Distribution of Lymphocyte Subsets in Healthy Human Immunodeficiency Virus-Negative Adult Ethiopians from Two Geographic Locales

Immunological values for 562 factory workers from Wonji, Ethiopia, a sugar estate 114 km southeast of the capital city, Addis Ababa, Ethiopia, were compared to values for 218 subjects from Akaki, Ethiopia, a suburb of Addis Ababa, for whom partial data were previously published. The following markers were measured: lymphocytes, T cells, B cells, NK cells, CD4⁺ T cells, and CD8⁺ T cells. A more in depth comparison was also made between Akaki and Wonji subjects. For this purpose, various differentiation and activation marker (CD45RA, CD27, HLA-DR, and CD38) expressions on CD4⁺ and CD8⁺ T cells were studied in 60 male, human immunodeficiency virus-negative subjects (30 from each site). Data were also compared with Dutch blood donor control values. The results confirmed that Ethiopians have significantly decreased CD4⁺ T-cell counts and highly activated immune status, independent of the geographic locale studied. They also showed that male subjects from Akaki have significantly higher CD8⁺ T-cell counts, resulting in a proportional increase in each of the CD8⁺ T-cell compartments studied: naïve (CD45RA⁺CD27⁺), memory (CD45RA⁻CD27⁺), cytotoxic effector (CD45RA⁺CD27⁻), memory/effector (CD45RA⁻CD27⁻), activated (HLA-DR⁺CD38⁺), and resting (HLA-DR⁻CD38⁻). No expansion of a specific functional subset was observed. Endemic infection or higher immune activation is thus not a likely cause of the higher CD8 counts in the Akaki subjects. The data confirm and extend earlier observations and suggest that, although most lymphocyte subsets are comparable between the two geographical locales, there are also differences. Thus, care should be taken in extrapolating immunological reference values from one population group to another.     

T-Cell Receptor Vβ Repertoire CDR3 Length Diversity Differs within CD45RA and CD45RO T-Cell Subsets in Healthy and Human Immunodeficiency Virus-Infected Children

The T-cell receptor (TCR) CDR3 length heterogeneity is formed during recombination of individual Vβ gene families. We hypothesized that CDR3 length diversity could be used to assess the fundamental differences within the TCR repertoire of CD45RA and CD45RO T-cell subpopulations. By using PCR-based spectratyping, nested primers for all 24 human Vβ families were developed to amplify CDR3 lengths in immunomagnetically selected CD45RA and CD45RO subsets within both CD4⁺ and CD8⁺ T-cell populations. Umbilical cord blood mononuclear cells or peripheral blood mononuclear cells obtained from healthy newborns, infants, and children, as well as human immunodeficiency virus (HIV)-infected children, were analyzed. All T-cell subsets from newborn and healthy children demonstrated a Gaussian distribution of CDR3 lengths in separated T-cell subsets. In contrast, HIV-infected children had a high proportion of predominant CDR3 lengths within both CD45RA and CD45RO T-cell subpopulations, most commonly in CD8⁺ CD45RO T cells. Sharp differences in clonal dominance and size distributions were observed when cells were separated into CD45RA or CD45RO subpopulations. These differences were not apparent in unfractionated CD4⁺ or CD8⁺ T cells from HIV-infected subjects. Sequence analysis of predominant CDR3 lengths revealed oligoclonal expansion within individual Vβ families. Analysis of the CDR3 length diversity within CD45RA and CD45RO T cells provides a more accurate measure of disturbances in the TCR repertoire than analysis of unfractionated CD4 and CD8 T cells.     

