Hypomethylation of glycine dehydrogenase promoter in peripheral blood mononuclear cells is a new diagnostic marker of hepatitis B virus-associated hepatocellular carcinoma

Background: Glycine dehydrogenase(GLDC) plays an important role in the initiation and proliferation of several human cancers. In this study, we aimed to detect the methylation status of GLDC promoter and its diagnostic value for hepatitis B virus-associated hepatocellular carcinoma(HBV-HCC). Methods: We enrolled 197 patients, 111 with HBV-HCC, 51 with chronic hepatitis B(CHB), and 35 healthy controls(HCs). The methylation status of GLDC promoter in peripheral mononuclear cells(PBMCs) was identified by methylation specific polymerase chain reaction(MSP). The mRNA expression was examined using real-time quantitative polymerase chain reaction(q PCR). Results: The methylation frequency of the GLDC promoter was significantly lower in HBV-HCC patients(27.0%) compared to that in CHB patients(68.6%) and HCs(74.3%)( P < 0.001). The methylated group had lower alanine aminotransferase level( P = 0.035) and lower rates of tumor node metastasis(TNM) Ⅲ/Ⅳ( P = 0.043) and T3/T4( P = 0.026). TNM stage was identified to be an independent factor for GLDC promoter methylation. GLDC mRNA levels in CHB patients and HCs were significantly lower than those in HBV-HCC patients( P = 0.022 and P < 0.001, respectively). GLDC mRNA levels were significantly higher in HBV-HCC patients with unmethylated GLDC promoters than those with methylated GLDC promoters( P = 0.003). The diagnostic accuracy of alpha-fetoprotein(AFP) combined with GLDC promoter methylation for HBV-HCC was improved compared with that of AFP alone(AUC: 0.782 vs. 0.630, P < 0.001). In addition, GLDC promoter methylation was an independent predictor for overall survival of HBV-HCC patients( P = 0.038). Conclusions: The methylation frequency of GLDC promoter was lower in PBMCs from HBV-HCC patients than that from patients with CHB and HCs. The combination of AFP and GLDC promoter hypomethylation significantly improved the diagnostic accuracy of HBV-HCC.

Peripheral blood RNA biomarkers can predict lesion severity in degenerative cervical myelopathy

Degenerative cervical myelopathy is a common cause of spinal cord injury,with longer symptom duration and higher myelopathy severity indicating a worse prognosis.While numerous studies have investigated serological biomarkers for acute spinal cord injury,few studies have explored such biomarkers for diagnosing degenerative cervical myelopathy.This study involved 30 patients with degenerative cervical myelopathy(51.3±7.3 years old,12 women and 18 men),seven healthy controls(25.7±1.7 years old,one woman and six men),and nine patients with cervical spondylotic radiculopathy(51.9±8.6 years old,three women and six men).Analysis of blood samples from the three groups showed clear differences in transcriptomic characteristics.Enrichment analysis identified 128 differentially expressed genes that were enriched in patients with neurological disabilities.Using least absolute shrinkage and selection operator analysis,we constructed a five-gene model(TBCD,TPM2,PNKD,EIF4G2,and AP5Z1)to diagnose degenerative cervical myelopathy with an accuracy of 93.5%.One-gene models(TCAP and SDHA)identified mild and severe degenerative cervical myelopathy with accuracies of 83.3%and 76.7%,respectively.Signatures of two immune cell types(memory B cells and memory-activated CD4^(+)T cells)predicted levels of lesions in degenerative cervical myelopathy with 80%accuracy.Our results suggest that peripheral blood RNA biomarkers could be used to predict lesion severity in degenerative cervical myelopathy.

Using Optical Tweezers to Study the Friction of the Red Blood Cells

In the last two decades the study of red blood cell elasticity using optical tweezers has known a rise appearing in the scientific research with regard to the various works carried out. Despite the various work done, no study has been done so far to study the influence of friction on the red blood cell indentation response using optical tweezers. In this study, we have developed a new approach to determine the coefficient of friction as well as the frictional forces of the red blood cell. This approach therefore allowed us to simultaneously carry out the indentation and traction test, which allowed us to extract the interfacial properties of the microbead red blood cell couple, among other things, the friction coefficient. This property would be extremely important to investigate the survival and mechanical features of cells, which will be of great physiological and pathological significance. But taking into account the hypothesis of friction as defined by the isotropic Coulomb law. The experiment performed for this purpose is the Brinell Hardness Test (DB).

