The T-cell receptor repertoires expressed by CD4+ and CD4- large granular lymphocytes derived from the same patients suggest the persistent action of an immune-mediated selection process

The lymphoproliferative syndrome with large granular lymphocytes (LGL) is an heterogeneous disorder of unknown etiology. The analysis of T-cell receptor (TCR) genes rearrangements has shown that, in most cases, the disease is associated with clonal proliferation of CD8+CD57+ LGL. However, the putative neoplastic nature of these expansions remains questionable because clonal proliferations of CD8+ cells have recently been found also in physiologic conditions. To obtain more precise information on the mechanisms responsible for LGL expansions, we decided to compare the molecular characteristics of TCRBV chains expressed by LGL with different phenotype and function, but derived from the same patients. To this end, we characterized, at the molecular level, the TCR repertoires of fractionated T-cell populations of two unusual patients with concurrent expansions of CD4+CD57+ and CD4-CD57+ LGL. Our results show that the dominant TCRBV chains expressed by the different CD4+ and CD4- LGL populations were strictly oligoclonal. However, the molecular characteristics of the dominant V-D-J rearrangements also imply that the selection of these clones was not due to a neoplastic event. Rather, our data suggest that these particular LGL proliferations can be ascribed to a chronic T-cell-mediated immune response that involves recognition by the engaged TCR of antigens that are not necessarily presented to immune system in the classical major histocompatibility complex-restricted pathway.     

IL-4 protects adult C57BL/6 mice from prolonged Cryptosporidium parvum infection: analysis of CD4+alpha beta+IFN-gamma+ and CD4+alpha beta+IL-4+ lymphocytes in gut-associated lymphoid tissue during resolution of infection

Resistance of adult C57BL/6 mice to severe Cryptosporidium parvum infection is dependent on CD4+alpha beta+ TCR lymphocytes. In this study, we demonstrated that treatment with anti-IFN-gamma mAb extended oocyst excretion 18 days longer, and anti-IL-4 mAb extended oocyst excretion at least 11 days longer than isotype control mAb treatment. Analysis of the specific activity of anti-IFN-gamma mAb present in treated mouse sera suggested that IFN-gamma may have a limited role in the resolution phase of infection. Changes were also documented in numbers of CD4+alpha beta+IFN-gamma+ and CD4+alpha beta+IL-4+ lymphocytes in Peyer's patches and intraepithelium of adult C57BL/6 mice during resolution of C. parvum infection. Resistance to initial severe infection was associated with CD4+alpha beta+IFN-gamma+ lymphocytes, and eventual resolution of infection was associated with CD4+alpha beta+IL-4+ lymphocytes. Analysis of cytokine expression following in vitro stimulation with C. parvum Ags during resolution of infection demonstrated consistent increases in CD4+alpha beta+IL-4+ lymphocytes, but not CD4+alpha beta+IFN-gamma+ lymphocytes. The relevance of CD4+alpha beta+IL-4+ lymphocytes in protection against C. parvum was then evaluated in C57BL/6 IL-4 gene knockout mice (IL-4(-/-)). Adult IL-4(-/-) mice excreted oocysts in feces approximately 23 days longer than IL-4(+/+) mice. Further, anti-IFN-gamma mAb treatment increased the severity and the duration of infection in IL-4(-/-) mice compared with those in IL-4(+/+) mice. Together, the data demonstrated that IFN-gamma was important in the control of severity of infection, and either IFN-gamma or IL-4 accelerated termination of infection. However, neither IL-4 nor IFN-gamma was required for the final clearance of infection from the intestinal tract of adult mice.     

