in domestic pigs.Type Th1 immune response may play an important role in induction of protective immunity.

Objective To investigate the significance of aldehyde dehydrogenase 1 (ALDH1)expression in appraising the prognosis of nasopharyngeal squamous carcinoma (NPC).Methods The expression of ALDH1 protein was detected by immunohistochemistry in human specimens obtained from 120 NPC patients without the history of radiotherapy or chemothetapy.Results A relatively high expression of ALDH1 were observed in 40.0 ％ (48/120) patients of NPC,the expression of ALDH1 in the edge of the cancer nests and stroma,particularly spindle cancer cells were strongly positive.The positive expression of ALDH1 was closely associated with patients' nasopharyngeal tumor size (P =0.011),lymphatic metastasis (P =0.005) and clinic stage (P =0.001),but not associated with their gender and age (both P ＞ 0.05).The Kaplan-Merier and Cox regression analysis indicated that ALDH 1 closely correlated with clinical progrosis of the patients.It showed that ALDH1 was an independent risk factor which may affect the prognosis of NPC patients.Conclusion The expression of ALDH1 protein closely correlate with clinic outcome of NPC,suggesting that ALDH1 is a risk factor for clinic progrosis of NPC patients.

Objective To study the protective effect of a mixed recombinant Bifidobacterium (Bb) vaccine of Taenia solium in piglets. Methods Healthy piglets of 40 days old were randomly divided into experimental group and control group, 4 in each group. Experimental group was given 1011 CFU rBb-TSO45W-4B vaccine and rBb-TSOL18 vaccine (in the ratio of 1 : 1). Control group was given Bb liquid medium (MRS). A total of two times of immunization were conducted, once for every two weeks. At different time points after immunization, the serum was separated from precaval vein blood to detect the level of IgG by enzyme linked immunosorbent assay (ELISA). Piglets were challenged with Taenia solium eggs on the 4th week after the last immunization and killed 3 months after infection. The cysticerci were separated to count and calculate the reduction rate of cysticerci. Blood from precaval vein was collected to separate serum and prepare peripheral blood lymphocytes (PBMC). The levels of IgG,IgG1 and IgG2a in serum sampl es and the levels of interleukin ( IL )-2 , interferon (IFN)-γ, IL-4 and IL-10 in PBMC culture supernatant were determined by ELISA. The level of PBMC proliferation was tested using methyltetrazolium (MTT) assay. Results In experimental group, the level of serum IgG increased from the 2nd to the 8th weeks after immunization, and reached the highest level on the 4th week after immunization (mg/L:270 . 64 ± 1 . 94 vs 207.74 ± 2.24, t=42.479, P<0.05). Reduction rate of cysticercus was 80.48%. Compared with control group, the levels of IgG and IgG2a in serum were significantly increased, while the level of IgG1 was significantly decreased (mg/L: 364.15 ± 11.52 vs 245.94 ± 8.81, 89.74 ± 1.13 vs 62.61 ± 0.84, 20.52 ± 1.00 vs 34.11 ± 0.65, t=16.303, 38.579, - 22.772, P < 0.05). The levels of IL-2 and IFN-γ in PBMC culture supernatant were significantly increased, while the levels of IL-4 and IL-10 were significantly decreased (ng/L:215.24 ± 3.31 vs 174.19 ± 2.14, 28.21 ± 0.27 vs 17.69 ± 0.28, 40.35 ± 0.34 vs 52.57 ± 0.29, 71.34 ± 0.36 vs 94.82 ± 0.45, t =20.839, 53.623,-54.743,- 81.266, P<0.05). The level of PBMC proliferation was significantly increased (0.620 ± 0.051 vs 0.242 ± 0.053, t=10.259, P<0.05). Conclusions It is concluded that the mixed rBb vaccine of Taenia solium might give piglets a certain protection. Th1 type immune response plays an important role in the protection.

Objective To dynamically observe humoral and cellular immune responses induced in mice by immunization with a recombinant BCG-TSOL18 vaccine of Taenia solium.Methods Totally 80 Kunming mice were divided into 4 groups by using random number table according to body mass, 20 mice in each group: rBCG-TSOL18 intraperitoneal injection group [mice were vaccinated with 5 × 106 colony forming units (CFU) recombinant BCG-TSOL18 vaccine of Taenia solium through intraperitoneal injection], rBCG-TSOL18 intragastric administration group(mice were vaccinated with 4 × 108 CFU recombinant BCG-TSOL18 vaccine of Taenia solium through intragastric administration), BCG control (mice were vaccinated with 5 × 106 CFU BCG through intraperitoneal injection), PBS control (mice were vaccinated with PBS through intraperitoneal injection).Zero, 2, 4, 6 and 8 weeks after immunization, eye blood was collected and serum w as separated.Levels of specific IgG and IgG2a were detected by enzyme linked immunosorbent assay (ELISA).Proliferation level of spleen lymphocytes was detected by CCK-8.Levels of interleukin-2 (IL-2) and IL-4 were determined by ELISA.Results The level of specific IgG in rBCG-TSOL18 intraperitoneal injection group and rBCG-TSOL18 intragastric administration group increased from 2 to 8 weeks, and reached the highest level by the 6th week (0.310 ± 0.022, 0.356 ± 0.026).Compared with 0 week in the same group, BCG and PBS control group of the same time periods (0.054 ± 0.005, 0.057 ± 0.006, 0.093 ± 0.014, 0.085 ± 0.010), there were statistically significant differences (all P ＜ 0.05).The level of specific IgG2a increased from 2 to 8 weeks, and reached the highest level by the 6th week (0.965 ± 0.031, 1.144 ± 0.049).Compared with 0 week in the same group, BCG and PBS control group of the same time periods (0.102 ± 0.014, 0.093 ± 0.012, 0.115 ± 0.012, 0.103 ± 0.013), there were statistically significant differences (all P ＜ 0.05).The proliferation level of spleen lymphocytes increased from 2 to 8 weeks, and reached the highest level by the 6th week (0.524 ± 0.032, 0.755 ± 0.016).Compared with 0 week in the same group, BCG and PBS control group of the same time periods (0.301 ± 0.018, 0.305 ± 0.020, 0.362 ± 0.033, 0.334 ± 0.027), there were statistically significant differences (all P ＜ 0.05).The levels of IL-2 and IL--4 in spleen lymphocyte culture supernatant increased from 2 to 8 weeks and from 4 to 6 weeks, respectively, reached the highest level by the 6th and 4th weeks, respectively [(1 665.41 ± 33.93), (1 785.11 ± 42.39)ng/L and (281.62 ± 23.79), (357.95 ± 42.57)ng/L].Compared with 0 week in the same group, BCG and PBS control group of the same time periods [(1 411.63 ± 20.54), (1 405.12 ± 21.42),(1 455.20 ± 25.03), (1 434.47 ± 17.47)ng/L and (190.17 ± 11.01), (196.96 ± 14.00), (200.51 ± 30.79), (189.64 ±40.90)ng/L], there were statistically significant differences (all P ＜ 0.05).These indexes mentioned above were statistically significantly different (all P ＜ 0.05) between rBCG-TSOL18 intragastric administration group and rBCG-TSOL18 intraperitoneal injection group.Conclusion The recombinant BCG-TSOL18 vaccine of Taenia solium might induce mice to produce humoral and cellular immune responses, and the effect of intragastric administration is better than that of intraperitoneal injection.

stinguish active tuberculosis and healthy cases with tuberculosis exposure history according SFC count.