Patients with severe trauma require an extremely timely treatment and transfusion plays an irreplaceable role in the emergency treatment of such patients. An increasing number of evidence-based medicinal evidences and clinical practices suggest that patients with severe traumatic bleeding benefit from early transfusion of low-titer group O whole blood or hemostatic resuscitation with red blood cells, plasma and platelet of a balanced ratio. However, the current domestic mode of blood supply cannot fully meet the requirements of timely and effective blood transfusion for emergency treatment of patients with severe trauma in clinical practice. In order to solve the key problems in blood supply and blood transfusion strategies for emergency treatment of severe trauma, Branch of Clinical Transfusion Medicine of Chinese Medical Association, Group for Trauma Emergency Care and Multiple Injuries of Trauma Branch of Chinese Medical Association, Young Scholar Group of Disaster Medicine Branch of Chinese Medical Association organized domestic experts of blood transfusion medicine and trauma treatment to jointly formulate Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients ( version 2024). Based on the evidence-based medical evidence and Delphi method of expert consultation and voting, 10 recommendations were put forward from two aspects of blood support mode and transfusion strategies, aiming to provide a reference for transfusion resuscitation in the emergency treatment of severe trauma and further improve the success rate of treatment of patients with severe trauma.

Objective To establish a mouse model treated with busulfan and to investigate its effects on the metabo-lism of spermatogonial stem cells(SSCs)of mouse testis.Methods C57BL/6J male mice with age of 8 weeks were injected with 10 mg/kg of busulfan intraperitoneally,then Thy1 positive cells were selected by immunomagnetic beads on day 0,day 5 and day 10 and followed by identification for purity and metabolomic analysis.Results The testis weight ratio decreased and the tissue structure of testis was damaged(P＜0.05).Based on the results of principal component analysis(PCA)and partial least squares discriminant analysis(PLS-DA),there were signifi-cant metabolic differences between the sample groups treated for 0 d,5 d and 10 d.A total of 89 differential metabolites were identified including glutathione(GSH),arginine and unsaturatedfatty acids(UFAs),and their important metabolic pathways involved glycerophospholipid metabolism,arginine and proline metabolism.Conclu-sions Affecting the specific metabolic pathway may result in obvious reproductive toxicity and lead to decrease of testicular weight as well as tissue structure damage in mice.Metabolomic analysis showed that the potential repro-ductive toxicity mechanism of SSCs may be related to the metabolic pathways such as lipid metabolism,arginine and proline metabolism.

Objective To evaluate availability and outcomes of using anterior half of the peroneus longus tendon (AHPLT) in knee ligament reconstruction as an autograft source.Methods From July 2007 to January 2008,100 patients with knee ligament injuries were recruited in this study.There were 33 males and 67 females aging from 16 to 62 years (mean,32.3 years).49 cases had undergone medial patellofemoral ligament reconstruction,19 cases multiligament reconstruction,18 cases double-bundle posterior cruciate ligament (PCL) reconstruction and 14 cases double-bundle anterior cruciate ligament (ACL) reconstruction.AHPLT was used as sole (49 cases) or part (51 cases) of reconstruction materials.One-incision or two-incision striping techniques were adopted to harvest AHPLT.Ligaments were fixed with screws.Post-operative assessments included Kujala knee score,Lysholm knee score,Marx knee score,International Knee Documentation Committee (IKDC) subjective evaluation form and objective evaluation grade,the Foot and Ankle Disability Index (FADI) and the American Orthopedic Foot and Ankle Society (AOFAS) scale.Results 92 cases were followed up for more than 24 months.Postoperative Kujala score,IKDC subjective score,Lysholm score and Marx score were improved significantly in all four groups of patients.According to IKDC objective evaluation grade,the number of patients reaching Grade A (normal) or Grade B (near-normal) in multiligament,PCL and ACL reconstruction were 17,15 and 12,with an excellent rate of 89.5％ (17/19),93.7％ (15/16) and 100％ (12/12),respectively.Preoperative and postoperative AOFAS scores were 97.4±2.0 and 97.2±1.6,respectively,while the FADI scores preoperatively and postoperatively were 96.8±2.2 and 96.9±2.5,respectively.These results had no statistical significance.No signs of peroneal nerve injury or peroneus longus tendon rupture was found.Conclusion It is acceptable to use AHPLT as an autograft due to its feasibility to harvest,good clinical outcome,and low rate of donor site morbidity at a minimum of two-year follow-up.