Clinical significance of peripheral blood immune cells in patients with gastric cancer after surgery

BACKGROUND Gastric cancer is one of the most common malignant tumors worldwide,and surgical resection is one of the main ways to treat gastric cancer.However,the immune status of postoperative patients is crucial for prognosis and survival,and immune cells play an important role in this process.Therefore,it is helpful to understand the immune status of postoperative patients by evaluating the levels of peripheral blood immune cells,especially total T cells(CD3+),helper T cells(CD3+CD4+),and suppressor T cells(CD3+CD8+),and its relationship to sur-vival.AIM To analyzed the immune cells in peripheral blood of patients with gastric cancer after surgery,detect the levels of total T cells,helper T cells and suppressor T cells.METHODS A total of 58 patients with gastric cancer who received surgical treatment were included in the retrospective study.Flow cytometry was used to detect the level of peripheral blood immune cells and analyze the correlation between total T cells,helper T cells and inhibitory T cells.To explore the relationship between these immune markers and patient survival.RESULTS The results showed that the levels of total T cells,helper T cells,and suppressor T cells changed in patients after gastric cancer surgery.There was a significant positive correlation between total T cells,helper T cells and suppressor T cells(r=0.35,P<0.01;r=0.56,P<0.01).However,there was a negative correlation between helper T cells and suppressor T cells(r=-0.63,P<0.01).Follow-up showed that the survival rate of patients in the high-level total T cell group was significantly higher than that in the low-level group(28.87±24.98 months vs 18.42±16.21 months).The survival curve shows that the curve of patients in the high-level group is shifted to the upper right,and that of the low-level group is shifted downward.There was no significant difference between the levels of helper T cells and suppressor T cells and patient survival time.CONCLUSION By detecting peripheral blood immune cells with flow cytometry,we can initially evaluate the immune status of patients after gastric cancer surgery and initially explore its relationship with patient survival.

Predictive value of red blood cell distribution width and hematocrit for short-term outcomes and prognosis in colorectal cancer patients undergoing radical surgery

BACKGROUND Previous studies have reported that low hematocrit levels indicate poor survival in patients with ovarian cancer and cervical cancer,the prognostic value of hematocrit for colorectal cancer(CRC)patients has not been determined.The prognostic value of red blood cell distribution width(RDW)for CRC patients was controversial.AIM To investigate the impact of RDW and hematocrit on the short-term outcomes and long-term prognosis of CRC patients who underwent radical surgery.METHODS Patients who were diagnosed with CRC and underwent radical CRC resection between January 2011 and January 2020 at a single clinical center were included.The short-term outcomes,overall survival(OS)and disease-free survival(DFS)were compared among the different groups.Cox analysis was also conducted to identify independent risk factors for OS and DFS.RESULTS There were 4258 CRC patients who underwent radical surgery included in our study.A total of 1573 patients were in the lower RDW group and 2685 patients were in the higher RDW group.There were 2166 and 2092 patients in the higher hematocrit group and lower hematocrit group,respectively.Patients in the higher RDW group had more intraoperative blood loss(P<0.01)and more overall complications(P<0.01)than did those in the lower RDW group.Similarly,patients in the lower hematocrit group had more intraoperative blood loss(P=0.012),longer hospital stay(P=0.016)and overall complications(P<0.01)than did those in the higher hematocrit group.The higher RDW group had a worse OS and DFS than did the lower RDW group for tumor node metastasis(TNM)stage I(OS,P<0.05;DFS,P=0.001)and stage II(OS,P=0.004;DFS,P=0.01)than the lower RDW group;the lower hematocrit group had worse OS and DFS for TNM stage II(OS,P<0.05;DFS,P=0.001)and stage III(OS,P=0.001;DFS,P=0.001)than did the higher hematocrit group.Preoperative hematocrit was an independent risk factor for OS[P=0.017,hazard ratio(HR)=1.256,95%confidence interval(CI):1.041-1.515]and DFS(P=0.035,HR=1.194,95%CI:1.013-1.408).CONCLUSION A higher preoperative RDW and lower hematocrit were associated with more postoperative complications.However,only hematocrit was an independent risk factor for OS and DFS in CRC patients who underwent radical surgery,while RDW was not.

