Interleukin 1: an important mediator of host resistance against Pneumocystis carinii.

The importance of endogenous interleukin 1 (IL-1) in resistance to Pneumocystis carinii infection was examined in a SCID mouse model. Naturally acquired pulmonary infection of P. carinii in SCID mice was completely cleared by reconstitution of the infected mice with immunocompetent spleen cells. IL-1 activity in the lung homogenate supernatant of these mice increased significantly after reconstitution and returned to baseline level after the clearance of P. carinii. Treatment of reconstituted SCID mice with 35F5, a monoclonal antibody against murine type I IL-1R almost completely inhibited the clearance of P. carinii. In contrast, treatment with control rat immunoglobulin G had no detectable effect. Further study revealed that for the complete clearance of P. carinii, IL-1 must be present at the early stage of immune responses induced by reconstitution, since clearance could be blocked by a single injection of 35F5 into SCID mice at 2 d, but not at either 8 or 13 d postreconstitution. Furthermore, pulmonary recruitment of neutrophils, macrophages, and lymphocytes was significantly inhibited in mice that received 35F5 treatment. These findings strongly suggest that, in reconstituted SCID mice, endogenous IL-1 is important in host resistance to P. carinii infection and that IL-1 may function early in the host response possibly by the recruitment of inflammatory cells into the lungs.

Role of cytokines (interleukin 1, tumor necrosis factor, and transforming growth factor beta) in natural and lipopolysaccharide-enhanced radioresistance.

Studies of radioresistance and radioprotection provide an excellent in vivo model for dissection of the pathophysiological role of cytokines. The availability of neutralizing antibodies to cytokines has made it possible to assess the contribution of cytokines to host defense and repair processes involved in radioresistance and radioprotection. Administration of anti-interleukin 1 receptor (IL-1R) antibody (35F5) or anti-tumor necrosis factor (TNF) antibody (TN3 19.12) reduced survival of irradiated CD2F1 mice. These results demonstrate conclusively that natural levels of IL-1 and TNF contribute to radioresistance of normal mice. Furthermore, the radioprotective effect of administered IL-1 was blocked not only with anti-IL-1R antibody but also with anti-TNF antibody. Similarly, the radioprotective effect of TNF was reduced with anti-IL-1R antibody. These data suggest that cooperative interaction of both cytokines is necessary to achieve successful radioprotection. Finally, when LPS was used as a radioprotector, the combined administration of anti-IL-1R and anti-TNF not only blocked the radioprotection with LPS, but actually revealed LPS to have a radiosensitizing effect. This effect may be due to induction of TGF-beta, since administration of this cytokine results in reduced survival of irradiated mice.

Human epidermal Langerhans cells express the high affinity receptor for immunoglobulin E (Fc epsilon RI).

It has been suggested that epidermal Langerhans cells (LC) bearing immunoglobulin E (IgE) may be involved in the genesis of atopic disease. The identity of the IgE receptor(s) on LC remained unclear, although it represents a crucial point in understanding cellular events linked to the binding of allergens to LC via IgE. In this report, we demonstrate that epidermal LC express the high affinity receptor for the Fc fragment of IgE (Fc epsilon RI) which has, so far, only been described on mast cells and basophils. Epidermal LC react with antibodies specific for the alpha subunit of the tetrameric (alpha, beta, 2 gamma) Fc epsilon RI. Specific transcripts for Fc epsilon RI alpha and Fc epsilon RI gamma were detected in LC and correspond to those of human basophils and of the human basophil cell line KU812. Furthermore, human basophils, KU812 cells, and LC express the putative beta subunit. Thus human LC express the complete structure of Fc epsilon RI. This finding opens new perspectives in the putative functional role of this structure on antigen-presenting cells.

Interleukin 12, interferon gamma, and tumor necrosis factor alpha are the key cytokines of the generalized Shwartzman reaction.

