Michael P. Everson

Potentiation of IL-2-induced T-cell proliferation by retinoids☆

We evaluated the capacity of retinoids to potentiate proliferative responses of murine T-cells to recombinant human interleukin 2 (rIL-2). Concanavalin A (Con A) prestimulated spleen cells responded in a dose-dependent manner to added rIL-2. All-trans-retinoic acid (RA) at 10(-8) M potentiated the proliferative response by fivefold at saturating levels of IL-2. In similar experiments, two closely related retinamides, all-trans-(phenyl)retinamide (PR) and N-(4-hydroxyphenyl)retinamide (4-HPR), also potentiated murine splenocyte rIL-2 responses. Potentiation of IL-2-induced proliferation was dose-responsive to the concentration of added retinoid with peak potentiation occurring at 10(-10) - 10(-8) M in the presence of 10 U/ml rIL-2. Significant potentiation was observed at retinoid concentrations as low as 10(-14) M. Fluorescence flow cytometry of the responding cells revealed that among L3T4+, Lyt-2+ or total T-cells, at 72 h following Con A stimulation, essentially all of the cells expressed IL-2 receptors (IL-2R). This apparently represents near maximum IL-2R expression and treatment of the cells with retinoids did not increase IL-2R expression at that time point. The potentiation of IL-2 responses by retinoids was also observed with IL-2-dependent HT-2 cells, 98% of which were IL-2R positive. HT-2 proliferative responses to rIL-2 were potentiated as much as fourfold by 10(-10) M RA. HT-2 proliferative responses to rIL-2 were potentiated by all three retinoids dose dependently. Significant potentiation was observed with as little as 10(-14) M retinoid. Retinoids in the absence of IL-2 induced no proliferative responses. These data suggest that retinoids can augment the capacity of IL-2 to induce T-cell proliferation using Con A-activated murine splenic T-cell blasts and a long-term-cultured T-cell line.

Lack of evidence of systemic inflammatory rheumatic disorders in symptomatic women with breast implants

&NA; Breast implants containing silicone have been used for approximately 30 years for breast augmentation or reconstruction. In general, the implants have been well tolerated and reports have indicated a high degree of patient satisfaction. Nonetheless, there have been anecdotal reports of patients with musculoskeletal complaints that have been attributed to silicone breast implants. To investigate this further, we prospectively examined 70 women with silicone breast implants who had complaints that they or their referring physicians thought were related to their implants. On clinical examination, the majority of the patients had fibromyalgia, osteoarthritis, or soft‐tissue rheumatism. One patient had rheumatoid arthritis, which predated her implants, and one had Sjögrens syndrome. Because many of our patients had myalgic symptoms, we further evaluated these patients by measuring circulating levels of soluble factors including interleukin‐6, interleukin‐8, tumor necrosis factor‐alpha, soluble intercellular adhesion molecule‐1, and soluble interleukin‐2 receptor, which have been previously found to be elevated in patients with inflammatory diseases. We found that the levels of these molecules in women with silicone breast implants were not different from those seen in normal subjects and were significantly less than those seen when examining chronic inflammatory disorders such as rheumatoid arthritis or systemic lupus erythematosus. In summary, our clinical and laboratory evaluation of symptomatic breast implant patients argues against an association of silicone breast implants with a distinctive rheumatic disease or a systemic inflammatory disorder. Given these findings and the clinical picture, it is our impression that most symptomatic women with silicone breast implants have well‐delineated noninflammatory musculoskeletal syndromes. Moreover, these data fail to support the concept that their symptoms are due to a systemic inflammatory response related to their implants.