Phenotypic and functional characterization of vaginal dendritic cells in a rat model of Candida albicans vaginitis

This study analyzes the phenotype of vaginal dendritic cells (VDCs), their antigenic presentation and activation of T-cell cytokine secretion, and their protective role in a rat model of Candida vaginitis. Histological observation demonstrated a significant accumulation of OX62⁺ VDCs in the mucosal epithelium of Candida albicans-infected rats at the third round of infection. We identified two subsets of OX62⁺ VDCs differing in the expression of CD4 molecule in both noninfected and Candida-infected rats. The OX62⁺ CD4⁺ subset of VDCs displayed a lymphoid cell-like morphology and expressed the T-cell antigen CD5, whereas the OX62⁺ CD4⁻ VDC subset exhibited a myeloid morphology and was CD5 negative. Candida infection resulted in VDC maturation with enhanced expression of CD80 and CD134L on both CD4⁺ and CD4⁻ VDC subsets at 2 and 6 weeks after Candida infection. CD5⁻ CD4⁻ CD86⁻ CD80⁻ CD134L⁺ VDCs from infected, but not noninfected, rats spontaneously released large amounts of interleukin-12 (IL-12) and tumor necrosis factor alpha, whereas all VDC subsets released comparable levels of IL-10 and IL-2 cytokines. Furthermore, OX62⁺ VDCs from infected rats primed naïve CD4⁺ T-cell proliferation and release of cytokines, including gamma interferon, IL-2, IL-6, and IL-10, in response to staphylococcal enterotoxin B stimulation in vitro. Adoptive transfer of highly purified OX62⁺ VDCs from infected rats induced a significant acceleration of fungal clearance compared with that in rats receiving naive VDCs, suggesting a protective role of VDCs in the anti-Candida mucosal immunity. Finally, VDC-mediated protection was associated with their ability to rapidly migrate to the vaginal mucosa and lymph nodes, as assessed by adoptive transfer of OX62⁺ VDCs labeled with 5 (and 6-)-carboxyfluorescein diacetate succinimidyl ester.     

Antigen-non-specific regulation centered on CD25+Foxp3+ Treg cells

CD4⁺CD25⁺Foxp3⁺ regulatory T cells (Tregs) are of special interest in immunology because of their potent inhibitory function. Many fundamental aspects of Tregs, including their antigenic profile, development and peripheral homeostasis, remain highly controversial. Here, we propose a Treg-centered antigen-non-specific immunoregulation model focused on the T-cell system, particularly on CD4⁺ T cells. The T-cell pool consists of naive T cells (Tnais), Tregs and effector T cells (Teffs). Regardless of antigen specificity, the ratio of the activated T-cell subsets (Treg/Teff/Tnai) and their temporal and spatial uniformity dictate the differentiation of Tnais. Activated Tregs inhibit the activation, proliferation, induction and activity of Teffs; in contrast, activated Teffs inhibit the induction of Tregs from Tnais but cooperate with Treg-specific antigens to promote the proliferation and activity of Tregs. In many cases, these interactions are antigen-non-specific, whereas the activation of both Tregs and Teffs is antigen-specific. Memory T-cell subsets are essential for the maintenance of adaptive immune responses, but the antigen-non-specific interactions among T-cell subsets may be more important during the establishment of the adaptive immune system to a newly encountered antigen. This is especially important when new and memory antigens are presented closely—both temporally and spatially—to T cells, because there are always baseline levels of activated Tregs, which are usually higher than levels of memory T cells for new antigens. Based on this hypothesis, we further infer that, under physiological conditions, Tregs in lymph nodes mainly recognize antigens frequently released from draining tissues, and that these self-reactive Tregs are commonly involved in the establishment of adaptive immunity to new antigens and in the feedback control of excessive responses to pathogens.     

Low Relative Frequencies of CD26+ CD4+ Cells in Long-Term Nonprogressing Human Immunodeficiency Virus Type 1-Infected Subjects

A broad antibody panel was used for immunophenotyping of human immunodeficiency virus type 1 (HIV-1)-infected patients who were long-term nonprogressors (LTNP). The LTNP were compared with patients in the early phase of infection and patients who had progressed to advanced immunodeficiency. Changes in CD8⁺ subset distribution were observed mainly at acquisition of HIV-1 infection, whereas CD4⁺ subset changes appeared during progression of HIV-1 infection. The decreasing levels of CD4⁺ cells were characterized by an increasing frequency of cells expressing the activation markers HLA-Dr and CD45RO but not the CD28 surface antigen. The LTNP exhibited significant changes compared to HIV-negative patients in almost all markers. Compared to patients in the early phase of infection, the only difference was a relatively lower frequency of CD4⁺ cells expressing CD26 among the LTNP. The results show that HIV-1-infected persons who have no signs of immunodeficiency despite many years of infection have an immunophenotypic pattern that is substantially different from that of noninfected persons. Despite the long duration of infection, the LTNP exhibit a pattern similar to that of newly infected persons, with the exception of lower expression of CD26 on CD4⁺ cells.     