The Shwartzman reaction is elicited by two injections of lipopolysaccharide (LPS) in mice. The priming LPS injection is given in the footpad, whereas the lethal LPS challenge is given intravenously 24 h later. The injection of interferon gamma (IFN-gamma) or interleukin 12 (IL-12) instead of the LPS priming injection induced the lethal reaction in mice further challenged with LPS. Antibodies against IFN-gamma when given together with the priming agent, prevented the lethal reaction in mice primed with either LPS, IL-12, or IFN-gamma. Antibodies against IL-12, when given together with the priming agent, prevented the lethal reaction in mice primed with either LPS or IL-12 but not with IFN-gamma. These results strongly suggest that LPS induces the release of IL-12, that IL-12 induces the production of IFN-gamma, and that IFN-gamma is the cytokine that primes macrophages and other cell types. Upon LPS challenge, the lethal Shwartzman reaction is induced by a massive production of inflammatory cytokines that act on the target sites already sensitized by IFN-gamma. If mixtures of TNF and IL-1 or mixtures of TNF and IFN-gamma are used to challenge mice previously primed with IFN-gamma or IL-12, mortality is induced. In the same conditions, the individual cytokines or a mixture of IL-1 and IFN-gamma do not replace the LPS challenge. When the mice are primed with LPS, the combination of TNF, IL-1, and IFN-gamma induced only a partial mortality incidence suggesting that the involvement of other LPS-induced factors.

Inhibition of interleukin 1 (IL-1) binding and bioactivity in vitro and modulation of acute inflammation in vivo by IL-1 receptor antagonist and anti-IL-1 receptor monoclonal antibody.

Recombinant human interleukin 1 receptor antagonist (IL-1ra) and 35F5, a neutralizing monoclonal antibody (mAb) to the type I mouse IL-1 receptor, were examined for their ability to bind to IL-1 receptors (IL-1Rs) on various types of mouse cells and to block immune and inflammatory responses to IL-1 in vitro and in mice. IL-1ra competed for binding of 125I-IL-1 alpha to type I IL-1R present on EL-4 thymoma cells, 3T3 fibroblasts, hepatocytes, and Chinese hamster ovary cells expressing recombinant mouse type I IL-1R. The IC50 values for IL-1ra binding (ranging from 2 to 4 ng/ml) were similar to those of IL-1 alpha. In contrast, IL-1ra bound with very low affinity (IC50 values ranging from 10 to 200 micrograms/ml) to cells expressing type II IL-1R, i.e., 70Z/3 pre-B cell line and polymorphonuclear leukocytes (PMN) derived from bone marrow and acute inflammatory exudates. The mAb 35F5 bound specifically to type I IL-1R; no inhibition of 125I-IL-1 alpha binding to cells having type II IL-1R was observed with very high concentrations of antibody. While neither IL-1ra nor 35F5 had intrinsic activity in bioassays using T helper D10.G4.1 cells and mouse thymocytes, both agents blocked the ability of IL-1 to stimulate proliferation of these cells. The effects of IL-1ra and 35F5 on acute inflammatory responses in mice were also evaluated. IL-1ra and 35F5 blocked the local accumulation of PMN after intraperitoneal injection of rIL-1 alpha. The response to IL-1 was inhibited when IL-1ra or 35F5 was administered simultaneously with or before administration of IL-1. IL-1ra and 35F5 also blocked PMN accumulation after intraperitoneal injection of lipopolysaccharide or proteose peptone, suggesting IL-1 is important in mediating responses to these agents. In addition, IL-1ra and 35F5 significantly blocked the ability of IL-1 to stimulate egress of PMN from bone marrow, to induce a transient neutrophilia, and to elevate serum levels of hepatic acute phase proteins, IL-6, and corticosterone. Thus, IL-1ra and 35F5 competitively inhibit the binding of IL-1 to the IL-1R on certain cell types. These two IL-1 receptor antagonists act to inhibit biological responses induced by IL-1 and other inflammatory agents.

Novel interleukin-2 receptor subunit detected by cross-linking under high-affinity conditions.

