Extracts of Cistanche deserticola

The senescence accelerated mouse prone 8 substrain (SAM-P8), widely accepted as an animal model for studying aging and antiaging drugs, was used to examine the effects of dietary supplementation with extracts of Cistanche deserticola (ECD) which has been used extensively in traditional Chinese medicine because of its perceived ability to promote immune function in the elderly. Eight-month-old male SAM-P8 mice were treated with ECD by daily oral administrations for 4 weeks. The results showed that dietary supplementation of 150 mg/kg and 450 mg/kg of ECD could extend the life span measured by Kaplan-Meier survival analysis in dose-dependent manner. Dietary supplementation of SAM-P8 mice for 4 weeks with 100, 500, and 2500 mg/kg of ECD was shown to result in significant increases in both naive T and natural killer cells in blood and spleen cell populations. In contrast, peripheral memory T cells and proinflammatory cytokine, IL-6 in serum, were substantially decreased in the mice that ingested 100 and 500 mg/kg of ECD daily. Additionally, Sca-1 positive cells, the recognized progenitors of peripheral naive T cells, were restored in parallel. Our results provide clear experimental support for long standing clinical observational studies showing that Cistanche deserticola possesses significant effects in extending life span and suggest this is achieved by antagonizing immunosenescence

Impaired thymic iNKT cell differentiation at early precursor stage in murine haploidentical bone marrow transplantation with GvHD

IntroductionEarly recovery of donor-derived invariant natural killer T (iNKT) cells are associated with reduced risk of graft-versus-host disease (GvHD) and overall survival. Patients with severe GvHD, however, had much slower iNKT cell reconstitution relative to conventional T cells.MethodsTo characterize the delay of iNKT cell reconstitution and explore its possible causes, we used a haploidentical bone marrow transplantation (haplo-BMT) mouse model with GvHD. We found the delayed recovery of thymic and peripheral iNKT cell numbers with markedly decreased thymic NKT1 subset in GvHD mice. The defective generation of thymic iNKT precursors with egress capability contributed to the reduced peripheral iNKT cells in GvHD mice. We further identified intermediate NK1.1- NKT1 precursor subpopulations under steady-state conditions and found that the differentiation of these subpopulations was impaired in the thymi of GvHD mice. Detailed characterization of iNKT precursors and thymic microenvironment showed a close association of elevated TCR/co-stimulatory signaling provided by double positive thymocytes and macrophages with defective down-regulation of proliferation, metabolism, and NKT2 signature in iNKT precursor cells. Correspondingly, NKT2 but not NKT1 differentiation was favored in GvHD mice.DiscussionThese data underline the important roles of TCR and co-stimulatory signaling in the differentiation of thymic iNKT subsets under transplantation conditions

Changes of cytokines during a spaceflight analog--a 45-day head-down bed rest.

Spaceflight is associated with deregulation in the immune system. Head-down bed rest (HDBR) at -6° is believed to be the most practical model for examining multi-system responses to microgravity in humans during spaceflight. In the present study, a 45-day HDBR was performed to investigate the alterations in human immune cell distributions and their functions in response to various stimuli. The effect of countermeasure, Rhodiola rosea (RR) treatment, was also examined. A significant decrease of interferon-γ (IFN-γ) and interleukin-17 (IL-17) productions by activated T cells, increase of IL-1β and IL-18 by activated B and myeloid cells were observed during HDBR. The upregulation of serum cortisol was correlated with the changes of IL-1 family cytokines. In addition, a significant increase of memory T and B cell and regulatory T cells (Treg) were also detected. The uptake of RR further decreased IFN-γ level and slowed down the upregulation of IL-1 family cytokines. These data suggest that for prolonged HDBR and spaceflight, the decreased protective T cell immunity and enhanced proinflammatory cytokines should be closely monitored. The treatment with RR may play an important role in suppressing proinflammatory cytokines but not in boosting protective T cell immunity

Homeostatic properties and phenotypic maturation of murine CD4+ pre-thymic emigrants in the thymus.

