DNA immunization utilizing a herpes simplex virus type 2 myogenic DNA vaccine protects mice from mortality and prevents genital herpes.

A gene transfer vector for DNA immunization was developed in which the promoter was derived from the murine muscle creatine kinase (MCK) gene; a gene expressed only in differentiated skeletal muscle. In vitro, we observed high-level, but unrestricted, gene expression from the cytomegalovirus (CMV) promoter unlike expression from the MCK promoter which was weak but restricted to myofibers. A myogenic DNA vaccine (MDV) that encoded the glycoprotein D gene from herpes simplex virus type-2 (HSV-2) was used to DNA immunize mice. MDV immunization resulted in virus specific immunity that protected HSV-2 infected mice from mortality and prevented the development of genital herpes. Therefore, we conclude that high-level gene expression or the use of a strong transcription unit was not a prerequisite for an efficacious DNA vaccine and the use of a nonviral tissue specific promoter could suffice.

Anti-interleukin-6 antibodies inhibit herpes simplex virus reactivation.

Herpes simplex viruses (HSVs) infect epithelial cells, become localized in neurons, and can reactivate in response to a variety of stimuli, including ultraviolet light and hyperthermia. The sequence of gene activation during viral replication is known, but the molecular linkage between exogenous stimuli and HSV reactivation has not been determined. It was hypothesized that interleukin (IL)-6 acts as a signal between exogenous stimuli and neurons, stimulating HSV reactivation from latency. Mouse corneas were infected with HSV-1, and ocular reactivation was induced 5-7 weeks later by thermal stress or corneal exposure to ultraviolet light. Anti-IL-6 monoclonal antibodies were administered to the latently infected mice 8-12 h before the reactivation stimulus. Treatment with anti-IL-6 antibodies resulted in significantly lower frequencies of ocular reactivation compared with those in mice treated with a control immunoglobulin. These results support the hypothesis that IL-6 plays a role in HSV reactivation from latency.

Administration of dehydroepiandrosterone sulfate retards onset but not progression of autoimmune disease in NZB/W mice.

NZB/W mice spontaneously develop an autoimmune disease characterized by the formation of anti-DNA antibodies and subsequent development of a fatal immune complex-mediated glomerulonephritis. Treatment of NZB/W F1 female mice with DHEAS, a precursor of DHEA, beginning at 2 months of age delayed the onset of autoimmune disease and prolonged survival. Animals treated with DHEAS beginning at 2 months of age had significantly lower anti-dsDNA serum antibody titers when compared to controls. Interestingly, DHEAS treatment had no effect on titers of anti-phosphatidylcholine (PtC) "natural" antibodies. Serum levels of IL-10, which increase with onset of disease, were also significantly reduced in mice treated with DHEAS beginning at 2 months of age. In contrast, if DHEAS treatment was started at 6 months of age, there was no effect on mortality rates. In addition, treatment of animals with DHEAS beginning at 6 months of age did not lower serum titers of anti-dsDNA and had no ameliorating effect on anti-PtC antibody production. Serum levels of IL-10 were also unaffected in mice treated with DHEAS beginning at 6 months of age. Together, these data suggest that parenteral administration of DHEAS is effective at delaying autoimmune disease and prolonging survival when given prior to the onset of symptoms. However, DHEAS treatment does not affect the course of disease when treatment begins after the onset of disease. We propose that DHEA(S) therapy used under similar conditions would not provide a clinically beneficial effect in the specific symptoms of immune complex-mediated glomerulonephritis.

The use of oral dehydroepiandrosterone sulfate as an adjuvant in tetanus and influenza vaccination of the elderly.

