Higher Percentage Of CD4 Research Articles

Kidney Rejection in HIV-Positive Recipients Is Associated With Immune Activation and Higher Frequency of Donor-Reactive CD8 Cells.

Background: HIV+ kidney transplant recipients have 3 times higher rejection rate when compared to HIV- recipients. Approximately half of those who have at least one rejection experienced a rejection episode within the first 30 days after transplant, suggesting that the enhanced rejection may be due to potent immune activation or preformed memory responses to donor alloantigens. Methods: We analyzed banked peripheral blood samples from HIV+ patients collected just before transplant and compared to HIV+ disease status-matched non-transplant and HIV- age-matched control subjects. We compared the total CD4+ cell counts in the blood and used multicolor flow cytometry to determine the frequency of effector and memory T cells. The frequencies of donor-alloantigen-reactive CD8+ T cells, conventional CD4+ T cells, and regulatory T cells were measured using an improved one-way mixed lymphocyte reaction assay. Results: CD4+ T cell counts in HIV+ pre-transplant patients were not significantly lower than HIV- pre-transplant patients, likely because of the preferential enrollment of healthiest HIV+ patients defined by the study entry eligibility criteria. HIV+ transplant patients had higher percentages of CD4+CD45RA-CCR7- effector cells when compared with age-matched HIV+ non-transplant and HIV- patients (27.2±2.3, 12.0±2.0, vs. 16.1±3.0; n=52, 10, 9; p=0.0023). However, no difference between rejector and non-rejector was noted. HIV+ rejectors had higher frequency of donor-reactive CD8+ T cells when compared to non-rejectors (4.46±0.79 vs. 2.26±0.58; n=8, 8; p=0.045); whereas frequencies of CD4+ conventional T cells and regulatory T cells were not significantly different between the rejectors and the non-rejectors. Conclusion: Our results thus far are consistent with the interpretation that HIV+ transplant recipients have more activated immune system. Higher frequency of donor-reactive CD8+ T cell pre-transplant may distinguish rejectors from non-rejectors.

Higher percentage of CD3⁺CD154⁺ T lymphocytes predicts efficacy of TNF-α inhibitors in active axial SpA patients.

The objective of this study was to evaluate which subtypes of T lymphocytes (CD3(+)CD28(+) and CD3(+)CD154(+)) could predict clinical efficacy after TNF-α inhibitor treatment in active axial SpA patients. Patients who fulfilled Assessment of SpondyloArthritis international Society (ASAS) criteria for axial SpA had a BASDAI of ≥40 mm. All patients received TNF-α inhibitor treatment for 12 weeks. ASAS20 was used to evaluate the effect of the treatment at week 12. We detected the percentage of CD3(+)CD28(+) and CD3(+)CD154(+) T lymphocytes on lymphocyte cells in the peripheral blood in patients and healthy controls. We evaluated whether the percentage of the above subtypes of T lymphocytes could predict clinical efficacy by ROC curve analysis. Fifty-eight healthy controls and 74 active axial SpA patients were included. Mean age was 26.28 ± 9.08 and 26.95 ± 8.13 years for healthy controls and patients, respectively (p = 0.767). The percentage of CD3(+)CD154(+) T lymphocytes was significantly higher in axial SpA patients than in healthy controls (1.62 ± 1.89 % vs 0.79 ± 0.52 %, p < 0.0005). At baseline, the percentage of CD3(+)CD154(+) T lymphocytes was significantly higher in HLA-B27((+)) patients than HLA-B27((-)) ones (HLA-B27(+) vs HLA-B27(-):1.77 ± 1.95 % vs 0.41 ± 0.27 %, p = 0.005). Compared with baseline, the percentage of CD3(+)CD154(+) T lymphocytes significantly decreased to 0.87 ± 0.49 % at week 12 (p < 0.0005). Moreover, we found higher percentage of CD3(+)CD154(+) T lymphocytes could predict clinical efficacy of SpA patients with TNF-α inhibitor treatment (AUC = 0.733, p = 0.014). High percentage of CD3(+)CD154(+) is over-expressed on lymphocytes in peripheral blood of active SpA patients and can be down-regulated by TNF-α inhibitor therapy. High-percentage of CD3(+)CD154(+) T lymphocytes may predict clinical efficacy of TNF-α inhibitor treatment in active axial SpA patients.