Interleukin-2 (IL-2) binds to both high- and low-affinity classes of IL-2 receptors on activated T lymphocytes. Only the high-affinity receptors are involved in receptor-mediated endocytosis and normally transduce the mitogenic signals of IL-2; however, the structural features distinguishing the high- and low-affinity receptors are unknown. When 125I-labeled IL-2 was chemically cross-linked to activated human T lymphocytes, two major bands were identified. First, as predicted, a 68- to 72-kilodalton band, consisting of IL-2 (15.5 kilodaltons) cross-linked to the IL-2 receptor (55 kilodaltons), was observed. Second, an unpredicted 85- to 92-kilodalton moiety was detected. This band was not present when IL-2 was cross-linked to transfected C127 cells, which exclusively express low-affinity receptors. The data presented are most consistent with the existence of a 70- to 77-kilodalton glycoprotein subunit (p70) which, upon associating with the 55-kilodalton low-affinity receptor (p55), transforms it into a high-affinity site. It is proposed that p55 and p70 be referred to as the alpha and beta subunits, respectively, of the high-affinity IL-2 receptor.

Recombinant interleukin-12 suppresses the synthesis of immunoglobulin E by interleukin-4 stimulated human lymphocytes.

Interleukin-12 is a recently discovered lymphokine displaying an array of in vitro activities suggesting a major role in protective immunity against infectious agents like viruses. This study provides evidence that IL-12 may also be implicated in the selection of the immunoglobulin isotypes. We show that picomolar concentrations of rIL-12 markedly inhibit the synthesis of IgE by IL-4-stimulated PBMC. The suppression of IgE is observed at the protein and at the mRNA levels, it is isotype specific, and it is abolished by neutralizing anti-IL-12 mAbs. IL-12 may suppress IgE synthesis by: (a) inducing the production of IFN-gamma, a known inhibitor of IgE synthesis and (b) by a novel mechanism which is IFN-gamma independent. The best evidence for this is from studies on IgE synthesis by IL-4-plus hydrocortisone-stimulated umbilical cord blood lymphocytes, which do not produce detectable amounts of IFN-gamma. In such cultures, rIL-12 inhibits IgE synthesis even in the presence of a large excess of neutralizing anti-IFN-gamma mAb.

Preferential localization of systemically administered radiolabeled interleukin 1alpha in experimental inflammation in mice by binding to the type II receptor.

Previously, we have shown that systemically administered radiolabeled interleukin 1alpha (IL-1alpha) accumulates preferentially in inflammatory foci in mice. Since inflammation is characterized by influx of leukocytes, which represent IL-1 receptor (IL-1R) positive cells, radiolabeled IL-1 may specifically localize in inflammation by binding to its receptors on infiltrated leukocytes. This hypothesis was tested in a series of studies in mice with acute focal inflammations. Evidence for specific IL-1-IL-1R interaction in induced inflammation was found: microscopic autoradiography revealed that 125I-IL-1alpha localized at the site of inflammatory cells with time; 125I-myoglobin, a similar-sized protein with no known interactions in vivo, was not retained in the inflammation. Furthermore, the uptake 125I-IL-1alpha in inflammatory tissue was significantly lower in neutropenic mice than in immunocompetent mice (0.05+/-0.004 vs. 0.65+/-0.06% ID/g at 48 h after injection, P < 0.0007). Moreover, the uptake of 125I-IL-1alpha at the inflammatory site could be blocked with the anti-IL-1R type II antibody 4E2. At 48 h after injection, the uptake with and without blocking the type II IL-1R was 0.13+/-0.01 and 0. 65+/-0.05% ID/g, respectively (P < 0.0001). These in vivo studies provide evidence that systemically administered radiolabeled IL-1alpha localizes in inflammatory tissue by specific receptor binding, predominantly by binding to the type II IL-1R.

Regulation of the human c-myc gene: 5' noncoding sequences do not affect translation.