Elderly individuals often exhibit a poorer immune response and shorter duration of immunity to vaccines than younger persons. Improvement in vaccine response has been demonstrated when administering the hormone dehydroepiandrosterone sulfate (DHEAS) as an adjuvant in animal trials. Two separate, randomized double-blinded vaccine trials were therefore conducted using DHEAS as an oral adjuvant in individuals age 65 or older. Sixty-six individuals were randomized to DHEAS, 50 mg po bid for 4 days, or a placebo capsule. Tetanus vaccination was given immediately before the fifth dose. At entry the level of protective antibody was age-dependent (P = 0.009), and by 28 days post-vaccination most individuals had protective levels of antibody, with no difference noted between treatment groups. In the second study, 67 individuals received placebo capsules or DHEAS immediately before and 24 h after influenza vaccination. The number of individuals who developed protective titers (> or = 1:40) was not different in the two groups. The mean log increase in HAI response was greater in the DHEAS group to all three vaccine components, although this did not achieve significance. Minimal side-effects of DHEAS administration were noted. Given the trend toward improved response in the elderly to influenza, larger trials using DHEA as an adjuvant in vaccines that are neoantigens may be indicated.

Induction of common mucosal immunity by hormonally immunomodulated peripheral immunization.

The study described in this report demonstrates that peripheral lymph nodes draining nonmucosal tissues can effectively serve as induction sites for the establishment of common mucosal immunity if the microenvironmental conditions are altered to mimic those normally present within mucosa-associated lymphoid tissues (e.g., Peyer's patches). Lymph node lymphocytes exposed in situ to the immunomodulatory influences of the hormone 1 alpha, 25-dihydroxy vitamin D 3 were found to produce less gamma interferon and interleukin-2 (IL-2) and far more IL-4, IL-5 and IL-10 than lymphocytes from control animals. When couples with vaccination with hepatitis B surface antigen (HBsAg), the hormone, immunomodulated switch from a peripheral lymph node phenotype to a Peyer's patch-like pattern promoted the induction of both a systemic and a common mucosal immune response. This was determined by the observed increased concentrations of serum anti-HBsAg antibody and by finding that anti-HBsAg secretory antibodies were detectable in urogenital, lachrymal, fecal and oral secretions only in the hormone-treated animals. In addition, specific antibody-secreting cells were detectable in the lamina propria of the lungs and small intestines of the hormone-treated animals subsequent to vaccination, indicating that the homing properties of antigen-specific B cells were being affected by the treatment procedure. The humoral and mucosal immune responses were further augmented if both 1 alpha, 25-dihydroxy vitamin D 3 and dehydroepiandrosterone were used together as hormonal immunomodulators. This novel immunization technique may afford new opportunities to effectively intervene in sexually transmitted diseases and other diseases caused by mucosal pathogens.

Problems and priorities for controlling opportunistic pathogens with new antimicrobial strategies; an overview of current literature.

An International Study Group on New Antimicrobial Strategies (ISGNAS) has been formed in response to the recognition that development of microbial resistance to antibiotics is becoming a serious, world-wide problem. The group met in 1993 for the first time to discuss the feasibility of developing rational alternatives to the use of antibiotics and prepared, as a result, a comprehensive overview of normal (physiological) mechanisms involved in the control of potentially pathogenic (oppotunistic) microorganisms. One objective of ISGNAS is to understand the conditions which allow opportunistic microbes present among the symbionts to cause an infection. There is a need for more coherent information concerning the habitat, growth requirements and host and pathogen properties which allow opportunistic pathogens to cause life-threatening infections. In particular, information is urgently being sought to understand the complexity of the interactions between the vast number of microbial species, and the interactions between the microbes and their host. Another goal is to inspire and enable basic and clinical research that will lead to the development of new therapies for regulating colonization, translocation and infection by opportunistic micro-organisms in patients during periods of decreased resistance. With a sufficient amount of knowledge of how healthy individuals keep opportunistic micro-organisms under control, it may become feasible for physicians to maintain host resistance and inter-microbial factors involved in the containment of opportunistic microbes. Therapies aimed at boostering natural resistance mechanisms will be of critical importance to individuals whose resistance has been compromised as a result of another clinical condition.

Nucleic acid vaccine encoding gD2 protects mice from herpes simplex virus type 2 disease.

