Cell Subsets In Peripheral Blood Research Articles

Analysis of selected T cell subsets in peripheral blood after exhaustive effort among elite soccer players.

IntroductionPhysical exercise induces developing of naïve T lymphocyte subsets into polarised effector ones by immune system. The aim of the study was to examine the influence of exhaustive effort on the selected Th cell subsets and inflammation markers among soccer players.Materials and methodsFourteen soccer players aged 18 (16-21) years old performed the progressive efficiency test on mechanical treadmill. Th and Tc memory lymphocytes’ subsets and selected cytokine concentrations pre-exercise, post-exercise and in recovery were analysed by flow cytometry.ResultsPhysical effort induced changes in Th cell percentages. Increase in recovery Treg and Th17 cell subsets’ percentages in comparison to pre-exercise values were observed (10.98 (9.83-14.07) vs. 3.95 (3.15-5.53), P < 0.001 in autumn; 10.58 (7.54-12.67) vs. 4.83 (3.73-6.81), P < 0.010 in spring for Treg and 29.21 (26.34-32.16) vs. 21.64 (18.48-25.76) in autumn; 27.15 (24.60-29.16) vs. 17.43 (15.83-19.77) in spring, both P < 0.010; for Th17, respectively). Increases in Th1 cell percentages post-exercise (31.86 (28.72-33.72) in autumn, 25.60 (21.50-29.19) in spring, both P < 0.010) and in recovery (34.64 (31.21-38.20) in autumn; 26.68 (25.17-28.07) in spring, both P < 0.001) compared to pre-exercise (22.70 (21.21-26.74) for autumn and 15.64 (14.38-19.63) for spring, respectively) were found. Interestingly, no changes in Th2 cell subsets were found. Post-exercise and recovery changes in IL-6, IL-8, TNF-α and IL-10 were also observed.ConclusionsIt seems that the given effort in the progressive test induced an anabolic effect being related not only with cytokine profile but also with CD4+ T cells’ differentiation and peripheral distribution.

Levels of regulatory T cells and invariant natural killer cells and their associations with regulatory B cells in patients with non-Hodgkin lymphoma.

Due to their immunoregulatory properties, several specialized cell subsets, including regulatory T (Treg), invariant natural killer T (iNKT) and regulatory B (Breg) cells, are involved in the pathogenesis of non-Hodgkin lymphoma (NHL). However, the interaction between various cells remains to be elucidated. The aim of the present study was to evaluate the levels of Treg, iNKT and Breg cell subsets and their interrelationships in the peripheral blood (PB) and bone marrow (BM) of patients with B-cell NHL who received rituximab-based regimens and achieved a complete remission. A total of 20 patients and 20 healthy age- and sex-matched controls were prospectively enrolled for investigation of Treg, iNKT and Breg cell subsets in PB and BM by flow cytometry and cell culture. Prior to administration of combination chemotherapy with rituximab, the patients had lower levels of Breg cells and, to a lesser degree, Treg cells, but not iNKT cells, in PB compared with controls. Compartmental differences in the levels of Treg and Breg cell subsets, but not iNKT cells, were observed between PB and BM, suggesting an increase in trafficking through the blood of these regulatory cell subsets to the marrow. Following complete remission, the levels of circulating Treg, iNKT and Breg cell subsets increased. The levels of Treg cells were not significantly associated with iNKT and Breg cell subsets, although negative correlations were observed. Taken together, these results may provide new insights into the potential role of regulatory cell subsets in patients with B-cell NHL. However, whether the observed differences between PB and BM may affect clinical outcomes requires further investigation.

Effect of emodin on T cell subsets in NOD mice with NaI‑induced experimental autoimmune thyroiditis.

