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Interleukin-10 (IL-10) is described as a cytokine that exerts immune downregulating actions. In this regard, our study indicates that IL-10 activity on human saphenous veins is coupled to nitric oxide (NO) release. We demonstrated this phenomenon by using in vitro real-time amperometric measurement of NO levels in explanted human saphenous veins after IL-10 exposure. IL-10-induced NO release can inhibit the adherence of monocytes (75.7 +/- 15 cells/600 microm2 of endothelial surface) and granulocytes (65 +/- 18 cells/600 microm2 of endothelial surface) from control values (250-300 cells/600 microm2 of endothelial surface; p < 0.005). This inhibition is directly sensitive to NO synthase inhibition. The specificity of the IL-10 effects is shown by its sensitivity to antibody. In vivo measurement of IL-10 levels during and after cardiopulmonary bypass surgery indicated that they are higher at 6 h after skin closure (1,400 pg/ml) compared with levels found during surgery (300 pg/ml). We surmise that the postsurgical increase of IL-10 levels may be an immunoregulatory attempt to downregulate the diffuse inflammation that has been shown to be associated with cardiopulmonary bypass surgery.

1. The effects of interleukin-2 (rhIL-2) and interleukin-4 (rhIL-4) were investigated on gamma-aminobutyric acid (GABA)-induced inward currents on isolated, identified neurons of Lymnaea stagnalis L. (Mollusca, Gastropoda) by using a concentration clamp technique. 2. It was shown that the interleukins modified the GABA-induced inward current in an opposite direction: rhIL-2 (2-100 U/ml) decreased the peak value of IGABA in a dose-dependent manner, whereas rhIL-4 (0.2-100 U/ml), on the contrary, potentiated it. Both types of modulation were partially or fully reversible. 3. The reversal potential of IGABA was not shifted by these cytokines. 4. The time-to-peak value and inactivation time constant of the gamma-aminobutyric acid (GABA)-induced current was decreased by rhIL-4. The modulatory effect of rhIL-4 was eliminated after conjugation of this cytokine with its antibody. 5. It appears that cytokines could play a role in regulating the neural excitability through GABA-erg mechanisms.

Human papillomaviruses (HPVs) are double-stranded, circular, epitheliotropic DNA viruses of which nearly 70 types have been identified. Specific HPV types exhibit a predilection to infect certain sites; however, occurrence is not unique or restricted to these sites. HPV typing may also be helpful in determining the oncogenic potential of HPV lesions. The most common HPV types, 6 and 11, are associated with benign mucosal lesions, whereas types 18, 16, 31, and 33 are thought to confer a high rate of malignant transformation. We describe a patient with both palmar verrucae and esophageal papillomatosis that proved to be HPV type 45 by polymerase chain reaction. HPV 45 has a high homology to HPV 18 and is a member of the relatively new "high-risk" mucosal HPV family in terms of cervical oncogenic potential. To our knowledge, HPV 45 has never been reported in cutaneous warts or esophageal lesions.

This is the first study documenting the induction of gamma-interferon (IFN-gamma) in human embryonic fibroblasts during human cytomegalovirus (HCMV) replication. Infection of cells with HCMV resulted in the consistent production of IFN-gamma RNA, as determined by RT-PCR and Northern blot analysis. Western blot analysis of cell lysates and immunoprecipitates from the cultural fluids of infected cells demonstrated the presence of IFN-gamma at the protein level. Induction of IFN-gamma required infectious HCMV, since high-dose ultraviolet inactivation of the virus stock eliminated IFN-gamma production. Further, IFN-gamma induction appears to be a late event in the virus replication cycle, since inhibition of HCMV DNA synthesis (e.g., phosphonoacetic acid) blocked the increase in IFN-gamma. Soluble factor(s) released from HCMV-infected cells apparently did not contribute to the induction of IFN-gamma, since virus stocks from which virus had been removed by sedimentation did not induce production of IFN-gamma. The appearance of IFN-gamma at late stages of HCMV infection and its elimination in the presence of an inhibitor (Actinomycin D) of RNA synthesis indicate a true transcriptional induction of this lymphokine at the RNA and protein levels. The significance of IFN-gamma production with regard to the replication and pathogenesis of HCMV in vitro and in vivo will require further investigation.