Application of neutrophil-lymphocyte ratio and red blood cell distribution width in diabetes mellitus complicated with heart failure

BACKGROUND Accumulating clinical evidence has shown that diabetes mellitus(DM)is a serious risk factor for cardiovascular disorders and an important factor for adverse cardiovascular events.AIM To explore the value of the combined determination of the neutrophil-lymphocyte ratio(NLR)and red blood cell distribution width(RDW)in the early diagnosis and prognosis evaluation of DM complicated with heart failure(HF).METHODS We retrospectively analyzed clinical data on 65 patients with type 2 DM(T2DM)complicated with HF(research group,Res)and 60 concurrent patients with uncomplicated T2DM(control group,Con)diagnosed at Zhejiang Provincial People’s Hospital between January 2019 and December 2021.The NLR and RDW values were determined and comparatively analyzed,and their levels in T2DM+HF patients with different cardiac function grades were recorded.The receiver operating characteristic(ROC)curves were plotted to determine the NLR and RDW values(alone and in combination)for the early diagnosis of HF.The correlation between NLR and RDW with the presence or absence of cardiac events was also investigated.RESULTS Higher NLR and RDW levels were identified in the Res vs the Con groups(P<0.05).The NLR and RDW increased gradually and synchronously with the deterioration of cardiac function in the Res group,with marked differences in their levels among patients with grade II,III,and IV HF(P<0.05).ROC curve analysis revealed that NLR combined with RDW detection had an area under the curve of 0.915,a sensitivity of 76.9%,and a specificity of 100%for the early diagnosis of HF.Furthermore,HF patients with cardiac events showed higher NLR and RDW values compared with HF patients without cardiac events.CONCLUSION NLR and RDW were useful laboratory indicators for the early diagnosis of DM complicated with HF,and their joint detection was beneficial for improving diagnostic efficiency.Additionally,NLR and RDW values were directly proportional to patient outcomes.

Development of Non-ABO Red Blood Cell Alloantibodies in Patient Undergoing Allogeneic Haematopoietic Stem Cell Transplant

Alloantibodies that are non ABO Alloimmunization to protein antigens happens only after exposure, in contrast to ABO isohaemagglutinins, which are present naturally, even in the absence of prior exposure. It is recognized that while non-ABO RBC antibodies are less common than ABO antibodies, they generate essentially the same issues that lead to unfavorable clinical results. If non-ABO alloantibodies are identified early on, these issues related complications may be avoided This call for an in-depth understanding of the recipient and donor’s ABO-Rh grouping, antibody screening, and the phenotype of certain antigens. Equally important, the temporal association time between transplantation and hemolysis can help identify the underlying mechanism of hemolysis and direct appropriate management. Therefore, for that, it is crucial to identify the etiology of post-HSCT anemia for prevention and therapy, in addition to a thorough grasp of the mechanism of anemia in non-ABO-incompatible HSCT and the temporal link between HSCT and anemia. Finding the cause of post-HSCT anemia is essential for prevention and therapy, in addition to a thorough grasp of the mechanism of anemia in non-ABO-incompatible HSCT and the temporal link between HSCT and anemia. Therefore, for that, it is crucial to identify the etiology of post-HSCT anemia. In this case report review, we would like to highlight the vital role of transfusion medicine services and stem cell clinical teams in paying particular attention to the clinical significance of non-ABO alloantibodies involved to avoid causing overt hemolysis of incompatible donor RBCs or delayed erythropoiesis. Considering the fact that some of the Haematopoietic stem cell transplant centers do not give an attention to the other non-ABO RBC antigens.

