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.

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.

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.

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.

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.

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

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.

Effect of Sinomenine on IL-8, IL-6, IL-2 Produced by Peripheral Blood Mononuclear Cells

The effect of Sinomenine on IL-8, IL-6, IL-2 and mIL-2R produced by peripheral blood mononuclear cells was investigated by using cell culture, radioimmunoassay and flow cytometry. It was showed that production of IL-8 and mIL-2R was inhibited, but the levels of IL-6 were enhanced by Sinomenine. Our results also demonstrated that Sinomenine did not have any effect on the production of IL-2. The study demonstrated that Sinomenine was able to regulate the production of cytokines. This may be one of the mechanisms by which Sinomenine works on rheumatoid arthritis.

Transplantation of autologous peripheral blood mononuclear cells in the subarachnoid space for amyotrophic lateral sclerosis:a safety analysis of 14 patients

There is a small amount of clinical data regarding the safety and feasibility of autologous peripheral blood mononuclear cell transplantation into the subarachnoid space for the treatment of amyotrophic lateral sclerosis.The objectives of this retrospective study were to assess the safety and efficacy of peripheral blood mononuclear cell transplantation in 14 amyotrophic lateral sclerosis patients to provide more objective data for future clinical trials.After stem cell mobilization and collection,autologous peripheral blood mononuclear cells（1 × 109） were isolated and directly transplanted into the subarachnoid space of amyotrophic lateral sclerosis patients.The primary outcome measure was incidence of adverse events.Secondary outcome measures were electromyography 1 week before operation and 4 weeks after operation,Functional Independence Measurement,Berg Balance Scale,and Dysarthria Assessment Scale 1 week preoperatively and 1,2,4 and 12 weeks postoperatively.There was no immediate or delayed transplant-related cytotoxicity.The number of leukocytes,serum alanine aminotransferase and creatinine levels,and body temperature were within the normal ranges.Radiographic evaluation showed no serious transplant-related adverse events.Muscle strength grade,results of Functional Independence Measurement,Berg Balance Scale,and Dysarthria Assessment Scale were not significantly different before and after treatment.These findings suggest that peripheral blood mononuclear cell transplantation into the subarachnoid space for the treatment of amyotrophic lateral sclerosis is safe,but its therapeutic effect is not remarkable.Thus,a large-sample investigation is needed to assess its efficacy further.

Lactobacilli,bifi dobacteria and E.coli nissle induce pro-and anti-inflammatory cytokines in peripheral blood mononuclear cells

AIM： To investigate whether the stimulation of peripheral blood mononuclear cells （PBMNC） with the cell debris and cell extraction of different probiotic strains is similar or species specific. METHODS： Three strains of bifidobacteria, 4 strains of lactobacilli, and E. coli nissle were sonicated and centrifuged in order to divide them into cell extract and cell debris. PBMNC were separated by density gradient and incubated for 36 h with either the cell debris or the cell extract of single strains of probiotic bacteria in doses from 10^2 to 10^8 CFU/mL. Cell supernatants were taken and interleukin （IL）-10, IL-1β, and tumor necosis factor （TNF）-α were determined by ELISA. RESULTS： Depending on the species super-family, the strains had different stimulation patterns. Except for both L. casei strains, the cell extract of bitTdobacteria and/actobacilli had less stimulating capacity than cell debris, whereas the cell extract of E. coli nissle had similar stimulating properties to that of the cell debris of the strain and significantly more stimulating capacity than that of bifidobacteria and lactobacilli. The cell debris of bifidobacteria stimulated more cytokine release than the cell debris of lactobacilli. The cell debris of lactobacilli did not have a stimulating capacity when lower concentrations were used. Neither cell extraction nor cell debris had an inhibitory effect on the production of the tested cytokines by stimulated PBMNC. CONCLUSION： The incubation of probiotic strains, which have been used in clinical trials for inflammatory diseases, with immunocompetent cells leads to different species specific reactions. High IL-10 response to cell debris of bifidobacteria and E. coli nissle can be found. This corresponds to positive effects of bihdobacteria and E. coli nissle in clinical trials for inflammatory bowel disease compared to negative outcomes obtained with lactobacilli.