Glycosphingolipids as Novel Targets for T-Cell Suppression by the B Subunit of Recombinant Heat-Labile Enterotoxin

Heat-labile enterotoxin subunit B (LTB) is a noncatalytic protein derived from Escherichia coli that binds to ganglioside GM₁, a glycosphingolipid on the surface of mammalian cells. In this study, the effects of recombinant LTB (rLTB) on murine lymphocytes were examined in vitro. T and B cells readily bound fluorescein isothiocyanate-labeled rLTB. CD8⁺ T cells bound twice as much as CD4⁺ T cells and B cells. Exposure of T-cell subsets and B cells to rLTB abrogated mitogen-driven proliferation. CD8⁺ T cells were more susceptible to rLTB than either CD4⁺ T cells or B cells. There were differences in the sensitivity of lymphocytes from various strains of mice to rLTB. This was attributed to qualitative and quantitative differences in the CD4⁺ T cells. rLTB induced apoptosis in both T-cell subsets, but the level was significantly higher in CD8⁺ T cells. Apoptosis peaked at around 8 h after exposure to rLTB and incubation at 37°C. Binding to ganglioside GM₁ was essential for suppression, since rLTB/G33D, a mutant which does not bind GM₁, failed to inhibit proliferation or induce apoptosis. Naive T cells, which were acutely sensitive to rLTB, became more resistant after activation. Conversely, activated T cells regained their sensitivity to rLTB when they reverted back to a resting state. A 1-h pulse with rLTB was sufficient to inhibit T-cell proliferation and cytotoxic-T-lymphocyte generation in primary mixed lymphocyte reaction cultures. CD8⁺ T cells were preferentially depleted in these cultures. rLTB also induced functional modifications in T cells as indicated by inhibition of gamma interferon secretion after polyclonal activation. Thus, rLTB may have immunomodulatory properties independent of its ability to induce apoptosis.     

Characterization of CD8+CD57+ T cells in patients with acute myocardial infarction

Although T cells are known to be involved in the pathogenesis of coronary artery disease, it is unclear which subpopulation of T cells contributes to pathogenesis in acute myocardial infarction (MI). We studied the immunological characteristics and clinical impact of CD8⁺CD57⁺ T cells in acute MI patients. The frequency of CD57⁺ cells among CD8⁺ T cells was examined in peripheral blood sampled the morning after acute MI events. Interestingly, the frequency of CD57⁺ cells in the CD8⁺ T-cell population correlated with cardiovascular mortality 6 months after acute MI. The immunological characteristics of CD8⁺CD57⁺ T cells were elucidated by surface immunophenotyping, intracellular cytokine staining and flow cytometry. Immunophenotyping revealed that the CD8⁺CD57⁺ T cells were activated, senescent T cells with pro-inflammatory and tissue homing properties. Because a high frequency of CD8⁺CD57⁺ T cells is associated with short-term cardiovascular mortality in acute MI patients, this specific subset of CD8⁺ T cells might contribute to acute coronary events via their pro-inflammatory and high cytotoxic capacities. Identification of a pathogenic CD8⁺ T-cell subset expressing CD57 may offer opportunities for the evaluation and management of acute MI.     

T-Cell-Mediated Immune Responses in Patients with Cutaneous or Mucosal Leishmaniasis: Long-Term Evaluation after Therapy