The discovery of the ability of the nervous system to communicate through "public" circuits with other systems of the body is attributed to Ernst and Berta Scharrer, who described the neurosecretory process in 1928. Indeed, the immune system has been identified as another important neuroendocrine target tissue. Opioid peptides are involved in this communication (i.e., neuroimmune) and with that of autoimmunoregulation (communication between immunocytes). The significance of opioid neuropeptide involvement with the immune system is ascertained from the presence of novel delta, mu, and kappa receptors on inflammatory cells that result in modulation of cellular activity after activation, as well as the presence of specific enzymatic degradation and regulation processes. In contrast to the relatively uniform antinociceptive action of opiate and opioid signal molecules in neural tissues, the presence of naturally occurring morphine in plasma and a novel mu3, opiate-specific receptor on inflammatory cells adds to the growing knowledge that opioid and opiate signal molecules may have antagonistic actions in select tissues. In examining various disorders (e.g., human immunodeficiency virus, substance abuse, parasitism, and the diffuse inflammatory response associated with surgery) evidence has also been found for the involvement of opiate/opioid signaling in prominent mechanisms. In addition, the presence of similar mechanisms in man and organisms 500 million years divergent in evolution bespeaks the importance of this family of signal molecules. The present review provides an overview of recent advances in the field of opiate and opioid immunoregulatory processes and speculates as to their significance in diverse biological systems.

Opioid peptides activate immunocytes and opiate alkaloids inhibit this activation in the mussel, Mytilus edulis. Here we present evidence that cells of another invertebrate, Leucophaea maderae, can be influenced in a similar way by the Met-enkephalin analogue D-Ala2-Met5-enkephalin (DAMA) and morphine. Effects of different signal molecules on Leucophaea hemocytes were evaluated by computer-assisted image analysis of their conformational state. A small percentage of the untreated cells were found to display spontaneous conformational changes after 25 min of incubation without pharmacological agents which was noted as a decrease in both circularity factor and shape factor values. Activation caused the cells to become elliptical, a feature that appears to be characteristic of Leucophaea immunocytes. Administration of DAMA induced a similar activation of most of the cells. After 30 min these DAMA-activated cells started to display distinct locomotory activity not seen in the controls. alpha-Melanocyte-stimulating hormone (MSH, 10(-7)) added to the incubation medium after DAMA-activation caused the cells to return to their original "rounded" conformation. In addition, the presence of immunoreactive interleukin (IL-1), adrenocorticotropin (ACTH) and tumor necrosis factor (TNF) in the hemolymph was demonstrated. These data suggest an interaction between both vertebrate-type immunological signal molecules and neuropeptides in the regulation of immunological cells in Leucophaea.

Anabolic androgenic steroids (AS) have recently been placed on the Food and Drug Administration's (FDA's) list of controlled substances, because of the adverse effects seen in athletes taking accelerated dosages in attempts to enhance performance. Reported deleterious effects on abusers include sterility, gynecomastia in males, acne, balding, psychological changes, and increased risks of heart disease and liver neoplasia. Considering the roles of the immune and neuroendocrine systems and their interactions in many of these pathologies, it is important to determine the effects of these derivitized androgens on this connection. Little is known in this respect. We therefore determined the effects of anabolic steroids on certain immune responses and their effects on the extrapituitary production of corticotropin by lymphocytes. We present evidence that (1) both 17-beta and 17-alpha esterified AS, nandrolone decanoate and oxymethenelone, respectively, significantly inhibited production of antibody to sheep red blood cells in a murine abuse model; (2) the control androgens testosterone and dehydroepian-drosterone (DHEA) or sesame seed oil vehicle had no significant effects on antibody production; (3) nandrolone decanoate and oxymethenelone directly induced the production of the inflammatory cytokines IL-1 beta and TNF-alpha from human peripheral blood lymphocytes but had no effect on IL-2 or IL-10 production; (4) control androgens had no direct cytokine inducing effect; (5) nandrolone decanoate significantly inhibited IFN production in human WISH and murine L-929 cells; and (6) nandrolone decanoate significantly inhibited the production of corticotropin in human peripheral blood lymphocytes following viral infection. These data indicate that high doses of anabolic steroids can have significant effects on immune responses and extrapituitary production of corticotropin. Furthermore, the mouse model should provide an effective means by which to study other deleterious effects of anabolic steroid abuse in humans.

