Increased IL-17 expression in degenerated human discs and increased production in cultured annulus cells exposed to IL-1ß and TNF-α

Abstract

IL-17 is expressed in a number of tissues including the intervertebral disc, where it exerts strong inflammatory properties. We evaluated IL-17 using immunolocalization in herniated and non-herniated human discs, IL-17 gene expression, and the production of IL-17 by annulus cells cultured in three dimensions in the presence of IL-1ß or TNF-α. There was no difference in the percentage of IL-17 positive cells in annulus or nucleus in herniated vs. non-herniated disc specimens. Molecular studies confirmed expression of IL-17 in disc tissue, with significantly increased expression in more degenerated discs; there was no difference in expression between herniated vs. non-herniated discs. Exposure to IL-1ß or TNF-α resulted in significantly greater production of IL-17. Our findings expand understanding of IL-17 production by disc cells and reveal the importance of non-canonical IL-17 production in the disc. Significantly greater expression of IL-17 in more degenerated discs adds to our understanding of the changes in disc cell function with advancing stages of disc degeneration.     

Comparative EPR study on high-spin ferric porphine complexes and cytochrome P-450 having rhombic character

Comparative EPR studies were made on two high-spin Fe(III) porphine model systems and mammalian liver microsomal cytochromes P-450, all of which exhibit approximately the same degrees of rhombicity in their EPR spectra. Comparison of g values and linewidths as a function of temperature, and of the microwave power saturation demonstrated that EPR characteristics of P-450 are more similar to the Fe(III) porphines having the thiolate axial ligand than in the other model systems, the mixed crystals of Fe(III) porphine with the corresponding free base porphine, in which no thiolate ligand is involved.There is, however, a discrepancy between P-450 and the model thiolates with respect to the size of the zero-field parameter D. These observations indicate that P-450 heme has essential structural features in common with thiolates but the Fe-S bond of P-450 may be modified from its normal orientation in model thiolates, probably as a result of the constraints imposed by the protein stucture.     

Palladium-catalyzed homogeneous coupling reactions of steroids with organostannanes

Direct and carbonylative coupling reactions of various steroid derivatives possessing iodo- and bromo-alkenyl moiety (17-iodo-androst-16-ene, 1, 17-bromoandrost-2,16-diene. 2, 17-iodo-4-aza-4-methylandrost-16-en-3-one, 3, 17-iodo-4-azaandrost-16-en-3-one, 4) with vinyltributylstannane and ethynyltributylstannane were carried out in the presence of various palladium catalysts. While carbonylation took place only with vinyltributylstannane, 17-vinyl-, and 17-ethynyl-Δ¹⁶ steroids were produced via direct coupling with vinyltributylstannane and ethynyltributylstannane, respectively. Activities of some catalysts based on Pd(0) and Pd(II) precursors were compared, and Pd(PPh₃)₄ was found to be superior to other complexes in most cases. In the coupling of 17-iodoandrost-16-ene with organostannanes Pd₂(dba)₃ + 8 AsPh₃ in situ catalyst was found to be even more effective.     

In vitro binding of labeled 20-methylcholanthrene to a specific cytosol-protein

In vitro binding kinetics and specificity of [20-³H]methylcholanthrene ([³H]MC) interaction with mouse epidermal cytosol in the presence or absence of microsomal metabolizing systems were investigated. After an incubation period of 30 roin at 22°, the samples were dialyzed, subjected to gel filtration, ultrafiltration, and analyzed by 7 % basic polyacrylamide gel electrophoresis. A major portion of the binding appeared in a single peak on the gel which had the same mobility as bovine or mouse serum albumin. In vitro competition by other hydrocarbons or by promoters for binding of MC to this cytosol receptor protein showed an impressive correlation to carcinogenic and promoting activities. Essentially 100% of the MC bound to cytosol without a microsomal metabolizing system was non-covalent. However, binding Of MC to cytosol in the presence of microsomes plus reduced NADPH was party covalent which has previously been reported by Grover and Sims and Meunier and Chaveau. When bovine (BSA) or mouse serum albumin (MSA) was incubated with radioactive MC in the presence of competing non-radioactive carcinogens or promoters, little or no inhibition of binding was found. The finding that both a powerful tumor promoter and a strong carcinogen are competitors for the specific MC binding to the cytosol receptor protein indicates that it may represent a critical interaction for the promoting stage of chemical carcinogenesis in mouse skin.     

Adrenocorticotropic hormone and dibutyryl adenosine cyclic monophosphate-mediated Ca2+ uptake by rat adrenal glands

The effect of adrenocorticotropic hormone and dibutyryl cyclic AMP on the uptake of ⁴⁵Ca²⁺ by the rat adrenal gland has been investigated. After injection of ⁴⁵Ca²⁺ and adrenocorticotropic hormone into rats, the adrenal ⁴⁵Ca²⁺ concentration was significantly enhanced 90 to 180 min following hormone administration. The rise in adrenal ⁴⁵Ca²⁺ content was accompanied by a marked increase of the serum corticosterone levels. During incubation of rat adrenal glands in the presence of ⁴⁵Ca²⁺, adrenocorticotropic hormone and dibutyryl cyclic AMP caused significant accumulation of adrenal ⁴⁵Ca²⁺ and increased corticosterone synthesis. The degree of stimulation of both adrenal ⁴⁵Ca²⁺ uptake and corticosterone synthesis by adrenocorticotropic hormone or dibutyryl cyclic AMP was dependent upon the concentration of calcium in the incubation medium and upon the amount of adrenocorticotropic hormone or dibutyryl cyclic AMP added. Theophylline mimicked the stimulatory effect of adrenocorticotropic hormone and dibutyryl cyclic AMP and increased the uptake of ⁴⁵Ca²⁺ by rat adrenal glands in vitro. Determination of calcium by atomic absorption spectroscopy showed that the adrenocorticotropic hormone-mediated adrenal ⁴⁵Ca²⁺ uptake was due to a net accumulation of calcium in the tissue and not only to an increased rate of exchange of extracellular ⁴⁵Ca²⁺ with the intracellular calcium pool. Adrenocorticotropic hormone-stimulated adrenal ⁴⁵Ca²⁺ uptake was not observed when steroidogenesis was inhibited with elipten. Both adrenocorticotropic hormone-mediated corticosterone synthesis and adrenal ⁴⁵Ca²⁺ uptake were abolished after treatment of rats with cycloheximide but not after treatment with actinomycin D, indicating that adrenal ⁴⁵Ca²⁺ uptake and steroidogenesis have similar requirements for de novo protein synthesis, but not RNA synthesis.     

The effect of antiestrogens on egg yolk protein synthesis and estrogen-binding to chromatin in the rooster liver

Nafoxidine and CI-628, two well known antiestrogenic compounds, reduce the stimulating effect of estradiol on the estrogen-binding capacity of the liver chromatin from roosters. In vitro both antiestrogens compete with [³H] estradiol for the binding sites on the liver chromatin. They inhibit the estrogen-induced synthesis of egg yolk proteins (vitellogenin) and fail to induce this estrogen-specific protein synthesis by themselves. They show the ability, however, to increase the estrogen-binding sites on the liver chromatin to some extent.