T-cell immune responses in patients with cutaneous leishmaniasis (CL) and mucosal leishmaniasis (ML) were studied during the active disease, at the end of therapy, and 1 to 17 years posttherapy (long-term follow-up). Lymphocyte proliferative responses, phenotypic characterization of CD4⁺ and CD8⁺ Leishmania-reactive T cells, and cytokine production were assayed. Patients with active ML and CL showed higher proportions of CD4⁺ than CD8⁺ T cells. In CL, the healing process was associated with a decrease of CD4⁺ and an increase of CD8⁺, leading to similar CD4⁺ and CD8⁺ proportions. This pattern was only seen in ML after long-term therapy. Long-term follow-up of patients with CL showed a positive CD4⁺/CD8⁺ ratio as observed during the active disease, although the percentages of these T cell subsets were significantly lower. Patients with CL did not show significant differences between gamma interferon (IFN-γ) and interleukin-5 (IL-5) production during the period of study. Patients with active ML presented higher IFN-γ and IL-5 levels compared to patients with active CL. IL-4 was only detected during active disease. Patients long term after cure from ML showed increasing production of IFN-γ, significant decrease of IL-5, and no IL-4 production. Two apparently beneficial immunological parameters were detected in tegumentary leishmaniasis: (i) decreasing proportions of CD4⁺ Leishmania-reactive T cells in the absence of IL-4 production associated with cure of CL and ML and (ii) decreasing levels of IL-5 long after cure, better detected in patients with ML. The observed T-cell responses maintained for a long period in healed patients could be relevant for immunoprotection against reinfection and used as a parameter for determining the prognosis of patients and selecting future vaccine preparations.     

Apoptosis in Cord Blood T Lymphocytes from Infants of Human Immunodeficiency Virus-Infected Mothers

Apoptosis continues to be controversial in human immunodeficiency virus (HIV)-induced pathogenesis. To investigate whether apoptosis occurs with HIV exposure with or without subsequent infection, levels of apoptosis were measured in cord blood lymphocytes (CBL) from seven newborns delivered to HIV-infected mothers and seven normal, unexposed newborns. Live cells were costained with antibodies to cell surface markers and the DNA dye 7-amino actinomycin D to immunophenotype apoptotic CBL subsets. Apoptosis was measured in fresh and cultured CBL in the presence and absence of CD3 T-cell receptor stimulation. Compared to the CD4⁺ CBL from HIV-unexposed newborns, CD4⁺ CBL from six HIV-exposed, noninfected newborns demonstrated significantly greater apoptosis after overnight culture even in the absence of CD3 stimulation. Compared to HIV-unexposed controls, CD8⁺ CBL from the six HIV-exposed newborns also demonstrated increased levels of apoptosis after overnight culture without stimulation. The one HIV-infected newborn in this study showed the highest levels of CD4⁺ and CD8⁺ apoptotic CBL. The data suggest that levels of apoptosis are increased in infants in association with HIV infection. Furthermore, CD4⁺ and CD8⁺ cord blood lymphocytes from HIV-exposed infants behaved differently than T lymphocytes from either normal, unexposed infants or an HIV-infected infant. These results suggest that exposure to HIV or HIV-induced factors increases the levels of apoptosis in CBL.     

Primary activation of antigen-specific naive CD4+ and CD8+ T cells following intranasal vaccination with recombinant bacteria

The primary activation of T-helper and T-cytotoxic cells following mucosal immunization with recombinant Streptococcus gordonii was studied in vivo by adoptive transfer of ovalbumin (OVA)-specific transgenic CD8⁺ (OT-I) and CD4⁺ (OT-II) T cells. A recombinant strain, expressing on the surface the vaccine antigen Ag85B-ESAT-6 from Mycobacterium tuberculosis fused to OVA T-helper and T-cytotoxic epitopes (peptides 323 to 339 and 257 to 264), was constructed and used to immunize C57BL/6 mice by the intranasal route. Recombinant, but not wild-type, bacteria induced OVA-specific CD4⁺ and CD8⁺ T-cell clonal expansion in cervical lymph nodes, lung, and spleen. OVA-specific CD4⁺ and CD8⁺ T-cell proliferation appeared first in cervical lymph nodes and later in the spleen, suggesting a possible migration of activated cells from the inductive site to the systemic district. A significant correlation between the percentages of CD4⁺ and CD8⁺ proliferating T cells was observed for each animal. The expression of CD69, CD44, and CD45RB on proliferating T lymphocytes changed as a function of the cell division number, confirming T-cell activation following the antigen encounter. These data indicate that intranasal immunization with recombinant S. gordonii is capable of inducing primary activation of naive antigen-specific CD4⁺ and CD8⁺ T cells, both locally and systemically.     