Telomerase Activity in Peripheral Blood Mononuclear Cells from Senile Patients with Pneumonia

To investigate the changes of the activity of telomerase in peripheral blood mononuclear cells （PBMCs） from senile patients with pneumonia, the telomerase activity was examined before and after the stimulation of phytohemagglutinin-M （PHA-M） in PBMCs from 10 control subjects （group A）, 12 non-senile patients with pneumonia （group B） and 9 senile patients with pneumonia （group C）. Also observed was the proliferative response of these PBMCs to PHA-M. The results showed that, both with or without the stimulation of PHA-M, the values of telomerase activity in PBMCs from group C patients （A values： pre-stimulation, 0.43±0.04; post-stimulation, 0. 63± 0. 03） were significantly lower than those in PBMCs from both group A patients （A values： prestimulation, 0. 65 ± 0. 05 ; post-stimulation, 1.26 ± 0. 13 ; P〈0. 001, respectively） and group B patients （A values： pre-stimulation, 0. 63±0. 03; post-stimulation, 0. 93± 0. 03; P〈0. 05, respectively）. The results of MTT test showed that the proliferative activity of PBMCs in group C patients （A value： 0. 35±0. 03） was also significantly lower than that in group A patients （A value： 0. 55±0. 04; P〈0. 05） and group B patients （A value= 0. 46±0.03; P〈0.05）. These results indicate that the telomerase activity decreases in senile patients with pneumonia, which may be one of the mechanisms for the weakened immune function in those patients.

Insensitivity of PI3K/Akt/GSK3 signaling in peripheral blood mononuclear cells of age-related macular degeneration patients

Our recent studies with cultured retinal pigment epithelium cells suggested that overexpression of interleukin 17 receptor C（IL-17RC）,a phenomenon observed in peripheral blood and chorioretinal tissues with age-related macular degeneration（AMD）,was associated with altered activation of phosphatidylinositide 3-kinase（PI3K）,Akt,and glycogen synthase kinase 3（GSK3）.We wondered whether or not altered PI3 K,Akt,and GSK3 activities could be detected in peripheral blood mononuclear cells（PBMC） obtained from AMD patients.In the patients＇ PBMC,absent or reduced serine-phosphorylation of GSK3α or GSK3β was observed,which was accompanied with increased phosphorylation of GSK3 substrates（e.g.CCAAT enhancer binding protein a,insulin receptor substrate 1,and TAU）,indicative of enhanced GSK3 activation.In addition,decreased protein mass of PI3K85α and tyrosinephosphorylation of PI3K50α was present in PBMC of the AMD patients,suggesting impaired PI3 K activation.Moreover,abnormally lowered molecular weight forms of Akt and GSK3 were detected in PBMC of the AMD patients.These data demonstrate that despite the presence of high levels of IL-17 RC,Wnt-3a and vascular endothelial growth factor,the PI3K/Akt/GSK3 signaling pathway is insensitive to these stimuli in PBMC of the AMD patients.Thus,measurement of PI3K/Akt/GSK3 expression and activity in PBMC may serve as a surrogate biomarker for AMD.

NOD2-and disease-specific gene expression profiles of peripheral blood mononuclear cells from Crohn's disease patients

AIM To investigate disease-specific gene expression profiles of peripheral blood mononuclear cells(PBMCs) from Crohn's disease(CD) patients in clinical remission.METHODS Patients with CD in clinical remission or with very low disease activity according to the Crohn's disease activity index were genotyped regarding nucleotidebinding oligomerization domain 2(NOD2),and PBMCs from wild-type(WT)-NOD2 patients,patients with homozygous or heterozygous NOD2 mutations and healthy donors were isolated for further analysis.The cells were cultured with vitamin D,peptidoglycan(PGN) and lipopolysaccharide(LPS) for defined periods of time before RNA was isolated and subjected to microarray analysis using Clariom S assays and quantitative realtime PCR.NOD2-and disease-specific gene expression profiles were evaluated with repeated measure ANOVA by a general linear model.RESULTS Employing microarray assays,a total of 267 genes were identified that were significantly up-or downregulated in PBMCs of WT-NOD2 patients,compared to healthy donors after challenge with vitamin D and/or a combination of LPS and PGN(P < 0.05;threshold:≥ 2-fold change).For further analysis by real-time PCR,genes with known impact on inflammation and immunity were selected that fulfilled predefined expression criteria.In a larger cohort of patients and controls,a disease-associated expression pattern,with higher transcript levels in vitamin D-treated PBMCs from patients,was observed for three of these genes,CLEC5 A(P < 0.030),lysozyme(LYZ;P < 0.047) and TREM1(P < 0.023).Six genes were found to be expressed in a NOD2-dependent manner(CD101,P < 0.002;CLEC5 A,P < 0.020;CXCL5,P < 0.009;IL-24,P < 0.044;ITGB2,P < 0.041;LYZ,P < 0.042).Interestingly,the highest transcript levels were observed in patients with heterozygous NOD2 mutations.CONCLUSION Our data identify CLEC5 A and LYZ as CD-and NOD2-associated genes of PBMCs and encourage further studies on their pathomechanistic roles.