Nucleic acid vaccinations with plasmids pWW65, containing the sequence for herpes simplex type 2 (HSV-2) gD2, and pRSVnt, lacking the gD sequence, were studied. Groups of mice were immunized with pWW65 alone, pWW65 plus 1,25-dihydroxyvitamin-D3 (D3), or pRSVnt. Clinical disease (vaginitis), serum and vaginal washing antibody levels, and vaginal washing virus titers were measured intravaginal HSV-2 challenge. No animals (0/10) in the pWW65 + D3 group, 6/10 animals in the pWW65 group, and 10/10 animals in the pRSVnt group developed severe disease by postchallenge day 13 (P<.001, P=.04 vs. pRSVnt). Virus titers in vaginal washings were significantly reduced in the pWW65 and pWW65+D3 groups versus the pRSVnt group (P<.001). Increasing levels of serum anti-gD2 antibodies were measured 2 and 6 days after challenge among animals in the pWW65 and pWW65+D3 groups but not among animals in the pRSVnt group. Vaccinations with a plasmid containing the gD2 gene are immunogenic and provide some protection from HSV-2-induced disease.

Th1/Th2-like immunity and resistance to herpes simplex labialis.

To investigate potential immunologic mechanisms of resistance to recurrent herpes simplex labialis, we assayed serum antibody titers and cultured peripheral blood mononuclear cell (PBMC) cytokine production among patients with a history of frequent episodes (H+S+), herpes simplex virus (HSV)-seropositive individuals without a history of herpes labialis (H-S+) and HSV-seronegative persons (H-S-). In addition, H+S+ patients were exposed to experimental ultraviolet radiation (UVR) on the lips and the immunologic assay results compared among those who developed experimental lesions and those who did not. H+S+ patients were found to have higher median serum titers of HSV antibody and trends to lower levels of HSV-specific PBMC IFN-gamma and IL-2 than H-S+ control patients (123 vs 66, P = 0.04; 424 vs 548 pg/ml, P = 0.08; 14 vs 26 pg/ml, P = 0.14, respectively). Correlation of the results with the occurrence of experimental lesions showed the inverse: the subgroup of H+S+ patients with UVR-induced lesions had lower titers of antibody and trends to higher levels of IFN-gamma and IL-2 than H+S+ patients who could not be induced (93 vs 149, P = 0.02; 501 vs 347 pg/ml, P = NS; 26 vs 11 pg/ml, P = NS, respectively). The size and duration of UVR-induced lesions showed positive correlations with IFN-gamma and IL-2 levels (r = 0.60-0.67, P = 0.02-0.04). Although the small number of patients limited the power of this study, the overall pattern of the findings suggests that a Th1-like cytokine response (IFN-gamma and IL-2 production) may be associated with resistance to naturally occurring episodes of herpes labialis. The development and severity of experimental UVR-induced herpes labialis appears to be regulated differently and may involve an immunopathologic mechanism.

Dehydroepiandrosterone reduces progressive dermal ischemia caused by thermal injury.

Progressive ischemia and necrosis of the skin following thermal injury are reduced by postburn administration of the steroid hormone dehydroepiandrosterone (DHEA). Thermally injured animals were provided with a subcutaneous injection of DHEA, or a related species of steroid hormone, at various times after burning. During the 96 hr following administration of the scald burn, tissue necrosis was closely monitored. Subcutaneous administration of DHEA at approximately 1 mg/kg/day achieved optimal protection against the development of progressive dermal ischemia. DHEA, 17 alpha-hydroxy-pregnenelone, 16 alpha-bromo-DHEA, and androstenediol each demonstrated, a similar level of protection. Other forms of steroids, including DHEA sulfate, androstenedione, 17 beta-estradiol, or dihydrotestosterone, exhibited no protective effect under the conditions tested. Additionally, intervention therapy with DHEA could be initiated up to 4 hr, but not 6 hr, after burn without a marked reduction in therapeutic benefit. Examination of the microvasculature of thermally injured dorsal skin suggested that postburn intervention with DHEA, either directly or indirectly, maintained a normal architecture in most of the dermal capillaries and venules within burn-exposed tissue. These findings suggest that systemic intervention therapy of burn patients with DHEA or a similar acting steroid hormone may be useful in preventing the progressive tissue destruction caused by progressive ischemia.