Immune responses to live and inactivated Nocardia seriolae and protective effect of recombinant interferon gamma (rIFN γ) against nocardiosis in ginbuna crucian carp, Carassius auratus langsdorfii

Looking into the fact that substantial mortality and morbidity is associated with intracellular Gram +ve bacterium, Nocardia seriolae infection, an effective vaccine against this pathogen is necessary to control the significant losses in aquaculture practices. Therefore, an attempt was made to evaluate the effect of live (sub-lethal) and inactivated (antigenic form) N. seriolae on cellular and humoral immunity in ginbuna crucian carp, Carassius auratus langsdorfii as well as the therapeutic potency of recombinant interferon gamma (rIFN γ) against N. seriolae infection. Effect of live and inactivated N. seriolae immunisation on the proliferation of CD4+ T cells, CD8α+ T cells and surface Ig M+ cells in peripheral blood leucocytes, spleen, head kidney and trunk kidney of ginbuna was studied after 1st, 3rd, 7th, 15th and 30th day post immunisation. The percentage of CD8α+ T cells in spleen and head kidney of ginbuna was significantly higher at 3rd day post immunisation. Similarly, surface Ig M+ cells level was found to increase in both live and inactivated N. seriolae immunised groups. On the contrary, high percentage of CD4+ T cells was observed in live N. seriolae immunised group in both the head and trunk kidneys at 30th day post immunisation. The humoral immune response to live and inactivated N. seriolae immunised ginbuna showed high antibody titre at 15th day post immunisation but the level declined subsequently in both the immunised groups. On challenge with virulent N. seriolae (1.2 × 108 CFU/ml), the relative percent survival was 62.5 and 75 in live and inactivated N. seriolae immunised groups, respectively. Furthermore, we have also studied the therapeutic potency of rIFN γ and found the possible involvement of IFN γ in resistance mechanism in fish. Administration of rIFN γ into ginbuna (at 10 μg/fish) one day before challenge study was found to protect ginbuna. The relative percent survival of ginbuna was 43.75 and 60 when challenged with 2 different doses of N. seriolae i.e., 1.2 × 108 CFU/ml and 5 × 107 CFU/ml, respectively. In summary, this study indicates that both forms of N. seriolae immunisation as well as rIFN γ indeed elicit an effective protective immunity which will help in designing suitable vaccine and/or adjunct therapy against N. seriolae infection in fish.

B16 cell lysates plus polyinosinic-cytidylic acid effectively eradicate melanoma in a mouse model by acting as a prophylactic vaccine.

Th1 antigen-specific T cells secrete interferon-γ, which is able to kill antigen-specific cancer cells and is helpful for cancer vaccines. The aim of the present study was to explore whether B16 cell lysates plus polyinosinic-cytidylic acid (polyI:C) can effectively inhibit the progression of melanoma in an animal model. In the present study, C57BL/6 mice were divided into three groups, with each group containing more than six mice. The groups of mice were immunized twice with B16 cell lysates plus polyI:C, B16 cell lysates, or phosphate-buffered saline only, respectively. The in vivo results demonstrated that splenocytes from the mice immunized with B16 cell lysates plus poly I:C contained higher percentages of CD3+CD8+ Tlymphocytes and CD3+CD4+ Tlymphocytes, which were detected by a fluorescence-activated cell sorter, and produced higher levels of antigen-specific splenocyte proliferation activity, as detected by MTT assay. The splenocytes from the mice immunized with B16 cell lysates in combination with polyI:C produced higher levels of interferon‑γ, as detected by quantitativepolymerasechainreaction and ELISA, as well as cytotoxic T lymphocyte activity when stimulated in vitro with B16 lysates. Additionally, subcutaneous immunization of the C57BL/6 mice with B16 cell lysates plus polyI:C conferred greater protection against tumor-forming B16 melanoma cells than that of the mice immunized with injection of B16 cell lysate alone. In conclusion, the cancer vaccine of B16 cell lysates plus polyI:C exerts potently protective effects that polarize responses toward Th1 and elicit antitumor immunity.

