CD4 Cell Subsets Research Articles

0442 Elucidating mechanisms using comparative molecular epidemiology: Immunologic alterations in workers exposed to trichloroethylene and formaldehyde

ObjectivesThere is a well-established connection between immune status and carcinogenesis, as an increased risk of cancer has been associated with a history of immunosuppressive medication use, with certain chronic infections...

Progranulin promotes tumour necrosis factor‐induced proliferation of suppressive mouse CD4+ Foxp3+ regulatory T cells

Progranulin (PGRN) is a pleiotropic growth factor with immunosuppressive properties. Recently, it was reported that PGRN was an antagonist of tumour necrosis factor (TNF) receptors, preferentially for TNFR2. However, we and others showed that TNF-TNFR2 interaction was critical for the activation and expansion of functional CD4(+) Foxp3(+) regulatory T (Treg) cells. We therefore examined the effect of PGRN on the proliferation of naturally occurring murine suppressive Treg cells induced by TNF. Consistent with our previous reports, TNF overcame the hyporesponsiveness of highly purified Treg cells to T-cell receptor stimulation. Furthermore, in the presence of interleukin-2, TNF preferentially stimulated proliferation of Treg cells contained in unfractionated CD4 cells. These effects of TNF on suppressive Treg cells were markedly increased by exogenous PGRN. TNF and TNFR2 interactions are required for this effect of PGRN, because the PGRN by itself did not stimulate Treg cell proliferation. The effect of PGRN on Treg cells was abrogated by antibody against TNFR2, and Treg cells deficient in TNFR2 also failed to respond to PGRN. Furthermore, PGRN also enhanced the proliferative responses of effector T cells to TNF, but to a lesser extent than that of Treg cells, presumably caused by the different levels of TNFR2 expression on these two subsets of CD4 cells. Hence, our data clearly show that PGRN promotes, rather than inhibits, the functional consequence of TNF-TNFR2 interaction on Treg cells.

B7-1/B7-2 blockade overrides the activation of protective CD8 T cells stimulated in the absence of Foxp3+ regulatory T cells

Although T cell activation has been classically described to require distinct, positive stimulation signals that include B7-1 (CD80) and B7-2 (CD86) costimulation, overriding suppression signals that avert immune-mediated host injury are equally important. How these opposing stimulation and suppression signals work together remains incompletely defined. Our recent studies demonstrate that CD8 Teff activation in response to cognate peptide stimulation is actively suppressed by the Foxp3(+) subset of CD4 cells, called Tregs. Here, we show that the elimination of Treg suppression does not bypass the requirement for positive B7-1/B7-2 costimulation. The expansion, IFN-γ cytokine production, cytolytic, and protective features of antigen-specific CD8 T cells stimulated with purified cognate peptide in Treg-ablated mice were each neutralized effectively by CTLA-4-Ig that blocks B7-1/B7-2. In turn, given the efficiency whereby CTLA-4-Ig overrides the effects of Treg ablation, the role of Foxp3(+) cell-intrinsic CTLA-4 in mitigating CD8 Teff activation was also investigated. With the use of mixed chimera mice that contain CTLA-4-deficient Tregs exclusively after the ablation of WT Foxp3(+) cells, a critical role for Treg CTLA-4 in suppressing the expansion, cytokine production, cytotoxicity, and protective features of peptide-stimulated CD8 T cells is revealed. Thus, the activation of protective CD8 T cells requires positive B7-1/B7-2 costimulation even when suppression by Tregs and in particular, Treg-intrinsic CTLA-4 is circumvented.

Heat Shock Proteins 60 and 70 Specific Proinflammatory and Cytotoxic Response of CD4+CD28nullCells in Chronic Kidney Disease

