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Objective To investigate the level of subpopulations of myeloid-derived suppressor cells (MDSCs) in elderly tumor-bearing mice versus elderly tumor-free mice,and to study the difference in immune suppressive functions between different subpopulations and their mechanisms.Methods A total of 20 healthy C57BL/6 elderly mice(aged 18-20 months) were randomly chosen to establish Lewis lung cancer models.The amount of monocytic-MDSCs (MO-MDSCs) and polymorphonuclear granulocytic-MDSCs(PMN-MDSCs) in tumor-free and tumor-bearing elderly mice was evaluated by using flow cytometry.MO-MDSCs and PMN-MDSCs were separated with Magnetic-Activated Cell Sorting (MACS) MicroBeads and their morphological characteristics were observed after May-Grunwald-Giemsa staining.The effects of MO-MDSCs and PMN-MDSCs on the proliferation of T cells were determined by Brdu-enzyme-linked immunosorbent assay(ELISA).And the immune suppressive mediators secreted by the subpopulations were detected by Real-time polymerase chain reaction(PCR).Results Compared to the tumor-free group,the proportion of MO-MDSCs in the spleen of tumorbearing group were increased [(12.44± 1.20) ％ vs.(38.42±3.66) ％,t=5.67,P＜0.001],while PMN-MDSCs were not [(10.34±0.68) ％ vs.(12.18±1.27) ％,t=2.21,P=0.09].The result of Brdu-ELISA showed that MO-MDSCs could suppress the proliferation of T cells [(0.30 ± 0.18) vs.(3.38±0.96),t=8.33,P＜0.001],while PMN MDSCs could not [(2.69±0.45)vs.(3.38±0.96),t =1.72,P=0.11].The result of PCR showed that as compared with PMN-MDSCs,Mo-MDSCs had the increased expression levels of arginase-1 (ARG-1),inducible nitric oxide synthase (iNOS),interleukin-10 (IL-10),interferon-γ(IFN-γ) (t =4.31,8.89,1.70,3.13,respectively,P ＜ 0.01 or 0.05),while the expression levels of interleukin-13 (IL-13),transforming growth factor-β(TGF-β) had no differences (t=4.94 and 2.75,P =0.39 and0.47).Conclusions MO-MDSCs are significantly increased in elderly Lewis lung cancer mice models.MO-MDSCs could mediate lung tumor evasion by suppressing the proliferation of T cells through highly expressing ARG-1,iNOS,IL-10 and IFN-γ.
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Objective To explore the immune property and possible mechanism of the CD71+ CD235a+ nucleated erythroid cells from peripheral blood in elderly,as compared with those of healthy young.Methods Peripheral blood obtained by venipuncture from healthy young(n=59,mean age=28 years) and elderly(n=78,mean age=68 years)donors were measured by flow cytometry to evaluate the frequency of the CD71+ CD235a+ cells in peripheral blood mononuclear cells (PBMC).In vitro assays,CD71+ CD235a+ cells were sorted by flow cytometry and T cells were sorted using CD4+T cell isolation kit,then the T cell proliferation assays were conducted by following groups:T cell + CFSE group;T cell +CFSE co-cultured with CD71+ CD235a+ cells from the young donors;T cell + CFSE co-cultured with CD71+ CD235a+ cells from the older donors.Real-time PCR were used to identify the expression of the immune cytokine secreted by CD71+ CD235a+ cells.Results (1)the results showed a significant increase in the percentage of CD71+ CD235a+ cells in the elderly compared with the young [(9.93± 2.95)％ vs (1.96 ± 1.16)％,t =3.37,P ＜ 0.001];(2)the CD71+ CD235a+ cells from the older donors could suppress the proliferation of the T cells (t =2.91,P＜ 0.05)while from the young group we could not observe this phenomenon(t =0.387,P＞0.05).(3)The results of Real-time PCR revealed that,compared with CD71+ CD235a+ cells from the young,CD71+ CD235a+ cells from the elderly expressed higher levels of Arg-2 (t =9.04,P＜0.01),IL-1β,IL-6 and TGF-β(t =4.51,5.46,6.92,all P＜0.05).Conclusions There are higher frequencies of CD71+ CD235a+ cells in aged samples than the young.And the CD71+CD235a cells from the elderly could secrete more Arg-2,IL-1β,IL-6 and TGF-β to suppress the T cell proliferation.
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Objective To explore the expression of negative costimulatory molecule B7-H1 of myeloid derived suppressor cells(MDSCs) and its roles in age-related immunosuppressive functions.Methods 36 C57BL/6 mice were divided into 2 groups:the elderly group(12-month-old,n=18) and the young group(4-8-week-old,n=18).The expressive level of negative costimulatory molecules of MDSCs in the two groups was measured by flow cytometry(FCM).Besides,MDSCs were sorted by using MDSC isolation kit and T cells were obtained by grinding lymph nodes.To explore the influence of B7-H1 on the MDSCs' immunosuppressive functions,T cell proliferation and intervention assay was conducted using the five following groups:T cells group,T cells + CFSE group [T cells with carboxyfluorescein diacetate succinimidyl ester(CFSE)],T cells +-CFSE cocultured with MDSCs from the young group,T cells + CFSE cocultured with MDSCs from the elderly group,and T cells + CFSE cocultured with MDSCs from elderly group and anti-B7-H1 blocking antibody.The effect of B7-H1 on T cell proliferation in MDSCs were observed.Results The expression level of B7-H1 of the MDSCs was significantly increased in the elderly group as compared with the young group(t=3.27,P＜0.05).T cell proliferation assay revealed that T cells were remarkably suppressed by MDSCs in elderly group as compared to the youth group(t=5.42,P＜0.05).The suppression of T cells by MDSCs was dramatically reversed by anti-B7-H1 blocking antibody in elderly group(t=8.28,P＜0.05).Conclusions B7-H1,a molecule expressed on the surface of MDSCs,plays a key role in the age-related immunosuppressive function of MDSCs.
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Objective To investigate the association between serum homocysteine (Hcy) level and oxidative stress in elderly patients with obstructive sleep apnea/hypopnea syndrome (OSAHS). Methods The 123 patients from January 2007 to June 2010 were recruited and divided into the elderly OSAHS group (n=55) and the non-elderly OSAHS group (n=68). One hundred healthy subjects were selected and divided into the elderly control group (n=52) and the non-elderly control group (n=48). The PSG variables were recorded and the levels of Hcy, malondialdehyde (MDA) and glutathione (GSH) were detected after sleep in all subjects. Serum Hcy level was measured by cyclophorase, MDA and GSH levels were measured by spectrophotometer. Results There were statistical differences in Hcy concentration between four groups (F=28.26, P=0.000). The Hcy concentration was higher in elderly OSAHS group than in other groups, and was higher in elderly control group than in non-elderly control group, and was higher in non-elderly OSAHS group than in non-elderly control group. There were statistical differences in MDA and GSH concentrations between the four groups (F=5.56, P=0.002;F=4.79, P=0.004). The concentrations of MDA and GSH were higher in elderly OSAHS group than in other groups. Multiple regression analysis indicated a statistically significant relationship of Hcy concentration with age, MDA, GSH and AHI (β=0.07, 0.50, 0.23 and 2.00). Conclusions The concentration of Hcy increases and oxidative stress enhances with aging, especially in the elderly patients with OSAHS. The mechanism of high level of serum Hcy in elderly patients with OSAHS may be the high oxidative stress.