Cambios en las Subpoblaciones de Linfocitos B y T en el Título de Anticuerpos Anti-Acuaporina-4 tras el Tratamiento de un Brote Agudo con Inmunoglobulinas y Rituximab

BackgroundThere is increasing evidence supporting that neuromyelitis optica (NMO) is an inflammatory humoral mediated disorder associated with NMO-IgG/AQP-4 antibodies. However, little is known about the subsets of B cells and T cells that contribute to the pathogenesis or therapy response. ObjectivesTo describe the clinical and immunological changes associated with intravenous immunoglobulins (IV-Igs) plus rituximab (RTX) in a patient with a severe acute attack of NMO and intrathecal synthesis of NMO-IgG/AQP-4, who previously did not respond to intravenous methylprednisolone and plasma exchange. MethodsWe sequentially analysed the levels of NMO-IgG/AQP-4 by immunohistochemistry, and B and T cells subsets by multiparametric flow-cytometry, in the CSF and peripheral blood (PB), before and alter IV-Igs plus RTX therapy. ResultsIn the CSF before treatment, and compared with PB, there was a higher percentage of CD4+ T cells and a lower percentage of CD8+ T cells and CD19+ B cells. After therapy, the percentage of CD4+ T cells remained high, and that of CD8+ T cells increased. The observed decrease in the percentage of CD19+ B cells was lower than in the PB. When the CSF was compared, it was found that the percentage of effector-memory and effector CD8+ T cells had increased after therapy, and that of IgM memory B cells and switched-memory B cells decreased. The observed changes paralleled the decrease of NMO-IgG/AQP-4 results to negative and the clinical improvement. ConclusionsOur findings confirm that, besides intrathecal humoral immune response against AQP4, B and T cell subsets are involved in the modulation of inflammation within and outside the central nervous system.

Evolution of peripheral blood T lymphocyte subsets after allogenic or autologous hematopoietic stem cell transplantation

With the aim to search for differences in T cell reconstitution after allogenic or autologous hematopoietic stem cell transplantation (HSCT), we characterized peripheral blood T-cell subsets by means of flow cytometry, in adult patients who had undergone either allogenic (n=23) or autologous (n=29) HSCT for the treatment of hematological malignancies. The patients were followed every 3 months for 21 months after HSCT.Compared to healthy controls (n=20 blood donors), the two transplanted groups displayed (i) a CD4 lymphopenia, (ii) a low percentage of naive T cells, (iii) high percentages of memory T cells and of activated T cells (HLA-DR+, CD25+) and high percentages of CD4 T cells with a high expression of CD25. The levels of TRECs (TCR rearrangement excision circles) were not significantly different between the two groups. In total, the differences of the nature and the speed of T lymphocyte reconstitution observed between the two patient groups were minor. This leads us to conclude that in allografted patients, lymphocyte activation as well as many other disturbances of subpopulations of peripheral blood lymphocytes are probably not related to the allogenicity of the graft, but are due to the expansion of T cells transfused with HSC and slow differentiation of T lymphocytes in the thymus progressively colonized by bone marrow-derived T-cell precursors.

Abstract W P100: Alterations of γδT Cell Subsets in Stroke Patients with White Matter Disease

Background: γδT cells have been shown in experimental models to play a role in the evolution of ischemic brain injury by increasing cerebral infarct volumes and neurological deficits. We have previously reported decreased levels of γδT cells in the subacute phase of human stroke. In this study we aimed to determine the functional significance of alterations in levels of γδT cells and other T cell subsets on the extent of ischemic brain injury. Methods: Blood was drawn from ischemic stroke patients within 24 hours of recruitment and stained with CD3 (PE-TR), γδTCR (PE), CD4 (FITC) and CD8 (PE-Cy5) monoclonal antibodies. γδT cells were also divided into functional subsets - CD4+ γδT, CD8+ γδT and null γδT cells. Percentages of circulating lymphocytes were measured by a fluorescent activated cell counter. The extent of acute ischemic injury was measured using lesion volume on diffusion weighted images by Image Processing Analysis and Visualization (MIPAV), while the extent of chronic ischemic injury which is manifested as white matter changes(WMC) was assessed using the Age Related White Matter Changes (ARWMC) rating score. Independent T-tests and non-parametric tests were used for between group comparisons. Results: Of the 41 patients recruited, the mean age was 66.8±13 years and 18 (43.9%) were men. Blood was drawn at a mean time of 47 ±30 hours after symptom onset. Compared to patients with absent-mild WMC (ARWMC score of 0-1), patients with moderate-severe WMC (ARWMC of 2-3) had higher percentages of CD8+ γδT cells (6.6% vs. 12.0%, p=0.005) and CD8+ T cells (22.2% vs. 28.5% p=0.036), and lower percentages of CD4+ T cells (71.0% vs. 63.5%, p=0.048) and null γδT cells (72.7% vs. 59.6%, p=0.03). However γδT percentages did not differ significantly. The volumes of acute ischemic stroke lesions did not correlate with γδT levels or with other T cell subsets. Conclusions: Alterations of T cell subsets, most notably, CD8+ γδT and CD8+ T cells were present in stroke patients with cerebral white matter disease. Their role in the formation of cerebral small vessel disease needs to be further studied.