The influence of untranslated 5' sequences on c-myc expression was compared by measuring the translational efficiencies of mRNAs which contain leaders derived from exon 1 or intron 1 of the human c-myc gene. Expression plasmids were constructed and introduced into COS cells, and the levels of c-myc mRNA and protein were examined. Our results show that mRNAs transcribed from constructs containing exon 1 or intron 1, which have different folding potential, are translated with approximately equal efficiencies. This suggests that the translation of c-myc mRNA is not controlled by secondary structure alone. In addition, we observed that transcripts in which exon 1 was deleted are not translated more efficiently, but are present at a higher steady-state level. Thus, this example provides evidence for possible control at the transcriptional level. Finally, since the c-myc product was produced in each of our test systems, the results suggest that this protein does not regulate its own transcription or translation via a specific interaction with c-myc exon 1 alone.

Production of human c-myc protein in insect cells infected with a baculovirus expression vector.

A cDNA fragment coding for human c-myc was inserted into the genome of the baculovirus Autographa californica nuclear polyhedrosis virus adjacent to the strong polyhedrin promoter. Insect cells infected with the recombinant virus produced significant amounts of c-myc protein, which constituted the major phosphoprotein component in these cells. By immunoprecipitation and immunoblot analysis, two proteins of 61 and 64 kilodaltons were detected with c-myc-specific antisera. The insect-derived proteins were compared with recombinant human c-myc-encoded proteins synthesized in Escherichia coli and Saccharomyces cerevisiae cells. The c-myc gene product was found predominantly in the nucleus by subcellular fractionation of infected insect cells.

Role of endogenous tumor necrosis factor alpha and interleukin 1 for experimental tumor growth and the development of cancer cachexia.

The aim of this study was to evaluate to what extent tumor necrosis factor alpha (TNF-alpha) and interleukin 1 may explain the development of experimental cancer cachexia. For this purpose, C57BL/6J mice bearing a transplantable low differentiated rapidly growing tumor were passively immunized every other day with rabbit or rat neutralizing immunoglobulins against either TNF-alpha (anti-TNF) or against an interleukin 1 receptor (anti-IL-1r). Anti-IL-1r in itself had no agonistic effect to the type I, T-cell/fibroblast IL-receptor. Tumor-bearing mice receiving either preimmune antiserum or nonimmune rat hybridoma IgG served as controls. Anti-TNF and anti-IL-1r inhibited tumor growth significantly, as measured by a lower wet and dry tumor weight at the end of 11 days of antiserum treatment (P less than 0.05). The acute phase response in tumor-bearing animals, measured as an increase in liver weight, hepatic RNA content, and increases in plasma concentrations of circulating IL-6, serum amyloid P, transferrin, complement (C3), and a decrease in plasma albumin, were unaffected by the specific antiserum treatments. Food intake, which declined significantly in pre/nonimmune injected tumor-bearing controls, was significantly improved in tumor-bearing animals immunized against TNF-alpha or the IL-1r. Whole body lipid content showed a trend to improvement in specifically immunized animals (P less than 0.07). The effects on whole body fat-free dry weight were insignificant, although numerically higher in specifically immunized tumor-bearing animals. The combination of anti-TNF and anti-IL-1r antiserum had no additive effects compared to single antiserum treatment suggesting that the two antibody treatments acted through a common mechanism. Cultured tumor cells, established from growing tumors, were sensitive to anti-TNF and anti-IL-1r, which both reduced tumor growth in vitro. This inhibitory effect by the antiserum could in part be reversed by the addition of recombinant IL-1 alpha and TNF alpha. We conclude that both TNF and IL-1 are involved in tumor growth and thus the progression of cancer cachexia. It seems as if the role of TNF and IL-1 was to promote tumor growth rather than restrict tumor growth in the present model. In this sense both TNF and IL-1 may act as tumor growth factors.

Antisera to the variable region of ras oncogene proteins, and specific detection of H-ras expression in an experimental model of chemical carcinogenesis.