Background. CD4+CD28null T cells are expanded in peripheral blood of patients with chronic kidney disease and associated with subclinical atherosclerosis. However, triggers for the oligoclonal expansion and activation of these cells are not clear. Methods. We investigated twenty-five stage V-IV chronic kidney disease (CKD) patients and eight healthy subjects (HC). Peripheral mononuclear cells were isolated and incubated with heat shock protein- (HSP) 60 and 70. CD4+CD28null and CD4+CD28+ cells were sorted by flowcytometry and antigen specific response was assessed by the mRNA and protein expression of interferon (IFN)-γ, perforin, and granzyme B using qRT-PCR and Elispot. Results. The basal mRNA expression of IFN-γ, perforin, and granzyme B in CD4+CD28null cells was higher in subjects with CKD compared to that in HC (P < 0.0001). Subjects with CKD also showed expression of IFN-γ, perforin, and granzyme B in the CD4+CD28+ subset, but this was much weaker than that seen in the CD4+CD28null population (P < 0.0001). We did not note the expression of these molecules at mRNA or protein level in either subset of CD4 cells in HC. After incubation with HSP60 and HSP70, CD4+CD28null cells showed increased expression at mRNA (P < 0.001) and protein level (P < 0.001). CD4+CD28+ cells also showed a weak increase in expression. No antigen-specific response was noted in HC. Conclusion. These data show that CD4+CD28null cells in subjects with CKD react with HSP60 and HSP70 by upregulating the expression of IFN-γ, perforin and granzyme B. Increased circulating level of HSP60 and HSP70 might play a role in initiation and/or progression of atherosclerosis in CKD subjects through perturbation of CD4+CD28null cells.

High CCR5 Density on Central Memory CD4+ T Cells in Acute HIV-1 Infection Is Mostly Associated with Rapid Disease Progression

CD4+ central memory T cells play a critical role in the pathogenesis of simian immunodeficiency virus disease, and the CCR5 density on the surface of CD4 T cells is an important factor in human immunodeficiency virus (HIV)-1 disease progression. We hypothesized that quantifying central memory cells and CCR5 expression in the early stages of HIV-infection could provide useful prognostic information. We enrolled two different groups of acute HIV-infected subjects. One group progressed to CD4 T cell numbers below 250 cells/µl within 2 years (CD4 Low group), while the other group maintained CD4 cell counts above 450 cells/µl over 2 years (CD4 High group). We compared the CCR5 levels and percentage of CD4 subsets between the two groups during the 1st year of HIV infection. We found no differences between the two groups regarding the percentage of naïve, central memory and effector memory subsets of CD4 cells during the 1st year of HIV-1 infection. CCR5 levels on CD4+ CM subset was higher in the CD4 Low group compared with the CD4 High group during the 1st year of HIV-1 infection. High CCR5 levels on CD4 central memory cells in acute HIV infection are mostly associated with rapid disease progression. Our data suggest that low CCR5 expression on CD4 central memory cells protects CD4 cells from direct virus infection and favors the preservation of CD4+ T cell homeostasis.

Eccrine squamous syringometaplasia associated with vemurafenib therapy

for genetic aberrations of the immune system that may play a role in predisposing individuals to developing MF. However, data on possible etiologic factors involved in the multistep carcinogenic mechanism are equivocal. In general, it is thought that MF represents a disease potentiated by chronic antigen stimulation resulting in clonally activated T cells with malignant potential. Infectious pathogens, both viral and bacterial, and chemical exposures have been implicated as possible inciting agents in MF; yet, evidence for causation was lacking. Skin-limited disease appears to be a result of clonal skin resident effector memory cells. Although the pathogenesis of MF remains unclear, associations between chronic dermatoses and MF have been described. Although our patient was not formally patch tested, it is plausible that an initial contact allergy from the sex hormones or chemical components of the contraceptive patch could have resulted in a robust, chronic dermatitis that enabled malignant transformation of a subset of CD4 cells. However, subsequent observations are necessary to validate this potential association between chronic dermatoses and MF.

CTLA-4-IG (ABATACEPT) therapy reduces t cell function for all defined cd4 cell subsets including regulatory T cells (TREG)

Background and objectivesRheumatoid arthritis (RA) is a complex autoimmune disorder primarily affecting peripheral joints where CD4+T cells are abundant at the site of inflammation. Regulatory T cells (Treg) are a...