Steroids as regulators of the mammalian immune response.

The mammalian immune system is multicellular in composition, and its proper function requires careful control over complex developmental pathways and many distinct types of effector responses. Numerous overlapping mechanisms of intercellular communication are needed to accomplish the tasks of proper regulation of the diverse cell types that constitute this essential protective system. One mechanism occurs by direct cell-to-cell contact through the interaction of membrane-associated molecules. Examples of this type of communication include the interaction that takes place between the antigen-specific T-cell receptor and the foreign peptides that are bound to major histocompatibility complex molecules, as well as costimulatory molecule interactions with their specific ligands expressed on antigen-presenting cells (e.g., CD28 and B7-1 or B7-2). A second mechanism occurs through the production, secretion, and activities of soluble mediators, collectively known as the cytokines. The cytokines are represented by a large and diverse group of molecules that are produced by a wide variety of cell types. Unique species of cytokines bind to specific membrane-associated receptors on target cells, inducing the activation of particular signal-transduction pathways. These processes subsequently lead to the diversity of cytokine-linked changes in cellular physiology. Some of the cytokines exert their influences in vivo via endocrine routes, although it is far more common for intercellular communication via cytokines to occur microenvironmentally via paracrine or autocrine pathways. The object of this review is to provide evidence supporting the concept that one mechanism for upstream regulation of cytokine production by immunocompetent cell types is controlled by the regulatory activities of various steroid hormones. Strain variation in susceptibility to infectious agents, the condition of immunosenescence, and the processes that control the development of common mucosal immunity are used as examples of immune mechanisms that may be under steroid hormone control.

The development of effective vaccine adjuvants employing natural regulators of T-cell lymphokine production in vivo.

Steroid hormones are important regulators of gene function in vivo. A number of naturally occurring species of steroid hormones are able to qualitatively and quantitatively influence the production of lymphokines by activated T cells in vitro. Similar mechanisms are probably also occurring naturally in vivo and could explain why mucosal and nonmucosal lymphoid organs harbor T cells having unique potentials for lymphokine production. It was established that the topical application of 1,25 dihydroxyvitamin D3 (1,25(OH)2D3) to normal mice changed the pattern of lymphokines produced by activated T cells isolated from the draining peripheral lymph nodes. The hormone-treated T cells produced a pattern of lymphokines similar to that normally found in Peyer's patches. Subcutaneous vaccination with a protein antigen, in a site afferent to 1,25(OH)2D3-manipulated lymph nodes, resulted in an enhanced serum antibody response and was uniquely capable of also stimulating a common mucosal immune response to the antigen as well. Common mucosal immunity was confirmed by demonstrating the presence of antigen-specific IgA and IgG responses in a number of mucosal secretions and by further establishing that antibody-secreting plasma cells had migrated to the lungs and small intestines of the hormone-treated and vaccinated animals. Additional experiments established that common mucosal immunity could also be induced in aged animals as long as the immune system of the vaccinated animals was functioning normally. This was accomplished by providing the aged animals with a dietary supplement of dehydroepiandrosterone sulfate (DHEAS). Previous studies by us have documented that aged animals provided with replacement levels of DHEAS, a natural steroid hormone whose endogenous production declines with advancing age, are able to mount normal systemic humoral and cellular immune response following subcutaneous vaccination with a variety of protein and polysaccharide antigens. The combination of supplemental DHEAS therapy with topical 1,25(OH)2D3 treatment at the time of vaccination provided the conditions needed to generate mucosal and systemic immune responses to inactivated influenza virus antigen by old animals.

Age- and microenvironment-associated influences by platelet-derived growth factor on T cell function.