Antisera were prepared in mice, rats and rabbits by immunization with peptides corresponding to regions of highest variability, located near the C-termini of four ras proteins. Two of these, H-ras (171-189) and K-rasB (171-186), react uniquely with H-ras and K-rasB gene products in immunoblots and immunoprecipitation reactions. Affinity-purified rabbit H-ras (171-189) antibody detects H-ras p21 in tissue culture cells and in tissue sections. Epithelial cells in normal mouse skin and cells in papillomas and carcinomas, in a mouse model system of chemical carcinogenesis in which mutational activation of H-ras occurs with high frequency, express high levels of H-ras p21 protein. These results suggest an hypothesis to explain the mechanism and preferential activation of particular ras loci in certain neoplasia.

In the mouse, the activation of the hypothalamic-pituitary-adrenal axis by a lipopolysaccharide (endotoxin) is mediated through interleukin-1.

The mechanisms through which endotoxin stimulates the hypothalamic-pituitary-adrenal axis are not well understood. In the studies reported here we tested the hypothesis that endotoxin increases plasma ACTH levels by releasing interleukin-1 (Il-1). Two experimental tools reported to interfere with the biological activity of IL-1 were used: antibodies directed against IL-1 receptors and alpha MSH. In a first series of experiments, adult male mice were injected with a lipopolysaccharide (LPS; 25 micrograms), antibodies against IL-1 receptor, alpha MSH (1-30 micrograms), or LPS and either IL-1 antibodies or alpha MSH. All treatments were administered ip. The endotoxin LPS caused a marked increase in plasma ACTH levels, measured 6 h later. Both alpha MSH and the Il-1 receptor antibodies, while having no effect by themselves, significantly (P less than or equal to 0.01) blocked LPS-induced ACTH release. In a second series of experiments, mice were injected ip with 500 ng recombinant human (rh) Il-1 alpha or 100 ng rhIl-1 beta in the presence or absence of alpha MSH (1-30 micrograms, ip). While not altering ACTH secretion induced by rhIl-1 alpha, 10-30 micrograms alpha MSH significantly (P less than or equal to 0.01) interfered with the effect of rhIl-1 beta. These results suggest 1) that endotoxin activates the hypothalamic-pituitary-adrenal axis through a mechanism involving the activation of interleukin-1 receptors; and 2) that the effect of rhIl-1 beta, but not -alpha, on ACTH secretion by the mouse can be partially blocked by alpha MSH.

Dual expression of p80 type I and p68 type II interleukin-I receptors on anterior pituitary cells synthesizing growth hormone.

Interleukin-1 alpha (IL-1 alpha) and IL-1 beta bind to either the p80 type I IL-1 receptor (IL-1RI) or the p68 type II IL-1R (IL-1RII) on both T and B lymphocytes. We and others have previously shown that the anterior pituitary gland also has specific high affinity binding sites for IL-1 alpha (Kd = approximately 1 nM) and expresses transcripts for both isoforms of the IL-1R. However, the identity of cells in the anterior pituitary gland that express the IL-1R and whether different populations of adenohypophyseal cells express different isoforms of the IL-1R remain unknown. Here we have used a combination of immunohistochemistry and histochemistry to localize IL-1RI and IL-1RII to specific target cells in the mouse anterior pituitary gland. Perfusion-fixed, paraffin-embedded sections of anterior pituitary gland were immunolabeled with well characterized monoclonal antibodies to either IL-1RI or IL-1RII and counterstained using a modified Gomori's method to document acidophils and basophils. Immunolabeling demonstrated that both IL-1RI and IL-1RII were abundantly expressed on a single population of anterior pituitary gland cells and that these cells could be classified on the basis of histochemical staining as a subpopulation of acidophils. The distribution, morphology, and proportion of cells immunolabeled for IL-1RI and IL-1RII were consistent with GH-synthesizing cells. To confirm this hypothesis, a modified indirect avidin-biotin complex, sequential peroxidase/alkaline phosphatase technique was used to label anterior pituitary gland cells with antibodies to IL-1RI followed by antibodies to IL-1RII, GH, PRL, or ACTH. The IL-1RI-positive cells predominately coexpressed IL-1RII and GH, but very little, if any, PRL or ACTH. These data establish that the predominant cell population in the murine anterior pituitary gland that constitutively expresses IL-1R stain as acidophils histochemically, is round to oval with dense granular cytoplasm and eccentric nuclei, synthesizes GH, and simultaneously expresses IL-1RI and IL-1RII isoforms.