There is general agreement that IFN-gamma is produced only by cells of immune origin (T-cells, NK cells, and recently macrophages). However, indirect evidence has suggested that undetectable, low levels of IFN-gamma produced by cells of non-immune lineage, such as the murine line L-929, enhanced the antiviral activity of IFNs-alpha and/or beta following induction by agents such as the double stranded RNA poly ICLC. Since L-929 cells were one of the prototypic cell lines for studying murine IFN induction and action, we felt that it would be important to validate this observation by detection of the mRNA for IFN-gamma. If confirmed, it might indicate a role for IFN-gamma in non-immune cells. The present investigations revealed that mouse L-929 fibroblasts produce IFN-gamma message following exposure to conventional IFN-alpha/beta inducers such as poly ICLC or Newcastle disease virus. In addition, we found that IFN-gamma itself will induce its own message. We further show that this is not a phenomenon isolated to transformed cells since we found that normal mouse embryo fibroblasts also produced the message, however in a constitutive fashion.

Kaposi's sarcoma (KS) accounts for more than 15% of AIDS-related malignancies. The etiology of KS is unresolved but is postulated to be multifactorial, involving viruses and overexpression of cellular growth factors and/or oncogenes. Recently, herpesvirus-like sequences (KSHV) were identified with high prevalence in AIDS-KS (AKS), endemic KS, and in classic KS biopsies (CKS). To confirm the presence and the prevalence of the KSHV sequences, 18 CKS and 13 AKS samples were tested using polymerase chain reaction (PCR) analysis. To our knowledge this is the highest number of CKS samples that has ever been included in a single study, and it is also important that the biopsies were obtained from different institutions and geographical locations. KSHV sequences were detected in 100% of the AKS samples and 72% of the CKS biopsies using PCR analysis. The presence of the unique KSHV sequences was confirmed by direct sequencing of representative PCR products obtained from AKS and CKS samples. Reverse transcriptase (RT)-PCR experiments showed that the KSHV sequences were transcribed to mRNA in both AKS and CKS samples. Our results confirm that the putative new herpesvirus-like agent is associated with both AKS and CKS and may have an etiological role in the pathogenesis of this malignancy.

Objective: Our purpose was to determine whether interleukin-10 is expressed in human placental tissue, which might imply a role for it in fetal allograft protection., Study Design: Detection of interleukin-10 messenger ribonucleic acid in human placental tissue and in human placental JAR cells by reverse transcription-coupled polymerase chain reaction was studied., Results: Interleukin-10 messenger ribonucleic acid was detected in human placental tissue from term mothers and in human placental JAR cells. Sequence analysis of the expected interleukin-10 complementary deoxyribonucleic acid fragment revealed 100% homology to authentic interleukin-10 complementary deoxyribonucleic acid., Conclusion: Our results indicated that human placental tissue from term mothers expressed high levels of interleukin-10 messenger ribonucleic acid, suggesting that cells that produce interleukin-10 and that are associated with the placenta may play a role in preventing rejection of the fetal allograft by the mother.

Herpesvirus-like DNA sequences (KSHV) have been reported to be associated with various forms of Kaposi's sarcoma (KS). To determine if KSHV was associated with other proliferative skin lesions from non-AIDS immunocompromised patients, 33 skin lesions (basal cell carcinomas, squamous cell carcinomas, actinic keratoses, verruca vulgaris, atypical squamous proliferations, and seborrhoeic keratosis) from 4 organ-transplant patients receiving immunosuppressive therapy were tested for KSHV by PCR. KSHV sequences were detected in 82% of these skin lesions. Our results suggest that KSHV is associated with lesions other than KS in non-AIDS immunocompromised patients, and may also be involved in the pathogenesis of the various forms of proliferative skin lesions from organ-transplant patients.

1. Recent data have shown that interleukin-10 (IL-10) is expressed and acts in mouse pituitary tumor cells and freshly isolated mouse pituitaries. 2. In this study, we show that poly(A+) RNA derived from normal human pituitary and hypothalamus expresses IL-10 message. 3. The majority of transcripts was likely from the pituitary and hypothalamus, and not from lymphocytes in the pituitary and hypothalamic vasculature, since both IL-10 and interferon-gamma mRNA levels, compared to equivalent amounts of RNA from peripheral blood lymphocytes, were much lower. 4. These results indicate that IL-10 may function in human neuroendocrine process as it does in the murine system, thus serving as an important signal molecule for bidirectional communication between the neuroendocrine and the immune systems.