Cytokine changes and embryo attachment in mouse endometrial cells following treated with peripheral blood mononuclear cells(PBMCs)expressing ectopic hCG,and hCG-activated PBMCs

Objective:To compare the effect of human chorionic gonadotropin(hCG)-producing peripheral blood mononuclear cells(PBMCs)and PBMCs activated by hCG in vitro and expressions of related immune genes in mouse implantation.Methods:hCG-producing PBMCs(transfected PBMC)and PBMCs activated by hCG in vitro were introduced into isolated mouse endometrial cells,while cell cultures were divided into four groups:the control,PBMC,transfected,and activated PBMC groups.The expression of studied genes(IL-1β,IL-6,Lif,and Vegf)was evaluated and blastocyst attachment on the cocultured cells(isolated endometrial cells and PBMC cells)was monitored in all four groups.Results:Data showed that expression decreased in the PBMC group compared to the treated PBMC(transfected and activated PBMCs)and increased in transfected PBMC compared to the activated PBMC.Attachment and migration of blastocysts were dramatically enhanced in the transfected PBMC group compared to the activated PBMC group(P<0.05).Conclusions:Use of hCG-producing PBMCs(transfected PBMC)has more influence on endometrial receptivity.

An Investigation on the Phenotype of Cultured Dendritic Cells from the Peripheral Blood of Patients with Breast Cancer

Objective To induce and culture the dendritic cells in the peripheral blood of breast cancer patients and research on their phenotype. Methods Mononuclear cells were isolated by Ficoll Hypaque centrifugation from 32 breast cancer patients peripheral blood. These cells were plated in six well culture plates(10 6/ml,2 ml/well) in RPMI 1640 medium supplemented with 10% heat inactivated fetal bovine serum,100 ng/ml GM CSF,20 ng/ml IL 4,and/or 20 ng/ml TNF α.Two hours later, nonadherent cells were gently removed and fresh medium was added. Cultured cells were analyzed by flow cytometry with fluorescence labeled monoclonal antibodies. Pictures of cultured and fluorescence stained cells were taken by confocal scanning microscope. Results The diameter of the cells was between 10 and 20 micron. Cells displayed a characteristic CD1a +,CD40 +,CD80 +, CD86 + and CD83 + phenotypes. All of these molecules were not specific for dendritic cells. CD1a and CD83 molecules could also be expressed on the surface of CD3 + T lymphocytes and CD19 + B lymphocytes, especially on activated lymphocytes. Conclusion The molecules of CD1a and CD83 are not specific phenotypes for dendritic cells. Currently, we still need to apply both cell morphology and costimulatory molecules such as CD40,CD80, and CD86 to the identification of dendritic cells.

Isolation of Fetal Nucleated Red Blood Cells from Maternal Blood

To find a simple, effective method of isolating fetal cells from maternal peripheral blood for prenatal diagnosis, 45 women were studied with their gestation being 6-14 weeks and age 21- 30 years. The fetal cells were isolated from maternal blood by using discontinuous density gradient centrifugation. Some of the isolated cells were made smear and counted under the microscope; others were used for predicting fetal sex by PCR amplification of Y chromosome specific DYZ1 gene. The major cells in the upper separation interface were lymphocytes and monocytes, with occasionally seen nucleated red blood cells (NRBC); while those in the middle separation interface were neutrocytes, with NRBC scattering. The ratio of NRBC/nucleated cells was 1. 98±0. 28× 10-5. There was no significant difference between the first and second trimester (P>0. 05). The amount of isolated fetal cells was sufficient for prenatal genetic diagnosis. Male pregnancy was correctly predicted in 10 out of 13 cases. It is concluded that the method of discontinuous density gradient centrifugation was of considerable importance in the development of non-invasive prenatal genetic diagnosis.

Enrichment of Fetal Nucleated Red Blood Cells by Multi-core Magnetic Composite Particles for Non-invasive Prenatal Diagnosis

A novel kind of multi-core magnetic composite particles, the surfaces of which were respectively mo- dified with goat-anti-mouse IgG and antitransferrin receptor（anti-CD71）, was prepared. The fetal nucleated red blood cells（FNRBCs） in the peripheral blood of a gravida were rapidly and effectively enriched and separated by the mo- dified multi-core magnetic composite particles in an external magnetic field. The obtained FNRBCs were used for the identification of the fetal sex by means of fluorescence in situ hybridization（FISH） technique. The results demonstrate that the multi-core magnetic composite particles meet the requirements for the enrichment and speration of FNRBCs with a low concentration and the accuracy of detetion for the diagnosis of fetal sex reached to 95%. Moreover, the obtained FNRBCs were applied to the non-invasive diagnosis of Down syndrome and chromosome 3p21 was de- tected. The above facts indicate that the novel multi-core magnetic composite particles-based method is simple, relia- ble and cost-effective and has opened up vast vistas for the potential application in clinic non-invasive prenatal diag- nosis.