Regulation of IL-2 gene expression by Siva and FOXP3 in human T cells

BackgroundSevere autoinflammatory diseases are associated with mutations in the Foxp3 locus in both mice and humans. Foxp3 is required for the development, function, and maintenance of regulatory T cells (Tregs), a subset of CD4 cells that suppress T cell activation and inflammatory processes. Siva is a pro-apoptotic gene that is expressed across a range of tissues, including CD4 T cells. Siva interacts with three tumor necrosis factor receptor (TNFR) family members that are constitutively expressed on Treg cells: CD27, GITR, and OX40.ResultsHere we report a biophysical interaction between FOXP3 and Siva. We mapped the interaction domains to Siva's C-terminus and to a central region of FOXP3. We showed that Siva repressed IL-2 induction by suppressing IL-2 promoter activity during T cell activation. Siva-1's repressive effect on IL-2 gene expression appears to be mediated by inhibition of NFkappaB, whereas FOXP3 repressed both NFkappaB and NFAT activity.ConclusionsIn summary, our data suggest that both FOXP3 and Siva function as negative regulators of IL-2 gene expression in Treg cells, via suppression of NFAT by FOXP3 and of NFkappaB by both FOXP3 and Siva. Our work contributes evidence for Siva's role as a T cell signalling mediator in addition to its known pro-apoptotic function. Though further investigations are needed, evidence for the biophysical interaction between FOXP3 and Siva invites the possibility that Siva may be important for proper Treg cell function.

Abstract 5525: Profiling counter-regulatory and cytotoxic immune pathways with cellular biomarkers in thyroid cancer patients

Abstract Clinical studies suggested an association between thyroid cancer outcome and patient immunologic function. Cytotoxic T-Lymphocyte Antigen 4 (CTLA4) and Programmed Death 1 (PD-1) are strong negative regulators of immune function whose aberrant function has been associated with poor outcomes in patients with solid malignancies. We carried out this study to compare the degree and patterns of expression of CTLA4 and PD-1 in peripheral circulating mononuclear cells in adult patients with thyroid cancer (N=21) compared with adult healthy controls (N=5). We used multiparameter flow cytometry to determine the absolute cell count per ml of blood for the cells of interest. Functional assays of immunologic competence of B-cell, T-cell as well as NK cell subpopulations was assessed by granzyme-B and interferon (IFN) gamma production following PMA/ionomycin stimulation in vitro using standard assay method. Differences in specific cell count between patients and healthy control was assessed by F-test statistics and p-value &amp;lt;0.05 was considered significant. There was no significant difference between cancer patients and healthy controls in the overall numbers of T-cell, B-cell, and NK cell subsets. Similarly, the proportion of CD4, CD8 and NK cell subsets with CTLA4, PD-1, IFN-gamma or Granzyme-B expression with and without PMA/ionomycin stimulation were not significantly different when comparing patients with thyroid cancer to healthy controls. Within the thyroid cancer patient population, however, the distribution of CD4 (+) T-cells, total B-cells, plasma cells, and PD ligand 1-expressing monocytes, macrophages and dendritic cells i.e. CD64 (+) PD-L1 (+) was significantly different between patient with active disease and those in remission. The Granzyme-B (+) CD4 (+) T-cell was significantly lower in patients in remission than in patients with active disease both with and without in vitro activation with PMA/ionomycin (0.08 vs. 0.19 per ml blood × 10⁁6; p=0.03). Intriguingly, functional profiling in a limited number of thyroid cancer patients revealed a markedly impaired ability of CD8 (+) T-cells to produce IFN-gamma and Granzyme B following PMA/ionomycin stimulation. Thus suggesting possible immune exhaustion in this subpopulation of thyroid cancer patients. Overall, our results show a trend toward impaired cytotoxic immune pathway in thyroid cancer patients with active disease burden when compared to patients in remission. A similar trend was observed in comparison to healthy subjects but the small number of control subjects limits statistical significance. Further elucidation of this finding using larger patient sample is clearly warranted and currently ongoing. (Supported by Career Development Award (NCI P50 CA128613) to TK Owonikoko and the Georgia Cancer Coalition Distinguished Scholar Award to DM Shin, SS Ramalingam, FR Khuri, and TK Owonikoko) Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5525. doi:10.1158/1538-7445.AM2011-5525

Terminally Differentiated CD8 Cells in HIV-Infected Children: HIV-GAG/POL Specificity and IFN-γ Production