Comparative Analysis of Human Cytomegalovirus-Specific CD4+ T-Cell Frequency and Lymphoproliferative Response in Human Immunodeficiency Virus-Positive Patients

Evaluation of human cytomegalovirus (HCMV)-specific T-helper immunity could contribute in optimizing anti-HCMV therapy in human immunodeficiency virus (HIV)-infected patients. Testin the lymphoproliferative response (LPR) is the standard technique used to evaluate T-helper response, but its use in the routine diagnostic laboratory setting can be problematic. The most promising new alternative technique is the determination of HCMV-specific CD4⁺ T-cell frequency by flow cytometry detection of intracellular cytokine production after short-term antigen-specific activation of peripheral blood mononuclear cells. HCMV-specific LPR and CD4⁺ T-cell frequency were compared in a group of 78 blood samples from 65 HIV-infected patients. The results showed concordance in 80.7% of samples. In addition, comparative analysis of sequential blood samples from 13 HIV-infected patients showed that while in about half of patients the T-helper HCMV-specific immune response remained stable during highly active antiretroviral therapy (HAART), in the other half declining levels of the HCMV-specific CD4⁺-mediated immune response were determined by either one or both assays. In conclusion, our data suggest that the determination of HCMV-specific CD4⁺ T-cell frequency can be considered a valuable alternative to the LPR test for the detection of HCMV-specific T-helper response in HIV-infected patients. It could facilitate wider screening of anti-HCMV T-helper activity in HIV-infected patients, with potential benefits for clinicians in deciding strategies of discontinuation or maintenance of anti-HCMV therapy.     

Preservation of Lymphocyte Immunophenotype and Proliferative Responses in Cryopreserved Peripheral Blood Mononuclear Cells from Human Immunodeficiency Virus Type 1-Infected Donors: Implications for Multicenter Clinical Trials

Human immunodeficiency virus type 1 (HIV-1) infection results in impaired immune function that can be measured by changes in immunophenotypically defined lymphocyte subsets and other in vitro functional assays. These in vitro assays may also serve as early indicators of efficacy when new therapeutic strategies for HIV-1 infection are being evaluated. However, the use of in vitro assays of immune function in multicenter clinical trials has been hindered by their need to be performed on fresh specimens. We assessed the feasibility of using cryopreserved peripheral blood mononuclear cells (PBMC) for lymphocyte immunophenotyping and for lymphocyte proliferation at nine laboratories. In HIV-1-infected patients with moderate CD4⁺ lymphocyte loss, the procedures of density gradient isolation, cryopreservation, and thawing of PBMC resulted in significant loss of CD19⁺ B cells but no measurable loss of total T cells or CD4⁺ or CD8⁺ T cells. No significant changes were seen in CD28⁻ CD95⁺ lymphocytes after cell isolation and cryopreservation. However, small decreases in HLA-DR⁺ CD38⁺ lymphocytes and of CD45RA⁺ CD62L⁺ were observed within both the CD4⁺ and CD8⁺ subsets. Fewer than 10% of those specimens that showed positive PBMC proliferative responses to mitogens or microbial antigens lost their responsiveness after cryopreservation. These results support the feasibility of cryopreserving PBMC for immunophenotyping and functional testing in multicenter AIDS clinical trials. However, small changes in selected lymphocyte subsets that may occur after PBMC isolation and cryopreservation will need to be assessed and considered in the design of each clinical trial.     

Most CD56+ Intestinal Lymphomas Are CD8+CD5− T-Cell Lymphomas of Monomorphic Small to Medium Size Histology