Chronic lymphocytic thyroiditis (CLT), also known as Hashimoto's thyroiditis, is an autoimmune disease in which the thyroid gland is gradually destroyed. To date, only a limited number of agents can effectively suppress thyroiditis development in CLT patients. The aim of the current study was to investigate the protective effect of emodin on experimental autoimmune thyroiditis (EAT) in mice, which is considered an excellent model for CLT. NaI was used to induce the EAT model in non-obese diabetic (NOD) mice. An ELISA method was employed to detect the TgAb level (thyroid inflammation) in the serum of the EAT mice. The T cell subsets in peripheral blood and spleen were detected by flow cytometry. The histopathological study revealed that the thyroid inflammatory cell infiltration was significantly reduced by emodin compared with the model group. In addition, ELISA assays indicated that the NaI-induced serum TgAb upregulation was dramatically revered by emodin. Moreover, the level of serum IFN-γ and the cell populations of CD3+CD4+IL-4+, CD3+CD4+ IFN-γ+, CD3+CD8+IL-4+, CD3+CD8+ IFN-γ+ T cells in peripheral blood monocytes and splenic lymphocytes were significantly increased by NaI in the model group compared with in the normal group. Nevertheless, this type of increase was markedly attenuated by emodin. To conclude, the EAT model was successfully established by treating NOD mice with NaI. Emodin indicated an inhibitory effect on the autoimmune response that was significantly different in EAT compared with control mice. Furthermore, the anti-inflammatory action of emodin on EAT mice may be mediated via the inhibition of the secretion of IFN-γ and the cell numbers of CD3+CD4+IL-4+, CD3+CD4+ IFN-γ+, CD3+CD8+IL-4+ and CD3+CD8+ IFN-γ+ T cells in the peripheral blood monocytes and splenic lymphocytes. Therefore, the data may offer valuable insight on the efficacy of treatment of CLT with emodin.

Endometrium infection by human herpesvirus-6A: implication in female idiopathic infertility

We first reported HHV-6A DNA presence in 43% of endometrial cells from women with idiopathic infertility, whereas no fertile control women harbored the virus1. Growing evidence confirms this implication1-6. We investigated the effect of HHV-6A infection on the immunological status of the endometrium. This is a case-control study. The sample size guarantees to achieve a 0.8 power with 5% alpha error in detecting differences between groups. Endometrial biopsies, uterine flushing and whole blood samples were collected during the implantation (mid secretory phase) windows in 67 idiopathic infertile women. The presence of HHV-6A infection was evaluated by DNA and mRNA analysis (U22, U42, U94 genes) on endometrial cellular subsets1. We analyzed the endometrial immunological status evaluating: i) the levels of immune-regulatory HLA-G and HLA-E molecules, innate inflammatory (IL-6, IL-8, TNF-alpha), Th1 pro-inflammatory (IFN-gamma, IL-1alpha, IL-1beta, IL-12) and Th2 anti-inflammatory (IL-4, IL-10) cytokines by multiplex ELISA in uterine flushing samples; ii) endometrial receptivity to cytotrophoblasts in an endometrial 3D in vitro model; iii) natural killer (NK) cells and regulatory T cells percentage and immune-phenotype in endometrial biopsies and peripheral blood by flow cytometry. We confirmed the presence of HHV-6A infection in a 40% of idiopathic infertile women. We observed increased levels of IFN-gamma, IL-1beta and IL-12 (p<0.001; Student t test), decreased levels of both IL-4 and IL-10 (p<0.001) in uterine flushing samples from HHV-6A positive women. HLA-G and HLA-E molecules were not expressed on the surface of endometrial epithelial cells from HHV-6A positive women. As a proof of principle, endometrial biopsis positive for HHV-6A infection presented a lower permittivity to cytotrophoblasts invasion. We observed a lower amount of endometrial (e)NK cells in the endometrium of women positive for HHV-6A infection compared with endometrium negative for HHV-6A (p<0.0001; Student T test). In particular, when we analyzed (e)NK subpopulations, we observed a lower percentage of CD56brightCD16- (e)NK cells in women positive for HHV-6A infection (p<0.0001). When we looked at peripheral blood cell subsets, we observed a decrease in CD4+CD25+CD127dim/- regulatory T cells in women positive for HHV-6A infection (p=0.016). The identification of an effect of HHV-6A infection on endometrial immune status opens new perspectives in idiopathic infertile women care management. In addition, it would be possible to select antiviral therapies as novel therapeutic approaches to those idiopathic infertile women characterized by the presence of endometrial HHV-6A infection, to increase their pregnancy rate.

