0.05). The improvement in total score of symptoms of the herb group was better than that of the western medicine group (P0.05). The blood glucose,HbA1c,and insulin of both groups was lowered after treatment (P

Objective To investigate the effects of exogenous glial cell derived neurotrophic factor (GDNF)on colon glial cells in slow transit constipation (STC ) rats,and to explore the optimal concentration of GDNF in order to provide evidence for intestinal neurotrophic therapy in the treatment of STC.Methods A total of 132 SD rats were divided into STC group and control group,66 rats in each group.STC rats were established by feeding with rhubarb.Six rats were randomly selected from either groups to verify whether STC model was successfully established.And the left 120 rats of two groups were randomly divided into six subgroups:STC group one to group six and control group one to group six,ten rats in each group,which were untreated,injected through tail vein with saline,and 0.001 ,0.010, 0.050,0.100 μg/L GDNF 2 mL respectively for one week.The expression of Sox-8 at protein level of either group were detected by Western blotting.Independent sample t test was performed for statistical analysis.Results After treated with 0.001 μg/L GDNF (STC group three),there was no significant
difference in expression level of Sox-8 between STC group three and STC group one (13.38 ±0.70 vs 13.39±0.45 ,t = 0.042,P = 0.969 ).After treated with 0.010 μg/L GDNF (STC group four),the difference in expression level of Sox-8 between STC group four and STC group three was significant (21 .11 ±2.56 vs 13.38±0.70,t=5 .040,P <0.01).After treated with 0.050 μg/mL GDNF (STC group five),the expression level of Sox-8 was higher than that in STC group four (31.86±1.57 vs 21.11±2.56,t=-6.198,P <0.01 ).The Sox-8 expression of untreated,saline treated,0.001 and 0.050 μg/L GDNF treated STC rats (STC group one,two,three and five)were lower than those of the corresponding control groups (t= 3.394,12.103,10.302,- 6.120,all P < 0.05 ).Conclusion Exogenous GDNF could increase Sox-8 expression in colon tissue of STC rats,an increase in the number of colon glial cells could repair enteric nervous system,and 0.050 μg/L was the optimal concentration.

Objective To Study on CXCR1/CXCR2 antagonist G31P anti-inflammatory reaction mediated by neutrophils.Methods Detect whether G31P can block chemotaxis of neutrophils induced by human IL-8 and inhibit the release of IL-8 by epithelia of segmental bronchus;establish HEK293 cell line transfected by pcDNA3.0-CXCR1 ,2,4 and detect the chemotaxis of IL-8 for HEK293 ;establish the experi-mental model of pneumonia induced by the P.aeruginosa,take count of the nucleated cells in the bronchoal-veolar lavage fluid(BALF),analyze myeloperoxidase(MPO) of lung tissue and observe the histopathology changing of it.Results G31P can inhibit the chemotaxis for neutrophils and transfected HEK293 cell line,inhibit the A549 releasing of inflammatory mediators;the proportion of neutrophils declines in G31P treat-ment group,pathology examination appears clear discrepancy.Conclusion G31P can block the chemotaxis of chemotactic factor with ELR+ CXC to neutrophils,block the combination of chemotactic factor with its re-ceptor CXCR2,block the CXCR2 on the surface of alveolar epithelia and vascular endothelial cells.Accordingly,neutrophils recruiting to topoinflammation can be prevented.