Platelet-derived growth factor (PDGF) is produced by numerous cell types in response to a variety of activation signals. Although the role of PDGF in cellular proliferation is well established, the immunomodulatory effects of this peptide growth factor are only now being delineated. We previously established that PDGF alters the profile of lymphokines produced by activated T cells obtained from the peripheral lymph nodes of adult mice. We now report that T cells residing in lymphoid organs that receive their major afferent lymphatic drainage from gut mucosa are relatively resistant to the effects of this growth factor. As the vast majority of peripheral T cells are in the recirculating T cell pool, these findings suggest that tissue-specific microenvironmental factors function to regulate the sensitivity of T cells to PDGF-mediated influences. Additional studies have determined that the normal process of aging is accompanied by a systemic loss in T cell responsiveness to PDGF. Although the T cells of immature (2-wk-old) and young adult mice are responsive to the immunomodulatory influences of PDGF, T cells of aged animals (> 100 wk) are relatively resistant to its effects. Some of the immune abnormalities noted to occur during the aging process appear to represent a consequence of the dysregulated production of IL-6. We therefore evaluated whether IL-6 was responsible for modifying the sensitivity of T cells to PDGF. Data presented herein demonstrate that IL-6 abrogates the ability of PDGF to modify lymphokine production by T cells after activation. Therapeutic measures capable of reducing spontaneous IL-6 production in aged mice restored the ability of their T cells to respond to PDGF, suggesting that the dysregulated production of IL-6 within the aged host interferes with the ability of PDGF to convey important microenvironmental information to T cells residing in peripheral lymphoid organs.

Immunomodulation in vivo by the Mycoplasma arthritidis superantigen, MAM.

Mycoplasma arthritidis produces a potent superantigen (MAM) that activates specific murine and human T lymphocytes to proliferate and secrete lymphokines. We show here that MAM also influences both T- and B-cell functions in vivo. Lymphocytes from mice injected with MAM exhibit a suppression of proliferative responses to MAM in vitro but only a partial suppression of responses to other mitogens. This T-cell anergy not only decreased contact sensitivity to dinitrofluorobenzene but also prolonged survival of skin transplants. In contrast, B-cell reactivity is increased following in vivo injection of MAM, as evidenced by enhanced antibody responses to sheep red blood cells and ovalbumin. Also, there is a marked decrease in the ability of splenocytes from MAM-injected mice to produce interleukin-2 (IL-2) but a marked increase in their ability to produce IL-4 and IL-6. The combined results suggest that MAM induces a lymphokine profile that favors activation of B-cell functions, with a resulting potential for triggering of autoimmune disease.

Altered regulation of IL-6 production with normal aging. Possible linkage to the age-associated decline in dehydroepiandrosterone and its sulfated derivative.

Normal aging in humans has been recently shown to be accompanied by reduced control over production of the multifunctional cytokine IL-6. This cytokine was reported to be quantitatively elevated in most serum samples obtained from "normal" elderly humans. In the present investigation, we report that IL-6 levels are elevated in serum samples obtained from aged mice, and its spontaneous production could also be easily detected in culture supernatants of unstimulated lymphoid cells obtained from aged, but not mature, adult donors. Spontaneous production of IL-6 was consistently observed in culture supernatants of lymphoid cells from both the spleen and mesenteric lymph nodes from aged donors, but was absent from supernatants derived from their peripheral lymph nodes. In aged mice, the reduced regulation of IL-6 production could be effectively prevented and/or reversed by supplementing aging animals with dehydroepiandrosterone sulfate, a steroid hormone whose endogenous production is known to decline with advancing age in all species tested. It was also established that serum obtained from old dehydroepiandrosterone sulfate-treated mice contained lower (normal) levels of serum amyloid P substance (an acute phase reactant), reduced levels of serum Ig (all classes and isotypes) and lower titers of tissue-specific autoantibodies than untreated aged controls. Therefore, a number of well described, age-related conditions, some of which could be contributing to the pathologic phenotype of old age, may actually represent secondary effects to this age-associated change in IL-6 production.

Reversal of the immunosenescent phenotype by dehydroepiandrosterone: hormone treatment provides an adjuvant effect on the immunization of aged mice with recombinant hepatitis B surface antigen.