CD8 cells are key to antiviral immunity and can be divided by phenotype into early (CD28+ CD27+), intermediate (CD28-CD27+) and terminally differentiated subsets (CD28- CD27-). Despite effective HAART there is an unexplained expansion of CD8+CD28-CD27-T cells in HIV-infected children. The cytokine production and specificity of this terminally differentiated CD8 T cell subset in chronic virus infection is unclear. PATIENTS, METHODS & RESULTS: In a cohort of 26 HIV-infected children the cytokine production of terminally differentiated CD8 cells was analyzed by intracellular staining and FACS analysis and was compared to children with chronic hepatitis B infection and to healthy children. The specificity of CD8 subsets was analyzed by staining with Gag/Pol tetramers in a cohort of 13 patients. We show that an increased production of interferon-γ in terminally and early/intermediate differentiated CD8 cell subsets after stimulation is specific for HIV-infection. The expanded population of terminally differentiated CD8+CD28-CD27- T cells does include HIV Gag/Pol specific T cells in adults but not in children. The expansion of terminally differentiated CD8 cells might be important for immunomodulation but in children it does not appear to play a role in HIV Gag and Pol specific immunity.

Regulatory T cells and chronic immune activation in human immunodeficiency virus 1 (HIV-1)-infected children

The function of CD4(+) T cells with regulatory activity (T(regs)) is the down-regulation of immune responses. This suppressive activity may limit the magnitude of effector responses, resulting in failure to control human immunodeficiency virus 1 (HIV-1) infection, but may also suppress chronic immune activation, a characteristic feature of HIV-1 disease. We evaluated the correlation between viral load, immune activation and T(regs) in HIV-1-infected children. Eighty-nine HIV-1-infected children (aged 6-14 years) were included in the study and analysed for HIV-1 plasmaviraemia, HIV-1 DNA load, CD4 and CD8 cell subsets. T(reg) cells [CD4(+)CD25(high)CD127(low) forkhead box P3 (FoxP3(high))] and CD8-activated T cells (CD8(+)CD38(+)) were determined by flow cytometry. Results showed that the number of activated CD8(+)CD38(+)T cells increased in relation to HIV-1 RNA plasmaviraemia (r = 0·403, P < 0·0001). The proportion of T(regs) also correlated positively with HIV-1 plasmaviraemia (r = 0·323, P = 0·002), but correlated inversely with CD4(+) cells (r = -0·312, P = 0·004), thus suggesting a selective expansion along with increased viraemia and CD4(+) depletion. Interestingly, a positive correlation was found between the levels of T(regs) and CD8(+)CD38(+)T cells (r = 0·305, P = 0·005), and the percentage of T(regs) tended to correlate with HIV-1 DNA load (r = 0·224, P = 0·062). Overall, these findings suggest that immune activation contributes to the expansion of T(reg) cells. In turn, the suppressive activity of T(regs) may impair effector responses against HIV-1, but appears to be ineffective in limiting immune activation.

Central inflammation versus peripheral regulation in multiple sclerosis

Th17 cells are a highly pro-inflammatory T-helper cell subset characterised by the expression of IL17. They have been implicated in a variety of allergic and autoimmune conditions. T-regulatory (Treg) cells, a subset of CD4 cells which express Foxp3, CD25, IL10 and TGFβ, can suppress the activity of Th17 cells. In this study, we show that the circulating levels of Th17 and Treg cells in peripheral blood are correlated; furthermore, the expression of the pro-inflammatory cytokine IL17 and the anti-inflammatory cytokine IL10 by CD4 cells are also correlated. However, we found no clear correlation between cerebrospinal fluid (CSF) IL10 and IL17 cytokine levels in MS, approaching a negative correlation at the time of relapse, and an overall negative correlation between CSF IL17 and TGFβ levels, suggesting a lack of central regulation of pro:anti-inflammatory balance in this demyelinating condition.

Increased Prevalence of IL-17-Producing Peripheral Blood Lymphocytes in Pre-eclampsia

Systemic inflammation is a dominant component in the pathogenesis of pre-eclampsia. Besides the imbalance of Th1 and Th2 cells, alterations of the prevalence of Th17 and regulatory T cells have also been suggested to contribute to inflammation. We aimed to describe the prevalence of these four CD4 lymphocyte subtypes in pre-eclampsia and normal pregnancy, along with that of IL-17-producing CD8 and NK cells. Twenty pre-eclamptic and 22 normal pregnant women were enrolled in this study. Using flow cytometry, we determined the prevalence of IL-17-producing cells among the CD4, CD8 and NK cell subsets. Furthermore, we measured the prevalence of CD4+ Tregs, and Th1/Th2 cells were characterized using cell surface chemokine receptor markers. We demonstrated that there is a shift not only in the Th1/Th2 but also in the Th17/Treg balance favouring skewness towards a pro-inflammatory status in pre-eclampsia. The proportion of CD8 and NK cells that express IL-17 was also higher in pre-eclampsia. The prevalence of IL-17-producing CD4, CD8 and NK cells is elevated in pre-eclampsia, indicating that both the innate and adaptive arms of the immune system are involved in the development of the exaggerated maternal systemic inflammation observed in this pregnancy-specific disorder.