Objective To investigate the synergistic therapy effects of B and T lymphocyte attenuator(BTLA) extracellular domain in combination with heat shock protein 70 (HSP70)-TC-1 antigen peptide complex on the mouse model of cervical cancer and the related immunological mechanisms. Methods(1)Detecting the BTLA and herpesvirus entry mediator (HVEM) gene expression in the tumor microenvironment after C57BL/6 mice were inoculated with TC-1 tumor cells by realtime PCR; BTLA,HVEM expression on tumor infiltrating lymphocytes cell surface were detected by flow cytometry (fluorescence intensity). (2) According to different treatments, tumor-bearing mice were divided into 5 groups, which was injected with pcDNA3. 1 (empty vector plasmid as control), psBTLA (vector plasmid which expresses BTLA extracellular domain), HSP70 (HSP70-TC-1 cell peptide complex), HSP70 +pcDNA3.1 or HSP70 + psBTLA, respectively. The weight of tumor was recorded. The expression of immunoregulatory genes in tumor microenvironment were detected. The change of lymphocyte amount and cytotoxicity were detected too; lymphocyte proliferation activity was measured by tritium thymidine incorporation assay; the concentration of interleukin (IL) 2 and interferon-γ(IFN-γ) in supernatants of spleen lymphocyte were measured by enzyme-linked immunosorbent assay (ELISA). Results (1) BTLA gene expression was gradually increased after tumor cells inoculation. The highest expression level was 2. 83 + 0. 35 at 14th day, which had statistical significance difference with the 7th day expression of 1.66±0. 25 (P < 0. 05). While HVEM mRNA expression did not change significantly (P > 0. 05). The 7th and 14th day after TC-1 cells inoculation, the average fluorescence intensity of BTLA expression on the surface of tumor infiltrating lymphocytes was 33.5 and 51.8, respectively, in which there was statistically significant difference (P <0. 05); while the difference of HVEM expression was not statistically significant (57. 2 vs 49. 3 ,P >0. 05). (2)The 28th day after inoculation, tumor inhibition rate of HSP70 + psBTLA group was 88%, which was significantly higher than other treatment groups (P <0. 05). The 28th day after TC-1 cells inoculation, combination therapy not only promoted IFN-γ and IL-2 gene (3. 12 + 0.71,3.20 + 0. 62)expression but also reduced transforming growth factor-β (TGF-β), Foxp3 and IL-10 expression (0. 25±0. 03,0. 19 +0. 03,0. 31 +0. 04;P <0. 05). It also promoted CD8+ T lymphocyte infiltration(52 +6)/high power field, cytotoxicity (65.5±2.4) %, proliferation (15.0 × 103 cpm) and cytokine IL-2 , IFN-γsecretion(824±51), (1096±112) pg/ml, which were all significantly higher than other groups (P <0. 05). Conclusion The effect of immunotherapy on tumor can be augmented by the combination of psBTLA which expresses extracellular domain of BTLA and HSP70-TC-1 tumor antigen peptide complex,which could improve the expression of the related immunoregulatory genes to establish a much better microenvironment in favor of anti-tumor immune response against the mice model of the cervix carcinoma.

Objective To explore the effect of blocking BTLA-HVEM (herpesvirus entry mediator-B and T lymphocyte attenuator) pathway on dendritic cell function and the related immunological mechanisms. Methods Murine BTLA extracellular domain eukaryotic expression vector psBTLA was constructed by gene recombination and transfected CHO by Lipofection method. Mouse bone marrow cells were induced to differentiate into DCs by GM-CSF plus IL-4. Expression of BTLA and HVEM on DCs was detected after HSPT0-TC-1 peptide complex stimulation by FACS. Expression of BT-1 and secretion of IL-12 were detected after HSP70-TC-1 peptide complex plus psBTLA transfected CHO culture supernatant stimulation on DCs. Pretreated DCs co-cultured with the same genetic background mouse splenocytes and lymphocytes proliferation and cytokine secretion were detected. Effect of psBTLA gene transfer in vivo on BT-1 expression of DCs and tumor growth on tumor-bearing mice was detected. Results Extracellular domain of murine BTLA was successfully constructed, psBTLA stable transfection CHO cells were obtained and expression of BTLA extracellular domain(sBTLA) was detected the in its culture supernatant. BTLA and HVEM expression of DCs were increased after stimulation by the antigen peptide complex. When DCs were treated with antigen peptide complex plus culture supernatant containing sBTLA, B7-1 expression and IL-12 secretion were increased. Co-cultured with splenocytes, lymphocytes proliferation and cytokine secretion, such as IL-2 and IFN-γ,, were also increased. Gene transfection with psBTLA in vivo promoted B7-1 expression on DCs and inhibited cervical cancer cells growth. Conclusion Blockade of BTLA-HVEM inhibitory pathway with sBTLA can further improve DCs function, activation of lymphocytes and promote antitumor immune response.