The expression of the natural killer (NK) cell marker CD56 has been reported to occur in NK cell lymphomas/leukemias and a small group of peripheral T-cell lymphomas but has not been studied extensively in primary intestinal non-B-cell lymphomas. Normal human jejunal intraepithelial lymphocytes (IELs) are mainly T-cell receptor (TCR)-αβ⁺CD3⁺CD8⁺CD5^low and include an ~15% fraction of CD56⁺ cells that could be the cells of origin for CD56⁺ intestinal T-cell lymphoma (ITL). To test this hypothesis, 70 cases diagnosed as ITL were immunophenotyped, and 15 CD56⁺ cases (21%) were identified. The majority of the CD56⁺ lymphomas was of monomorphic small to medium-sized histology, shared the common phenotype βF1^±CD3ε/cyt⁺CD8⁺CD4⁻CD5⁻CD57⁻TIA-1⁺ and had clonally rearranged TCR γ-chain genes. In contrast, the CD56⁻ lymphomas were mainly composed of pleomorphic medium and large cells or had a morphology most consistent with anaplastic large-cell lymphoma and were mostly CD8⁻. These findings suggest that the majority of CD56⁺ intestinal lymphomas are morphologically and phenotypically distinct T-cell lymphomas most likely derived from activated cytotoxic CD56⁺CD8⁺ IELs. Some overlapping histological and clinical features between CD56⁺ and CD56⁻ ITLs indicate that the former belong to the clinicopathological entity of ITL. The consistent expression of cytotoxic-granule-associated proteins introduces ITL (both CD56⁺ and CD56⁻) into the growing family of usually aggressive extranodal lymphomas of cytotoxic T-cell and NK-cell derivation. In contrast to putative NK-cell lymphoma of the sinonasal region, intestinal NK-cell lymphoma seems to be very rare.     

Dysregulated Synthesis of Intracellular Type 1 and Type 2 Cytokines by T Cells of Patients with Cutaneous T-Cell Lymphoma

Mycosis fungoides (MF) and Sezary syndrome (SS) are the two main clinical entities of cutaneous T-cell lymphoma (CTCL). As the disease progresses from MF to SS, a switch from a type 1 (interleukin [IL]-2 and gamma interferon [IFN-γ]) to a type 2 (IL-4) cytokine production profile occurs. Although roles for type 1 and type 2 cytokines in the pathogenesis of CTCL have been proposed, the cellular origins of these cytokines are unclear. Using flow cytometry to identify individual T-cell subsets, we studied cytokine synthesis by the T cells of 13 patients with SS and 12 with MF and 9 hematologically healthy donors. Upon activation with phorbol 12-myristate 13-acetate (PMA), the numbers of T cells synthesizing IL-2 were similar for all study groups. Whereas the predominant T-cell producing IL-2 in healthy donors and in those with MF was CD7⁺, in patients with SS, it was CD7⁻. Although the number of IL-4⁺ CD4⁺ T cells was low for all study groups, there was a significantly higher number of IL-4⁺ CD8⁺ T cells in patients with MF than in those with SS or healthy donors. There was a decline in the number of IFN-γ-producing T cells in CTCL donors compared to that in healthy donors. More importantly, there was a significant decrease in the number of IFN-γ-producing T cells with disease progression from MF to SS. The inability of these T cells to synthesize IFN-γ may have prognostic value in CTCL, since it may be responsible for the progression of the disease from MF to SS.     

Effect of Photopheresis on Lymphocyte Population in Children with Newly Diagnosed Type 1 Diabetes

In recent years photopheresis has been claimed to be an effective form of immunomodulation. It has also been shown to have an effect on the disease process at the onset of type 1 diabetes. In a double-blind, placebo-controlled randomized study, we analyzed if the effect of photopheresis in children with newly diagnosed diabetes is related to changes in the balance of lymhocyte populations. We also analyzed if lymphocyte subsets were related to recent infection, mild or aggressive disease manifestations, heredity, or gender. Nineteen children received active treatment with photopheresis, while 21 children received sham pheresis (placebo group). No influence of a history of previous infection, heredity, or certain clinical parameters on lymphocyte subsets was found. At the onset of type 1 diabetes, girls showed a higher proportion and a larger number of T cells (CD3⁺) and T-helper cells (CD4⁺) and a higher proportion of naïve CD4⁺CD45RA⁺ cells. In the placebo group, an increase in the number of subsets with the activated phenotype in both the CD4 (CD29⁺) and the CD8 (CD11a⁺) compartments was noted during the course of the study. These changes did not occur in the photopheresis group. No relation between lymphocyte subsets and clinical outcome was found 1 year after the treatment with photopheresis. In conclusion, we found no major effect of photopheresis on lymphocyte populations in a group of children with newly diagnosed type 1 diabetes. However, in the placebo group the proportions of activated CD4 and CD8 cells increased over time. Since these changes did not occur in the actively treated group, our findings suggest that photopheresis may have some suppressive effects.     