Expression and localization of p80 and p68 interleukin-1 receptor proteins in the brain of adult mice.

The biological effects of interleukin-1 (IL-1) are mediated by two distinct receptors, the p80 type I IL-1 and p68 type II IL-1 receptor proteins (IL-1RI and IL-1RII, respectively), both of which have been recently co-localized to the growth hormone synthesizing cells of the adenohypophysis. Previous studies have shown that IL-1 can bind to specific structures in the central nervous system, but the distribution of IL-1RI and IL-1RII proteins in the adult mouse brain has not been reported. Here we have used immunohistochemistry to study the expression, distribution and cellular localization of both isoforms of the IL-1 receptor proteins in the adult mouse brain. Using a combination of processing techniques (AMeX fixation and cryosectioning), we have immunolabeled brain sections for each isoform of the IL-1R. Both isoforms are expressed in the CNS, particularly in neuronal soma of the granular layer of the dentate gyrus and pyramidal cells of fields CA1-CA4 of Ammon's horn of the hippocampus, in epithelial cells of the choroid plexus and ependymal layer, and in neuronal soma of Purkinje cells of the cerebellum. The IL-1RII isoform, but not IL-1RI, is expressed in specific neuronal soma and proximal cell processes of neurons of the paraventricular gray matter of the hypothalamus. These immunohistochemical data directly demonstrate the neuronal expression of both IL-1R proteins in situ. The distribution and cellular localization of IL-1R proteins in the CNS provide a molecular basis for understanding reciprocal interactions between the immune system and the brain.

Human interleukin-1 alpha is chemotactic for normal human keratinocytes.

Interleukin-1 (IL-1) has been shown to have mitogenic and chemotactic properties for a variety of cell types includes keratinocytes. These studies suggested that keratinocytes possess receptors for IL-1. In this study, the chemotactic properties of IL-1 for keratinocytes were confirmed and IL-1 receptors were demonstrated on keratinocytes using a radio receptor assay. Crosslinking studies with IL-1 alpha identified two major bands of Mr 97 kDa and 133 kDa. Thus, keratinocytes possess high affinity IL-1 receptors and respond to IL-1 by directed migration.

Visceral ischemia-reperfusion injury promotes tumor necrosis factor (TNF) and interleukin-1 (IL-1) dependent organ injury in the mouse.

Acute visceral ischemia and subsequent reperfusion injury, which accompanies the surgical repair of a thoracoabdominal aorta aneurysm, is associated with high rates of morbidity and mortality. The purpose of the present study was to determine whether endogenous tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 (IL-1) production contributes to organ dysfunction in animals subjected to visceral ischemia secondary to 30 min of supraceliac aortic occlusion. C57BL6/j mice were treated with either a TNF binding protein (TNF-bp-10 mg/kg) or an anti-IL-1 receptor type 1 antibody (150 micrograms) 2 h prior to 30 min of supraceliac aortic occlusion. An additional group of mice received 30 min of infrarenal aortic occlusion to determine the contribution of lower torso ischemia-reperfusion injury to the changes seen following supraceliac aortic occlusion. Visceral organ ischemia for 30 min produced by supraceliac aortic occlusion followed by 2 h of reperfusion produced measurable TNF-alpha in 38% of untreated mice, but TNF-alpha was undetectable in both sham-operated mice and following infrarenal aortic occlusion. After 2 h of reperfusion, lung myeloperoxidase levels were significantly elevated in the mice experiencing visceral ischemia-reperfusion compared with either a sham operation or infrarenal ischemia-reperfusion (11.6 +/- 1.3 U/g vs. 3.4 +/- .2 U/g and 3.7 +/- 1.0 U/g, respectively, p < .05). Pretreatment with TNF-bp and anti-IL-1 antibody decreased lung neutrophil recruitment (7.2 +/- 1.2 U/g and 4.6 +/- 1.1 U/g) and capillary membrane permeability changes in mice following visceral ischemia-reperfusion. The present study demonstrates that brief (30 min) clinically relevant visceral ischemia produces TNF-alpha and IL-1 dependent lung injury.