A naturally antagonist of human IL-23 inhibits the development and function of human Th17 cells (51.4)

Abstract The Th17 subset of CD4 cells is critical to inflammation. Secretion of IL-17 by Th17 cells drives inflammation in human diseases and their mouse models, including IBD, psoriasis and arthritis. The differentiation of mature Th17 cells depends upon stimulation in the presence of IL-6, TGF-β and IL-21 and the induction of RORγt, but their phenotype is stabilised by IL-23, which is essential to Th17 stability and function. Initial induction of Th17 cells can be antagonised by IL-4 or IFN-γ, but mechanisms through which terminal differentiation can be inhibited have not been identified. We found that HuIL23Rα chain mRNA transcripts exist that lack exon-9 (“Δ9”); these are translated to a truncated receptor form containing the entire external domain of the wild-type protein. Here, we show that this soluble variant of the HuIL23Rα chain antagonises IL-23 signalling and Th17 maturation. The variant is secreted and present at low levels in the blood. This soluble form represents 10% of HuIL23Rα chain mRNA. It binds IL-23 in solution and inhibits the phosphorylation of STAT3 caused by IL-23. In experiments designed to differentiate human Th17 cells in vitro, the Δ9 form inhibits the production of Th17-associated cytokines, including IL-17A and IL-17F. Δ9 does not bind IL-12; thus it is a specific inhibitor of IL-23 and a modulator of Th17 cells. Our results indicate that this soluble form of HuIL23Rα likely functions to regulate Th17 activity.

Restoring the Functional Immunogenicity of Chronic Lymphocytic Leukemia Using Epigenetic Modifiers.

Abstract 5116 BackgroundChronic lymphocytic leukemia (CLL) is a malignancy arising from immune cells (B-lymphocytes) endowed with intrinsic antigen-presenting capabilities. Such a function however is lost during malignant transformation and CLL cells are well known for their inability to process and present antigens to the T-cell arm of the immune system. Instead, malignant CLL cells elicit a vast array of immune regulatory mechanisms conducive to T-cell dysfunction and immunosupression. Recent evidence suggests that DNA methylation inhibitors (DNMTi) and histone deacetylase inhibitors (HDACi) can have lasting cancer-specific effects on the expression of highly immunogenic cancer-testis antigens (CTAs) as well as modulating costimulatory and major histocompatability molecule (MHC) surface expression. MethodsTo investigate the potential efficacy of a combined therapy of DNMTi (5-aza-2'-deoxycytidine) and HDACi (LAQ) in human CLL we performed a gene expression microarray profile and further confirmed the expression of CTAs. Furthermore we investigated phenotypic alterations due to changes in costimulatory molecules (CD40, 80, and 86), and MHC molecules (HLA-A, B, C, and HLA-DR) in both cell lines and primary cells from patients with CLL. To confirm that alterations in molecule surface expression correlated to more potent immune stimulation we characterized the formation of the immune-synapse between the CLL antigen presenting cell and healthy T-lymphocytes in treated and untreated samples using confocal microscopy and flow conjugation assay. To demonstrate functional significance we subjected treated and untreated CLL cell lines to mixed lymphocyte proliferation assays with CD4, CD8, and NK cell subsets. ResultsAs expected we demonstrate by gene expression profile and RT-PCR that many highly immunogenic CTAs (SSX family, MAGE family, NY-ESO-1, GAGE-2 and NXF2) are significantly upregulated after treatment with an HDACi and DNMTi in combination. Interestingly, preliminary data showed interferon gamma mRNA upregulation. Consistent with this information, our data also indicates that a more proinflammatory signaling pathway is developed between the CLL B-cell and the T-lymphocyte after combination treatment demonstrated by upregulated surface expression of CD86, 80, 40, HLA-A,B,C, and HLA-DR as well as the formation of significantly more, robust, numerous, and organized actin-mediated immune-synapses, changes which were not duplicated using either drug alone. Functionally, only the combined HDACi/DNMTi treatment of CLL cells led to increased allogenic T and NK cell proliferation and cytotoxicity. ConclusionsTaken together these data indicate that combined HDACi/DNMTi may benefit current antigen specific vaccine strategies for CLL by inducing the expression of a highly antigenic CTAs, increasing CLL costimulatory capacity, repairing immunological synapse, and functionally increasing the proinflammatory status of the CLL APC. DisclosuresPinilla:Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Bristol-Myers Squibb: Research Funding; exelixis: Research Funding.

