AIM: To investigate the effects of deep-frozen treatment on the immune response of bone-patellar tendon-bone(BPB) xenograft rejection.METHODS: A muscle pocket model was used to study the immune response of rat to deep-frozen treated BPB xenograft from guinea pig,autograft and fresh xenograft served as controls.The expression of T-cell surface activation antigen CD25 in the peripheral blood and the morphological changes of the implants were used to measure the immune response.RESULTS: The expression of CD25 in CD4+,CD8+ cells greatly increased 3 d after fresh BPB xenograft,the obvious difference to that of autograft(P

Objective To investigate the cellular immune response to the recombinant DNA vaccine CRT120/HPV6bE7 in mice. Methods The recombinant encoding HPV6bE7, linked with CRT120, was constructed in pcDNA3.1 eukaryotic vector with the report gene GFP. The pcDNA3.1-GFP -HPV6bE7 was transfected into B16 cells by a lipofectamine kit. The C57BL/6 mice were vaccinated with recombinant DNA plamids. The T-cell phenotype in the peripheral blood lymphocytes of the immunized mice was measured by flow cytometry. The CTL activity of the lymphocytes from the spleens and lymph nodes, and the levels of IL-2 and IFN-? were analyzed. Results The constructed recombinant plasmid CRT120/HPV6bE7 was analyzed. Positive transfected cell clones were established and could stably express the target gene HPV6bE7. Compared with HPV6bE7, CRT120/HPV6bE7 plasmid enhanced to a greater extent CD8+ T-lymphocyte differentiation, the number of TCR?? T-lymphocytes and the levels of IL-2 and IFN-? (all P

Objective To construct four expression plasmids, pcDNA3.1-GFP/HPV6bE6, pcDNA3.1-GFP/HPV6bE7, pcDNA3.1-GFP/HPV11E6, pcDNA3.1-GFP/HPV11E7 and their transfected murine cell lines. Methods The Four recombinant expression plasmids comprising HPV6bE6,HPV6bE7,HPV11E6 and HPV11E7 linked with GFP, respectively, were constructed and transfected to B16 cells by lipofectamine kit. Positive clones were selected by G418 and observed by fluorescent microscopy and identified by RT-PCR. Results The four constructed recombinant plasmids were authenticated by restriction enzyme digestion and DNA sequencing. Under the fluorescent microscope, the green fluorescence could be observed in cytoplasm and nucleus of four transfected B16 cell lines. The RNA extracted from positively transfected clones resistant to G418 were analyzed by RT-PCR, which demonstrated the presence of four expected fragments. Conclusions The transfected murine cell lines B16 can express HPV6bE6,HPV6bE7,HPV11E6 and HPV11E7 gene. These transfected cell lines can be further transplanted to mice in order to investigate the biological properties and immunological mechanisms of these genes in vivo.

Objective To summarize clinical experience of one-station therapy for infected seriouslyischemic diabetic foot.Methods The clinical data of 15 patients (15 diseased limbs in total) with infected seriously-ischemic diabetic foot,who were admitted to authors' hospital during the period from June 2015 to April 2016 to receive treatment,were retrospectively analyzed.For all patients,one-station sequential therapy was carried out,which included endovascular revascularization (EVR) to open occluded vessel,surgical debridement and closed negative pressure wound drainage and antiseptic moisturizing wound dressing.The healing rate of infected wound and the limb salvage rate were evaluated.Results The 15 patients included 10 males and 5 females,with a median age of 77 years old.Lower extremity angiography showed that multiple segmental lesions of lower limb were detected in 13 patients and simple leg lesions in 2 patients.According to TASC Ⅱ update classification,leg artery disease of grade D was observed in 13 patients and artery disease of grade C in 2 patients.After EVR therapy,at least one branch of leg arteries was reopened in 14 limbs.Intact arterial arch of pedal-plantar loop (PPL) was seen in 6 patients,semi-arterial arch in 7 patients,and absent of arterial arch in 2 patients.After surgical debridement,the wound was washed by using negative pressure wound therapy (NPWT) device as well as serf-made washing equipment.The time to control wound infection was (7.85±2.84) days.After discharge,the patients were followed up every 3-4 days,at the same time wound dressing exchange with antibacterial moisturizing sulfadiazine silver lipid hydrogel was conducted.Wound healing was achieved in 12 patients,and the mean healing time was (3.70±0.87) months.The wound failed to heal in 3 patients,among them below knee amputation had to be performed in 2 patients (13.3％,both patients showed absent of arterial arch of PPL),and the remaining one patient died of cardiovascular event.Statistically significant difference in PPL pathological changes existed between wound healing group and wound un-healing group (P=0.006 7).Conclusion The treatment of infected seriouslyischemic diabetic foot is rather complicated.Being one-station therapy,the sequential managements,which include EVR,NPWT device together with washing equipment and use of antibacterial moisturizing wound dressing,can effectively increase the blood supply to the affected limb,shorten the time to control infection and lower amputation rate.Therefore,one-station therapy should be regarded as the preferred method for infected seriously-ischemic diabetic foot.