P007 Purity and yield optimization of EasySep human whole blood CD34 + cell isolation kit

Aim Relapse of the underlying hematological malignant disease after stem cell transplantation remains a major cause of mortality. Decline of CD34 + cell subset donor chimerism is an early marker of relapse before it can be detected in unfractionated peripheral blood or other cell subsets. One of the main challenges in performing CD34 + cell donor chimerism is obtaining pure CD34 + cell due to very low frequency of this cell population in peripheral blood. In this report, we describe our experience in optimizing the EasySep Human Total CD34 + isolation protocol to maximize the yield without compromising the purity of this rare cell subpopulation. Methods CD34 subset is isolated from 6 ml of buffy coat using EasySep Human Total CD34 + Kit, StemCell Technologies. CD15+, CD3+, CD19+, and CD34 + subsets are isolated by immunomagnetic positive selection in series from buffy coat. First, CD15 + subset is isolated using 1 ml buffy coat. The remaining 5 ml sample is used to obtain pure lymphocyte using Lymphocyte Enrichment Kit. CD3 + subset is isolated and followed by CD19 + subset isolation from supernatant of CD3 + subset isolation. Supernatant of CD19 + subset isolation was treated with RosetteSep cocktail provided in the kit and 10% whole blood mixture. CD34 + subset was then isolated from RosetteSep cocktail treated supernatant. Lineage-specific subsets were measured in flow cytometry. Results CD34 + purity was confirmed via flow cytometry. Conclusions The optimized method improved the efficiency of vendor recommended protocols by using less starting blood sample and producing higher yield and purity. Download high-res image (307KB) Download full-size image

Vγ9Vδ2 T cells proliferate in response to phosphoantigens released from erythrocytes infected with asexual and gametocyte stage Plasmodium falciparum

Vγ9Vδ2 T cells, the dominant γδ T cell subset in human peripheral blood, are stimulated by phosphoantigens, of which (E)-4-Hydroxy-3-methyl-but-2-enyl pyrophosphate, is produced in the apicoplast of malaria parasites. Cell-free media from synchronised Plasmodium falciparum asexual ring, trophozoite, and schizont stage-cultures of high purity as well as media from ruptured schizont cultures, all stimulated Vγ9Vδ2 T cell proliferation, as did media from pure gametocyte cultures, whereas media from uninfected erythrocytes cultures did not. The media from ruptured schizont cultures and all the asexual and gametocyte stage cultures contained only background iron levels, suggesting that all erythrocyte haemoglobin is consumed as the parasites develop and supporting that the phosphoantigens were released from intact parasitized erythrocytes. The Vγ9Vδ2 T cell-stimulating agent was not affected by freezing, thawing or heating but was sensitive to phosphatase treatment, confirming its phosphoantigen identity. In summary, phosphoantigens are released from parasitised erythrocytes at all developmental blood stages.

Distinct Changes of BTLA and HVEM Expressions in Circulating CD4+ and CD8+ T Cells in Hepatocellular Carcinoma Patients.

BTLA/HVEM (B and T lymphocyte attenuator/herpes virus entry mediator) pathways play a critical role in T cell suppression in tumor. However, its dynamic changes in different T cell subsets in peripheral blood and their clinical significance are largely unclear in cancer patients. In the current study, we showed distinct changes of BTLA and HVEM expressions on peripheral blood CD4+ and CD8+ T cells in patients with hepatocellular carcinoma (HCC); BTLA expression were significantly upregulated on circulating CD4+ but not CD8+ T cells. In sharp contrast, the levels of HVEM expression were significantly downregulated on circulating CD8+ but not CD4+ T cells. A strong positive correlation between BTLA expression on circulating CD4+ T cells and BTLA expression on autologous CD8+ counterparts was observed in healthy donors but absent in HCC patients. More importantly, we found that blockade of the BTLA/HVEM pathway increased IFN-γ production in both circulating CD4+ and CD8+ T cells. Collectively, our data suggested that the BTLA/HVEM pathway contributes to peripheral T cell suppression in HCC patients, and BTLA/HVEM may serve as attractive targets for HCC immunotherapy.