Characterization of circulating CD4+ CD8+ lymphocytes in healthy individuals prompted by identification of a blood donor with a markedly elevated level of CD4+ CD8+ lymphocytes

During flow cytometric analysis of lymphocytes from healthy donors, we identified a donor (donor A) with 22% CD4+ CD8+ cells (versus values of < 4% for 65 other controls). To determine if CD4+ CD8+ cells from donor A and other controls were similar, we first defined the phenotypic profile of control CD4+ CD8+ cells. Enriched CD4+ CD8+ cell populations for 10 controls were prepared by a two-step positive selection scheme with anti-CD4-coated magnetic beads and anti-CD8-coated culture flasks; the selected population averaged 69% CD4+ CD8+ cells and 31% CD4+ CD8- cells. For all 10 controls, two subsets of CD4+ CD8+ cells, CD4dim CD8bright and CD4bright CD8dim, were observed. Phenotypic profiles of these two CD4+ CD8+ subsets were defined by pairing anti-CD8 with other monoclonal antibodies, and the profiles were compared with each other and with those of CD4+ CD8-, CD4- CD8bright, and CD4- CD8dim cells. CD8bright and CD4bright CD8dim cells differed in their proportions of CD62-L+ cells and in their levels of CD11a and CD2 expression. Both CD4+ CD8+ subsets resembled CD4+ CD8- cells in CD45RA, CD45RO, and CD25 expression; the comparable CD- CD8+ cells in CD62-L expression; and CD4- CD8bright cells in CD11b, CD11b, CD16/56, and CD28 expression. CD38 expression in both CD4+ CD8+ subsets was decreased compared with those of other cell subsets. Whereas control CD4+ CD8+ cells averaged 33% CD4dim CD8bright, CD4+ CD8+ cells from donor A were > 90% CD4dim CD8bright. Donor A CD4dim CD8bright cells exhibited proportional decreases in CD25 and CD62-L expression and increases in CD11b and CD54 expression compared with those of control CD4dim CD8bright cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immunity to Cryptosporidium muris infection in mice is expressed through gut CD4+ intraepithelial lymphocytes

The role of gut intraepithelial lymphocytes (IEL) in immunity to cryptosporidial infection was investigated with a murine infection model involving Cryptosporidium muris. Oocyst shedding was monitored in severe combined immunodeficiency (SCID) mice infected with C. muris following intravenous injection of mesenteric lymph node (MLN) cells or intestinal IEL from BALB/c donor mice which were naive or previously infected with C. muris. SCID mice receiving no lymphoid cells developed chronic infections and excreted large numbers of oocysts until the end of the experiment. SCID mice injected with IEL from immune animals, however, were able to overcome the infection, and furthermore, these animals produced fewer oocysts and recovered sooner than ones which received IEL or MLN cells from naive BALB/c donors. Similar levels of protection were obtained in SCID mice injected with either 2 X 10(6) IEL or MLN cells from immune donor mice. Depletion of CD4+ cells from immune IEL, however, abrogated the ability to transfer immunity to SCID mice, while depletion of CD8+ cells only marginally reduced the protective capacity of immune IEL. Finally, control SCID mice which received no lymphocytes had < or = 1% CD4+ cells in the IEL from the small intestine, whereas the IEL from SCID mice recovered from infection, as a result of injection with immune IEL, contained 15% CD4+ cells. Thus, the ability to control C. muris infection correlated with the presence of the protective CD4+ cells in the gut epithelium.     

Cross-linking of the CD40 ligand on human CD4+ T lymphocytes generates a costimulatory signal that up-regulates IL-4 synthesis

Although there is good evidence that the induction of IL-4 synthesis in CD4+ T lymphocytes is favored by Ag presentation by B cells and not macrophages, the precise molecular signals provided by B cells to T cells that enhance IL-4 synthesis are not clear. To examine this issue, we established an APC-independent system to activate highly purified T cells and induce cytokine synthesis, using immobilized mAbs against several T cell surface molecules, including CD3, CD28, and the CD40 ligand (CD40L). The counter-receptors for all three of these molecules are expressed on B cells, and include CD40, which is expressed primarily on B cells, but also on dendritic cells and thymic epithelium. We found that IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28, whereas ligation of CD3/TCR and CD28 in the absence of CD40L triggering resulted in little or no IL-4 synthesis. CD40L costimulation greatly enhanced IL-4 synthesis both in T cells from normal nonallergic adult subjects as well as in naive T cells from cord blood. Furthermore, we demonstrated that IL-4 synthesis was optimally enhanced when the strength of the CD3/TCR signal was limiting, while IL-4 synthesis was inhibited when CD3/TCR stimulation was maximal. These studies confirm that IL-4 synthesis can be induced in normal T lymphocytes in the absence of exogenous IL-4, and demonstrate that CD40L costimulation is of fundamental importance in regulation of IL-4 production. In addition, these findings provide a mechanism by which B cells preferentially enhance IL-4 synthesis in T cells at low Ag concentrations.     

