Identification of negative and positive regulatory elements in the human renin gene.

Renin gene expression is tissue-specific and under complex hormonal control. To investigate which DNA elements are involved in the control of human renin gene expression, we performed transient DNA transfer experiments with renin-chloramphenicol acetyltransferase fusions. We have mapped a complex arrangement of positive and negative control sequences in the 5' flanking region of the human renin gene.

Identification of cis-acting sequences responsible for phorbol ester induction of human serum amyloid A gene expression via a nuclear factor kappaB-like transcription factor.

We have analyzed the 5'-flanking region of one of the genes coding for the human acute-phase protein, serum amyloid A (SAA). We found that SAA mRNA could be increased fivefold in transfected cells by treatment with phorbol 12-myristate 13-acetate (PMA). To analyze this observation further, we placed a 265-base-pair 5' SAA fragment upstream of the reporter chloramphenicol acetyltransferase (CAT) gene and transfected this construct into HeLa cells.

The immune response of BALB/c mice to influenza hemagglutinin: commonality of the B cell and T cell repertoires and their relevance to antigenic drift.

An extensive analysis of the class II (I-Ad)-restricted T cell repertoire for influenza hemagglutinin (HA) of the H3 subtype, elicited by natural infection, has shown that majority of CD4+ memory T cell clones focus on antibody-binding regions of HA, sites B and E, and are sensitive to the residue substitutions that have occurred in these regions during antigenic drift. The proliferative responses of CD4+ clones to synthetic peptides have identified T cell epitopes within site B, HA1 177-199 and HA1 182-199, and site E. HA1 56-76.

The structural requirements for class II (I-Ad)-restricted T cell recognition of influenza hemagglutinin: B cell epitopes define T cell epitopes.

A majority of I-Ad-restricted CD4+ clones elicited by influenza X31 (H3N2) virus infection, recognize a synthetic peptide of hemagglutinin (HA) corresponding to an antibody binding region of the HA1 subunit (site B: HA1 177-199). The structural requirements for class II-restricted T cell recognition were investigated by determining the proliferative responses of representative CD4+ clones to truncated HA1 peptides and synthetic peptide analogues. Two distinct T cell epitopes were identified and CD4+ clones, specific for either determinant, were sensitive to the same single amino acid substitutions in synthetic peptides at HA1 193 S----N or HA1 198 A----E, that had featured in antigenic drift and abrogated antibody binding to native HA.

Cotransmitters: differential effects of serotonin (5-HT)-depleting drugs on levels of 5-HT and TRH and their receptors in rat brain and spinal cord.

Following codepletion of endogenous serotonin (5-HT, greater than 90%) and thyrotropin-releasing hormone (TRH, 66%) by neonatal treatment with the serotonergic neurotoxin, 5,7-dihydroxytryptamine (DHT), a 33% (n = 12, P less than 0.01) increase in specific TRH receptor binding was observed in adult rat spinal cord (SC) homogenates. A 20-21% increase in TRH receptors was also observed in the medulla/pons (MP) (n = 12, P less than 0.

Ovariectomy permits progesterone to increase the binding of [3H]spiperone to the anterior pituitary gland in estrogen-primed rats.

The binding of the dopamine (DA) D2 antagonist spiperone to DA receptors in the anterior pituitary gland was evaluated in intact and ovariectomized (OVX) estrogen-primed rats in which circulating levels of progesterone (P) were varied. Nembutal (PB; 35 mg/kg, ip) was administered at 1130 h to intact proestrous animals to prevent the LH-stimulated release of ovarian P. Two hours later some of these intact rats were QVX to remove the endogenous ovarian steroids.

The specificity of the human mineralocorticoid receptor: clinical clues to a biological conundrum.