Unraveling stem cell and progenitor subsets in autologous grafts according to methods of mobilization: implications for prediction of hematopoietic recovery

Background aimsIn the autologous setting, granulocyte colony-stimulating factor (G-CSF) (G), or, when failing, G plus plerixafor (G+P), are common regimens for mobilization of stem cells into peripheral blood. To delineate mobilization effects on graft composition and hematopoietic recovery, we compared contents of stem cells and progenitor cells in products of G+P- and G patients. Paired samples of G+P patients and prior insufficient G mobilization were available for analyses. MethodsSubset analyses of grafts were performed by flow cytometry and myeloid colony-forming assay. In search of new markers to ascertain graft quality, we determined the fractions of aldehyde dehydrogenase bright (ALDHbr) cells. ResultsG grafts contained higher percentages of CD34+ cells, CD34+CD38- cells, and committed progenitors (CD34+CD38+) compared with G+P grafts. A detailed characterization of the mobilized CD34+ cell subset showed higher percentages of CD38– among the CD34+ cells of the G+P group (P = 0.032). In contrast, the CD34+ cell subset in G grafts was characterized by a higher percentage of ALDHbr cells (P < 0.0001). Studying engraftment and day +100 graft function the G and G+P transplanted patients were comparable with respect to neutrophils, whereas in platelets they differed. In the prediction of engraftment and hematopoietic recovery, the dose of infused ALDHbr cells correlated best to both platelet (r = 0.565, P = 0.002) and neutrophil reconstitution (r = 0.366, P = 0.06). ConclusionsBesides showing dissimilar distributions of CD34+CD38– cells and progenitors in G and G+P grafts, this study further designated ALDHbr as a promising marker in determination and prediction of graft quality and hematopoietic recovery.

CD8+DR+ T-Cells and C3 Complement Serum Concentration as Potential Biomarkers in Thrombotic Antiphospholipid Syndrome

Purpose. To assess complement factors and T lymphocyte activation subset abnormalities in patients with thrombotic antiphospholipid syndrome (APS) as potential biomarkers for development of clinical complications. Methods. We assessed C3, C4, factor B concentrations (nephelometry), complement haemolytic functional activity (CH100, radial immune diffusion), and the activation status of CD4+ and CD8+ T-cells (three-colour flow cytometry) in patients with thrombotic APS. Antiphospholipid (aPL) positive patients without APS-related clinical criteria, systemic lupus erythematosus (SLE) patients, and healthy individuals were evaluated as controls. A clinical followup was performed to assess the potential relationship between the immunological parameters and development of APS-related complications. Results. Lower concentrations of C3 and higher levels of CD8+DR+ cells were risk factors for development of APS-related complications during followup, including rethrombosis and neuropsychiatric symptoms. Patients with diagnosed thrombotic APS had significantly lower levels of C3, C4, and CH100 as well as higher percentages of activated CD4+DR+ and of CD8+DR+ T-cells than healthy controls but similar to that observed in autoimmune disease controls. Conclusion. Lower C3 and C4 complement levels and higher percentages of CD8+DR+ T-cells were observed in thrombotic APS patients. The potential role of these abnormalities as biomarkers of clinical outcome warrants further evaluation in a multicenter study.