Epstein-Barr virus DNA loads in the peripheral blood cells predict the survival of locoregionally-advanced nasopharyngeal carcinoma patients

Objective:Circulating cell-free Epstein-Barr virus(EBV)DNA has been shown to be a valuable biomarker for population screening and prognostic surveillance for nasopharyngeal carcinoma(NPC).Despite important insights into the biology of persistence,few studies have addressed the clinical significance of cell-based EBV-DNA loads in peripheral blood cells(PBCs).Methods:A prospective observational cohort study was conducted involving 1,063 newly diagnosed,locoregionally-advanced NPC patients at Sun Yat-sen University Cancer Center from 2005 to 2007.Cox regression analysis was conducted to identify the association of PBC EBV DNA loads to overall survival(OS)and other prognostic outcomes.Prognostic nomograms were developed based on PBC EBV DNA loads to predict survival outcomes for NPC patients.Results:After a median follow-up of 108 months,patients with higher PBC EBV-DNA loads had significantly worse OS[hazard ratio(HR)of medium,medium-high,and high vs.low were 1.50,1.52,and 1.85 respectively;Ptrend<0.001].Similar results were found for progression-free survival and distant metastasis-free survival.The concordance index of the prognostic nomogram for predicting OS in the training set and validation set were 0.70 and 0.66,respectively.Our data showed that the PBC EBV DNA load was an independent and robust survival biomarker,which remained significant even after adjusting for plasma EBV DNA loads in a subset of 205 patients of the cohort(HR:1.88;P=0.025).Importantly,a combination of PBC EBV DNA load and plasma EBV DNA load improved the predicted OS.Conclusions:The EBV-DNA load in PBCs may be an independent prognosis marker for NPC patients.

Culture of Porcine Peripheral Blood Monocyte-derived Dendritic Cells in vitro

[Objective] This study aimed to establish an in vitro culture model for porcine peripheral blood monocyte-derived dendritic cells （MoDCs）. [Method] Fresh peripheral blood mononuclear cells （PBMCs） were separated from pig, and precursor dendritic cells were obtained by adherence method. The dendritic cells were treated by recombinant porcine granulocyte-monocyte colony stimulating factor （rpGM-CSF） and recombinant porcine interleukin-4 （rplL-4） together, and lipopolysaccharide （LPS） respectively. The cells in different time periods were collected. The morphology of the collected cells was observed by scanning electron microscopy; the expression of surface molecules and phagocytic ability to FITC-dextran were detected by flow cy- tometry; and the stimulating ability for allogeneic T cells was detected by mixed lymphocyte reaction. [Result] The DCs suffering maturation induction in vitro showed typical dendritic morphology; compared with those of DCs untreated by LPS, the cell surface expression of CDla, CD80, CD86, SLAII and CD172a of DCs treated by LPS was significantly increased, the phagocytic ability was reduced slightly, and the stimulating ability for allogeneic T cells was enhanced to some extent. [Conclusion] An in vitro culture method was successfully established for porcine MoDCs in this study, laying a foundation for further study on the role of porcine MoDCs in immunoregulation and anti-virus infection.

Value of red blood cell distribution width in prediction of diastolic dysfunction in cirrhotic cardiomyopathy