Association between Tuberculin Skin Test Reactivity, the Memory CD4 Cell Subset, and Circulating FoxP3-Expressing Cells in HIV-Infected Persons

Lack of reactivity to the tuberculin skin test (TST) is widely observed in individuals with advanced human immunodeficiency virus type 1 (HIV-1) infection. Biopsy specimens from the TST reaction site and from skin not infiltrated with purified protein derivative were obtained from 15 HIV-1-infected and 23 uninfected persons who did not have active tuberculosis and who were from a community in which the incidence of tuberculosis was very high. Histologic sections (size, 8 mum) were immunohistochemically stained for CD4, CD8, CD28, CD45RA, CD45RO, CD62L, CD1a, human leukocyte antigen (HLA)-DR, granulysin, interferon-gamma, and FoxP3 and were analyzed by single-cell in situ digital imaging. Peripheral blood mononuclear cells were analyzed using a fluorescence-activated cell sorter. Biopsy specimens obtained from TST-reactive skin of HIV-1-infected persons demonstrated fewer CD4(+) T cells at the TST site (P = .36) but more HLA-DR(+) T cells (P = .037) than did such biopsy specimens obtained from HIV-1-uninfected persons. Among HIV-1-infected persons, the total number of cells (P = .008) and numbers of CD45RO(+) memory T cells (P = .003) were significantly higher in TST-reactive persons than in TST-unreactive persons. For HIV-1-infected persons, TST induration was inversely correlated with the numbers of FoxP3(+) T cells in the blood (P = .026) but was unrelated to the number of circulating CD4(+) T cells. For HIV-1 infected persons, the TST depends on memory T cells and is more strongly associated with the numbers of circulating FoxP3(+)CD4(+) T cells than with the total number of CD4(+) T cells.

Cathepsin W expressed exclusively in CD8 + T cells and NK cells, is secreted during target cell killing but is not essential for cytotoxicity in human CTLs

Cathepsin W (CatW, lymphopain) is a putative cysteine protease with restricted expression to natural killer (NK) cells and CD8(+) T cells and so far unknown function and properties. Here, we characterize in detail, the regulation of human CatW during T-cell development in response to different stimuli and its functional involvement in cytotoxic lymphocyte effector function. Western blots and real time polymerase chain reaction of sorted, unstimulated, and stimulated cell subsets (thymocytes, T cells, NK cells) and their culture supernatants were used to study regulation and expression of CatW. Primary CD8(+) T cells and short-term T-cell lines were transfected with small interfering RNA to study the involvement of CatW in effector function such as target cell killing and interferon-gamma production. Levels of CatW expression correlate closely with cytotoxic capacity both during development and in response to factors influencing cytotoxicity. Furthermore, CatW is secreted during specific target cell killing. However, knockdown of CatW expression by small interfering RNA neither influences target cell killing nor interferon-gamma production. Despite being expressed in the effector subset of CD8(+) and NK cells and of being released during target cell killing, our functional inhibition studies exclude an essential role of CatW in the process of cytotoxicity.

Spontaneous proliferation of memory (CD45RO+) and naive (CD45RO-) subsets of CD4 cells and CD8 cells in human T lymphotropic virus (HTLV) infection: distinctive patterns for HTLV-I versus HTLV-II.