Human mesenchymal stromal cells enhance the immunomodulatory function of CD8+CD28? regulatory T cells

One important aspect of mesenchymal stromal cells (MSCs)-mediated immunomodulation is the recruitment and induction of regulatory T (Treg) cells. However, we do not yet know whether MSCs have similar effects on the other subsets of Treg cells. Herein, we studied the effects of MSCs on CD8⁺CD28⁻ Treg cells and found that the MSCs could not only increase the proportion of CD8⁺CD28⁻ T cells, but also enhance CD8⁺CD28⁻T cells'' ability of hampering naive CD4⁺ T-cell proliferation and activation, decreasing the production of IFN-γ by activated CD4⁺ T cells and inducing the apoptosis of activated CD4⁺ T cells. Mechanistically, the MSCs affected the functions of the CD8⁺CD28⁻ T cells partially through moderate upregulating the expression of IL-10 and FasL. The MSCs had no distinct effect on the shift from CD8⁺CD28⁺ T cells to CD8⁺CD28⁻ T cells, but did increase the proportion of CD8⁺CD28⁻ T cells by reducing their rate of apoptosis. In summary, this study shows that MSCs can enhance the regulatory function of CD8⁺CD28⁻ Treg cells, shedding new light on MSCs-mediated immune regulation.     

Immunophenotyping of Peripheral Blood Lymphocytes in Saudi Men

Flow cytometry is an important tool for the diagnosis and follow-up of immunodeficiency patients, as well as for pateints with leukemia and lymphoma. Lymphocytes and their subsets show variations with race. The aim of this study was to establish reference ranges for lymphocytes and their subsets in an Saudi adult population by using flow cytometry. Blood samples obtained from 209 healthy Saudi men were used for this study. All blood donors were between 18 and 44 years old. Lymphocytes and their subsets were analyzed by flow cytometry, and the absolute and percentage values were calculated. We investigated the expression of T-cell markers (CD3, CD4, and CD8), B cells (CD19), and natural killer cells (CD16 and CD56). The absolute and percent values of each cell subset were compared with published data from different populations by using the Student t test. Reference ranges, each expressed as the mean ± the standard deviation, were as follows: leukocytes (6,335 ± 1759), total lymphocytes (2,224 ± 717), CD3 cells (1,618 ± 547), CD4 cells (869 ± 310), CD8 cells (615 ± 278), CD19 cells (230 ± 130), and CD3-CD16⁺/CD56+ cells (262 ± 178). The CD4/CD8 ratio was 1.6 ± 0.7. Our results for B cells, CD4 cells, and CD8 cells and for the CD4/CD8 ratio fell in between the reported results for Ethiopian and Dutch subjects. Our results were also different from previously reported findings in an Saudi adult population that showed no increase in CD8 T cells. We thus establish here the reference ranges for lymphocytes and their subsets in a large cohort of Saudi men. The CD8 cell count was not abnormally high, as previously reported, and fell in between previous results obtained for African and European populations.     

Interleukin-4-promoted T helper 2 responses enhance Nippostrongylus brasiliensis-induced pulmonary pathology

The role of CD4⁺ T-cell interleukin-4 (IL-4) receptor alpha (IL-4Rα) expression in T helper 2 (TH2) immune responses has not been defined. To examine this role, we infected CD4⁺ T-cell IL-4Rα knockout (KO) mice with the parasitic nematode Nippostrongylus brasiliensis, which induces strong host TH2 responses. Although N. brasiliensis expulsion was not affected in CD4⁺ T-cell IL-4Rα KO mice, the associated lung pathology was reduced. Infected CD4⁺ T-cell IL-4Rα KO mice showed abrogation of airway mucus production. Furthermore, CD4⁺ T-cell IL-4Rα KO mouse lungs contained reduced numbers of lymphocytes and eosinophils. Restimulation of pulmonary region-associated T-cell populations showed that TH2 cytokine responses were disrupted. Secretion of IL-4, but not secretion of IL-13 or IL-5, from mediastinal lymph node CD4⁺ T cells was reduced in infected CD4⁺ T-cell IL-4Rα KO mice. Restimulation of tissue-derived CD4⁺ T cells resulted in equivalent levels of IL-4 and IL-13 on day 7 postinfection (p.i.) in control and CD4⁺ T-cell IL-4Rα KO mice. By day 10 p.i. the TH2 cytokine levels had significantly declined in CD4⁺ T-cell IL-4Rα KO mice. Restimulation with N. brasiliensis antigen of total lung cell populations and populations with CD4⁺ T cells depleted showed that CD4⁺ T cells were a key TH2 cytokine source. These data demonstrated that CD4⁺ T-cell IL-4 responsiveness facilitates eosinophil and lymphocyte recruitment, lymphocyte localization, and TH2 cytokine production in the allergic pathology associated with N. brasiliensis infections.     