BACKGROUND Clinical diagnosis of cirrhotic cardiomyopathy(CCM) often encounters challenges of lack of timeliness and disease severity, with the commonly positive indicator usually associated with advanced heart failure.AIM To explore suitable biomarkers for early CCM prediction.METHODS A total of 505 eligible patients were enrolled in this study and divided into four groups according to Child-Pugh classification: Group Ⅰ, Class A without CCM(105 cases);Group Ⅱ, Class A with CCM(175 cases);Group Ⅲ, Class B with CCM(139 cases);and Group Ⅳ, Class C with CCM(86 cases). Logistic regression and receiver operating characteristic(ROC) curve analyses were performed to determine whether red blood cell distribution width(RDW) was an independent risk factor for CCM risk. The relationships between RDW and Child-Pugh scores, Model for End-Stage Liver Disease(MELD) scores, and N-terminal pro-brain natriuretic peptide(NT-proBNP) were analyzed by Pearson correlation analysis.RESULTS A constant RDW increase was evident from Group Ⅰ to Group Ⅳ(12.54 ± 0.85, 13.29 ± 1.19, 14.30 ± 1.96, and 16.25 ± 2.13, respectively). Pearson correlation analysis showed that RDW was positively correlated with Child-Pugh scores(r = 0.642, P < 0.001), MELD scores(r = 0.592, P < 0.001), and NT-proBNP(r = 0.715, P < 0.001). Furthermore, between Group Ⅰ and Group Ⅱ, RDW was the only significant index(odds ratio: 2.175, 95% confidence interval [CI]: 1.549-3.054, P < 0.001), and it reached statistical significance when examined by ROC curve analysis(area under the curve: 0.686, 95%CI: 0.624-0.748, P < 0.001).CONCLUSION RDW can serve as an effective and accessible clinical indicator for the prediction of diastolic dysfunction in CCM, in which a numerical value of more than 13.05% may indicate an increasing CCM risk.

Comparison of the conventional tube and erythrocyte-magnetized technology in titration of red blood cell alloantibodies

BACKGROUND Erythrocyte alloantibodies are mainly produced after immune stimulation,such as blood transfusion,pregnancy,and transplantation,and are the leading causes of severe hemolytic transfusion reactions and difficulty in blood grouping and matching.Therefore,antibody screening is critical to prevent and improve red cell alloantibodies.Routine tube assay is the primary detection method of antibody screening.Recently,erythrocyte-magnetized technology(EMT)has been increasingly used in clinical practice.This study intends to probe the application and efficacy of the conventional tube and EMT in red blood cell alloantibody titration to provide a reference for clinical blood transfusion.AIM To investigate the application value of conventional tube and EMT in red blood cell alloantibody titration and enhance the safety of blood transfusion practice.METHODS A total of 1298 blood samples were harvested from blood donors at the Department of Blood Transfusion of our hospital from March 2021 to December 2022.A 5 mL blood sample was collected in tubing,which was then cut,and the whole blood was put into a test tube for centrifugation to separate the serum.Different red blood cell blood group antibody titers were simultaneously detected using the tube polybrene test,tube antiglobulin test(AGT),and EMT screening irregular antibody methods to determine the best test method.RESULTS Simultaneous detection was performed through the tube polybrene test,tube AGT and EMT screening irregular antibodies.It was discovered that the EMT screening irregular antibody method could detect all immunoglobulin G(IgG)and immunoglobulin M(IgM)irregular antibodies,and the results of manual tube AGT were satisfactory,but the operation time was lengthy,and the equipment had a large footprint.The EMT screening irregular antibody assay was also conducted to determine its activity against type O Rh(D)red blood cells,and the outcomes were satisfactory.Furthermore,compared to the conventional tube method,the EMT screening irregular antibody method was more cost-effective and had significantly higher detection efficiency.CONCLUSION With a higher detection rate,the EMT screening irregular antibody method can detect both IgG and IgM irregular antibodies faster and more effectively than the conventional tube method.

Effect of IL-18 on peripheral blood mononuclear cells of chronic hepatitis B and hepatitis B virus DNA released by HepG2.2.15 cell lines