To prepare recombinant human retinol binding protein 4 (RBP4) by using the baculovirus expression system and to detect its immunogenicity, the fusion DNA fragment of secretory signal peptide SS64 and human RBP4 gene was subcloned into a baculovirus transfer vector pFastBac-dual(pFBd), and the corresponding recombinant transfer plasmid was transformed into E. coli strain DH10bac, after transposition recombinant shuttle bacmid was screened out. The logarithmic phase Sf9 cells were transfected with the recombinant bacmid and then the recombinant baculovirus containing hRBP4 expression box were generated. After amplification of recombinant baculovirus, the recombinant baculovirus seeds were obtained. To express human RBP4, logarithmic phase Sf9 cells were infected with the virus seeds and SDS-PAGE and Western blotting were used to detect and identify the expression. Finally, to prepare a batch of RBP4 protein, logarithmic phase Sf9 cells in suspension culture were infected with recombinant baculovirus seeds and the supernatant was harvested after 120 hours post-infection for purification. Finally for preparation of polyclonal antibody and evaluation of immunogenicity, the recombinant hRBP4 from insect cells and from E. coli were immunized rabbits. Restriction enzyme digestion and sequencing confirmed that the recombinant baculovirus transfer plasmid was constructed correctly, and subsequently recombinant RBP4-bacmid was generated successfully. SDS-PAGE and Western blotting analysis suggested that human RBP4 protein was highly expressed in Sf9 cells with the molecular weight of approximately 23 kDa. The recombinant RBP4 protein could be secreted into the medium efficiently, and the expression level was calculated amount of 100 mg/L. Finally the rabbit antiserum was harvested after recombinant RBP4 immunization, therein the titer of antiserum against baculovirus recombinant RBP4 is 1:100 000 whereas the titer of antiserum against E. coli recombinant RBP4 is only 1:10 000. Overall, human RBP4 was high efficiently expressed successfully with good antigenicity in baculovirus system, and high affinity antiserum was obtained. A solid foundation was laid for the next step of the preparation of human serum RBP4 detection kit.
Animals ; Baculoviridae ; genetics ; Blotting, Western ; Cloning, Molecular ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; Genetic Vectors ; Humans ; Immune Sera ; Insecta ; Rabbits ; Recombinant Proteins ; biosynthesis ; immunology ; Retinol-Binding Proteins, Plasma ; biosynthesis ; immunology ; Sf9 Cells ; metabolism ; Transfection

Objective To investigate the effects of IL-12 coexpression level on antigen expression and immune responses induced by HBsAg DNA vaccination. Methods DNA vaccine plasmid pHBV carrying codon-optimized preS2-S gene of reference sequence CHN-HBV07-C in China was constructed. Three DNA vaccine plasmids pHBV-12i, pHBV-12l and pHBV-12h were also constructed by subcloning three different IL-12 expression cassettes with various expression strengths to plasmid pHBV, respectively. Expression levels of IL-12 and HBsAg in vaccine plasmid-tranfected 293T cells were measured by quantitative ELISA. DNA vaccines were administered intramuscularly to BALB/c mice and HBsAg-specific cellular immune responses were determined by IFN-γ ELISPOT. HBsAg-specific antibodies were tested by Chemiluminescence Quantitative Immunoassay. Results The HBsAg expression level in 293T cells was 70 ng/ml when transfected by plasmid pHBV without IL-12 expression cassette, and the HBsAg level was 18 ng/ml when transfected by plasmid pHBV-12l carrying low-level IL-12 expression cassette, whereas the HBsAg level was only 6 ng/ml when transfected by plasmid pHBV-12h carrying high-level IL-12 expression cassette.Results of DNA vaccination revealed that HBsAg-specific humoral and cellular immune responses were significantly decreased in mice administering vaccine pHBV-12h carrying high-level IL-12 expression cassette. Although HBsAg-specific antibody responses in mice inoculated with pHBV-12l were also decreased when compared with those in pHBV-vaccinated mice without IL-12 expression, the HBsAg-specific cellular immune responses were significantly increased. Conclusion High-level coexpression of IL-12 may suppress the expression of HBsAg, Whereas modest coexpression of IL-12 significantly enhanced the HBsAg-specific T cell responses induced by DNA vaccination. Therefore, it is so important to balance the expression between adjuvant and antigen to enhance the immune response.