Functional characterization of porcine CD4+CD8+ extrathymic T lymphocytes

The porcine immune system is unique in that the expression of CD4 and CD8 antigens defines four subpopulations of resting, extrathymic (CD1-) T lymphocytes. In addition to CD4-CD8+ and CD4+CD8- T lymphocytes, CD4-CD8- and CD4+CD8+ lymphocyte subpopulations are prominent in blood as well as in lymphoid tissues. In the present study, a functional comparison was made between CD4+CD8- and CD4+CD8+ T lymphocyte subpopulations. In a primary in vitro immune response against alloantigenic stimulator cells, both subpopulations proliferated without significant differences in their reactivity. Different results were obtained when analyzing the antigen-specific functions of the two CD4+ subpopulations in a secondary response against recall viral antigen; these experiments were performed with T lymphocytes from pseudorabies virus-immunized pigs. The proliferative response against viral antigens could be assigned to the CD4+CD8+ subpopulation, whereas the CD4+CD8- subpopulation remained nonreactive. Further analyses of the virus-specific in vitro immune response revealed a major histocompatibility complex (MHC) class II restricted helper T lymphocyte reaction involving CD4 but not CD8 molecules as restriction elements. Taken together, these results demonstrate that only the extrathymic CD4+CD8+ T lymphocyte subpopulation of swine contains MHC class II-restricted antigen-specific memory T helper cells.     

Reciprocal regulation of CD30 expression on CD4+ T cells by IL-4 and IFN-gamma

Prior studies have implicated CD30 as a marker for Th2 cells, but the mechanism that underlies this correlation was unknown. We show here that CD30 was expressed on activated CD4+ T cells in the presence of IL-4. In the absence of endogenously produced IL-4, however, even Th2 lineage cells lost CD30 expression. Thus, CD30 is not an intrinsic marker of Th2 cells, but is inducible by IL-4. CD30 was also found to be down-regulated by IFN-gamma. Committed Th1 effector cells do not express CD30, although differentiating Th1 lineage cells temporarily express CD30. The transient expression of CD30 on differentiating Th1 lineage cells was mainly the result of endogenously produced IL-4 induced by IL-12. Culture of IL-12-primed cells under conditions that reverse the phenotype (Ag plus IL-4) resulted in two cell populations based upon their ability to express CD30. One population responded to IL-4 upon restimulation and became a CD30-positive, Th0-like cell population, while the other remained CD30 negative and synthesized only IFN-gamma. Thus, CD30 expressed on CD4+ T cells reflected the ability of CD4+ T cells to respond to IL-4.     

IL-10 impacts autoimmune diabetes via a CD8+ T cell pathway circumventing the requirement for CD4+ T and B lymphocytes