Effects of neonatal surgical castration and immunocastration in male pigs on blood T lymphocytes and health markers

Surgical castration in pig husbandry is criticized for welfare reasons. Thus, it is necessary to evaluate alternative ways of rearing male pigs, such as entire or immunocastrated animals. Immunocastration is a vaccination directed against gonadotropin-releasing hormone (GnRH) to suppress the production of sexual hormones. This study aimed at investigating the effects of these two methods of castration in comparison with intact male pigs on blood T-lymphocyte subsets and function, the immunoglobulin (Ig) response to an influenza vaccine and health markers during sexual development. A total of 70 animals were allocated to three experimental groups: entire (E), surgically castrated at 5 to 6 days of age (SC), and immunized against GnRH at 3 and 4 months of age (IC). Blood samples were collected at 3, 4 and 5 months. At slaughter, global health status and body and spleen weights were measured. Results showed that SC male pigs had fewer blood lymphocytes than E pigs at 4 and 5 months (P<0.05), whereas IC pigs did not differ significantly from E pigs. The percentages of CD3+, CD3+CD4+ and CD3+CD8+ lymphocytes were not altered by treatment (P>0.1). Compared with E pigs, the SC pigs had a higher percentage of CD3+CD4+CD8+ cells at 4 months, whereas the IC pigs had a higher percentage at 5 months (P<0.05). Regarding γδT cells, SC pigs had a lower percentage than E pigs at 4 and 5 months (P<0.05), whereas IC pigs did not differ significantly from E pigs at any age. However, there were no consequences on T-lymphocyte proliferation and total IgG or anti-influenza Ig. At slaughter, relative spleen weight was decreased in IC pigs, whereas pneumonia score was decreased in SC pigs relatively to E pigs. Overall, no clear functional consequences of either method on commercial pig immune abilities were demonstrated, but more investigations are required to ascertain this conclusion.

Characterization of CD4 and CD8 T cell responses in MuSK myasthenia gravis

Muscle specific tyrosine kinase myasthenia gravis (MuSK MG) is a form of autoimmune MG that predominantly affects women and has unique clinical features, including prominent bulbar weakness, muscle atrophy, and excellent response to therapeutic plasma exchange. Patients with MuSK MG have predominantly IgG4 autoantibodies directed against MuSK on the postsynaptic muscle membrane. Lymphocyte functionality has not been reported in this condition. The goal of this study was to characterize T cell responses in patients with MuSK MG. Intracellular production of IFN-gamma, TNF-alpha, IL-2, IL-17, and IL-21 by CD4+ and CD8+ T cells was measured by polychromatic flow cytometry in peripheral blood samples from 11 Musk MG patients and 10 healthy controls. Only one MuSK MG patient was not receiving immunosuppressive therapy. Regulatory T cells (Treg) were also included in our analysis to determine if changes in T cell function were due to altered Treg frequencies. CD8+ T cells from MuSK MG patients had higher frequencies of polyfunctional responses than controls, and CD4+ T cells had higher IL-2, TNF-alpha, and IL-17. MuSK MG patients had a higher percentage of CD4+ T cells producing combinations of IFN-gamma/IL-2/TNF-gamma, TNF-alpha/IL-2, and IFN-gamma/TNF-alpha. Interestingly, Treg numbers and CD39 expression were not different from control values. MuSK MG patients had increased frequencies of Th1 and Th17 cytokines and were primed for polyfunctional proinflammatory responses that cannot be explained by a defect in CD39 expression or Treg number.

CD22-Targeted Chimeric Antigen Receptor (CAR) T Cells Containing The 4-1BB Costimulatory Domain Demonstrate Enhanced Persistence and Superior Efficacy Against B-Cell Precursor Acute Lymphoblastic Leukemia (ALL) Compared To Those Containing CD28