BACKGROUND: Interleukin-18 (IL-18), a pro-inflamma- tory cytokine that induces interferon-γ (IFN-γ) production in T cells and natural killer cells, plays a critical role in the T-lymphocyte helper type 1 ( Th1) response. This study was designed to explore the effect of IL-18 on peripheral blood mononuclear cells ( PBMCs) derived from chronic hepatitis B (CHB) and on hepatitis B virus (HBV) DNA released by HepG2.2.15 cell lines, which were transfected with hepatitis B virus gene in vitro. METHODS: PBMCs isolated from 25 healthy people and 25 patients with CHB were stimulated with HBcAg and IL-18 of various concentrations for 72 hours. The levels of IFN-γ in the supernatants of cultured PBMCs were determined by ELISA. After the stimulation of IL-18 of various concentra- tions, PBMCs derived from one patient were co-cultured for 96 hours with HepG2. 2. 15 cells which had been cul- tured for 24 hours, and then the supernatants were collected by centrifugation and used for HBV DNA quantitative as- say. RESULTS: When PBMCs were stimulated by HBcAg and IL-18 at various concentrations, the levels of IFN-γ in the supernatants of CHB groups were much higher than those in normal control groups, at 0.2 ng/ml: t =11.70, P< 0.01; at 1.0 ng/ml: t =16.19, P<0.01; and at5.0 ng/ml: t =20.12, P <0.01. In the CHB groups, the levels of IFN-γ in the supernatants of PBMCs stimulated by HBcAg alone were lower than both those stimulated by HBcAg and EL-18 at various concentrations and those stimulated by HBcAg and EL-18 (5.0 ng/ml) together with EL-12 (mild: t = 2.20, P<0.05; moderate; t=2.97, P<0.05; severe; t = 0.66, P >0.05). The content of HBV DNA in the superna- tant of co-cultivation of HepG2. 2. 15 cells and PBMCs without stimulated materials was higher than that stimula-ted by HBcAg and EL-18 at various concentrations of HBc- Ag and IL-18 together with IL-12/IFN-α1lb. CONCLUSION: DL-18 can induce IFN-γ secretion and pro- bably play a key role in the modulation of both innate and adaptive immunity. It has implications in improving im- munoregulatory effect and increasing the ability of immune cells to kill cells infected by virus.

Transplantation of mesenchymal stem cells from human umbilical cord versus human umbilical cord blood for peripheral nerve regeneration

BACKGROUND： Mesenchymal stem cells （MSCs） appear to be a good alternative to Schwann cells in the treatment of peripheral nerve injury. Fetal stem cells, like umbilical cord blood （UCB） and umbilical cord （UC） stem cells, have several advantages over adult stem cells. OBJECTIVE： To assess the effects of UC-derived MSCs （UCMSCs） and UCB-derived MSCs （UCBMSCs） in repair of sciatic nerve defects. DESIGN, TIME AND SETTING： A randomized controlled animal experiment was performed at the laboratory of Department of Oral and Maxillofacial Surgery, Seoul National University Dental Hospital, from July to December 2009. MATERIALS： UCMSCs were provided by the Research Institute of Biotechnology, Dongguk University. UCBMSCs were provided by the Laboratory of Stem Cells and Tumor Biology, College of Veterinary Medicine, Seoul National University. Dulbecco＇s modified Eagle＇s medium （DMEM） was purchased from Gibco-BRL, USA. METHODS： Seven-week-old Sprague-Dawley rats were randomly and evenly divided into three groups： DMEM, UCBMSCs, and UCMSCs. A 10-mm defect in the left sciatic nerve was constructed in all rats. DMEM （15 μL） containing 1×10^6 UCBMSCs or UCMSCs was injected into the gap between nerve stumps, with the surrounding epineurium as a natural conduit. For the DMEM group, simple DMEM was injected. MAIN OUTCOME MEASURES： At 7 weeks after sciatic nerve dissection, dorsal root ganglia neurons were labeled by fluorogold retrograde labeling. At 8 weeks, electrophysiology and histomorphometry were performed. At 2, 4, 6, and 8 weeks after surgery, sciatic nerve function was evaluated using gait analysis. RESULTS： The UCBMSCs group and the UCMSCs group exhibited similar sciatic nerve function and electrophysiological indices, which were better than the DMEM group, as measured by gait analysis （P 〈 0.05）. Fluorogold retrograde labeling of sciatic nerve revealed that the UCBMSCs group demonstrated a higher number of labeled neurons; however, the differences were not significant. Histomorphometric indices were similar in the UCBMSCs and UCMSCs groups, and total axon counts, particularly axon density （P 〈 0.05）, were significantly greater in the UCBMSCs and UCMSCs groups than in the DMEM group. CONCLUSION： Transplanting either UCBMSCs or UCMSCs into axotomized sciatic nerves could accelerate and promote sciatic nerve regeneration over 8 weeks. Both treatments had similar effects on nerve regeneration.

