Adriamycin as an inhibitor of 11 beta-hydroxylase activity in adrenal cortex mitochondria

Inhibition of 11 beta-hydroxylase activity was observed to be due to the interaction of adriamycin with adrenal cortex mitochondria. The inhibition of the enzyme was uncompetitive, with an apparent Ki of 100 microM, and was dependent upon the concentration of the drug and the time of incubation. Adriamycin increased the oxygen consumption of these mitochondria. EPR studies showed that adriamycin was reduced to a free radical semiquinone which served to shuttle electrons to oxygen, leading to an impairment in the reduction of cytochrome P450. It is suggested this may be the mechanism for the inhibitory effect of the drug on 11 beta-hydroxylase activity.     

Catalytic activity of cytochrome oxidase and cytochrome c in apolar solvents containing phospholipids and low amounts of water

Cytochrome c and cytochrome oxidase, in bovine heart submitochondrial particles and in their purified forms, were transferred to a ternary system that contained phospholipids (10 mg/ml toluene), the apolar solvent toluene, and water at concentrations of 13-15 microliters (high water) and 3 microliters (low water) per milliliter of toluene. When the enzymes were transferred back to an all water system, they exhibited full catalytic capacity. In the low water ternary system, cytochrome c could be reduced by ascorbate introduced via inverted micelles. Also in this system, cytochrome oxidase was reduced by ascorbate and cytochrome c but its oxidation was highly impaired. Data on the kinetics of reduction by ascorbate of cytochrome c and cytochrome oxidase under these conditions are presented. Cytochrome oxidase reduced in the organic solvent by ascorbate failed to form a complex with CO, but formed a complex with cyanide introduced via inverted micelles. The oxidized and the ascorbate-reduced cytochrome oxidase-cyanide complex exhibited a trough at 415 nm and a peak at 433 nm. The extent and rate of formation of the cyanide complex were higher with the reduced form of cytochrome oxidase. To achieve protein-protein interactions (cytochrome c-cytochrome oxidase) in the ternary system, it was necessary to extract the two proteins together. There was no functional interaction when they were extracted separately and mixed. In the high water ternary system reduced cytochrome oxidase was not detected, and it oxidized ascorbate at a higher rate than in the low water system; however, this rate was several orders of magnitude lower than in aqueous media.     

A consensus sequence of plant hemoglobins

From the study of plant hemoglobin protein and gene sequences, a consensus sequence was constructed and analysed by computer methods; analysis comprised the amino acid composition, hydropathy profile, and match degree with other plant and non-plant hemoglobin sequences. Resulting consensus sequence shows the main features of hemoglobin genes and proteins in plants.     

TRIIODOTHYRONINE—Clinical Effects in Patients with Suboptimal Response to Other Thyroid Preparations

Thirty patients with evidence of hypometabolism or a clinically related condition were given triiodothyronine after suboptimal response to thyroxin or desiccated thyroid. DL- and L-isomers of triiodothyronine were used and compared. Thirteen patients (43.3 per cent) were improved and 17 (56.6 per cent) were unimproved or became worse. Side effects occurred in 9 of 34 trials (26 per cent), and could not always be eliminated by decreasing the dosage. The highest percentage of good response occurred in a small group (four of five) who received a combination of desiccated thyroid or thyroxin with supplemental triiodothyronine. Despite careful analysis of the data, no basis was found on which to predict which patients would receive benefit from the triiodothyronine. However, the occasional improvement, sometimes dramatic, suggested that a therapeutic trial with triiodothyronine in difficult or unresponsive cases of hypometabolism or hypothyroidism is justified.     

Respiratory electron transfer activity in an asolectin-isooctane reverse micellar system.

Bovine heart submitochondrial particles (SMP) were solubilized in an asolectin isooctane reverse micellar system and the functionality of the respiratory chain was tested by spectroscopic and amperometric techniques. Electron transfer rate supported by NADH was very slow as evidenced by the low cytochrome reduction levels attained over long incubation periods. In the presence of KCN, NADH caused 34% and 12.5% reduction of the cytochromes aa3 and c, respectively, and negligible reduction of cytochrome b. Supplementation of the system with menadione rose the NADH-dependent reduction of all the cytochromes to levels that were close to the total content. However, no measurable O2 uptake activity took place in the presence of NADH plus menadione, or with ascorbate (or NADH) plus TMPD reducing systems. Therefore, it is suggested that in the organic medium, electron transfer from NADH to O2 is arrested at the terminal oxidase step. Cytochrome oxidase reduced by ascorbate (or NADH) plus TMPD seems to be trapped in its half reduced state (ie, a2+ a3(3+)). Although it is poorly reactive with O2, it can transfer electrons back to cytochrome c and TMPD. The electron transfer block to O2 was overcome when PMS was used instead of TMPD. This seems to be due to the recognized capacity of PMSH2 to carry out simultaneous reduction of both a CuA and a3 CuB redox centers of cytochrome oxidase. The cytochrome oxidase reaction in the organic solvent was highly sensitive to KCN (Ki 1.9 microM) and showed bell-shaped kinetics towards the PMS concentration and a sigmoidal response to water concentration, reaching its maximal turnover number (18 s-1) at 4 mM PMS and 1.1% (v/v) water.(ABSTRACT TRUNCATED AT 250 WORDS)     

The Ras guanine nucleotide exchange factor RasGRF1 promotes matrix metalloproteinase-3 production in rheumatoid arthritis synovial tissue

Introduction  

Fibroblast-like synoviocytes (FLS) from rheumatoid arthritis (RA) patients share many similarities with transformed cancer cells, including spontaneous production of matrix metalloproteinases (MMPs). Altered or chronic activation of proto-oncogenic Ras family GTPases is thought to contribute to inflammation and joint destruction in RA, and abrogation of Ras family signaling is therapeutic in animal models of RA. Recently, expression and post-translational modification of Ras guanine nucleotide releasing factor 1 (RasGRF1) was found to contribute to spontaneous MMP production in melanoma cancer cells. Here, we examine the potential relationship between RasGRF1 expression and MMP production in RA, reactive arthritis, and inflammatory osteoarthritis synovial tissue and FLS.