ABSTRACT
<p><b>OBJECTIVE</b>To investigate the number of peripheral blood CD5(+) B cells and their ability of secreting IL-10 in patients with immune thrombocytopenia (ITP).</p><p><b>METHODS</b>Peripheral blood lymphocytes were isolated from 57 pre-treated, 40 post-treated ITP patients and 25 controls using Ficoll-Hypaque density centrifugation and then stained with PE-CD5/FITC-CD19 for flow cytometric analysis. After 24-hour culture, lymphocytes were stained with APC-IL-10 for intracellular cytokine detection. ELISA assay was employed to determine IL-10 concentration in supernatants.</p><p><b>RESULTS</b>The percentage and absolute number of CD5(+) B cells in peripheral blood from pre-treated ITP patients were significantly higher than that from normal controls (3.75 ± 2.37)% vs (2.10 ± 1.08)%, P < 0.01; (6.29 ± 5.77)× 10(7)/L vs (3.06 ± 1.90)× 10(7)/L, P < 0.01. CD5(+) B cells expressed more intracellular IL-10 than other lymphocyte subsets both in ITP patients and normal controls. The percentages of IL-10(+) cells within CD5(+) B cells in pre-treated ITP patients and normal controls were (29.51 ± 20.73)% and(15.90 ± 9.58)%, respectively(P < 0.01). Intracellular mean fluorescence intensity (MFI) of IL-10 in CD5(+) B cells was 27.95 ± 13.99 in pre-treated patients, which was significantly higher than that in controls (P < 0.01). In contrast, IL-10 concentration in supernatants was (173.05 ± 102.50) ng/L in pre-treated ITP group, which was lower than that (230.61 ± 76.96) ng/L in controls. In patients who achieved remission, the number of CD5(+) B cells decreased to level comparable to normal controls. While intracellular IL-10 MFI of CD5(+) B cells in post-treated ITP patients remained as high as in pre-treated ones, the IL-10 concentration in supernatants increased to level similar to controls.</p><p><b>CONCLUSION</b>The significantly increased number of CD5(+) B cells and accumulated IL-10 in CD5(+) B cells suggested impaired IL-10 secretion in ITP patients. The number and the ability of secreting IL-10 of CD5(+) B cells could be restored after effective treatments in patients with ITP.</p>
Subject(s)
Adult , Aged , Female , Humans , Male , Middle Aged , Young Adult , B-Lymphocytes , Allergy and Immunology , Metabolism , CD5 Antigens , Metabolism , Case-Control Studies , Interleukin-10 , Blood , Purpura, Thrombocytopenic, Idiopathic , Blood , Allergy and ImmunologyABSTRACT
This study examined the effects of a recombinant adenovirus Ad-PTEN-EGFP on the proliferation of A549 cells,a human lung carcinoma cell line,in vitro and on the growth of the implanted tumors in the nude mice in vivo,explored the underlying mechanisms and evaluated the in vitro transfection efficiency of Ad-PTEN-EGFP into A549 cells.The expression of Ad-PTEN-EGFP in the A549 cells was determined.The proliferation and the apoptosis rates of the A549 cells with Ad-PTEN-EGFP transfection or not was detected by MTT and flow cytometry.Ad-PTEN-EGFP at different doses was injected intratumorally to the tumor-bearing mice induced by the A549 cells.Tumor sizes were measured on an altemate day.After all the mice were sacrificed,the implanted tumors were removed for routine histological examination,weight test,HE staining and immunohistochemical staining.The expressions of Bax,P16 and P53 in the tumor tissues and those of caspase-3,CD34and VEGF in the mouse sera were detected.Tumor cell apoptosis was measured by TUNEL method.The results showed that the vitality of the A549 cells after transfection with Ad-PTEN-EGFP declined.The expression of green fluorescent protein was observed under fluorescent microscope.The transfection rate was in excess of 50%.The mRNA and protein expression of PTEN in the transfected cells was confirmed.The proliferation rate of the transfected cells was significantly decreased when compared with that of the non-transfected cells(P<0.05).The number of the apoptosis cells was increased in the transfected cells(P<0.05).The models of implanted tumors were successfully established by injection of the A549 cells in the flank of Balb/c nude mice.Administration of Ad-PTEN-EGFP to the tumor-bearing nude mice resulted in a suppression of tumor growth.There were statistically significant differences in the tumor weight and tumor volume between the Ad-PTEN-EGFP-treated group and the control groups(P<0.05).In contrast to those in the control groups,tumor tissues in the Ad-PTEN-EGFP-treated group were shown to have typical extensive vacuolar degeneration and massive hemorrhagic necrosis.Apoptotic bodies were also observed in the tumor cells.The expressions of Bax,caspase-3 and P16 were increased(P<0.05)while those of CD34,VEGF and P53 decreased(P<0.05)in the Ad-PTEN-EGFP-treated group.It is concluded that Ad-PTEN-EGFP could induce the apoptosis of the A549 cells and inhibit their proliferation.And it could also substantially suppress the tumor growth in the tumor-bearing nude mice and induce apoptosis of the tumor cells as well.These findings carry significant implications for adenovirus vector-based PTEN gene therapies for lung cancers.
ABSTRACT
.01). Spearman rank correlation analysis demonstrated a positive correlation between the expression of COX-2 and p53 (r=0.113, P=0.421). It was concluded that COX-2 can increase the expression of p53 protein, therefore suppressing apop-tosis.