This study sought to establish whether administration of dehydroepiandrosterone (DHEA) or its sulfate derivative to aged mice could effectively correct the immunosenescent phenotype. Supplemental DHEA sulfate and topical DHEA fully corrected the age-associated dysregulated production of T cell lymphokines by cells from all of the different lymphoid organs tested. Either DHEA or DHEA sulfate supplementation promoted enhanced antibody responses against recombinant hepatitis B surface antigen (rHBsAg) by the aged recipients when incorporated directly into the vaccine. When DHEA was provided either topically or was incorporated directly into vaccine, vigorous primary and secondary antibody responses were detected in the aged mice given a single administration of DHEA, regardless of the mode of administration. It was also established that DHEA treatment could enhance specific antibody responses to rHBsAg in aged animals that had previously not been effectively immunized by conventional vaccination procedures.

Adaptive immune responses during murine pregnancy: pregnancy-induced regulation of lymphokine production by activated T lymphocytes.

OBJECTIVE: We hypothesized that the lymphokine production by splenocytes and decidual lymphocytes would be altered because of changes in immunoregulation during pregnancy. STUDY DESIGN: Splenocytes and decidual lymphocytes were isolated from syngeneic and allogeneic pregnant mice at different times of gestation. The lymphocytes (10(7) cells/ml) were stimulated with anti-CD3 antibody, and culture supernatants were assayed for several lymphokines, including interleukin-2, interferon-gamma, interleukin-4, interleukin-6, granulocyte-macrophage colony-stimulating factor, and interleukin-3. Statistical analysis was by analysis of variance or paired t test. RESULTS: Activated splenocytes produced significantly less interleukin-2 and more interleukin-4, interleukin-6, and interleukin-3 as murine pregnancy advanced. Production of interferon-gamma and granulocyte macrophage colony-stimulating factor by activated splenocytes peaked in the first 8 to 14 days of pregnancy. Stimulated decidual lymphocytes produced modest amounts of interleukin-6, granulocyte-macrophage colony-stimulating factor, and interleukin-3 during pregnancy but no interleukin-2, interferon-gamma, or interleukin-4. Similar results were found for both syngeneic and allogeneic matings. CONCLUSIONS: Our findings indicate that splenocyte lymphokine production favors interleukin-4 production over interleukin-2 production. This finding suggests that antibody production would be enhanced and cytotoxic cellular immune responses inhibited during pregnancy. These changes occurred regardless of mating partner, suggesting that the specific antigenic stimulus during normal pregnancy does not regulate lymphokine production. Activated splenocytes and decidual lymphocytes were found to differ in their capacity to produce lymphokines, indicating that the decidua constitutes a distinct and unique immunologic microenvironment.

Administration of dehydroepiandrosterone to burned mice preserves normal immunologic competence.

Burned individuals display a reduced ability to elicit cellular and humoral immune responses and a depression in the vitro production of certain T-cell lymphokines. Treatment of burned mice with 100 micrograms of dehydroepiandrosterone within 1 hour after injury resulted in preserving a completely normal capacity to produce T-cell-derived lymphokines and to generate cellular immune responses. In addition, dehydroepiandrosterone-treated thermally injured mice demonstrated an above-normal ability to resist an induced infection with the intracellular pathogen, Listeria monocytogenes. Dehydroepiandrosterone-treated animals also did not exhibit the sustained plasma levels of interleukin 6 that normally accompany thermal injury and infection. Because of its antiglucocorticoid effects and positive immunoregulatory influences, we believe dehydroepiandrosterone to be a beneficial form of therapy for thermally injured individuals.

Natural regulators of T-cell lymphokine production in vivo.

The mammalian immune system possesses the intrinsic capacity to evoke a wide variety of functionally distinct effector mechanisms following stimulation by a particular antigenic substance. Such diversity in available responses is absolutely essential to the immunocompetent host, which must continually deal with a diverse set of potential pathogens within its ever-changing environment. The development of appropriate types of immune responses, therefore, represents a highly dynamic process that requires that an equivalent consideration be given to a large array of components, any one of which is capable of modulating the final outcome. While the nature and complexity of the antigen(s), plus the intracellular or extracellular mode of presentation, provide specificity and some selection to the developing process, the route of antigen entry, as well as the physiological status of the host at the time of antigen insult, also contribute significantly to the formation of any immune response. The overall objective of this article is to introduce the concept that platelet-derived growth factor (PDGF) (either preformed or synthesized in response to stimulation), plus a number of steroid hormones (some of which are end-organ metabolized at local tissue sites), can all play significant roles in the genesis of immunologic responses in vivo.