Objective To investigate the safety and feasibility of endovascular stent angioplasty in treating symptomatic stenosis of middle cerebral artery.Methods Endovascular angioplasty with coronary stents was performed in 27 patients with symptomatic stenosis of middle cerebral artery.The clinical results were reviewed and analyzed.Results Of the total 27 patients,successful placement of the coronary stents Was achieved in 24.Angiography immediately after the procedure showed that the stenotic degree of the diseased artery was markedly decreased from preoperative (80±19)%to postoperative (8±4)%,the improvement was very obvious.Percutaneous transcatheter angioplasty had to be employed in two cases because of the failure of stent placement.A mean follow-up period of 18 months was carried out.During the following up period no transient cerebral ischemia attack occurred in 25 patients and no newly-developed cerebral infarction in region fed by the responsible vessels occurred either.Re=irrigation cerebral hemorrhage was seen in one patient,which occurred three hours after the placement of the stent.In one case the placed stent fell off and immigrated into the siphon of internal carotid artery,and the displaced stent Was took out later with a catching apparatus.In another case re-stenosis occurred six months after the stenting.Conclusion Percutaneous endovaacular stent angioplasty is a safe and effective treatment for symptomatic stenosis of middle cerebral artery,although its long-term results need to be further evaluated.

Objective The aim of the study WSfl to establish the method using CDl27 as the new biomarker to identify regulatory T cells(Treg cells)and apply the CD 127 to detect the Treg cells in patients with gastric cancer.Methods The phenotypes of Treg cells were analyzed using five-color flow cytometry method Foxp3.FITC/CD127-PE/CD4-PerCP/CD25-APC/CD3-PC7.The mRNA and protein expression of Foxp3 in isolated CD+4 CD25high CD127-/low Treg cells were detected.The relationships between Foxp3 and CD127 protein expression in CD4+ T cells from aduh human peripheral blood were investigated.PBMCs,Ascltes,turnor-infihration lymphocyte and tumor-draining lymph nodes in 35 patients with gastric cancer and PBMCs in 20 normal healthy donors were evaluated for the proportion of Treg ceils,as well as the percentage ot the total CD +4 cells.Results CD4+ CD25 high CD-127 low ceils expressed the high Foxp3 in protein level(87.1%)and mRNA level.Within the CD4+CD+25 population,there was a significant correlation between Foxp3 and the CD127low phenotype(r=0.985,P<0.01).Compared with healthy olunteers,patients with gastric malignancies had a higher proportion of CD4+4 Cdhigh 25 CD-low127 cells in peripheral blood(t=2.542,P<0.05).The Dereentages of Treg cells were more abundant in ascites(t=2.357,P<0.05),TIL(t=6.174,P<0.01) and tumor-draining lymph nodes(t=5.481,P<0.01)of individuals with gastric cancer than that in their blood.There were significant differences in the prevalence of Treg ceils between the early and advanced disease stages in gastric cancer[(6.04±2.31)%in stageⅠ+Ⅱ VB(10.16±2.29)% Ⅲ+Ⅳ,t=2.473,P<0.05].Conclusions The CD127 biomarker can be used to selectively enrich human Treg cells for in vitro functional studies.The populations of CD4+ CD high25 CD127-/low Treg cells increased ith tumor stage in individuals with gastric cancer.