Objective: The trauma of cardiopulmonary by-pass (CPB) in cardiac surgery results in a whole body diffuse inflammatory response characterized in part by hyperstimulation of leukocytes. Partially this is due to an increase in the release of biological response modifiers such as cytokines, as noted by the immunocyte stimulatory actions of cell-free plasma obtained postoperatively from CPB patients. The present study was conducted to determine whether CPB plasma induced immunocyte hyperstimulation can be prevented with naturally occurring immune inhibitory substances, specifically, interleukin (IL)-10 and/or morphine., Experimental Design: Controlled in vitro study of the application of drugs to naive immunocytes to block the exitation caused by CPB-plasma., Setting: University-based tertiary care hospital., Patients: Plasma was obtained from ten patients undergoing CPB. Eligibility included admission for elective cardiac surgery, which no chronic illnesses or acute processes., Interventions: Monocytes and granulocytes were pretreated with IL-10 and/or morphine before exposure to plasma obtained from patients undergoing CPB, as CPB-plasma would stimulate naive monocytes and granulocytes in a manner similar to that previously reported in CPB-patients., Measures: Computer-assisted microscopic image analysis, measuring cellular conformational and velocity changes, was used to evaluate the effect of treatment on the immunocytes response to stimulation with CPB-plasma., Results: Pretreatment of cells with IL-10 and/or morphine significantly diminished the hyperstimulation induced by CPB-plasma in a concentration-dependent manner. In contrast, when the cells were initially or simultaneously exposed to CPB-plasma, IL-10 and/or morphine had no effect.

Tumor necrosis factor (TNF) is a monokine produced primarily by macrophages. TNF has a number of activities including direct lysis of certain transformed cells and induction of antiviral activity. One of the protoypical transformed cell lines used for studying TNF cytolysis is murine L-929 cells. Because of the lysis, TNF has not been shown to have antiviral activity in these cells. Since retinoic acid (RA) induces a normal phenotype in the L-929 cells, we sought to determine if their conversion to a normal phenotype would 1) render them insensitive to the cytolytic effect and 2) allow for the development of an antiviral state. We present evidence that both the cis- and trans- forms of RA and to a lesser extent, the RA precursor beta-carotene, can inhibit recombinant human TNF cytolytic activity in mouse L-929 cells. However, blockage of the cytolytic activity does not allow development of an antiviral state.

The effect of interleukin-4 (IL-4) was studied on voltage-gated Ca-current of the neuron RPeD1 of Lymnaea stagnalis L. (Mollusca, Basommatophora), using two-microelectrode voltage-clamp method. It was found that: 1. The neuron RPeD1 possessed a high voltage-activated Ca-current characterized by -40 mV activation threshold and reaching its maximum at +5+5(-)+10 mV. The activation time constant of the current was found to be a monnexponential function of the membrane potential. Currents were almost completely inactivated within 200-300 ms. 2. IL-4 (10-200 U/ml) uniformly and reversibly increased the peak value of Ca-current in a dose- and time-dependent manner (at concentrations 75, 150 and 200 U/ml the potentiation was 5.3 +/- 1.8% (n = 3), 5.5 +/- 1. (n = 3) and 14.6 +/- 7.4% (n = 4), respectively. 3. No changes in the membrane resistance and holding current were observed during IL-4 application. 4. The time constants of activation and inactivation were not affected by IL-4. The maximal conductance as the only kinetic parameter was increased under the influence of IL-4. 5. The affect in modulation of HVA Ca-current caused by IL-4 was additive between the doses of 10-15o U/ml but saturated at concentrations > 200 U/ml. 6. IL-4 antiserum was a potent inhibitor for enhancement of Ca-current by IL-4.