Spontaneous lymphocyte proliferation (SLP) in vitro is a characteristic feature of about 50% of individuals infected with HTLV-I or HTLV-II. Both CD4 cells and CD8 cells contribute to SLP in HTLV-I infection, whereas SLP in HTLV-II infection is usually restricted to CD8 cells. In this study, we asked if SLP was restricted to the memory (CD45RO+) cell subset of CD4 and CD8 cells in HTLV infection. Purified CD4 and CD8 cells were separated into CD45RO+ and CD45RO- populations by a modified panning technique, and spontaneous proliferation (SP) of the cell subsets was assessed. For all five HTLV-I-infected persons whose mononuclear cell cultures were SLP+, only CD45RO+ cells, but not CD45RO- cells, within CD4 and CD8 subsets showed SP. In contrast, five of six SLP+ HTLV-II+ individuals showed SP in both the CD45RO+ and the CD45RO- subsets of CD4 cells, and 10 of 12 SLP+ HTLV-II+ individuals showed SP of both the CD45RO+ and CD45RO- subsets of CD8 cells. Polymerase chain reaction studies showed that proviral genome was generally present in both CD45RO+ and CD45RO- subsets of CD4 and CD8 cells, regardless of HTLV type and SP activity. These findings show that SP of both CD4 and CD8 cells in HTLV-I infection is usually restricted to CD45RO+ memory cells, whereas in HTLV-II infection, both CD45RO+ memory and CD45RO- naive subsets of CD4 and CD8 cells may exhibit SP. It thus appears that HTLV-I infection and HTLV-II infection exhibit distinctive dysregulatory effects on memory and naive T cell subpopulations.

High serum levels of soluble CD8 in insulin-dependent diabetes.

In type 1 (insulin-dependent) diabetes mellitus (IDDM) CD8+ T cells represent the majority of lymphocytes which infiltrate the pancreatic islets during beta cell destruction. Soluble CD8 antigen (sCD8) has been shown to correlate with CD8 cell subset activation. In this study we measured by ELISA sCD8 levels in sera from: 33 newly diagnosed IDDM patients; 29 type 1 diabetics with duration of disease more than 1 year; 37 healthy siblings of IDDM patients; 19 healthy controls. Sera from both groups of IDDM patients and from healthy siblings exhibited soluble CD8 mean levels significantly higher than controls (P = 0.0001, P < 0.003, P < 0.03 respectively). Soluble CD8 levels above the normal range (mean +/- 2 s.d. of controls) were found in a percentage of newly diagnosed subjects (54.5%) significantly higher than in subjects with a long-standing duration of disease (6.9%, P < 0.0005) and healthy siblings (16.2%, P < 0.002). Our results suggest that the raised levels of soluble CD8 near to diabetes onset may indicate the activation of CD8+ T cells probably responsible for the autoimmune beta cell destruction.

Cytometric profiling in multiple sclerosis uncovers patient population structure and a reduction of CD8low cells

As part of a biomarker discovery effort in peripheral blood, we acquired an immunological profile of cell-surface markers from healthy control and untreated subjects with relapsing-remitting MS (RRMS). Fresh blood from each subject was screened ex vivo using a panel of 50 fluorescently labelled monoclonal antibodies distributed amongst 56 pools of four antibodies each. From these 56 pools, we derived an immunological profile consisting of 1018 'features' for each subject in our analysis using a systematic gating strategy. These profiles were interrogated in an analysis with a screening phase (23 patients) and an extension phase (15 patients) to identify cell populations in peripheral blood whose frequency is altered in untreated RRMS subjects. A population of CD8(low)CD4(-) cells was identified as being reduced in frequency in untreated RRMS subjects (P = 0.0002), and this observation was confirmed in an independent sample of subjects from the Comprehensive Longitudinal Investigation of MS at the Brigham & Women's Hospital (P = 0.002). This reduction in the frequency of CD8(low)CD4(-) cells is also observed in 38 untreated subjects with a clinically isolated demyelination syndrome (CIS) (P = 0.0006). We also show that these differences may be due to a reduction in the CD8(low)CD56(+)CD3(-)CD4(-) subset of CD8(low) cells, which have a natural killer cell profile. Similarities between untreated CIS and RRMS subjects extend to broader immunological profiles: consensus clustering of our data suggests that there are three distinct populations of untreated RRMS subjects and that these distinct phenotypic categories are already present in our sample of untreated CIS subjects. Thus, our large-scale immunophenotyping approach has yielded robust evidence for a reduction of CD8(low)CD4(-) cells in both CIS and RRMS in the absence of treatment as well as suggestive evidence for the existence of immunologically distinct subsets of subjects with a demyelinating disease.