Parkinson\u2019s disease patients have a complex phenotypic and functional Th1 bias: cross-sectional studies of CD4+ Th1/Th2/T17 and Treg in drug-na\xefve and drug-treated patients

BackgroundParkinson’s disease (PD) affects an estimated 7 to 10 million people worldwide, and only symptomatic treatments are presently available to relieve the consequences of brain dopaminergic neurons loss. Neuronal degeneration in PD is the consequence of neuroinflammation in turn influenced by peripheral adaptive immunity, with CD4+ T lymphocytes playing a key role. CD4+ T cells may however acquire proinflammatory phenotypes, such as T helper (Th) 1 and Th17, as well as anti-inflammatory phenotypes, such as Th2 and the T regulatory (Treg) one, and to what extent the different CD4+ T cell subsets are imbalanced and their functions dysregulated in PD remains largely an unresolved issue.MethodsWe performed two cross-sectional studies in antiparkinson drug-treated and drug-naïve PD patients, and in age- and sex-matched healthy subjects. In the first one, we examined circulating Th1, Th2, Th17, and in the second one circulating Treg. Number and frequency of CD4+ T cell subsets in peripheral blood were assessed by flow cytometry and their functions were studied in ex vivo assays. In both studies, complete clinical assessment, blood count and lineage-specific transcription factors mRNA levels in CD4+ T cells were independently assessed and thereafter compared for their consistency.ResultsPD patients have reduced circulating CD4+ T lymphocytes, due to reduced Th2, Th17, and Treg. Naïve CD4+ T cells from peripheral blood of PD patients preferentially differentiate towards the Th1 lineage. Production of interferon-γ and tumor necrosis factor-α by CD4+ T cells from PD patients is increased and maintained in the presence of homologous Treg. This Th1-biased immune signature occurs in both drug-naïve patients and in patients on dopaminergic drugs, suggesting that current antiparkinson drugs do not affect peripheral adaptive immunity.ConclusionsThe complex phenotypic and functional profile of CD4+ T cell subsets in PD patients strengthen the evidence that peripheral adaptive immunity is involved in PD, and represents a target for the preclinical and clinical assessment of novel immunomodulating therapeutics.

CD8 T Effector Memory T Cells as Predictive Biomarker of Severe Acute Rejection in Lung Transplantation

Severe acute rejection (AR) after lung transplantation (LT) increases the risk of bronchiolitis obliterans syndrome with subsequent impairment in patient and graft survival. The identification of biomarkers to minimize AR rate and improve long term graft survival should be a priority in transplant field. Previously, the CD8 effector memory T (TEM) cells have been identified as a potential biomarker of acute rejection in patient awaiting for a lung transplantation(1). The aim of the study was to correlate CD8 TEM with severity of acute rejection after LT. Material and Methods 76 patients with end-stage lung disease were recruited from 2010 in our institution and prospectively followed after LT. Flow cytometry and monoclonal antibodies to identify CD8 T cell subsets in peripheral blood were used. Protocol biopsy at 21 day post transplantation was performed to identify acute rejection. Results and Discussion A total of 41 (54%) of LT recipients had acute rejection and 9 of them were severe (grade A3 or A4). Before transplant, the patients with severe acute rejection had significantly higher CD8 TEM/mm3 than no AR group (median 328.5; IQR: 187.2-507.6) vs (median 70.2; IQR: 49.3-123.8); p=0.027, whereas the levels of CD8 TEM in mild AR group are reduced compared with severe AR (median 95.4; IQR: 45.8 – 189.8); p=0.0748. The CD8 TEM levels pre-LT are increased in those with subsequent severe AR. Conclusion Monitoring the levels of CD8 TEM prior lung transplantation could identify patients at risk of suffering severe acute rejection. This biomarker should be validated in larger multicentre studies before be used in clinic. Reference San Segundo, D et al. PLoS One. 2013 Nov 13;8(11):e80601

ZAP-70 expression is associated with increased CD4 central memory T cells in chronic lymphocytic leukemia: cross-sectional study