Objective To investigate the populations of CD+4CD25high Foxp3+ regulatory T cells(Treg cells)and mRNA expression of Foxp3 in peripheral and the marginal region of tumor from patients with gastric cancer,and to evaluate the prevalence of Treg cells from patients with gastric cancer in relation to the TNM stages.Methods PBMC,Ascites,tumor-infiltration lymphocyte and tumor-draining lymph nodes in 47 patients with gastric cancer(TNM I11,Ⅱ18,Ⅲ10,Ⅳ8)and PBMC in 20 normal healthy donors were evaluated for the proportion of Treg cells,as a percentage of the total CD4+ cells,by flow cytometric analysis.Levels of mRNA for Foxp3 were measured witll a real-time quantitative PCR Results The percentage of CD4+CD25high Foxp3+ Treg cells in PBMC for cases of gastric cancer were significantly higher than those for healthv donors(P<0.05).The percentage of Treg cells were more abundant in ascites(P< O.05),1rIL(P<0.01)and tumor-draining lymph nodes(P<0.01)of individuals with gastric cancer than in their blood.And the Foxp3 mean fluorescent intensity (MFI) of Treg cells in TIL and tumor draining lymph nodes with gastric caners were significantly higher than PBMC and ascites(P<0.05).The proportion 0f Treg cell in patients of gastric cancers with stage I,II,Ⅲ,Ⅳ were(5.72±1.95)%,(6.45± 2.23)%,(9.58±3.13)%,(11.70±2.30)%,respectively.There were significant correlation between Drevalenee of Treg cells and disease stages in gastric cancer(r=0.784,P<0.01).Foxp3 mRNA levels were higher in PBMC from the group with gastric cancers than in the group with healthy donors.Foxp3 mRNA levels were correlated with TNM stages(r=0.623,P<0.05).Conclusions Our results suggest that the populations of CD4+CDhigh25 Foxp3+Treg cells and Foxp3 mRNA levels in patients with gastric cancer are signifieantlv higher in comparison to those in control donors.In addition,the expression of Fox：p3 mRNA increased with tumor stage.The increased prevalence of Treg cells may be one of the explanations for impaired cell-mediated immunity in cancer bearing hosts.

Objective To summarize the experiences of "three holes and one hook in the end(TOE)" laparoscopic cholecystectomy(LC) in 1 260 cases and to investigate the operation procedures,technical points and the prevention of complications.Methods The data of 1 260 patients suffering from chronic calculous cholecystitis,acute calculous cholecystitis,atrophic cholecystitis,gallbladder polyps etc.,who were admitted to this hospital and treated by TOE from March 1999 to March 2008 were included and analysed retrospectively in this study.Results One thousand two hundred and sixty of cases were cured,including 1 252 cases of succeeding LC(99.37%),conversions to open in 8 cases,no death,no bile duct injury,with intraoperative hemorrhage in 3 cases,umbilicus infection in 2 cases,gallbladder fossa hydrops in 3 cases,with operation time for 8-60 min(mean 38.5 min) and hospitalization for 3-7 d(mean 5 d) after surgery.During the follow up of 1 002 cases for 1 to 7 years(mean 3.5 years),there were no complications such as bile fistula,bile duct stricture,residual stones of biliary duct,etc..Conclusion TOE is worthy of application and promotion for the excellent effectiveness,few complications,rapid recovery and safety.

Scaffolds provide a three-dimensional environment that is desirable for inducing and promoting the production of cartilaginous tissue.Ideally the scaffold should:① have directed and controlled degradation;② promote cell viability,differentiation,and extracellular matrix production;③ adhere and integrate with the surrounding native cartilage;④ span and assume the size of the defect;⑤ provide mechanical integrity depending on the defect location;⑥ have no toxicity,no immunogenicity or inflammatory induction;⑦ allow for the diffusion of nutrients and waste products.To date,a wide range of natural and synthetic materials have been investigated as a scaffold for cartilage repair.Based on the morphology and structure,these materials can be divided into hydrogel,sponge,fiber mesh and so on.Natural polymers that have been explored as bioactive scaffolds for cartilage engineering include:alginate,agarose,fibrin,hyaluronic acid,collagen,gelatin,chitosan,chondroitin sulfate,and cellulose.Synthetic polymers currently explored for cartilage repair include poly(?-hydroxy esters),polyethylene glycol,poly(NiPAAm),poly(propylene fumarates),and polyurethanes.