IL-10 is essential for an early phase of diabetes in nonobese diabetic (NOD) mice, but later becomes protective against its development. The mechanism by which IL-10 mediates the pathway to diabetes in these mice is unknown. Herein, we dissected the cellular and costimulation requirements for diabetes in transgenic (tg) NOD mice that expressed IL-10 in their pancreatic islets (IL-10-NOD mice). We found that IL-10 alone did not cause diabetes because the offspring (IL-10-NOD-scid mice) from back-crosses of IL-10-NOD mice with NOD-scid mice had no diabetes. Moreover, these IL-10-NOD-scid mice were free of lymphocytic infiltration. Treatment of IL-10-NOD mice with depleting anti-CD4 mAb or control mAb had no effect on diabetes. Surprisingly, depletion of CD8+ T cells by treatment with the corresponding mAb inhibited diabetes without attenuating insulitis, demonstrating a critical role for CD8+ T cells in the disease process. Interestingly, B cell-deficient IL-10-NOD mice readily developed diabetes with kinetics and incidence similar to those observed in wild-type mice, demonstrating that B lymphocytes as APCs were not required in the disease process. Administration of anti-CD40 ligand (CD40L) mAb did not prevent disease, indicating that CD40/CD40L costimulation is not required for diabetes in IL-10-NOD mice. Immunization of IL-10-NOD mice with CFA or heat-shock protein 65, known to block diabetes in NOD mice, had no effect on their diabetes. We demonstrate that IL-10 contributes early to the pathology of diabetes via a CD8+ T cell pathway, eliminating the requirement for B lymphocytes and CD40-CD40L costimulation. Our findings provide a mechanism for the participation of IL-10 in the early development of diabetes.     

Processing and release of IL-16 from CD4+ but not CD8+ T cells is activation dependent

IL-16 is synthesized as a precursor molecule of 68 kDa (pro-IL-16) that is processed by caspase-3, a member of the IL-1 converting enzyme (ICE) family. This cleavage results in a 13-kDa carboxy terminal peptide, which constitutes the bioactive secreted form of IL-16. We have previously reported constitutive IL-16 mRNA expression and pro-IL-16 protein in CD4+ and CD8+ T cells. Although bioactive IL-16 protein is present in unstimulated CD8+ T cells, there is no bioactive IL-16 present in CD4+ T cells. Along these lines, unstimulated CD8+ T cells contain active caspase-3. In the current studies we investigated the regulation of IL-16 protein and mRNA expression in CD4+ T cells and determined the kinetics of secretion following stimulation of the TCR. CD4+ T cells release IL-16 protein following antigenic stimulation, and this release is accelerated in time by costimulation via CD28. However, CD3/CD28 costimulation did not alter IL-16 mRNA appearance or stability in either CD4+ or CD8+ T cells. The secretion of bioactive IL-16 from CD4+ T cells correlated with the appearance of cleavage of pro-caspase-3 into its 20-kDa active form. Thus, resting CD8+ T cells contain active caspase-3 that is capable of cleaving pro-IL-16, whereas CD4+ T cells require activation for the appearance of active caspase-3. The mechanism of release or secretion of bioactive IL-16 is currently unknown, but does not correlate with cellular apoptosis.     

Donor cell induced CD69 expression and intracellular IL-2 and IL-4 production by peripheral blood lymphocytes isolated from kidney transplant recipients

Flow cytometry assays, which measure CD69 activation and intracellular cytokine production, have been used to measure peripheral blood lymphocyte (PBL) responses to in vitro antigen exposure. In the present study, we show that, in healthy individuals and immunosuppressed kidney transplant recipients, CD69 expression and intracellular cytokine production by peripheral blood T cells compare favorably to thymidine uptake as a measure of PBL response to alloantigen in mixed leukocyte culture (MLC). Heparinized whole blood from 23 healthy individuals was incubated for 24-48 h with 3rd party allogeneic monocytes; blood from twelve kidney transplant recipients was incubated with monocytes from their kidney donor and with monocytes from unrelated individuals. The percentage of T cells expressing surface CD69 or intracellular IL-2 or IL-4 was determined by 3-color flow cytometry. We identified 5 donor-specific response patterns in our kidney transplant group. One transplant recipient was hyporesponsive; his cells did not express CD69 or produce IL-2 in response to either donor or 3rd party allogeneic cells. All other transplant recipients expressed CD69 and IL-2 in response to 3rd party allogeneic cells. Two had no response to donor cells (donor-specific hyporesponsiveness), three had donor-specific anergy (CD69 expression without cytokine production in response to donor cells), five had a donor-specific Thl response (CD69 expression and IL-2 production in response to donor cells), and one had a donor-specific Th2 response (CD69 expression and IL-4 but not IL-2 production in response to donor cells). Rapid measures of donor-specific hyporesponsiveness such as CD69 activation antigen expression and intracellular cytokine production may prove valuable in monitoring lymphocyte function and aid in the long-term management of kidney transplant recipients.     