A mechanism of retinoid potentiation of murine T-cell responses : early upregulation of interleukin-2 receptors

The capacity of retinoids to amplify the proliferative response of BALB/c lymphocytes to concanavalin A (Con A)2 in the presence of exogenous interleukin-2 (IL-2) and the induction of IL-2 receptors (IL-2R) on L3T4+ and Lyt-2+ T-cells was evaluated. Preincubation with Con A for 8 h in the presence of retinoids resulted in a greater than two-fold increase in spleen cell proliferative response to Con A plus rIL-2 over the following 72 h relative to the response of cells preincubated with Con A alone. Peak potentiation of IL-2 responses occurred over a pharmacologic range of retinoic acid (RA) concentration (10(-10)-10(-8) M) in the presence of 20 U/ml rIL-2. This potentiation of the response to IL-2 was likewise observed after 8 h prestimulation with Con A with splenic T-cells enriched by passage over nylon wool. Preincubation of the spleen cells with Con A plus RA without the subsequent addition of IL-2 resulted in a proliferative response that was potentiated nearly to the level of the response produced by subsequent addition of IL-2 to Con A-activated cells. Preincubation of the cells with Con A in the presence of RA produced a true synergy with IL-2; the resulting increase in response was greater than the sum of the increases produced by RA or IL-2 alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Pharmacological and toxicological properties of arotinoids SMR-2 and SMR-6 in mice☆

Studies were conducted to define primary pharmacological and toxicological properties of two arotinoids, SMR-2 and SMR-6, in male B6D2F1 mice. Mice were gavaged daily for up to 22 days with retinoids in corn oil (0.1, 0.2, or 0.4 mg/kg day SMR-2 or SMR-6 or 2.5, 10, or 30 mg/kg all-trans-retinoic acid as a reference control). Toxicological and biochemical endpoints were assayed after 8, 15, and 22 days. At toxic doses, i.e., those inducing weight loss, morphological changes were observed in skin, lymph nodes, spleen, bone marrow, liver, thymus, forestomach, adrenal, bone, and testes. Biochemical alterations included elevated serum alkaline phosphatase, corticosterone, and interleukins-1, -2, and -3. Additional immune alterations included increased responsiveness of spleen cells to both thymus-dependent and thymus-independent mitogens and increases in the total number of B cells in the spleen. At doses not inducing weight loss, target organ effects included the appearance of plasma cells and infiltration of polymorphonuclear cells in lymph nodes; myeloid cell hypercellularity in bone marrow; hematopoiesis in spleen; subacute inflammation in forestomach; and periportal cytoplasmic vacuolization in liver. At the low doses, SMR-2 resulted in decreased responsiveness of spleen cells to mitogens and SMR-6 caused increased responsiveness. SMR-6 also increased interleukin-1 and-2 production at low doses. Biochemical effects included reduced activities of liver aryl hydrocarbon hydroxylase (AHH) and soluble brain protein kinase C. Overall, the results suggest that leukopoiesis and reduced liver AHH and reduced soluble protein kinase C activities are the primary and initial pharmacological and toxicological effects of retinoids.

Leukocyte transfusion-associated granulocyte responses in a patient with X-linked hyper-IgM syndrome

X-linked hyper-IgM syndrome (XHIM) is a severe congenital immunodeficiency caused by mutations in CD154 (CD40 ligand, gp39), the T cell ligand for CD40 on B cells. Chronic or cyclic neutropenia is a frequent complicating feature that heightens susceptibility to severe infections. We describe a patient with a variant of XHIM who produced elevated levels of serum IgA as well as IgM and suffered from chronic severe neutropenia. Eight of ten leukocyte transfusions with cells from a maternal aunt, performed because of mucosal infections, resulted in similar episodes of endogenous granulocyte production. Transfection studies with the mutant CD154 protein indicate that the protein is expressed at the cell surface and forms an aberrant trimer that does not interact with CD40. The data suggest that allogeneic cells from the patients aunt, probably activated T cells bearing functional CD154, may interact with CD40+ recipient cells to produce maturation of myeloid precursors in the bone marrow.

