Frequency and expression of inhibitory markers of CD4(+) CD25(+) FOXP3(+) regulatory T cells in patients with common variable immunodeficiency.

Common variable immunodeficiency (CVID) is the most symptomatic primary antibody deficiency associated with recurrent infections and chronic inflammatory diseases as well as autoimmunity. CD4(+) CD25(+) FOXP3(+) regulatory T cells (Tregs) are critical T cell subsets for maintaining self-tolerance and regulation of immune response to antigens thus play a pivotal role in preventing autoimmunity. Thirty-seven CVID patients and 18 age-/sex-matched controls were enrolled. Peripheral blood mononuclear cells (PBMCs) were obtained from both groups, and the percentage of Tregs was calculated using flow cytometry method. The mRNA expression of Tregs' surface markers cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) and glucocorticoid-induced tumour necrosis factor receptor (GITR), which are associated with Tregs' inhibitory function, was compared between patients and controls by quantitative real-time PCR TaqMan method. The results revealed that the frequency of Tregs was significantly lower in CVID patients than normal individuals (P < 0.001). In addition, CVID patients with autoimmunity were found to have markedly reduced proportion of Tregs compared to those cases without autoimmune diseases (P = 0.023). A significant difference was seen in factor forkhead box P3 (FOXP3) expression between CVID patients and controls (P < 0.001). The mRNAs of CTLA-4 and GITR genes were expressed at lower levels in CVID patients compared to control group (P = 0.005 and <0.001, respectively). Our findings showed reduced proportion of Tregs in CVID patients together with downregulation of FOXP3 protein and diminished expression of inhibitory Tregs' markers. It could be concluded that all of these changes may be responsible for cellular immune dysregulation observed in these patients especially those with autoimmune manifestation.

Effects of single administration of morphine on G-protein mRNA level in the presence and absence of inflammation in the rat spinal cord.

Antinociceptive potency of opioids is greater against various noxious stimuli in animals with peripheral inflammation. Opioid agonists stimulate activation of G-protein-coupled receptor. Changes in the resting levels of G-protein subtypes could have an effect on intracellular signalling pathways. The present study was designed to investigate the effects of analgesic morphine treatment on the level G-protein subunits mRNA in the presence and absence of inflammation. Our results showed that the carrageenan administration increased G-protein subunits. Administration of analgesic dose of morphine alone and in the presence of inflammation induced different alterations in the levels of G-protein mRNA. Taken together, the results obtained using real time RT-PCR suggested that G-protein genes expression levels following the acute administration of morphine between animals with and without inflammation could influence, at least in part, analgesic responsiveness.

Inhibition of phosphodiestrase 9 induces cGMP accumulation and apoptosis in human breast cancer cell lines, MCF-7 and MDA-MB-468.

OBJECTIVES: Phosphodiesterase 9 (PDE9) is a major isoform of phosphodiesterase hydrolysing cGMP and plays a key role in proliferation of cells, their differentiation and apoptosis, via intracellular cGMP signalling. The study described here was designed to investigate expression, activity and apoptotic effect of PDE9 on human breast cancer cell lines, MCF-7 and MDA-MB-468. MATERIALS AND METHODS: Activity and expression of PDE9 were examined using colorimetric cyclic nucleotide phosphodiesterase assay and real-time RT-PCR methods respectively; cGMP concentration was also measured. MTT viability test, annexin V-FITC staining, Hoechst 33258 staining and caspase3 activity assay were used to detect apoptosis. RESULTS: Treatment of both cell lines with BAY 73-6691 lead to reduction in PDE9 mRNA expression, PDE9 cGMP-hydrolytic activity and elevation of the intracellular cGMP response. BAY 73-6691 significantly reduced cell proliferation in a dose- and time-dependent manner and caused marked increase in apoptosis through caspase3 activation. CONCLUSION: Our results revealed that BAY 73-6691 induced apoptosis in these breast cancer cell lines through the cGMP pathway. These data suggest that BAY 73-6691 could be utilized as an agent in treatment of breast cancer.

Serum levels of interleukin (IL)-10, IL-17, transforming growth factor (TGF)-ß1, and interferon-γ cytokines and expression levels of IL-10 and TGF-ß1 genes in renal allograft recipients after donor bone marrow cell infusion.

BACKGROUND: Cytokine storm generated by an alloimmune response after transplantation can lead to either graft survival or rejection. The aim of this study was to evaluate the serum levels of interleukin (IL)-10, IL-17, transforming growth factor (TGF)-ß1, and interferon (IFN)-γ and expression levels of IL-10 and TGF-ß1 in renal allograft recipients with or without donor bone marrow cell infusion (DBMI). METHODS: We retrospectively followed 28 living unrelated kidney recipients, including 14 with and 14 without DBMI infusion for 2 years. Also, 14 healthy subjects were included as a normal control group. PBMC gene expression analysis for mRNA levels of IL-10 and TGF-ß1 cytokines relative to ß-actin as a reference gene was performed using quantitative fluorescence real-time polymerase chain reaction at the end of 2 years posttransplantation. Also, serum levels of IL-10, TGF-ß1, IFN-γ, and IL-17 in the 3 groups were measured by enzyme-linked immunosorbent assay at the same time. RESULTS: Both patient groups showed increased gene expression and serum content of IL-10 compared with normal controls. The expression levels were only significant between control patients and normal subjects (P=.02). Serum levels of IFN-γ and IL-17 were higher in untreated patients compared with normal controls (P=.03 and P=.07, respectively). DBMI patients showed significantly lower levels of serum TGF-ß1 and IL-17 compared with normal subjects (P=.05 and P=.06, respectively). Also, infused patients showed a positive correlation between circulating levels of IL-17 and IL-10 (r=0.692; P=.006), and an inverse correlation between serum creatinine and TGF-ß1 levels (r=-0.580; P=.03). CONCLUSION: The decreased levels of inflammatory cytokines besides IL-10 with increased TGF-ß1 levels and better allograft function with improved clinical outcomes were observed among infused patients, possibly indicating immunomodulatory effects of this approach in kidney allograft patients.