Coinfection with Heligmosomoides polygyrus fails to establish CD8+ T-cell immunity against Toxoplasma gondii

CD8⁺ T-cell immunity is important for long-term protection against Toxoplasma gondii infection. However, a Th1 cytokine environment, especially the presence of gamma interferon (IFN-γ), is essential for the development of primary CD8⁺ T-cell immunity against this obligate intracellular pathogen. Earlier studies from our laboratory have demonstrated that mice lacking optimal IFN-γ levels fail to develop robust CD8⁺ T-cell immunity against T. gondii. In the present study, induction of primary CD8⁺ T-cell immune response against T. gondii infection was evaluated in mice infected earlier with Heligmosomoides polygyrus, a gastrointestinal worm known to evoke a polarized Th2 response in the host. In the early stage of T. gondii infection, both CD4 and CD8⁺ T-cell responses against the parasite were suppressed in the dually infected mice. At the later stages, however, T. gondii-specific CD4⁺ T-cell immunity recovered, while CD8⁺ T-cell responses remained low. Unlike in mice infected with T. gondii alone, depletion of CD4⁺ T cells in the dually infected mice led to reactivation of chronic infection, leading to Toxoplasma-related encephalitis. Our observations strongly suggest that prior infection with a Th2 cytokine-polarizing pathogen can inhibit the development of CD8⁺ T-cell immune response against T. gondii, thus compromising long-term protection against a protozoan parasite. This is the first study to examine the generation of CD8⁺ T-cell immune response in a parasitic nematode and protozoan coinfection model that has important implications for infections where a CD8⁺ T-cell response is critical for host protection and reduced infection pathology.     

Decreased Levels of CD54 (ICAM-1)-Positive Lymphocytes in the Peripheral Blood in Untreated Patients with Active Juvenile Dermatomyositis

Significant abnormalities are observed in the peripheral blood of juvenile dermatomyositis (JDM) patients with active disease. In this study, we confirm that there is a significant increase in the relative percentage of B lymphocytes in the peripheral blood of a group of untreated children with newly diagnosed active JDM compared to healthy children (P < 0.0001). In order to investigate if properties intrinsic to B cells contributed to their relative increase in JDM, the percentage of B cells expressing activation markers (CD23, CD25, CD54, and CD69) was measured and compared to pediatric controls. Compared to healthy children less than 10 years of age (not significantly different from the JDM group), the JDM patients had an increase in the proportion of lymphocytes expressing CD19 (B cells; P = 0.0017) and decreases in the percentage of lymphocytes that were CD3⁻ CD16⁺ and/or CD56⁺ (NK cells; P = 0.01) and CD3⁺ CD8⁺ (T suppressor/cytotoxic cells; P = 0.02). There were no significant differences in any of the B-cell activation markers assessed. Of note, the percentage of CD54⁺ non-B lymphocytes (i.e., T cells and NK cells expressing CD54) was significantly lower in the JDM patients (25% ± 5%) than in the “age-related” healthy control group (43% ± 4%; P = 0.013). These results suggest the following for untreated children with active JDM: (i) the increase in the percentage of peripheral blood B cells is not due to intrinsic B-cell activation, and (ii) CD54/ICAM-1⁺ non-B cells, CD8⁺ T cells, and NK cells are being removed from circulation and may be participating in the pathophysiology of the disease.