Proliferative response of human CD4+ T lymphocytes stimulated by the lectin jacalin

The Gal beta(1-3)GalNAc-binding lectin jacalin is known to specifically induce the proliferation of human CD4+ T lymphocytes in the presence of autologous monocytes and to interact with the CD4 molecule and block HIV-1 infection of CD4+ cells. We further show that jacalin-induced proliferation is characterized by an unusual pattern of T cell activation and cytokine production by human peripheral blood mononuclear cells (PBMC). A cognate interaction between T cells and monocytes was critical for jacalin-induced proliferation, and human recombinant interleukin (IL)-1 and IL-6 did not replace the co-stimulatory activity of monocytes. Blocking studies using monoclonal antibodies (mAb) point out the possible importance of two molecular pathways of interaction, the CD2/LFA-3 and LFA-1/ICAM-1 pathways. One out of two anti-CD4 mAb abolished jacalin responsiveness. Jacalin induced interferon-gamma and high IL-6 secretion, mostly by monocytes, and no detectable IL-2 synthesis or secretion by PBMC. In contrast, jacalin-stimulated Jurkat T cells secreted IL-2. CD3- Jurkat cell variants failed to secrete IL-2, suggesting the involvement of the T cell receptor/CD3 complex pathway in jacalin signaling. IL-2 secretion by CD4- Jurkat variant cells was delayed and lowered. In addition to CD4, jacalin interacts with the CD5 molecule. Jacalin-CD4 interaction and the proliferation of PBMC, as well as IL-2 secretion by Jurkat cells were inhibited by specific jacalin-competitive sugars.     

CD30+ lymphocytes in chronic gingivitis from HIV-positive patients: a pilot study

Th2 type lymphocytes are characterized by high expression of CD30 glycoprotein. Increased serum levels of CD30 and Th2 IL-4 producing T-cells are found during AIDS progression. Since HIV-positive patients are more susceptible to periodontal disease, quantitative analysis of positive cells for the CD30 receptor in chronic gingivitis of both HIV-infected and non-infected patients (NSG) would help to clarify the immunoregulation of HIV-associated periodontal diseases. The purpose of this study was to evaluate CD30+ lymphocytes in gingival biopsies from sites exhibiting chronic gingivitis on HIV-positive patients (CG-HIV) and NSG. A biotin-streptavidin amplified system was used for identification of the CD30 receptor. The results demonstrated increased proportions of Th2 cells in CG-HIV as compared to NSG. Additional studies are necessary to understand the importance of these cells to the biological activity or inactivity of the disease.     

IL-2 and IL-7 differentially induce CD4-CD8- alpha beta TCR+NK1.1+ large granular lymphocytes and IL-4-producing cells from CD4-CD8- alpha beta TCR+NK1.1- cells: implications for the regulation of Th1- and Th2-type responses