Relationship between Single Nucleotide Polymorphism in TNF-α Gene Promoter Region and Inhibitory Effects of Triptolide on TNF-α Production in Peripheral Blood Mononuclear Cells of Healthy Humans

The relationship between tumour necrosis lactose （TNF-α） gene polymorphism and inhibitory effects of triptolide on TNF-α production from peripheral blood mononuclear cells （PBMC） of healthy humans was investigated. Genomic DNA from 41 healthy people was typed for TNF-α- 308 polymorphism by allele-specific polymorphism chain reaction （AS-PCR）. The TNF-α concentration in the supernatant was measured by ELISA. The results showed that the production of TNF-α from TNF-α -308 non-G/G genotype PBMC was higher than that from TNF-α-308 G/G genotype PBMC after stimulated by LPS. Triptolide could lower the production of TNF-α from G/ G genotype PBMC, but had no effect on the level of TNF-α from non-G/G genotype PBMC. It was concluded that TNF-α gene polymorphism was related to the TNF-α production from triptolide-inhibited PBMC culture in healthy humans.

Expression of Heme Oxygenase-1 in the Peripheral Blood Mononuclear Cells from Asthmatic Patients

Summary： To explore the expression of heme oxygenase-1 （HO-1） in the peripheral blood mononuclear cells （PBMCs） and its relationship with pulmonary ventilation function in asthmatic patients, 18 asthmatic patients and 18 healthy subjects were selected. HO-1 protein and mRNA levels in PBMCs were measured by immunohistochemical staining and reverse transcription-polymerase chain reaction （RT-PCR）, respectively. Blood carbon monoxide Hb （COHb）, serum total IgE and pulmonary ventilatory function were observed. Our results showed that the percentage of cells positive for immunohistochemical staining of HO-1 were significantly higher in asthmatic patients （41.72±7.44） % than that in with healthy subjects （10.45±4.36）% （P〈0.001） and the optical density of PBMC HO-1 mRNA was higher in asthmatic patients （26.05±4. 14） than that in healthy subjects （10.82±4.26） （P〈0.001）. The relation analysis showed that PBMC HO-1 protein and mRNA levels had significantly negative relation with FEV, %, PEFR, MEFR50 , respectively （r=-0.51-0.89, P〈0.05-0. 001, respectively） and a positive relation with COHb and serum total IgE （r=0.48-0. 85, 0.05-0. 001, respectively）. It is concluded that the expression of PBMC HO-1 protein and mRNA increased significantly in asthmatic patients, and HO-1 may play a significant role in the pathogenesis of asthma. The expression of HO-1 may bear a relation with severity of asthma.

Application of Circulating Tumor Cells in Peripheral Blood in Judging the Prognosis of Patients with Renal Cancer and Related Indexes of Blood Coagulation

Objective: To investigate the value of the number of circulating tumor cells (CTC) in peripheral blood in the prognosis and coagulation-related indicators of patients with renal cancer. Methods: 65 patients with renal cell carcinoma (RCC) confirmed pathologically were divided into CTC positive group and CTC negative group according to the CTC count (5 pcs/3.5 ml). Compare the age, gender, tumor location, TNM (clinical stage), pathological grade, tissue type, lymph node metastasis, distant metastasis, prognosis and prothrombin time (PT), fibrinogen (FIB), partial coagulation of the two groups of patients The correlation between the results of zymogen time (APTT) and D-dimer (DD) and the number of CTC. Results: There were significant differences in TNM, lymph node metastasis, and distant metastasis between the two groups (P < 0.05). The number of CTC in patients was correlated with FIB and D-D levels (P < 0.05). Conclusion: The number of CTC in patients with renal cell carcinoma is correlated with some clinical phenotypes (TNM, lymph node metastasis, distant metastasis) and some coagulation indexes (FIB, D-D), and can jointly predict the prognosis of renal cancer.