We have developed a novel method to study the subtype-specific expression of interferon-alpha (IFN-alpha) in the mouse system: we synthesized and used consensus oligonucleotide primers to allow simultaneous polymerase chain reaction (PCR) amplification of multiple murine IFN-alpha gene sequences. In addition, a set of subtype-specific oligomer probes were designed and used to distinguish between IFN-alpha genes that differ by only a few bases. The consensus primers, corresponding to two regions highly conserved among murine IFN-alpha subtypes, were used in reverse transcription and PCR amplification of total cellular RNA, isolated from IFN-gamma-treated murine L-929 cells, to yield a fragment of the anticipated approximately 520-bp size. Southern analysis of the amplified product using an internal consensus oligomer probe confirmed specific amplification of murine IFN-alpha gene(s). Subtype-specific oligonucleotide probes indicate that IFNs-alpha 1, -alpha 2, and -alpha 5 are present following IFN-gamma treatment, whereas IFN-alpha 4 remains virtually absent. Our results indicate that the expression of specific IFN-alpha subtypes may be subject to complex regulation, dependent upon inducing agents and cell types involved, and countless other factors. The procedure described here represents a novel method for studying the subtleties of the murine IFN-alpha mRNA expression.

1. Interleukin-10 (IL-10) has a wide range of activities in the immune system such as modulation of interferon-gamma (IFN-gamma) and antibody production. The neuropeptide hormone corticotropin (ACTH) has similar activities, suggesting that a bidirectional communication mechanism operates between the immune and the neuroendocrine system involving these two substances. 2. Murine pituitary tumor cells (AtT-20) were found to produce up to 3 ng/ml of IL-10. 3. Pituitary cell corticotropin production was enhanced by IL-10 treatment. 4. IL-10 induced the production of ACTH in mouse splenocytes. 5. Authenticity of pituitary-derived IL-10 was shown by the demonstration of identical nucleic acid sequences of reverse-transcribed, polymerase chain reaction amplified fragments of cDNA obtained from murine splenocytes, a murine pituitary tumor cell line, and freshly isolated murine pituitaries.

The effects of a synthetic peptide fragment of human immunodeficiency virus gp120 (HIV gp120) on opioid (D-ala2-D-met5 enkephalinamide; DAMA) and interleukin-1 (IL-1) induced chemotactic responses in human granulocytes and monocytes and invertebrate (Mytilus edulis) immunocytes were studied. Both DAMA and IL-1 increased the velocity of cell migration from both species and the response is chemotactic (e.g. directed). Non-treated control cells move randomly or not at all. The addition of gp120 to DAMA or IL-1 treated human granulocytes or monocytes results in a slower movement which is chemokinetic (loss of directionality or random) in nature. A similar phenomenon occurs in the invertebrate immunocytes. If gp120 alone is added, it inhibits the movement of spontaneously active human granulocytes and Mytilus edulis immunocytes. In contrast, it stimulates chemokinesis of spontaneously active human monocytes. These responses occur immediately after addition of the peptide. Based on experiments with the selective calcium channel antagonist nimodipine, it appears that the gp120 causes its effects by irreversible binding to a calcium channel. Our results suggest a universal inhibitory mechanism is occurring since the invertebrate immunocytes must recognize HIV gp120 peptide to result in this effect, possibly through a CD4 or other type of surface determinant.

1. Mytilus pedal ganglion contains a small population of glial cells that are immunopositive for interleukin-1 alpha. Positively stained fibers can also be seen in the neuropil of these sections. 2. The marine worm Nereis diversicolor also exhibits positive neural immunostaining for interleukin-1 alpha. 3. Both organisms contain hemocytes that contain immunoactivity for interleukin-1 alpha. The study suggests interleukin-1 alpha to be an ancient cytokine given its presence in organisms that evolved significantly earlier than mammals.

1. Corticotropin-releasing factor (CRF) appears to be a potentially important signal molecule in both vertebrate and invertebrate neuroimmune and autoimmunoregulatory processes. It appears to mimic the effects of alpha-melanocyte stimulating hormone (MSH) but has a longer duration of action. 2. alpha-Helical CRF, a specific inhibitor of CRF, antagonizes CRF-induced cellular immunosuppression but is ineffective in altering MSH-induced immunosuppression. 3. Both human and Mytilus immunocytes appear to have specific CRF receptors. 4. In another experiment, both CRF and MSH antagonize tumor necrosis factor stimulation of immunocytes. Again, alpha-helical CRF antagonizes only CRF activity, further suggesting the presence of a separate CRF receptor on these cells.