Abstract B cell precursor acute lymphoblastic leukemia (BCP-ALL) remains a leading cause of death from childhood cancers despite survival rates exceeding 80%. Antibody-based CAR-engineered T cells can recognize and eliminate tumors by binding directly to a surface antigen independent from MHC restriction. CAR immunotherapy against BCP-ALL has demonstrated impressive responses and sustained remission in clinical trials targeting CD19. However, some patients receiving the CD19 CAR T cells relapse with a CD19 negative leukemia. Thus, additional CAR targets are needed. CD22 is a Siglec family lectin consisting of 7 extracellular Ig domains that is expressed on the cell surface from the pre-B cell stage of development through mature B cells and is expressed on most B cell hematologic malignancies. We previously generated a second-generation (CD3-Zeta + CD28 costimulatory domain) anti-CD22 CAR derived from a membrane proximal epitope binding scFv (m971-28z) with potent activity in vivo (Haso W et al, Blood 2013). In clinical trials T cells expressing CD19-targeted CAR with 4-1BB costimulatory domains on CD19 CARs show prolonged persistence. To improve long-term persistence of the CD22 CAR, we re-engineered our CAR vector to include a 4-1BB signaling domain (m971-BBz). In vitro data using m971-BBz improved proliferation and expansion compared to m971-28z especially when lower concentrations of IL2 were included in the culture media. When no IL2 was added to the media only the 4-1BB containing CAR expanded. No difference in killing was detected in in vitro cytotoxicity assays. We next evaluated anti-tumor activity and persistence in the NSG mouse model engrafted with the NALM6-GL cell line on day 0 and treated with CAR T cells on day 3 to directly compare the efficacy of m971-28z and m971-BBz modified T cells activated with either OKT3 or anti-CD3/CD28 beads. m971-BBz outperformed m971-28z in terms of in vivo anti-tumor activity and long-term persistence. This effect was only detected when anti-CD3/CD28 beads were used for T cell expansion. OKT3-activated cells failed to persist and demonstrated inferior antitumor activity compared to bead-expanded T cells irrespective of the costimulatory domain and despite a higher percentage of CD8 T cells with significantly better cytotoxicity in vitro. Interestingly, early peripheral blood numbers of CAR T cells in recipients of bead-expanded products demonstrated a predominance of CD4+CAR T cells consistent with preinfusion CD4/CD8 ratios. At later time points this ratio decreased with a predominance of CD8+CAR T cells. In mice receiving m971-28z CAR the CD8+CAR T cells failed to persist resulting in leukemic relapse. Furthermore, direct comparison to the CD19 CAR (FMC63-BBz) in vivo showed that the anti-CD22 CAR (m971-BBz) has equivalent activity. We conclude that anti-CD3/CD28 bead-activated T cells modified to express an anti-CD22 CAR with a 4-1BB costimulatory domain demonstrates potent antitumor activity with long-term leukemic control and offers a promising therapeutic option for pediatric ALL. Disclosures: No relevant conflicts of interest to declare.

The association between CD4+ CD25+ regulatory T cells and non-small cell lung carcinoma

Lung cancer is the second most common malignancy in the USA, and it is the leading cause of cancer-related deaths among men and women. Regulatory T cells (Tregs) in peripheral blood play crucial roles in suppressing antitumor immune responses in cancer patients. A defect in the immune response may contribute to tumor growth. This study was conducted to test the hypothesis that regulatory T cell lymphocytes might contribute to immune dysfunction in non-small cell lung carcinoma (NSCLC) patients, with a specific aim of determining its role in the development and progression of NSCLC. CD4+ CD25+ Treg cell enumeration by flow cytometry was performed for 30 NSCLC patients both preoperatively and postoperatively, together with 20 controls. Higher percentages of CD4+ CD25+ Treg cells were found in NSCLC patients both pre- (11.24 ± 5.34) and postoperatively (8.75 ± 4.66) in comparison to the control group (2.16 ± 0.97); this difference was statistically significant (p < 0.001). From this study we concluded that the CD4+ CD25+ Treg population increased significantly and correlated with clinical staging in NSCLC patients: healthy donor < stage I < stage II < stage III whether preoperatively (r = 0.5, p = 0.005) or postoperatively (r = 0.4, p = 0.008). Treg cells decreased significantly after surgical removal of the tumors, from 11.24 ± 5.34 to 8.75 ± 4.66; the difference was statistically significant (p < 0.001). From this study we concluded that CD4+ CD25+ Treg lymphocytes increased in NSCLC patients; their percentage correlates with tumor staging, and it decreases following surgical removal of the tumor. Thus, manipulation of these regulatory T cells could lead to new strategies for the treatment of cancer.