After a tightly regulated developmental program in the thymus, "mature" single positive (SP) thymocytes leave the thymus and enter the periphery. These newly arrived recent thymic emigrants (RTEs) are phenotypically and functionally immature, and will complete a dynamic maturation in the peripheral lymphoid organs before being licensed to be resident naïve T cells. To study the early events occurring in the RTE maturation process, we identified the phenotype of CD4(+) pre-RTEs, a population of CD4(+) SP thymocytes that have acquired the thymus egress capability. Compared to peripheral naïve T cells, CD4(+) pre-RTEs displayed superior survival capability in lymphoreplete mice and faster proliferation under lymphopenic condition. The differences in Bcl2/Bim expression and/or heightened IL-7 signaling pathway may account for the pre-RTEs' better responsiveness to homeostatic signals. Qa2, the expression of which indicates the phenotypic maturation of SPs and RTEs, was found to be upregulated in CD4(+) pre-RTEs in thymic perivascular space. Migratory dendritic cells that surround this region contribute to Qa2 expression in pre-RTEs. The dendritic cell-driven Qa2 induction of CD4(+) pre-RTEs is independent of MHC class II and Aire molecules

Immune cell subset analysis by flow cytometry.

<p>(A) The analysis of immune cell subsets from peripheral blood. Lymphocytes were first gated from forward and side scatter. CD3⁺CD56^- cells were then gated as conventional T cells. Within this population, naïve T cells were CD4⁺CD45RO^- or CD4-CD45RO^- (or termed as CD8⁺ naïve T cells); memory T cells were CD4⁺CD45RO⁺ or CD4^-CD45RO⁺ (or termed as CD8⁺ memory T cells). NKT, CD3⁺CD56⁺ and NK, CD3^-CD56⁺. The expression of CD19 was used to analyze B cells. Within CD19⁺ cells, CD21⁺CD27^- cells were naïve B cells; CD21^-CD27^- were tissue-like memory B cells; and CD27⁺ were classical memory B cells. The CD27⁺ cells were further divided into CD21⁺CD20⁺ resting memory B, CD21^-CD20⁺ activated memory B; and CD21^-CD20^- plasmablasts. (B) The analysis of Treg cells in PBMCs. Lymphocytes were stained with CD4, CD127, CD25, and Foxp3. The CD25 and Foxp3 expression of CD4⁺CD127^- cells was shown. (C) The analysis of iTreg differentiation in vitro. PBMCs were stimulated with anti-CD3 and anti-CD28 in the presence of TGF-β1 and IL-2 for 5 days. The cells were then stained with antibodies and the percentages of CD4⁺CD25⁺Foxp3⁺ cells were calculated. The plot showed the results of one representative volunteer at 5 different time points. The non-polarized control was stimulated with anti-CD3 and anti-CD28 for 5 days without the addition of polarizing cytokines.</p

Changes of T cell-derived cytokines during HDBR.

<p>(A) Scheme of the 45-day HDBR of -6° and the time of sample collections. (B) Changes of T cell-derived cytokines. PBMCs were stimulated by anti-CD3 and anti-CD28 antibodies for 2 days. The supernatants were analyzed by cytometric bead array for cytokines IFN-γ, IL-17A, IL-2, TNF-α (B) and TGF-β1, IL-10 (C). The average level of each cytokine at each time point was shown. The statistical significance between any two time points within the control group was calculated by two-tailed paired Student <i>t</i> test. The following terminology is used to denote <i>p</i> values calculated by Student <i>t</i> test: *<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.005.</p

Changes of IL-1β and IL-18 productions and serum cortisol level during HDBR.

<p>PBMCs samples were stimulated with CpG for 3 days (A) and LPS for 2 days (B). The supernatants were then collected and the concentrations of IL-1β and IL-18 were measured. The serum cortisol levels during HDBR were measured (C) and its correlations with CpG-stimulated IL-1β and IL-18 production were determined (D).</p

Production of TGF-β1 by immune cells.

<p>PBMCs were stimulated by antibodies against immunoglobulin or CpG for 3 days, or LPS for 2 days. The supernatants were examined for TGF-β1 concentration.</p

The <i>Rhodiola rosea</i> treatment results in further decreased IFN-γ secretion and slowed down the increase of IL-1 family proinflammatory cytokines.

<p>(A) The comparison of IFN-γ secretion between the control and RR treated groups. (B) The changes of IL-1β and IL-18 production in response to CpG in RR treated group. (C) The comparison of IL-1β and IL-18 production upon LPS stimulation between the control and RR treated groups. The average percentage of each cell populations at each time point was shown, together with the standard deviation. The statistical significance between the control and RR-treated groups within any single time point was calculated by two-tailed Student <i>t</i> test. The following is used to denote <i>p</i> values: *<i>p</i><0.05, **<i>p</i><0.01, ***p<0.005. The placebo group was shown as filled circle; RR-treated group as open square.</p