Effector functions of CD4-CD8- double negative (DN) alpha beta TCR+ cells were examined. Among mouse DN alpha beta TCR+ thymocytes, NK1.1+ cells expressing a canonical V alpha 14/J alpha 281 TCR but not NK1.1- cells produce IL-4 upon TCR cross-linking and IFN-gamma upon cross-linking of NK1.1 as well as TCR. Production of IL-4 but not IFN-gamma from DN alpha beta TCR+NK1.1+ cells was markedly suppressed by IL-2. Whereas V alpha 14/J alpha 281 TCR+ cells express NK1.1+, these cells are not the precursor of DN alpha beta TCR+NK1.1+CD16+B220+ large granular lymphocytes (LGL). IL-2 induces rapid proliferation and generation of NK1.1+ LGL from DN alpha beta TCR+NK1.1- but not from DN alpha beta TCR+NK1.1+ cells. LGL cells exhibit NK activity and produce IFN-gamma but not IL-4 upon cross-linking of surface TCR or NK1.1 molecules. In contrast to IL-2, IL-7 does not induce LGL cells or NK activity from DN alpha beta TCR+NK1.1- cells but induces the ability to produce high levels of IL-4 upon TCR cross-linking. Our results show that DN alpha beta TCR+ T cells have several distinct subpopulations, and that IL-2 and IL-7 differentially regulate the functions of DN alpha beta TCR+ T cells by inducing different types of effector cells.     

The CD7- T cell subset represents the majority of IL-5-secreting cells within CD4+CD45RA- T cells

Absence of CD7 is a stable phenotype in a subset of normal human T cells. Most circulating CD7- T cells express the CD4CD45RO+CD45RA- memory phenotype. We analysed CD4+CD45RA- peripheral blood lymphocytes that were separated into CD7+ and CD7- for their in vitro cytokine secretion in response to different stimuli. The CD4+CD7- subpopulation was found to secrete significantly higher levels of IL-5 compared with the CD4+CD7- subset upon stimulation with ionomycin/phorbol myristate acetate (PMA) plus anti-CD28 MoAbs. In contrast to IL-5 secretion, IL-4 and interferon-gamma (IFN-gamma) secretion was not significantly different in CD7+ and CD7- T cells upon stimulation in vitro. The data indicate that the CD4+CD7- T cell represents the majority of IL-5-secreting cells within the population of CD4+CD45RA- memory T cells. Since CD4+CD7- T cells were found to be enriched in various skin lesions associated with eosinophilic infiltration, the results of our study support the hypothesis that skin-infiltrating CD7- T cells are one of the major sources of IL-5 responsible for the development of eosinophilic inflammation in certain skin diseases.     

Increased in vitro induced CD4+ and CD8+ T cell IFN-gamma and CD4+ T cell IL-10 production in stable relapsing multiple sclerosis

Multiple sclerosis (MS) is presumed to be a T-cell mediated chronic inflammatory disease of the central nervous system. Investigators previously demonstrated increased IFN-gamma (pro-inflammatory) and IL-10 (counterregulatory anti-inflammatory) in MS. The balance of pro-inflammatory and counterregulatory anti-inflammatory cytokines may be important in the stabilization of disease activity. Purified CD4+ and CD8+ T cells from patients with clinically definite, stable relapsing MS (RRMS) were stimulated by anti-CD3 mAb or Con A for 48 hours and cytokine supernatants analysed for production of IL-2, IL-6, IFN-gamma, TNF-alpha (potential pro-inflammatory) and IL-4, IL-10, and TGF-beta (potential counterregulatory anti-inflammatory). Con A activated CD4+ and CD8+ T cell proinflammatory cytokine IL-2 secretion, CD4+ T cell IL-6 secretion, CD4+ and CD8+ T cell TNF-alpha secretion and CD8+ T cell IFN-gamma secretion was decreased significantly in RRMS subjects compared to controls. CD3 activated CD4+ and CD8+ T cell IL-6 secretion and CD4+ T cell TNF-alpha secretion was significantly decreased in MS subjects compared to controls. In contrast, there was increased CD3-induced IFN-gamma in both CD4+ and CD8+ T cells and counterregulatory anti-inflammatory CD3-induced IL-10 secretion in CD4+ T cells in RRMS compared to controls. These data suggest that an equilibrium of a pro-inflammatory (IFN-gamma) and a counterregulatory anti-inflammatory (IL-10) cytokine may define stable clinically definite early RRMS.     