Antipsychotics preserve telomere length in peripheral blood mononuclear cells after acute oxidative stress injury

Antipsychotics may prolong or retain telomere length,affect mitochondrial function,and then affect the metabolism of nerve cells.To validate the hypothesis that antipsychotics can prolong telomere length after oxidative stress injury,leukocytes from healthy volunteers were extracted using Ficoll-Histopaque density gradient.The mononuclear cells layer was resuspended in cell culture medium.Oxidative stress was induced with hydrogen peroxide in cultured leukocytes.Four days later,leukocytes were treated with aripiprazole,haloperidol or clozapine for 7 days.Real-time PCR revealed that treatments with aripiprazole and haloperidol increased the telomere length by 23%and 20%in peripheral blood mononuclear cells after acute oxidative stress injury.These results suggest that haloperidol and aripiprazole can reduce the damage to telomeres induced by oxidative stress.The experiment procedure was approved by the Ethics Committee of Faculty of Medicine of the University of Sao Paulo(FMUSP/CAAE approval No.52622616.8.0000.0065).

HLA Class I Expressions on Peripheral Blood Mononuclear Cells in Colorectal Cancer Patients

Objective： To investigate the expression change of human leukocyte antigen （HLA） class I on human peripheral blood mononuclear ceils （PBMCs） at both mRNA and protein levels, and to evaluate its roles in the development of colorectal cancer （CRC）. Methods： In the present study, 50 patients with CRC, 35 patients with benign colorectal lesion and 42 healthy volunteers were enrolled. Expression levels of HLA class I mRNA and protein were determined using real-time quantitative reverse transcription PCR （RT-PCR） and flow cytometry analysis, respectively. Results： The expression levels of HLA class I mRNA and proteins were not influenced by age and gender. The relative ratios of HLA class I mRNA were 0.99±0.27 in healthy controls, 0.76±0.19 in benign patients, and 0.48±0.21 in CRC patients. Mean fluorescence intensities of HLA class I were 145.58±38.14 in healthy controls, 102.05±35.98 in benign patients and 87.44±34.01 in CRC patients. HLA class I on PBMCs was significantly down-regulated at both mRNA and protein levels in patients with stage III and IV CRC. CRC patients with lymph node metastasis also showed a decreased HLA class I expression at protein level. Conclusion： HLA class I expressions on PBMCs are associated with staging of CRC and lymph node metastasis. Monitoring the expression of HLA class I on PBMCs may provide useful information for diagnosis and metastasis judgement of CRC.

IN VITRO STUDY ON CRYOPRESERVATION OF PERIPHERAL BLOOD STEM CELLS WITH UNCONTROLLED FREEZER

Objective: To evaluate the effects of cryopreservation of APBSCs in ?80°C un-controlled freezer and liquid nitrogen, and to search for the efficient combined cryoprotectant and the highest cell concentration for cryopreservating peripheral blood stem cell (PBSC). Methods: To compare the effect of three combined cryoprotectants, evaluation of in vitro proliferative capacity by colony-forming unit-Granulocyte-macrophage (CFU-GM) and burst-forming Unit-erythroid (BFU-E) assay, immunophenotyping for CD34+/CD38? cells by FCM, and viability assessment by trypan blue exclusion were performed. Results: The cryoprotectant 10%DMSO+HES+auto-Plasma resulted in the highest recovery rates. CFU-GM and BFU-E were (78.7±9.8)%, (69.8±14.1)%, respectively. The recovery rates of CFU-GM and BFU-E in A/C groups were (68.3±6.2)%/ (65.8±7.2)% and (63.4±9.7)%/ (60.4±10.5)%, respectively. The recovery rates of CD34+/CD38? cells and cell viability with the three combined reagents were 90% and 80%, respectively. Conclusion: 5%DMSO+ HES+auto-PLASMA is an ideal combined cryoprotectant for cryopreserving PBSC. The cell concentrations may be cryopreserved at 4×108 ml?1.