BackgroundAlthough chronic lymphocytic leukemia is basically a B cell disease, its pathophysiology and evolution are thought to be significantly influenced by T cells, as these are probably the most important interaction partner of neoplastic B cells, participating in their expansion, differentiation and survival. Chronic lymphocytic leukemia B cells may also drive functional and phenotypic changes of non-malignant T cells. There are few data about the association between memory T cells and prognosis, especially related to ZAP-70, a common reliable surrogate of the gold standard chronic lymphocytic leukemia prognostic markers. ObjectiveThe aim of this study was to investigate whether the expression of ZAP-70 in chronic lymphocytic leukemia patients is associated with abnormal patterns of the distribution of naïve and memory T cells related to crosstalk between these cells. MethodsIn this cross-sectional, controlled study, patients with chronic lymphocytic leukemia were compared with healthy blood donors regarding the expression of ZAP-70 and the distribution of naïve and memory T cell subsets in peripheral blood as measured by flow cytometry. ResultsZAP-70 positive patients presented an increased frequency and absolute number of central memory CD4+ T cells, but not CD8+ T cells, compared to ZAP-70 negative patients and age-matched apparently healthy donors. ConclusionsBecause central memory CD4+ T cells are located in lymph nodes and express CD40L, we consider that malignant ZAP-70-positive B cells may receive beneficial signals from central memory CD4+ T cells as they accumulate, which could contribute to more aggressive disease.

Switched Memory B Cells Are Increased in Oligoarticular and Polyarticular Juvenile Idiopathic Arthritis and Their Change Over Time Is Related to Response to Tumor Necrosis Factor Inhibitors

To investigate whether abnormalities in B cell subsets in patients with juvenile idiopathic arthritis (JIA) correlate with clinical features and response to treatment. A total of 109 patients diagnosed as having oligoarticular JIA or polyarticular JIA were enrolled in the study. B cell subsets in peripheral blood and synovial fluid were analyzed by flow cytometry. Switched memory B cells were significantly increased in patients compared to age-matched healthy controls (P < 0.0001). When patients were divided according to age at onset of JIA, in patients with early-onset disease (presenting before age 6 years) the expansion in switched memory B cells was more pronounced than that in patients with late-onset disease and persisted throughout the disease course. In longitudinal studies, during methotrexate (MTX) treatment, regardless of the presence or absence of active disease, the number of switched memory B cells increased significantly (median change from baseline 36% [interquartile range {IQR} 15, 66]). During treatment with MTX plus tumor necrosis factor inhibitors (TNFi), in patients maintaining disease remission, the increase in switched memory B cells was significantly lower than that in patients who experienced active disease (median change from baseline 4% [IQR -6, 32] versus 41% [IQR 11, 73]; P = 0.004). The yearly rate of increases in switched memory B cells was 1.5% in healthy controls, 1.2% in patients who maintained remission during treatment with MTX plus TNFi, 4.7% in patients who experienced active disease during treatment with MTX plus TNFi, and ~4% in patients treated with MTX alone. Switched memory B cells expand during the disease course at a faster rate in JIA patients than in healthy children. This increase is more evident in patients with early-onset JIA. TNFi treatment inhibits this increase in patients who achieve and maintain remission, but not inthose with active disease.

Age-associated distribution of normal B-cell and plasma cell subsets in peripheral blood

Humoral immunocompetence develops stepwise throughout life and contributes to individual susceptibility to infection, immunodeficiency, autoimmunity, and neoplasia. Immunoglobulin heavy chain (IgH) isotype serum levels can partly explain such age-related differences, but their relationship with the IgH isotype distribution within memory B-cell (MBC) and plasma cell (PCs) compartments remains to be investigated. We studied the age-related distribution of MBCs and PCs expressing different IgH isotypes in addition to the immature/transitional and naive B-cell compartments. B-cell and PC subsets and plasma IgH isotype levels were studied in cord blood (n=19) and peripheral blood (n=215) from healthy donors aged 0 to 90years by using flow cytometry and nephelometry, respectively. IgH-switched MBCs expressing IgG1, IgG2, IgG3, IgA1, and IgA2 were already detected in cord blood and newborns at very low counts, whereas CD27+IgM++IgD+ MBCs only became detectable at 1 to 5months and remained stable until 2 to 4years, and IgD MBCs peaked at 2 to 4years, with both populations decreasing thereafter. MBCs expressing IgH isotypes of the second immunoglobulin heavy chain constant region (IGHC) gene block (IgG1, IgG3, and IgA1) peaked later during childhood (2-4years), whereas MBCs expressing third IGHC gene block immunoglobulin isotypes (IgG2, IgG4, and IgA2) reached maximum values during adulthood. PCs were already detected in newborns, increasing in number until 6 to 11months for IgM, IgG1, IgG2, IgG3, IgA1, and IgA2; until 2 to 4years for IgD; and until 5 to 9years for IgG4 and decreasing thereafter. For most IgH isotypes (except IgD and IgG4), maximum plasma levels were reached after PC and MBC counts peaked. PC counts reach maximum values early in life, followed by MBC counts and plasma IgH isotypes. Importantly, IgH isotypes from different IGHC gene blocks show different patterns, probably reflecting consecutive cycles of IgH isotype switch recombination through life.