Accumulation of Effector Memory CD8+ T Cells in Nasal Polyps

T lymphocytes are prevalent in sinus mucosa and are implicated in chronic rhinosinusitis (CRS) pathogenesis. However, the major T-cell subpopulations, helper (CD4+) and cytotoxic (CD8+), have not been adequately examined in CRS. This study was designed to characterize human sinus mucosa and peripheral blood (PB) CD4+ and CD8+ T cells and their level of differentiation in CRS with nasal polyps (NPs), CRS without NPs, and control patients. A prospective study was performed. Percentages of CD4+ and CD8+ T cells and their levels of differentiation were analyzed in sinus mucosa and PB by flow cytometry. Cell populations were defined as naive, central memory, effector memory, and effector T cells using cell surface markers CD45RA, CD62L, and CD27. The influence of coexisting allergy, sinus eosinophilic mucus (EM), and culture results were examined. In all patients, sinus mucosa had a lower percentage of CD4+ and a higher percentage of CD8+ T cells compared with PB. However, CRS with NPs (n = 86) had a significantly higher percentage of mucosal CD8+ T cells compared with CRS without NPs (n = 40) in control (n = 13) patients (p < 0.0001). Effector memory T cells were increased in sinuses compared with PB in all patients; however, the percentage of effector memory CD8+ T cells was greatest in CRS with NP mucosa (p = 0.002). Surprisingly coexisting allergy or culture results did not influence the mucosal T-cell phenotype. CRS with NP patients with sinus EM had a significantly higher percentage of mucosal CD8+ T cells. Sinus mucosa in CRS with NPs is characterized by a significant enrichment of CD8+ T cells and a relative deficiency of CD4+ T cells. The majority of NP CD8+ T cells had a terminally differentiated, mature, effector memory phenotype, which raises the question, whether these cells are pathogenic or appear as a consequence of inflammation, independent of the presence of allergy or positive microbial culture.

Enhancement of antimycobacterial Th1-cell responses by a Mycobacterium bovis BCG prime-protein boost vaccination strategy

Tuberculosis is a major global health problem, and the only available vaccine Bacille Calmette-Guérin (BCG) is not sufficiently effective against the disease. It is extremely urgent to develop novel vaccine approaches. Previous research demonstrated that there were several Regions of Difference (RD1-16) between the substrains of BCG and Mycobacterium tuberculosis or Mycobacterium bovis. The ORFs Rv1769 and Rv1772 are located in the RD14 deletions and have not been major targets of study. However, some studies have demonstrated that the two genes (Rv1769 and Rv1772) are excellent T cell antigens, which might induce an immune response. What kind of role these ORFs might play in anti-mycobacterial immunity, however, is still unknown. In our research we used the BCG prime-protein boost strategy to immunize BALB/c mice and evaluated its immunogenicity. Our data suggest that our novel BCG-P+PRO69 vaccine could elicit the most long-lasting and strongest Th1 type cellular immune responses. This response is characterized by a strong antibody response, the proliferation rate of splenocytes, a high percentage of CD4+ and CD8+ T cells and high levels of IFN-γ in antigen-stimulated splenocyte cultures. These results indicate that prime-boost is a potent strategy and the protein of gene Rv1769 is a potential antigen or subunit vaccine to TB for further study.

The Correlation Between the Subsets of Tumor Infiltrating Memory T Cells and the Expression of Indoleamine 2,3-Dioxygenase in Gastric Cancer

Although many researchers have concentrated on the mechanism of T cell anergy resulting from over-expression of Indoleamine 2,3-dioxygenase (IDO), it remains unclear what alterations will developed in memory T cells (Tm) under over-expression of IDO. Immunohistochemical staining for IDO expression in gastric cancer tissues (n=50) was carried out. Tumor-infiltrating memory Tm cells were counted by flow cytometry. The association between IDO expression and the subsets of tumor infiltrating Tm are discussed. The higher IDO expressions were significantly associated with deeper invasion (P=0.016) and higher frequencies (P=0.038) of lymph node metastasis. The lower tumor-infiltrating CD4+Tm were significantly associated with the advanced clinical stage (P=0.026) and lymph node metastasis (P=0.016). The lower percentages of CD8+Tm were significantly related to undifferentiated histological type (P=0.042) and lymph node metastasis (P=0.037). However, the lower percentage of CD8+Tem was significantly correlated to differentiated histological type (P=0.017), lower frequencies of lymph node metastasis (P=0.014), and earlier clinical stage (P=0.008). The higher IDO expression patients had significantly lower percentages of CD4+Tm (P=0.012) and CD8+Tm (P=0.033). Nevertheless, it was confirmed that the higher level of IDO expression correlated with higher percentages of CD8+Tm cells in univariate and multivariate analysis (P=0.011). IDO over-expression and Tm in tumor microenvironments were correlated with the disease stage and histological type of gastric cancer. Higher IDO expression was related to the lower percentage of CD4+Tm and CD8+Tm, whereas the higher level of IDO expression related with a higher percentage of CD8+Tem.

GW24-e1722 FOXP3 demethylation as a means of identifying quantitative defects in regulatory T cells in acute coronary syndrome

ObjectivesThe contribution of regulatory T cells (Tregs) to the pathogenesis of acute coronary syndrome (ACS) remains poorly understood. One core obstacleis the lack of Treg-specific markers. A highly conserved CpG...