Objective To investigate the protective effect of sulodexide (SDX) on oxidized low density lipoprotein (ox-LDL) induced damage to human umbilical vein endothelial cell (HUVEC),and to discuss its mechanism.Methods By using CCK-8 method,the ox-LDL intervention HUVEC dose and the concentration of SDX were determined.The reactive oxygen species (ROS) assay kit was used to verify the protective effect of SDX on HUVEC.Real time fluorescent quantitation-polymerase chain reaction (RT-PCR) was employed to test the endothelial nitric oxide synthase (eNOS) and caveolin-1 mRNA expression;immunoblot assay was adopted to check the protein expression of phosphorylated eNOS (p-eNOS) and caveolin-1.The ability of cell migration was assessed by Transwell assay.Results Stimulated by 100 μg/ml concentration of ox-LDL,the cell viability of HUVEC decreased significantly (P＜0.01).After adding 125 LRU/ml concentration of LDX,the cell viability of HUVEC was remarkably improved (P＜0.01) and the production of ROS was strikingly decreased (P＜0.01).SDX could down-regulate the expression of caveolin-1 (P＜0.05) and up-regulate the expression of eNOS mRNA and p-eNOS (P＜0.05) for ox-LDL-damaged HUVEC,and markedly improve the migration ability of damaged HUVEC (P＜0.01).Conclusion By regulating the caveolin-1/eNOS signal route,SDX can improve impaired HUVEC cell migration ability,thus,to protect endothelial cells.

Objective To summarize the medical rescue of Chinese Government Medical Team (Chongqing) in Nepal earthquake region in order to explore the work pattern of transnational medical rescue,and improve the rescue efficiency.Methods From the experience about the post-earthquake medical rescue of Chinese Government Medical Team (Chongqing) in Nepal in 2015,several aspects were worthy to summarize such as the establishment of medical team,the layout of camp site,the work algorithm and process,with the analysis of injury feature and outcome of treatment.Results Under the setting of efficient organization and rational assignment of professional work,special working rules,the mutual transfer treatment and multi-disciplinary treatment were employed for 737 emergency patients.Of them,128 patients were hospitalized (including 63 patients completely recovered,56 patients were of clinical improvement,and 9 patients were critically ill transferred to other hospital for advanced treatment),and post-traumatic complication occurred in 48 cases without death.In addition,148 operations were carried out successfully.Conclusions The earthquake disaster has specific feature such as suddenness,a host of casualties and poor rescue conditions,and overseas rescue is with the presence of the language barrier,the difference in living habit,and the lack of coordination with local rescue system,therefore,rationally assigning personnel and resources and establishing work pattern with flexibility,orderly and good communication are the key to promote the efficiency of transnational medical rescue for the injured patients in earthquake region.

Objective To screen and identify the predicted epitopes of synthesized HLA-A*0201restricted CTL derived from HPVll E7 antigen.Methods Five HPVll E7 CTL epitope peptides and terramers consisting of HLA-A*0201 were selected by way of computer and synthesized by Sanquin company,including HPVllE7 7-15(TLKDIVLDL),15-23(LQPPDPVGL),47-55(PLTQHYQIL),81-89(DLLLGTLNI)and 82-90(LLLGTLNIV).These peptides binding to human peripheral blood-derived DCs were tested for their ability to activate T cells isolated from peripheral blood lymphocytes of HLA-A*0201 healthy individuals.the number of specific tetramer+CD8+T cells by flow cytometry,the level of the section of IFN-γ by ELISA,and the ability of the CTL to kill the target cells were observed.Results The immature DCs could be fully activated by all the five HPV11 E7 peptides.Peptide-loaded mature DCs were able to stimulate the epitope-specific T cells responses in vitro.An increased frequency(P<0.05)of T ceils specific for the E7 7-15 epitope compared to other epitopes of HPV11E7.The epitope-specific CTL of E7 7-15 induced by the activated DCs specifically killed HPV11E7 expressing 293 cell line,and in a ratio of 50:1,the specific cytolytic activity was the strongest than the others(P<0.05).Conclusion DCs loaded with HPV11 E7 7-15(TLKDIVLDL)peptide can induce highly effective and specific ectogenic processed epitopespecific CTL responses in vitro.This peptide may be the candidate for development of CTL based vaccine in the treatment of HPV infeetions.

OBJECTIVETo investigate the protective effects of hepatic fibrosis against a lethal dose of acetaminophen (APAP) and its underlying mechanisms using a carbon tetrachloride (CCl4)-induced mouse model of fibrosis.

METHODSThe experimental model of hepatic fibrosis was established by intraperitoneal injection of CC14 (in mineral oil), twice a week for 6 weeks; mice given a 6-week course of mineral oil injections served as normal controls. At the end of fibrosis induction, the expmimental (Fib group) and control (Norm group) mice were challenged with APAP (1 g/kg). Sera and liver tissues were harvested for analyses.To assess tolerance of the normal and fibrotic mice to the lethal dose of APAP, the survival rate,serum alanine aminotransferase (sALT) levels and hepatic histopathological changes were compared before and after the acute APAP challenge.HMGB 1 expression was analyzed by immunohistochemistry.One-way ANOVA test and Newman-Keuls test were used in statistical analysis.