Short-term administration of recombinant human erythropoietin decreases B cell number in human peripheral blood

ObjectivesThere are conflicting results on the influence of recombinant human erythropoietin (rHuEPO) administration to lymphocytes, especially to B cells. MethodsWe analyzed peripheral white blood cell (WBC) subsets in patients who received one bolus administration of rHuEPO. 119 autologous blood donors were enrolled in this study. Fourty-nine out of them were treated with rHuEPO. Blood samples were obtained before the first phlebotomy and one week later before the second one. By flow cytometry, we measured the numbers of WBC, lymphocytes, dendritic cells, CD4+ T cells, CD8+ T cells, natural killer (NK) cells, B cells, monocytes, and neutrophils, further details of B cell subsets. ResultsIn the EPO-treatment group, absolute numbers of lymphocytes, especially CD8+ T cells, NK cells, and B cells, significantly decreased after rHuEPO administration. In B cell subsets, absolute numbers of naïve B cells and IgD−CD27− B cells significantly decreased. Other B cell subsets, such as transitional B cells, memory B cells, and marginal zone B cells, also showed a decreasing trend. ConclusionThese findings suggest that a single administration of rHuEPO can influence human immune system via reduction of B cell number in peripheral blood.

Analysis of CD4+ T cell subsets in the peripheral blood of patients with progressive non-segmental vitiligo

Objective To explore the immune status of CD4+ T cell subsets in the peripheral blood of patients with progressive non-segmental vitiligo (NSV) . Methods Mononuclear cells were isolated from the peripheral blood of 30 patients with progressive NSV and 30 healthy controls by using Ficoll density gradient centrifugation. Flow cytometry was performed to determine the proportions of CD4+ T cell subsets. Enzyme-linked immunosorbent assay (ELISA) was conducted to detect serum levels of cytokines secreted by CD4+ T cell subsets. Results The proportions of Th1 and Th17 cells in the peripheral blood were significantly higher in the patients with progressive NSV than in the healthy controls (Th1 cells: 8.68% ± 4.54% vs. 5.64% ± 3.83%, P 0.05) . The patients with progressive NSV also showed significantly higher ratios of Th1/Treg and Th17/Treg compared with the healthy controls (Th1/Treg ratio: 1.32 ± 0.23 vs. 2.38 ± 0.28, P 0.05) , while the serum level of IL-17A was significantly higher in the patients than in the healthy controls (23.08 ± 5.80 pg/mL vs. 8.66 ± 1.83 pg/mL, P < 0.05) . Conclusions Effector Th1 and Th17 cells are the dominant CD4+ T cell subsets in the peripheral blood of patients with progressive NSV, and the imbalance between the effector and suppressor (Treg) CD4+ T cell subsets may be one of the factors for autoimmune response in patients with vitiligo. Key words: Vitiligo; CD4-positive T-lymphocytes; T-lymphocyte subsets; Cytokines; Autoimmunity

Peripheral White Blood Cell Subsets in Metastatic Colorectal Cancer Patients Treated with Cetuximab: The Potential Clinical Relevance.