Mytilus edulis, a marine bivalve mollusc, contains hemocytes that share certain properties with cells of the human monocyte-macrophage lineage. Because of this and previously reported similar responses to opioids between Mytilus hemocytes and human granulocytes, we determined whether Mytilus might also possess monokines or an immune monokine-like network. We chose to study two monokines, interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF). We found that Mytilus hemocytes respond to recombinant human IL-1 and TNF in a fashion similar to human granulocytes. Furthermore, we show that as in the human system, it appears that IL-1 mediates at least part of its effects through immunoreactive (ir) TNF in Mytilus. Finally, we show that Mytilus hemolymph contains ir IL-1 and TNF. To our knowledge, this is the first demonstration of these substances in this system.

We review evidence that an immune and nervous system linkage exists in invertebrates similar to that shown in vertebrates. Immunoreactive cytokines appear to play a role in this interaction. We have demonstrated that M. edulis, a marine bivalve mollusc, reacts to the vertebrate monokines interleukin-1, -6 and TNF. We have also demonstrated endogenous immunoreactive interleukin-1, -6 and tumour necrosis factor in M. edulis hemolymph, immunocytes and pedal ganglia. Our findings suggest an important common signalling mechanism that has been conserved over 500 million years of evolution.

The immune and neuroendocrine systems communicate and maintain homeostasis through various mechanisms, including the use of common signal and recognition molecules and the use of similar processes. This type of integrated network has profound effects on the onset and outcome of certain disease states, including endotoxic shock, in which a cascade of mediators influence the pathophysiologic responses. We have found that some of the common signal molecules shared between the immune and neuroendocrine systems are the peptide hormones adrenocorticotropin (ACTH) and endorphins (END). Our investigations have shown that these molecules are produced in vitro by cells of the immune system treated with various stimuli, including immunological stimuli such as bacterial lipopolysaccharide (LPS; endotoxin), virus infection (Newcastle virus; NDV), and the more classical neuroendocrine stimuli corticotropin-releasing hormone (CRH). We have proposed that the production of END by the peripheral immune system contributes to the pool of opioid peptides associated with the pathophysiology of endotoxic shock. Lymphocytes from LPS-sensitive C3HeB/FeJ mice but not LPS-resistant C3H/HeJ mice produce END and ACTH both in vitro and in vivo after treatment with LPS. Purification of the in vitro produced LPS-induced END from B-lymphocyte spleen cells followed by injections into both LPS-sensitive and -resistant mice elicits changes in body temperature and respiration rate. The spleen cells from the LPS-sensitive mice process ACTH and END differently depending on the stimulus for induction and the cell type in which the processing takes place. CRH or virus induce ACTH 1-39 and beta-END, whereas inductions with LPS yield major products of ACTH 1-22 to 1-26 and gamma-END, products that are for the most part unique to the immune system. We have shown that LPS induces a novel protease that functions optimally at pH 5 to cleave ACTH 1-39 into ACTH 1-22 to 1-26. This enzyme is present in LPS, but not mock or CRH-induced B cells from LPS-sensitive mice. The LPS-resistant mice did not possess this enzyme and therefore produced only the high-molecular-weight pro-opiomelanocortin (POMC)-like molecule. The inability to produce ACTH and END, presumably by their inability to process the precursor, may account, in part, for their lack of response to the LPS. The POMC peptides also may play an indirect role in orchestrating the pathophysiologic response, since both ACTH and END were shown to induce tumor necrosis factor (TNF). Our data strongly suggest that lymphocyte POMC peptides ACTH and END are important mediators in the overall response to endotoxin.

The ability of interferons (IFN) to exert a systemic effect following their oral administration was evaluated. One systemic effect of parenteral interferon administration has been shown to be a suppression of the number of peripheral white blood cells both in man and in mouse models. Using the mouse model of peripheral white blood cell suppression, the relative systemic effects of orally and subcutaneously administered interferons were determined. Murine IFN-beta, murine IFN-gamma and cross-reactive recombinant human IFN-alpha A/D were examined. The oral administrations of each of the three interferons were found to cause a dose-dependent suppression of the peripheral white blood cell counts. Significant levels of suppression were seen with as little as 5 units/day of murine IFN-beta and with 500 units/day of recombinant human IFN-alpha A/D and murine IFN-gamma. The dose-response curves obtained with orally administered interferons were much more shallow than those obtained with subcutaneously administered interferons. The results demonstrate that oral administration of interferons can provide a significant systemic effect. Further, the results support the possibility that the oral administration of interferons may have therapeutic potential.