RESULTSThe fibrotic liver was tolerant to the lethal dose of APAP,as evidenced by:(1) significantly higher survival rate in the Fib ± APAP group (80% vs. Norm+APAP group: 0%); (2) markedly lower sALT levels in the Fib+APAP group (6 437 ± 1 913 U/L vs. 12 456 ± 3 441 U/L), P=0.022; (3) remarkably well-preserved liver architecture in the Fib+APAP group.Immunohistochemical analysis showed high HMGB1 expression and cytoplasmic translocation in the Norm+APAP group,which was absent in the Fib+APAP group.

CONCLUSIONSCCl4-induced liver fibrosis protects mice against lethal dose of APAP, Possibly by a mechanism involving inhibition of the cytoplasmic translocation of HMGB1.

Objective:To explore the oxidative stress of cerebral white matter lesion (WML) and normal-appearing white matter (NAWM) with in vivo proton exchange rate (k ex) MRI on relapse-remitting multiple sclerosis (RRMS) patients. Methods:Clinical and imaging data of 37 patients (case group) with RRMS patients of Tongji Hospital of Tongji Medical College of Huazhong University of Science and Technology were analyzed retrospectively from November 2018 to November 2021, including 11 males and 26 females aged 18-41 (29±7) years. Another 22 age-matched healthy volunteers (control group) were recruited for the same period, including 4 males and 18 females aged 23-44 years with a median age of 25 (24, 28.25) years. All subjects received conventional MR protocols and chemical exchange saturation transfer imaging. The manifestation of WML on the k ex map and T 1WI images were assessed while the k ex values of WML, NAWM and normal white matter (NWM) of control group were quantitatively evaluated. Student′s t test was used to compare the k ex difference of WML and NAWM in the case group, NAWM in the case group and NWM in the control group, low-signal and isosignal WML in T 1WI. Spearman rank correlation was used to analyze the correlation of the k ex values of WML with patients′ expanded disability status scale (EDSS) score. Results:A total of 272 WML were found in the 37 RRMS patients, and 25.4% (69/272) were T 1-hypointense. The k ex value of WML in the case group [(932±108) s -1] was higher than that of NAWM [(771±26) s -1], and the difference was statistically significant ( t=8.95, P<0.001); the k ex value of NAWM in the case group [(771±26) s -1 ] was higher than that of NWM [(745±26) s -1] in the control group, and the difference was statistically significant ( t=3.96, P<0.001). The k ex value [(1 039±110) s -1] of WML with low signal at T 1WI was higher than that of WML with equal signal [(895±79) s -1], with a statistically significant difference ( t=9.78, P<0.001). Correlation analysis showed that the k ex value of WML in the case group was positively correlated with the EDSS score ( r=0.54, P<0.001). Conclusions:The elevated k ex values of WML and NAWM reflect the cerebral oxidative stress of RRMS patients and are positively correlated to the severity of tissue damage, which suggests the role of oxidative stress in RRMS lesion formation and brain atrophy.

Objective To observe the alterations of macular vascular density and the area of foveal avascular zone (FAZ) in branch retinal vein occlusion (BRVO) eyes.Methods A retrospective case-control study. Forty-five patients with unilateral BRVO and macular edema were enrolled in this study. Optical coherence tomography angiography (OCTA) was performed on the BRVO and fellow eyes. The scanning region in the macular area was 3 mm×3 mm. Macular vascular density and FAZ area in the superficial and deep retinal capillary plexi were measured in all eyes. The values of macular vascular density and FAZ area between BRVO eyes and fellow eyes, affected sector and unaffected sector were compared.Results The mean overall vascular density measured in the entire scan was lower in BRVO eyes compared with fellow eyes in both the superficial and deep capillary plexus (t=14.186, 9.468;P<0.05). The reduce degree of vascular density in the deep capillary plexus (7.65％) was higher than that in the superficial plexus (7.27％). In the superficial plexus, the vascular density was lower in the affected sector and the unaffected sector of the BRVO eyes compared with the corresponding sector in the fellow eyes (t=15.386, 9.435;P<0.05). The FAZ area enlarged in the BRVO eyes compared with the fellow eyes in the superficial capillary plexus and in the deep capillary plexus (t=3.216, 5.119;P<0.05). The degree of enlargement of FAZ area in the deep capillary plexus (0.19 mm2) was higher than that in the superficial plexus (0.11 mm2).Conclusions In eyes with BRVO, quantitative OCTA measurements confirm that vascular density decreased and FAZ area enlarged in the superficial and deep capillary plexi. The reduce degree of vascular density and enlargement degree of FAZ area in the deep capillary plexus are higher than those in the superficial plexus.