It was demonstrated that cetuximab-induced tumor regression is based on the effects exerted by immune cells included mainly in the innate immune response. Therefore, the focus of this study was to explore the alterations in the percentages of CD16+, and/or CD56+ lymphocytes, which are comprised of NK cells, and minority of CD56+CD3+ cells, in patients with metastatic colorectal cancer before or 2 months after the treatment with cetuximab-based regimens associated with the response to therapy. The changes in the percentages of lymphocytes and granulocytes in these patients were evaluated as well. We enrolled 50 patients with wild-type KRAS metastatic colorectal cancer. Disease progression was observed in 11/50 patients (non-responders), while other patients achieved partial response or stable disease (responders). Control groups included up to 72 healthy individuals. A significant decrease in the percentages of CD56+ and CD16+CD56+ lymphocytes together with a significant decrease in the percentage of lymphocytes and an increase in the ratio of granulocyte to lymphocyte percentages were observed in patients with metastatic colorectal cancer before therapy, compared with those in the healthy individuals. In contrast to those in the responders, the percentage of CD16+ lymphocytes in the overall white blood cell pool was shown to be significantly decreased in the non-responders, together with a significantly decreased percentage of lymphocytes, a significantly increased percentage of granulocytes, and an increased ratio of granulocyte to lymphocyte percentages before treatment compared with those in the healthy controls. Two months after the initiation of the treatment, significantly decreased percentages of CD16+, CD56+, and CD16+CD56+ lymphocytes were observed in patients, compared with those determined in the healthy controls. The same changes in the amounts of circulating immune cells were also observed in the responder subgroup, but the percentages of CD16+, CD56+, and CD16+CD56+ lymphocytes 2 months after treatment in the non-responder group did not differ significantly in comparison with healthy individuals. Considerable alterations of immune cell percentages observed in patients with metastatic colorectal cancer with disease progression indicate that the assessment of peripheral white blood cell architecture before treatment initiation may be clinically relevant.

Analysis of Peripheral B Cell Subsets in Patients With Allergic Rhinitis.

PurposeRecent evidence suggests that B cells can both promote and inhibit the development and progression of allergic disease. However, the characteristics of B cell subsets in patients with allergic rhinitis (AR) have not been well documented. This study aimed to analyze the characteristics of B cell subsets in the peripheral blood of AR patients.MethodsForty-seven AR patients and 54 healthy controls were enrolled in this study, and the B cell subsets in peripheral blood of all subjects were analyzed by flow cytometry. Moreover, the serum total immunoglobulin E (IgE) and IgE concentrations secreted into the cultured peripheral blood mononuclear cells (PBMCs) were measured by using enzyme-linked immunosorbent assay.ResultsWe found the peripheral blood of AR patients contained higher percentages of memory B cells, plasma cells, and CD19+CD24hiCD27+ regulatory B cells (Bregs) than those of age-matched healthy controls (P<0.05), while the percentages of naïve B cells and CD19+CD24hiCD38hi Bregs were significantly lower in AR patients than in healthy individuals (P<0.05). In addition, the serum total IgE and IgE concentrations secreted into the cultured PBMCs were elevated in AR patients than in the healthy controls (P<0.05).ConclusionsOur findings indicate that AR patients were characterized by increase in terminally differentiated memory B cells or plasma cells and decreases in CD19+CD24hiCD38hi Breg cells in the peripheral blood.

Decline in peripheral blood NKG2D+CD3+CD56+ NKT cells in metastatic colorectal cancer patients.

Colorectal cancer (CRC) is one of the main causes of cancer deaths in the world. This cancer can be divided into non-metastatic and metastatic CRC stages. CD3+CD56+ NKT cell subsets are a minor T cell subset in peripheral blood and conduct the killing of tumor cells in direct manner. Little is obvious about levels and surface markers of these cells such as NKG2D in different cancers, especially in CRC. We included 15 non-metastatic (low-grade), 11 non-metastatic (high-grade), 10 metastatic colorectal cancer patients and 18 healthy controls. The percentages of CD3+CD56+ NKT cells and NKG2D+CD56+ NKT cells from samples were analyzed by flow cytometry in peripheral blood mononuclear cells (PBMCs) of samples. We found that there was a significantly lower number of NKG2D+CD3+CD56+ cells in peripheral blood of patients with metastatic colorectal cancer compared with normal controls (77.53 ± 5.79 % vs 90.74 ± 9.84 %; p<0.01). The fact that frequency of NKG2D+CD56+ NKT cells was significantly lower in patients with metastatic colorectal cancer compared to healthy controls strengthens the hypothesis that NKT cells can play a substantial role in the protection against human colorectal cancer, and this opens up avenues for novel studies about elucidating the other aspects of tumor surveillance in CRC progression and immunotherapy (Tab. 2, Fig. 2, Ref. 46).

Association of imbalance of effector T cells and regulatory cells with the severity of asthma and allergic rhinitis in children.