The synthetic analog of methionine enkephalin, [D-Ala2-Met5]-enkephalin, when administered in vitro to Mytilus edulis ganglia and hemocytes and human peripheral blood lymphocytes, induces the formation of an immunoreactive interleukin-1-like molecule. Additionally, immunoreactive interleukin-1 (IL-1) activity has been found in Mytilus nervous tissue. The stimulatory actions of the extracted immunoreactive IL-1 on Mytilus hemocytes can be antagonized by an IL-1 antibody demonstrating the specificity of the substance. The evidence suggests that the nervous system, via an opioid-IL-1 relationship, can communicate with the immune/defense system through these similar signal molecules. Furthermore, the results indicate that an interleukin-like molecule must have evolved earlier than previously thought.

Mytilus edulis hemocytes have similarities with vertebrate monocyte/macrophages. We have recently shown that they respond to human TNF and IL-1. We tested the possibility that Mytilus hemocytes produce similar substances in response to LPS. We show that Mytilus hemocytes respond to LPS in a fashion similar to vertebrate monocytes and macrophages and that these responses are inhibited by antibodies to TNF and/or IL-1. These findings are demonstrated both in vitro and in vivo.

The modulatory effects of interleukin-1 (IL-1), an immunotransmitter, and FMRFamide, a molluscan neuropeptide, were studied on identified neurons of Helix pomatia L. (Mollusca, Gastropoda) by using the method of two microelectrode voltage clamp. IL-1 and FMRFamide uniformly decreased the voltage-activated inward current (ICa), while the voltage-dependent outward potassium current (Inet K) increased. IL-1 and FMRFamide were shown to use the same cellular targets as used by the low molecular weight neurotransmitters.

Our views of interferon production and action have evolved from the simple to the complex. We review evidence that interferon-gamma (IFN-gamma) may induce a portion of its antiviral activity through the induction of another interferon. This is shown by demonstrating IFN-alpha in the supernatant fluids of IFN-gamma-treated mouse cells and showing that, under certain conditions, the antiviral activity of IFN-gamma in mouse and human cells can be reduced by antibody to IFN-alpha or IFN-beta, respectively. Induction of mouse spleen, bone marrow, and peritoneal exudate cells also results in their production of IFNs-alpha and/or -beta. In addition, new preliminary data indirectly suggests the presence of IFN-gamma in poly(IC):poly(LC)-treated mouse cells. In both of these systems, the maximal antiviral activity appears to develop as a consequence of the induction of a second interferon.

Tumor necrosis factor (TNF) potentiates the antiviral activity of interferon-gamma (IFN-gamma) directly and through induction of IFN-beta in HEp-2 cells. Antibody to IFN-beta inhibits a large component of the TNF and IFN-gamma synergy and immunoreactive IFN-beta can be detected in the supernatant fluids of TNF-treated HEp-2 cells. Furthermore, the kinetics of development of this synergy and its susceptibility to inhibition by antibody to IFN-beta is demonstrated.

The finding that interferon-gamma (IFN-gamma) may require two rounds of protein synthesis to induce the antiviral state raises the possibility that this IFN may not be directly antiviral. We, therefore, examined the possibility that IFN-gamma induces one or both of the other IFNs (alpha and/or beta) which in turn induce the antiviral state. Evidence is presented showing that under certain conditions a large portion of IFN-gamma's antiviral activity in mouse L-929 cells is mediated by its induction of IFN-alpha based on the findings that: 1) the antiviral activity of IFN-gamma in cells at low densities can be blocked by poly and monoclonal antibody to IFN-alpha and, 2) IFN-alpha can be demonstrated in the supernatant fluids of IFN-gamma treated cells. This report raises the possibility that a major antiviral mechanism of IFN-gamma is via induction of IFN-alpha in the mouse system. If the majority of the antiviral activity of IFN-gamma is via induction of other IFNs, then the role and mechanism of IFN-gamma might have to be reevaluated.