Human mesenchymal stem cells creating an immunosuppressive environment and promote breast cancer in mice

Human mesenchymal stem cells (hMSC) can home to tumor sites and promote tumor growth. The effects of hMSC on tumor growth are controversial and involvement of hMSC in tumor immunology has not been adequately addressed. Therefore, we investigated whether injection of hMSC affects tumor appearance, growth and metastasis, and anti-tumor immunity in an experimental animal model of metastatic breast cancer. Injection of hMSC in BALB/c mice bearing mammary carcinoma promoted tumor growth and metastasis, which was accompanied by lower cytotoxic activity of splenocytes, NK cells and CD8+ T cells in vitro. Tumor-bearing mice that received hMSC had significantly lower percentages of CD3+NKp46+ NKT-like, higher percentages of CD4+Foxp3+ T cells, increased serum levels of Th2 and decreased serum levels of Th1 cytokines, and significantly higher number of CD4+ cells expressing IL-10. These results demonstrate that immunosuppressive environment created by hMSC promoted breast tumor growth and metastasis in mice.

Intracellular cytokine expression in T cells from patients with chronic lymphocytic leukemia

Functional disorders of T lymphocytes play an essential role in abnormal immune response in patients with chronic lymphocytic leukemia. The aim of this study was to assess the profile of cytokines expressed by T cells derived from patients with CLL. We have demonstrated that the intracellular levels of IL-2, IL-4, IFN-γ, TNF, IL-6 and IL-10 were significantly higher in T cells of CLL patients than in healthy donors. Moreover, the percentages of CD4+/CD3+/TNF+, CD4+/CD3+/IFN-γ+, and CD4+/CD3+/IL-2+ cells were significantly higher in ZAP-70-positive patients compared with ZAP-70-negative ones. Likewise, significantly higher percentages of CD4+/CD3+/TNF+, CD4+/CD3+/IFN-γ+ cells were observed in CD38-positive than in CD38-negative cases. What is more, there was a significant difference in median percentage of CD3+/CD4+ cells expressing TNF, IL-4, IFN-γ, IL-2 or IL-6 between patients carrying the 11q22.3 deletion and/or the 17p13.1 deletion and patients without these genetic aberrations. Our results confirm the functional disorders of T cells in CLL and their influence on the clinical course of the disease.

FOXP3 demethylation as a means of identifying quantitative defects in regulatory T cells in acute coronary syndrome

ObjectiveThe contribution of regulatory T cells (Tregs) to the pathogenesis of acute coronary syndrome (ACS) remains poorly understood. One core obstacle is the lack of Treg-specific markers. A highly conserved CpG enriched element in forkhead box P3 intron 1 (FOXP3 i l) is unmethylated only in Tregs, and measuring the unmethylation of FOXP3 i l can be used to identify the role of Tregs in clinical diseases. This study investigated whether analyzing the demethylation status of FOXP3 i 1 is a more reliable means than using Treg-specific surface markers in ACS. Methods and resultsWe evaluated circulating Tregs percentages on different levels including cell frequencies (CD4+CD25hiFOXP3+Tregs and CD4+CD25hiCD45+naïve Tregs) or FOXP3 mRNA, FOXP3 i 1 demethylation status and related cytokine secretion in 89 patients with ACS and 35 controls. FOXP3 i 1 demethylation assay showed that the amount of Tregs in ACS patients was significantly reduced than that in controls (p = 0.0005). However, flow cytometry analysis did not identify any reduction of CD4+CD25hiFOXP3+Tregs in ACS patients. Notably, younger patients had higher percentage of CD4+CD25hiFOXP3+Tregs but decreased percentage of CD4+CD25hiCD45+naïve Tregs than either controls or older patients. Furthermore, a DNA hypomethylation agent increased the amount of CD4+CD25hiFOXP3+Tregs and Tregs related cytokine IL-10 and suppressed the production of pro-inflammatory cytokine interferon-γ by inducing FOXP3 i 1 demethylation in vitro. ConclusionsA quantitative defect of Tregs, suggestive of decreased peripheral tolerance, could be a potential hallmark of ACS disease. Targeting FOXP3 i l demethylation might elevate the inhibitory activity of Tregs in ACS.