The presence of a dehydroepiandrosterone-specific receptor binding complex in murine T cells.

We have investigated the ability of dehydroepiandrosterone (DHEA) to alter the production of interleukin-2 (IL-2) and to bind to a specific binding complex in antiCD3 epsilon activated T cells. Binding activity correlated with the presence of a specific DHEA binding complex in the cytosol and nuclei of DHEA-responsive T-cell hybridomas, as well as in CD4+ and CD8+ cells isolated from peripheral lymph nodes of normal mice. Scatchard analysis determined that intact lymphocytes and cytosolic fractions contained high affinity binding for [3H]DHEA (approx. 2.6 nM) with 1000-7000 binding sites existing per cell. Five of the T-cell hybridomas tested both responded to DHEA treatment with increased production of IL-2 and also contained specific high affinity [3H]DHEA binding. Four additional T-cell hybridomas were found to contain no specific [3H]DHEA binding and were also unresponsive to DHEA influences on IL-2 production. Sucrose density gradients demonstrated a 3-4s [3H]DHEA binding complex in high salt and a 7-8s binding complex in low salt. Specific binding was inhibited by preincubation of the cytosol fractions with either trypsin or chymotrypsin, or by heating to 60 degrees C for 1 h (less than 15% of control). [3H]DHEA binding was unaffected by preincubation of the cytosol fractions with ribonuclease, deoxyribonuclease, or phospholipase A. The DHEA-protein complexes bound to DNA-cellulose with the amount of binding being slightly increased by preincubation at 25 degrees C as compared to 4 degrees C. As expected, [3H]DHEA binding was inhibited by the addition of unlabeled DHEA, but was also modestly inhibited by dihydrotestosterone and cortisol. Binding of DHEA was unaffected by progesterone, dexamethasone, estradiol, androsterone, DHEAS, and beta-etiocholanolone at all concentrations tested. DHEA was incapable of inhibiting the binding of [3H]DHT to the androgen receptor or [3H]dexamethasone to the glucocorticoid receptor. Collectively, these findings suggest that murine T cells contain a specific DHEA receptor. We believe that DHEA is a steroid hormone that is directly involved in the regulation of IL-2 production by both normal and some T-cell hybridomas.

Decidual cell biosynthesis of interleukin-6: regulation by inflammatory cytokines.

Intrauterine infection is an important cause of preterm labor and delivery and is characterized by increased production of inflammatory cytokines by gestational tissues. We have evaluated the biosynthesis of the inflammatory cytokine, interleukin-6 (IL-6), by human decidua and its regulation by other cytokines essential to the inflammatory process. We found that decidual cells secrete small amounts of IL-6 in the presence of growth medium supplemented only with 10% fetal calf serum. Interleukin 1 (alpha and beta) and tumor necrosis factor (TNF) all induced a significant concentration-dependent stimulation of IL-6 production by decidual cells. Treatment of decidual cells with actinomycin D or cycloheximide abrogated the increase in IL-6 production induced by IL-1 beta. Northern blot analysis of cultured decidual cells revealed an increase in IL-6 messenger RNA (mRNA) over time in response to IL-1 beta. These data indicate that IL-1 beta stimulates an increase in IL-6 mRNA and protein production, reflecting either direct gene activation or stabilization of IL-6 mRNA. The concentration range tested (0.1 to 10 ng/mL) of each cytokine is within the range of values found in the amniotic fluid of women destined to deliver preterm due to infection of gestational tissues. Our data suggest that IL-6 is produced by human decidua in response to inflammation and, in conjunction with other inflammatory mediators, may play a role in the pathophysiology of preterm labor due to infection.