Objective To assess the bone tunnel area at different times and sites of the tunnel after the anterior cruciate ligament(ACL) reconstruction in rabbits using Micro-CT.Methods Fifteen rabbits were performed ACL reconstruction using semitendinosus tendon autograft and randomly allocated into 3 groups and killed at 3,6,and 12 weeks after the operation.All samples undertook the micro-CT scanning(using SkyScan 1176,Bruker,U.S.A.) and were analyzed the areas of bone tunnels of femur and tibia after the 3-demension image rebuilding.For each tunnel,the area of the entrance,middle and exit of the tunnel were measured 3 times respectively and compared.Results The average area of the femoral tunnel did not change significantly with time,being 4.84 mm2,4.57 mm2 and 4.46 mm2 at 3,6 and 12 weeks after the operation(P=0.99).At the very beginning,the femoral tunnel area at the entrance was the biggest,while that of the middle was the smallest.Six weeks after the operation,significant differences were observed between the femoral tunnel area at the entrance and middle,as well as that between the exit and middle(P=0.0011,P=0.0106);However,12 weeks after the operation,significant differences were observed only between that at the entrance and middle(P=0.0227).The average tibial tunnel area increased significantly at 6 weeks(6.577 mm2) and decreased at 12 weeks(3.103 mm2) after the operation(P=0.0005).Moreover,no significant differences were observed in the average tibial tunnel area at different time points and sites(P＜0.05).At different sites,the average tibial tunnel area expanded at 6 weeks,and then declined at 12 weeks after the operation.Conclusion The bone tunnel area changes with time after the ACL reconstruction,first increasing followed by decreasing in the average tibial tunnel area.The femur and tibial tunnel have significant differences in the tunnel area at different sites,which change differently with time.The bone tunnel expansion after the anterior cruciate ligament reconstruction can be comprehensively measured repeatedly at different sites.

Ojective To explore the effect of the hydroxyapatite(HAp)and gelatin(Gel)coating on the healing of the polyethylene terephthalate(PET)artificial ligament.Methods The artificial ligaments were divided into a PET group with a pure PET surface and a PET/HAp/Gel group coated with HAp and Gel.Both coatings were observed using the scanning electron microscope(SEM).Forty-eight male New Zealand rabbits were randomly divided into two groups and underwent anterior cruciate ligament reconstruction,before two kinds of artificial ligaments were implanted respectively.Four weeks and 8 weeks after the operation,the rabbits were sacrificed,and histological hematoxylin and eosin (HE)staining as well as the biomechanical examination were performed.Results HAp/Gel coating was found depositing on the surface of PET artificial ligaments.Histological HE staining showed a thick fibrous connective tissue forming at the graft-host bone interface 4 weeks postoperatively,and the interface width of both groups were narrowed,with significantly more shrinking in the PET/HAp/Gel coating group.And new bone tissues were found in the interface of PET/HAp/Gel group 8 weeks after the operation.The biomechanical examination found significant differences in the failure load between the PET(46.16 ± 2.88 N) and PET/HAp/Gel group(71.32 ± 3.92 N)8 weeks after the surgery(P=0.0021).And 4 weeks and 8 weeks after the surgery,significant differences were found in the stiffness between the PET group and the PET/HAp/Gel group(11.06 ± 1.14 N/mm vs 16.20 ± 1.17 N/mm,P=0.0199;14.37 ± 0.88 N/mm vs 24.35 ± 1.35 N/mm,P=0.0008).Conclusion HAp/Gel coating can enhance the osteogenesis of PET artificial ligaments,promoting the new bone formation at the graft-host bone interface and herein strengthening the graft-host bone healing.

Objective To explore the residual level of FPMs in indoor dust samples in Shenzhen from 2020 and 2021, and to analyze its temporal distribution characteristics. Methods In the present study, indoor dust samples (n=193) from residential buildings in Shenzhen. were collected to analyze the temporal variation characteristics of FPMs. Ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was applied to determine the concentrations of FP and its four major metabolites (fipronil-sulfone, fipronil-sulfide, fipronil-desulfinyl, and fipronil-amide; abbreviated as FP-SFO, FP-SFI, FP-DES, and FP-AM) in the samples. The sum of the concentrations of FP and its four metabolites was represented as additive mass concentration (ΣFPMs). Additionaly, Wilcoxon test was performed to determine the temporal distribution differences of FPMs’ concentrations. Results From 2020 to 2021, the concentration of ΣFPMs for the in door dust samples in Shenzhen ranged from 0.51 to 4 415 ng/g (median: 18.8ng/g). FP, FP-SFO AND FP-SFI were the major target analytes in the sample with detection rates of 90.60%，86.20% and 75.40%, respectively. The detection rates of other metabolites were low (≤ 44.3%). Analysis of the temporal variation trend of FPMs’ concentrations showed that there was no significant difference in the levels of ΣFPMs between warm season（spring and summer）and cold season（autumn and winter）in the indoor dust samples from 2020 to 2021(2.38 vs 2.84ng/g , P > 0.05). However , the concentrations of FP-SFI and ΣFPMs in the indoor dust samples collected from 2021 showed an significantly increasing trend compared with 2020（1.02 vs 1.89 , 17.80vs. 20.10 ng/g , P < 0.05). Conclusion From 2020 to 2021 , the detection level of FPMs in indoor dust in Shenzhen is relatively high and shows an upward trend , with no obvious seasonal difference. However, whether the residual level of FPMs in indoor dust poses a risk to human health needs further study.