Nonsensitized murine lymphocytes produced mouse interferon when cocultured with foreign and transformed cells. Neutralization tests revealed that this interferon was of the alpha/beta type. The B lymphocyte appeared responsible for the bulk of the interferon production as determined by cell "panning," complement-mediated depletion, and immunofluorescence experiments.

Human milk was found to contain chemokinetic agents for human blood monocytes. The chemokinetic agents were whey proteins that were inactivated by heating at 56 degrees C for 20 min or treatment with trypsin. Three peaks of chemokinetic activity less than 60 kD in size were found by gel filtration chromatography. The chemokinetic activity of each peak obtained by gel filtration was partially blocked by polyclonal rabbit antibodies to recombinant human tumor necrosis factor-alpha (TNF-alpha). TNF-alpha, or a protein that immunologically cross-reacted with it, was also detected in human milk by blockage of the cytotoxicity of human milk by anti-TNF-alpha. Such proteins or others that elicit the release of TNF-alpha from the target cells may be responsible for the enhanced motility of human milk macrophages, and it is possible that they may alter the immunologic or metabolic activities of the alimentary tract of the recipient infant.

The possible chemokinetic effect of human tumor necrosis factor alpha (TNF-alpha) upon human monocytes was investigated by using a type 1 collagen gel system. Human recombinant TNF-alpha was found to be chemokinetic, and concentrations of that cytokine as low as 0.7 pg/microliters enhanced the movement of those leukocytes. Furthermore, we explored whether endogenous TNF-alpha stimulates monocyte movement by incubating blood monocytes with a TNF-alpha-stimulating agent, human recombinant interleukin 1, with or without antibodies to human TNF-alpha. Human recombinant interleukin 1 enhanced the motility of blood monocytes, and that effect was blocked by rabbit polyclonal antibodies to human TNF-alpha. Furthermore, the movement of blood monocytes no exposed to exogenous cytokines was decreased after incubation with anti-TNF-alpha. Thus, endogenous as well as exogenous TNF-alpha is a chemokinetic agent for human monocytes.

Experiments were performed to determine the effects of hydrazine sulfate (HS), a potential anti-cachexia agent, on tumor necrosis factor activities (cachectin/TNF) in vitro. We present evidence that HS significantly inhibits the lytic activity of TNF on L-929 cells, that HS has no direct effect on TNF itself, and that the minimum amount of time for maximum inhibition of TNF activity to occur after HS treatment is between 1 and 4 h. In addition to HS's effect on the cytolytic activity, we also determined its effect on the antiviral activity of TNF. We found that HS greatly potentiates TNF antiviral activity, while having no significant antiviral activity itself over a range of concentrations, that the potentiation was likely between HS and TNF-induced interferon-B1, and was maximal following 4 and 8 h of treatment with HS. Although the lytic activity of TNF has not been directly correlated with its cachectic activities, these studies provide evidence for an effect of HS on cachectin and its role in the wasting process. Furthermore, a rationale is provided for use of HS in conjunction with TNF for prevention and/or treatment of viral infection.

The finding that monocytes possess specific ACTH receptors led us to determine the effect of ACTH on monokine production. Here, we report that ACTH 1-39 will induce TNF-alpha from the adherent fraction of peripheral blood leukocytes. In addition, we also found that ACTH 1-39 will potentiate IFN-gamma's induction of TNF-alpha from these cells. Since previous reports showed that ACTH inhibits IFN-gamma's activation of macrophages to a cytocidal state, our data raise questions concerning the relative role of TNF-alpha in macrophage mediated cytolysis.

Retinoic acid (RA) inhibited transformed mouse L-929 and human WISH cell induction of interferon alpha/beta production by nonsensitized mouse spleen cells. The RA effect was both time and dose dependent and acted in near physiologic concentrations. The results suggest that the effect is due to a modulation of a previously described transformed cell surface associated glycoprotein IFN inducer.

Ia antigens were previously shown to be B-cell receptors for transformed cell surface glycoproteins (TCSG). Ia antigen recognition of TCSG subsequently initiated interferon production. These findings have been exploited to highly purify TCSG from mouse L (H-2k) cells in one step by Iak antigen Sepharose affinity chromatography. These results point to the highly specific recognition of TCSG by Ia antigens and the B-cells on which they reside. The implications and applications of these studies are discussed.