FACS-Sorted Spleen and Peyer’s Patch Dendritic Cells Induce Different Responses in Th0 Clones

Helper T cells in mice have been divided into two functionally distinct subsets based upon the pattern of cytokines secreted,1,2 with Th type 1 (Th l) cells producing IL-2 and IFN-γ upon activation and predominantly mediating cell-mediated immunity (CMI), and Th type 2 (Th2) cells producing IL-4, IL-5, IL-6, and IL-10 upon activation and primarily mediating humoral immunity by providing T cell help via cytokines for Ig isotype and subclass responses.1–4 For T cell activation, dendritic cells (DC) are the most potent inducers of primary in vivo T cell responses and are the principle in vitro stimulators of naive T cell activation in both mice and humans.5,6 Previous studies showed that DC from different lymphoid tissues could possess similar7 or different8 functions. These studies supported the notion that PP DC induce preferential IgA production (predominantly found at mucosal sites) and SP DC support IgM production by controlling the cytokines produced by T cells in mucosal and systemic tissues, respectively. Recent studies suggest that cytokines present during primary activation of naive T cells play a role in determining the pattern of cytokines produced during subsequent antigenic challenge.9 Perhaps more important to these stimulatory events are the APC or accessory cells (AC) resident in the local tissues that can stimulate initial cytokine secretion in this inductive milieu. In this regard, the present studies have addressed tissue specificity of the AC component of T cell activation using a single type of AC, namely, DC isolated from two different anatomical sites. We asked whether SP DC and PP DC, isolated under identical conditions, induce production of similar levels of T cell-derived cytokines that are involved in regulation of immune responses.

Divergent T-cell cytokine profiles induced by dendritic cells from different tissues.

Dendritic cells (DC) were originally described by Steinman and Cohn in 1973. 1, 2 Their methods of splenic DC isolation have involved the flotation of murine spleen (SP) cells on dense bovine serum albumin (BSA) gradients and differential glass or plastic adherence of SP cells. Nonadherent Peyer’s patch (PP) DC were isolated in our laboratory by Spalding and coworkers in 1983. 3 These methods employed flotation of PP cells on dense BSA gradients, clustering of DC within the buoyant PP population with periodate-modified SP T cells, dissociation of clusters, and final BSA flotation of enriched PP DC.

Characterization of 5B12.1, A Monoclonal Antibody Specific for IL-6

A monoclonal antibody (MAb) specific for interleukin-6 (IL-6) was generated by fusing SP2/0 cells with spleen cells from a mouse immunized with rat spleen cell derived plasmacytoma growth factor (rat PCT-GF). This MAb inhibited the growth of an IL-6-sensitive murine plasmacytoma clone, MD90, in the presence of the immunogen, rat PCT-GF. More interesting, however, this MAb demonstrated species cross-reactivity by neutralizing murine (recombinant and P388D1 cell line-derived) and human (recombinant) IL-6. IL-6 neutralization activity was also established in other IL-6 bioassays, such as the proliferation of spleen cells, plasmacytoma T1165, and a B-cell hybridoma 7TD1. IL-6 neutralization was overcome partially by increasing the concentration of PCT-GF. The MAb had no effect on PCT-GF-independent plasmacytoma KI81 proliferation. Plastic petri dish-bound MAb removed rmIL-6 activity. These results suggest that this MAb specifically binds IL-6 and neutralizes bioactivity of various PCT-GF, rmIL-6, and rhIL-6.

Dendritic Cells Regulate Development of Alloantigenic and Mitogenic THl Versus TH2 Responses

Previous studies indicate that different T helper cells may regulate humoral and cell-mediated immunity to foreign antigens. This regulation may be controlled in part by cytokines produced by different helper T cell (TH) populations, i.e., TH1 cells that produce IL-2 and IFN-γ, and TH2 cells that produce IL-4, IL-5, IL-6, and IL-10.1,2 However, the induction and regulation of these distinct TH subsets and their derived cytokines in vivo is still under investigation. It remains to be elucidated whether a given antigen presenting cell may induce similar or divergent cytokine production profiles or patterns in its regulation of cell-mediated versus humoral immunity. The studies presented here have addressed the possibility that DC regulate these divergent immune responses through induction of divergent T cell-derived cytokine production profiles.