Data from in vitro studies show that the mineralocorticoid receptor has an equal affinity for cortisol and aldosterone. This contrasts with the well known selectivity of aldosterone for binding to tissues such as the kidney despite the much higher circulating levels of glucocorticoids. This paper puts forward the hypothesis that the enzyme 11 beta-hydroxysteroid dehydrogenase (11 beta-OHSD) is responsible for determining this selectivity.

Negative control elements and cAMP responsive sequences in the tissue-specific expression of mouse renin genes.

The 5' flanking regions of the mouse renin genes (Ren1d and Ren2d) contain putative negative control and cAMP responsive elements. Sequence analysis shows additionally that these putative control elements in the Ren2d gene are interrupted by a 160-base-pair insertion. To document the functions of these elements, we isolated these regions and fused them to the reporter gene chloramphenicol acetyltransferase (CAT), which was linked upstream to a thymidine kinase (TK) promoter (pUTKAT1).

Functional human renin promoter in transfected cells: evidence for cell-specific expression.

Expression of the human renin gene is regulated in a tissue-specific manner, but study of this regulation has been limited by a lack of suitable cell lines that simulate endogenous control. In order to characterize the regulation of renin gene expression, the 5' flanking region (892 base pairs) from the human renin gene was linked to the chloramphenicol acetyltransferase gene and was introduced into multiple human cell lines by calcium phosphate precipitation or electroporation methods to assess transcriptional control. The human renin promoter was active when transfected into cultured human choriocarcinoma cells (JEG-3) and rat vascular smooth muscle cells, but it was not active in many other cloned cell types.

Increased tumor necrosis factor and IL-1 beta gene expression in the kidneys of mice with lupus nephritis.

Because TNF and IL-1 can initiate immunologic and inflammatory events alone or synergistically, a local increase in the levels of one or both of these cytokines in vivo may cause irreparable tissue damage. The purpose of this study was to evaluate local TNF and IL-1 beta gene expression in vivo in the kidneys of MRL-Ipr mice with autoimmune lupus nephritis. TNF mRNA was detected in the renal cortex of MRL-Ipr mice but was not present in the cortex of normal congenic MRL-++ or C3H/FeJ mice.

Localisation of 11 beta-hydroxysteroid dehydrogenase--tissue specific protector of the mineralocorticoid receptor.

In vitro the mineralocorticoid receptor is non-specific and does not distinguish between aldosterone and cortisol. In vivo certain tissues with this receptor are aldosterone selective (eg, kidney and parotid) whereas others with the same receptor are not (eg, hippocampus and heart). Experiments in rats showed that 11 beta-hydroxysteroid dehydrogenase (which converts cortisol to cortisone in man and corticosterone to 11-dehydrocorticosterone in the rat) was much more highly concentrated in aldosterone-selective tissues than in non-selective tissues.

Molecular biology of the renin-angiotensin system.

This paper reviews the molecular biology of the renin-angiotensin system. The renin gene structure is analyzed in detail, including an examination of the putative regulatory regions. The combined action of these regulatory sequences would result in the complex, tissue-specific expression and regulation observed in vivo.

A monoclonal antibody raised against a synthetic peptide representative of part of the amino acid sequence of rat immunoglobulin E detects thermally induced changes in that region of the IgE molecule.

A mouse monoclonal antibody (mAb) has been produced by conventional cell fusion methods against a synthetic peptide, p123, representative of a portion of the CH4 domain of rat immunoglobulin E (IgE). This monoclonal antibody was reactive with both peptide and purified rat IgE (p.rat IgE) by indirect enzyme immunosorbent assay (ELISA), and its binding to p.

Dexamethasone-suppressible hyperaldosteronism: studies on overproduction of 18-hydroxycortisol in three affected family members.

We report a newly diagnosed family in which a father and his two sons were found to be hypertensive and to have the rare familial condition dexamethasone-suppressible hyperaldosteronism (DSH). All three patients became normotensive on dexamethasone treatment alone and have been successfully maintained on low doses of the drug for 6 months since diagnosis. Each of the patients had extremely high plasma and urinary concentrations of the recently discovered steroid 18-hydroxycortisol, which were more than ten times higher than the upper normal limit.