Analysis of CD4^+CD25^+ Regulatory T Cells and Foxp3 mRNA in the Peripheral Blood of Patients with Asthma

The changes of CD4^＋CD25^＋ regulatory T cells （CD4^＋CD25^＋ Treg） and Foxp3 mRNA in peripheral blood mononuclear cells （PBMCs） from patients with asthma were investigated in order to elucidate the possible roles of CD4^＋CD25^＋ Treg in the development of asthma. The peripheral blood samples were collected from 29 healthy controls （normal control group） and 78 patients with asthma which included 30 patients in exacerbation group, 25 patients in persistent group, and 23 patients in remission group. By using flow cytometry and RT-PCR, the CD4^＋CD25^＋ Treg ratio and Foxp3 mRNA in PBMCs were detected. The CD4^＋CD25^＋ Treg ratio and Foxp3 mRNA in PBMCs of exacerbation and persistent groups were lower than that of remission and normal control groups （P〈0.05）. Although the CD4^＋CD25^＋ Treg ratio and Foxp3 mRNA of remission group were also lower than that of normal control group, there was no significant difference between them （P〉0.05）. As compared with persistent group, exacerbation group had lower CD4^＋CD25^＋ Treg ratio and Foxp3 mRNA （P〈0.05）. It was indicated that the decrease of CD4^＋CD25^＋ Treg ratio and its function in PBMCs may be responsible for pathogenesis of asthma.

Expression of Toll-like Receptor 9 in Peripheral Blood Mononuclear Cells from Patients with Different Hepatitis B and C Viral Loads

The aim of the present study was to investigate the expression of toll-like receptors （TLR） 9 in peripheral blood mononuclear cells （PBMC） of patients with chronic hepatitis B and C with different virus copies. The study group included 90 patients （60 with chronic hepatitis B, and 30 with chronic hepatitis C）, and 20 healthy people served as control group. The protein and mRNA levels of TLR9 were detected by using flow cytometry and real-time PCR. The serum viral copies of HBV and HCV were measured in all patients, and the correlation between HBV-DNA copies or HCV-RNA copies and the TLR9 expression was analyzed. Our results demonstrated that HBV or HCV infection led to a decreased expression of TLR9 mRNA and protein compared to the control group （P〈0.05）. The TLR9 protein and mRNA levels were negatively correlated with serum viral copies of HBV and HCV （r=-0.632, r=-0.909, P〈0.01）. It was concluded that TLR9 mRNA and protein are down-regulated in PBMC of HBV-infected or HCV-infected patients, and they are negatively correlated with serum viral copies and play an important role in detecting viral replication of HBV and HCV.

Dynamic changes of HBV DNA in serum and peripheral blood mononuclear cells of chronic hepatitis patients after lamivudine treatment

AIM： To study the dynamic changes of hepatits B virus （HBV） DNA in serum and peripheral blood mononuclear cells （PBMCs） of patients after lamivudine therapy. METHODS： A total of 72 patients with chronic HBV infection were included in this study. All patients were confirmed to have the following conditions： above 16 years of age, elevated serum alanine amonotransferase （ALT）, positive hepatitis B e antigen （HBeAg）, positive HBV DNA in serum and PBMCs, negative antibodies against HAV, HCV, HDV, HEV. Other possible causes of chronic liver damages, such as drugs, alcohol and autoimmune diseases were excluded. Seventy-two cases were randomly divided into lamivudine treatment group （n = 42） and control group （n = 30）. HBV DNA was detected both in serum and in PBMCs by fluorescence quantitative polymerase chain reaction （PCR）, during and after lamivudine treatment. RESULTS： In the treatment group, HBV DNA became negative both in serum and in PBMC, of, 38 and 25 out of 42 cases respectively during the 48 wk oflamivudine treatment, the negative rate was 90.5% and 59.5% respectively. In the control group, the negative rate was 23.3% and 16.7% respectively. It was statistically significant at 12, 24 and 48 wk as compared with the control group （P 〈 0.005）. The average conversion period of HBV DNA was 6 wk （2-8 wk） in serum and 16 wk （8-24 wk） in PBMC.CONCLUSION： Lamivudine has remarkable effects on HBV replication both in serum and The inhibitory effect on HBV DNA in PBMCs than that in serum inhibitory in PBMCs. is weaker