This study investigated the frequency and absolute numbers of different subsets of peripheral blood cells, including CD4+ T cells and CD19+ interleukin (IL) 10+ B regulatory cells (Breg) and their potential association with clinical laboratory measurements in children diagnosed with simple asthma or asthma plus allergic rhinitis (AR). The frequency and numbers of peripheral blood CD4+ interferon (IFN) gamma+ T-helper (Th) type 1, CD4+ IL-4+ Th2, CD4+ IL-17A+ Th17, CD4+CD25+ Forkhead Box P3+ regulatory T cells (Treg) and CD19+ IL-10+ Bregs in 22 children with asthma, 17 children with asthma and AR, and 25 healthy controls were determined by flow cytometry. The levels of serum cytokines were determined by enzyme-linked immunosorbent assay. In comparison with healthy controls, patients with asthma alone had significantly increased numbers of Th1, Th2, and Th17 cells, and their cytokines but decreased numbers of Tregs and Bregs, and the cytokines IL-10 and transforming growth factor beta 1. This imbalance between effector and regulatory cells and their cytokines further increased in patients with asthma and AR. The ratios of percentage effector T cells (Th1 + Th2 + Th17) to regulatory cells (Treg + Breg) were positively correlated with fractional exhaled nitric oxide but negatively correlated with forced expiratory volume in 1 second in patients with asthma or asthma plus AR. The imbalance of effector T cells and regulatory cells contributed to the pathogenesis of airway inflammation of asthma and AR in children.

Increased proportion of a CD38highIgD+ B cell subset in peripheral blood is associated with clinical and immunological features in patients with primary Sjögren's syndrome

We investigated the correlation between the increased proportion of peripheral B cell subsets and clinical and immunological features in primary Sjögren's syndrome (pSS). We found that the proportion of CD19+ B cells was significantly increased in pSS as compared with HC and was correlated with serum IgG levels. Moreover, in vitro IgG production by CD19+ B cells was significantly increased in pSS and was positively and significantly correlated with serum IgG levels. FACS analysis revealed that the proportions of peripherally CD38highIgD+ B cells and CD38highIgD− B cells were significantly increased in pSS. In addition, the proportion of CD38highIgD+ B cells positively correlated with ESSDAI scores and serum levels of IgG, anti-Ro/SSA and anti-La/SSB antibodies while that of CD38highIgD− B cells showed no correlation with these parameters. Our data suggest that increased proportion of CD38highIgD+ B cells in pSS is involved in IgG overproduction including autoantibodies, and correlates with disease progression.

Birth weight and postnatal microbial exposures predict the distribution of peripheral blood leukocyte subsets in young adults in the Philippines

The immune system not only provides protection against infectious disease but also contributes to the etiology of neoplastic, atopic, and cardiovascular and metabolic diseases. Prenatal and postnatal nutritional and microbial environments have lasting effects on multiple aspects of immunity, indicating that immune processes may play important roles in the developmental origins of disease. The objective of this study is to evaluate the association between birth weight and the distribution of leukocyte (white blood cell) subsets in peripheral blood in young adulthood. Postnatal microbial exposures were also considered as predictors of leukocyte distribution. Participants (n=486; mean age=20.9 years) were drawn from a prospective birth cohort study in the Philippines, and analyses focused on the following cell types: CD4 T lymphocytes, CD8 T lymphocytes, B lymphocytes, natural killer cells, monocytes, granulocytes. Higher birth weight was a strong predictor of higher proportion of CD4 T lymphocytes (B=0.12, s.e.=0.041, P=0.003), lower proportion of CD8 T lymphocytes (B=-0.874, s.e.=0.364, P=0.016), higher CD4:CD8 ratio (B=1.964, s.e.=0.658, P=0.003), and higher B lymphocytes (B=0.062, s.e.=0.031, P=0.047). Measures of microbial exposure in infancy were negatively associated with proportions of B lymphocytes and granulocytes, and lower CD4:CD8 ratio. Leukocytes are the key regulators and effectors of innate and specific immunity, but the origins of variation in the distribution of cell type across individuals are not known. Our findings point toward nutritional and microbial exposures in infancy as potentially important determinants of immune-phenotypes in adulthood, and they suggest that leukocyte distribution is a plausible mechanism through which developmental environments have lasting effects on disease risk in adulthood.