Signaling through a CD3 gamma-deficient TCR/CD3 complex in immortalized mature CD4+ and CD8+ T lymphocytes

The biologic role of each CD3 chain and their relative contribution to the signals transduced through the TCR/CD3 complex and to downstream activation events are still controversial: they may be specialized or redundant. We have immortalized peripheral blood CD4+ and CD8+ T lymphocytes from a human selective CD3 gamma deficiency using Herpesvirus saimiri. The accessibility of the mutant TCR/CD3 complex to different Abs was consistently lower in immortalized CD8+ cells when compared with CD4+ cells, relative to their corresponding CD3 gamma-sufficient controls. Several TCR/CD3-induced downstream activation events, immediate (calcium flux), early (cytotoxicity and induction of surface CD69 or CD40L activation markers or intracellular TNF-alpha) and late (proliferation and secretion of TNF-alpha), were normal in gamma-deficient cells, despite the fact that their TCR/CD3 complexes were significantly less accessible than those of controls. In contrast, the accumulation of intracellular IL-2 or its secretion after CD3 triggering was severely impaired in gamma-deficient cells. The defect was upstream of protein kinase C activation because addition of transmembrane stimuli (PMA plus calcium ionophore) completely restored IL-2 secretion in gamma-deficient cells. These results suggest that the propagation of signals initiated at the TCR itself can result in a modified downstream signaling cascade with distinct functional consequences when gamma is absent. They also provide evidence for the specific participation of the CD3 gamma chain in the induction of certain cytokine genes in both CD4+ and CD8+ human mature T cells. These immortalized mutant cells may prove to be useful in isolating cytosolic signaling pathways emanating from the TCR/CD3 complex.     

IL-16 activates the SAPK signaling pathway in CD4+ macrophages

IL-16 has been reported as a modulator of T cell activation and was shown to function as chemoattractant factor. The chemotactic activity of IL-16 depends on the expression of CD4 on the surface of target cells, but the intracellular signaling pathways are only now being deciphered. This report describes IL-16 as an additional activator of the stress-activated protein kinase (SAPK) pathway in CD4+ macrophages. Treatment of these cells with recombinant expressed IL-16 leads to the phosphorylation of SEK-1, resulting in activation of the SAPKs p46 and p54. IL-16 stimulation also leads to the phosphorylation of c-Jun and p38 MAPK (mitogen-activated protein kinase), without inducing MAPK-family members ERK-1 and ERK-2. Interestingly, the IL-16-mediated activation of SAPKs and p38 MAPK in macrophages alone induces no detectable apoptotic cell death. These observations suggest specific regulatory functions of IL-16 distinct from the proinflammatory cytokines TNF-alpha and IL-1beta.     

Antigen-specific B cells preferentially induce CD4+ T cells to produce IL-4

The bone marrow microenvironment influences whether a given B cell proliferates, differentiates, or undergoes apoptosis. In this report, we demonstrate that apoptosis of primary murine B lymphocyte precursors can be regulated either positively or negatively by stroma. Several stromal lines that support lymphocyte outgrowth suppressed the spontaneous apoptosis of pre-B cells by as much as 90%. Direct contact with stromal cells more effectively protected lymphocytes than did stromal cell-CM or a collection of recombinant cytokines. In contrast, one unique stromal cell clone actually induced lymphocyte apoptosis, and a second line appeared inert. A survey of adherent cell lines suggested that expression of life-sparing molecules is widespread but not ubiquitous. Experiments with neutralizing Abs to CD44, vascular cell adhesion molecule-1 (VCAM-1), CD9, intercellular adhesion molecule-1 (ICAM-1), or ICAM-2 suggested that these interaction molecules do not deliver short-term survival signals to B cell precursors. Of particular interest, direct interaction with lymphocyte-supportive stromal cells minimized the negative regulatory effects of IL-1alpha, and a glucocorticoid, but not IFN-beta or PGE2. These results demonstrate that the effect of negative regulators depends upon the context in which these signals are presented. As molecules that influence B lymphopoiesis are better defined, it will be important to consider the role of each in combination with other stimuli.