Evidence for multiple sites of interaction between IL-12 and its receptor.

We have previously described the identification of a protein, now designated IL-12R beta 1, that binds 125I-huIL-12 with a Kd of about 10 nM, corresponding to the low affinity 125I-huIL-12 binding sites seen on PHA-activated human lymphoblasts. Using expression cloning techniques, we have recently identified an additional IL-12-binding protein subunit, IL-12R beta 2, which binds 125I-huIL-12 with a Kd of about 5 nM when expressed alone in COS-7 cells. Coexpression of IL-12R beta 1 and IL-12R beta 2 in COS-7 cells results in formation of two classes of 125 I-huIL-12-binding sites with Kds of about 50 pM and 5 nM. Mouse IL-12 p40 subunit homodimer (mo(p40)2) blocked 125I-huIL-12 binding to human IL-12R beta 1, but did not inhibit binding to human IL-12R beta 2. In contrast, anti-huIL-12 monoclonal antibody 20C2, which does not block 125I-huIL-12 binding to human IL-12R beta 1, completely blocked binding to human IL-12R beta 2. These results demonstrate that two classes of IL-12 inhibitors, one that primarily blocks IL-12/IL-12R beta 1 interaction (e.g., mo(p40)2), and one that primarily blocks IL-12/IL-12R beta 2 interaction (e.g., 20C2), can be identified.

Regulation of myosin isoenzyme composition in fetal and neonatal rat ventricle by endogenous thyroid hormones.

The effect of thyroid hormone on the expression of ventricular isomyosins V1, V2, and V3 was studied in fetal and neonatal rats. Between 15 and 21 days gestation, V3 accounts for 80-90% of fetal ventricular myosin. After birth, there is a rapid transition from the fetal V3 isotype to an equal mixture of V1 and V3 at 3 days, and to 100% V1 at 3 weeks of age. The endogenous serum levels of thyroxine (T4) and triiodothyronine (T3) increase from trace amounts in the fetus to adult levels at 2-3 weeks of age; this increase correlates with the maximal expression of V1 during the same period. Expression of the V1 isomyosin can be eliminated in the neonatal rat if endogenous thyroid hormone synthesis is suppressed by propylthiouracil (PTU) treatment. In the PTU-treated rats, V3 is the only isomyosin synthesized between 1 and 30 days of age. In fetal ventricle, the amount of V1 is also decreased but not completely eliminated by PTU treatment. Conversely, the relative amount of V1 can be increased in the fetal ventricle by increasing the fetal serum concentrations of T4 and T3 to adult physiological levels. In these fetal ventricles, V1 represents greater than 85% of the total myosin. Likewise, the expression and accumulation of V1 could be stimulated in ventricles of PTU-treated, 12-day-old rats by administration of pharmacological or physiological doses of T3. Within 4 to 8 h after an initial dose of T3, V1 accumulates to 5-10% of the ventricular myosin, and by 72 h comprises 60-80% of the myosin. These results indicate that endogenous thyroid hormone induces the synthesis of ventricular heavy chain alpha, which as a dimer forms the V1 isomyosin, or plays a permissive role for the continued synthesis of heavy chain alpha in ventricles of fetal and neonatal rats.