Structural and functional basis for GABAA receptor heterogeneity.

When gamma-aminobutyric acid (GABA), the major inhibitory neurotransmitter in vertebrate brain, binds to its receptor it activates a chloride channel. Neurotransmitter action at the GABAA receptor is potentiated by both benzodiazepines and barbiturates which are therapeutically useful drugs (reviewed in ref. 1).

Novel and enhanced IL-1 gene expression in autoimmune mice with lupus.

IL-1 is a pleiotropic factor encoded for by at least two genes, alpha and beta, and capable of eliciting a broad set of immunologic and inflammatory events. MRL/MP-lpr (MRL-lpr) mice are an appealing model for studies of renal injury inasmuch as disease in this strain is spontaneous, rapid, predictable, and regulated by the lpr gene. Infiltration of macrophages and the proliferation of the glomerular mesangial cells are prominent features of renal disease.

Fine specificity of murine class II-restricted T cell clones for synthetic peptides of influenza virus hemagglutinin. Heterogeneity of antigen interaction with the T cell and the Ia molecule.

We have previously demonstrated diversity in the specificity of murine, H-2k class II-restricted, T cell clones for the hemagglutinin (HA) molecule of H3N2 influenza viruses and have mapped two T cell determinants, defined by synthetic peptides, to residues 48-68 and 118-138 of HA1. In this study we examine the nature of the determinant recognized by six distinct P48-68-specific T cell clones by using a panel of truncated synthetic peptides and substituted peptide analogs. From the peptides tested, the shortest recognized were the decapeptides, P53-62 and P54-63, which suggests that the determinant was formed from the 9 amino acids within the sequence 54-62.

Comparative inhalation toxicity of nickel sulfate to F344/N rats and B6C3F1 mice exposed for twelve days.

Groups of F344/N rats and B6C3F1 mice were exposed to aerosols of nickel sulfate hexahydrate (NiSO4.6H2O) 6 hr/day for 12 days to determine the short-term inhalation toxicity of this compound. Target exposure concentrations were 60, 30, 15, 7, 3.

Mapping of the human renin transcription start site: evidence for a single functional promoter.

The 5' flanking region of the human renin gene contains two putative promoter sequences (TATA boxes), named P1 and P2. These are located in positions -77 to -71 and -29 to -23 respectively, each followed by a possible translational start site (AUG). In order to identify whether these sequences are functional in the kidney, we employed the RNAse protection assay.

Guanine nucleotide regulation of receptor binding of thyrotropin-releasing hormone (TRH) in rat brain regions, retina and pituitary.

Guanine nucleotides inhibited the specific binding of [3H](3-Me-His2)thyrotropin-releasing hormone ([3H]MeTRH) to receptors for TRH in washed homogenates of rat anterior pituitary gland in a dose-related manner. The order of potency (at 100 and 500 microM final) was Gpp(NH)p (a stable analog of GTP) greater than GTP much greater than GDP much greater than cGMP (with the adenine nucleotides being inactive) in the pituitary and various brain regions. Gpp(NH)p at 1 mM caused 17-35% inhibition of [3H]MeTRH binding to different tissues including the pituitary, hypothalamus, retina and nucleus accumbens.

Mineralocorticoid activity of liquorice: 11-beta-hydroxysteroid dehydrogenase deficiency comes of age.

The sodium retention associated with liquorice ingestion has been thought to be due to a direct mineralocorticoid effect, despite the fact that it does not seem to occur in patients or animals with severe adrenal insufficiency. This study in seven normal subjects given liquorice showed that sodium retention is associated with a significant change in cortisol metabolism indicating inhibition of 11-beta-hydroxysteroid dehydrogenase (11 beta-OHSD). Congenital deficiency of this enzyme produces a syndrome of apparent mineralocorticoid excess.