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B Faris

Publications and source records attributed to B Faris.

At least 37 records · Page 2Linked to original sources
PubMed1988

Elastin mRNA levels and insoluble elastin accumulation in neonatal rat smooth muscle cell cultures.

Insoluble elastin accumulation, elastin mRNA translational efficiencies, and elastin mRNA levels were evaluated in cultures of neonatal rat aortic smooth muscle cells grown for several days in consecutive passages. When the products of in vitro translation were immunoprecipitated with an anti-alpha-elastin antibody, a single 79,000-Da protein was obtained. Northern blot analysis also indicated an elastin mRNA species corresponding to approximately 4.2 kilobases. Insoluble elastin accumulation increased in cells cultured for 7-21 days in first through fourth passages, while with one exception, relative levels and translational activity of elastin mRNA decreased with time in culture. The data indicated that a simple relationship between elastin accumulation and elastin mRNA levels was not evident.

Animals↗
PubMed1988

Glycosaminoglycan content in neonatal rat aortic smooth muscle cell cultures.

In the present study the biosynthesis of glycosaminoglycans (GAGs) by neonatal rat aortic smooth muscle cells in culture was studied. Heparan sulfate (HS) was the predominant GAG of the cell layer accounting for 32-49% of the total GAGs depending on the time in culture. The presence of low sulfated chondroitin sulfate (LSC) in aortic smooth muscle cell cultures is reported here for the first time. The effect of ascorbate on the synthesis and accumulation of these macromolecules resulted in a relative increase of C4S and DS in the cell layer. In contrast, the distribution of the GAGs which were secreted into the medium was not significantly effected by the addition of ascorbate. While HS was always found to be a minor component, the other GAGs were present in about equal concentrations. The total GAG accumulation in the medium was much greater (91-97%) than that of the cell layer (3-9%) indicating that the cells are synthesizing relatively large amounts of GAGs, although incorporation of these macromolecules into the extracellular matrix was consistently low.

Animals↗
PubMed1988

Cadmium-chloride-induced air-space enlargement with interstitial pulmonary fibrosis is not associated with destruction of lung elastin. Implications for the pathogenesis of human emphysema.

To determine whether lung elastin is lost during the evolution of cadmium-induced air-space enlargement with pulmonary fibrosis, the lung elastin of 5- to 7-day-old golden Syrian hamster pups was radiolabeled by giving [3H]valine. At maturity, a single intratracheal instillation of 0.5 ml of 0.025% CdCl2 solution was given. Lung mechanics, histologic examination, and biochemistry were studied 5, 10, 21, 42, 105, and 180 days after the cadmium treatment. The animals developed fibrosis and air-space enlargement with decreased lung volumes, compliance, and forced expiratory flow; their functional residual capacity was increased. The total lung collagen and total lung elastin were increased, but there was no loss of radiolabel in lung elastin. We conclude that CdCl2-induced air-space enlargement with pulmonary fibrosis is not accompanied by loss of neonatally formed lung elastic fibers. We hypothesize that air-space enlargement with fibrosis represents a stereotyped response of the lung to fibrosing injuries, which we hypothesize is due to forces from more fibrotic and atelectatic areas causing overdistension of less abnormal air spaces. The air-space enlargement of fibrosing human diseases such as sarcoidosis and eosinophilic granuloma may have a similar basis. Evidence is reviewed that human centrilobular emphysema may be a form of focal air-space enlargement with interstitial fibrosis; there may be mechanisms in addition to elastase-antielastase imbalance that cause human emphysema.

Animals↗
PubMed1988

Repair of protease-damaged elastin in neonatal rat aortic smooth muscle cell cultures.

The objective of this study was to investigate the elastin repair process in the rat aortic smooth muscle cell culture after proteolytic injury. Although little studied in vivo, elastin repair is thought to occur through a sequential process involving enzymatic removal (debridement) of damaged fibers followed by synthesis of tropoelastin, its subsequent processing, and eventual incorporation into new insoluble elastin. A second repair mechanism of proteolytically damaged elastin in a culture system is reported here. Repair in this system relates directly to restoration of resistance to elastin solubilization by hot alkali. As expected, severe injuries were observed with porcine pancreatic elastase (PPE). Using PPE, only 6% of the elastin, relative to control, was resistant to hot alkali immediately after elastase treatment. 4 wk later, resistance to hot alkali had increased dramatically to a mean of 90%. Repair took longer after injury with 75 micrograms of PPE as compared with 50 micrograms of PPE. The limited elastic fiber proteolysis induced by either human neutrophil elastase or porcine trypsin was repaired in culture within 2 wk. Elastin that had been radiolabeled with [3H]lysine 4-5 wk before injury was converted from a hot NaOH-susceptible to a NaOH-resistant elastin fraction during recovery from PPE injury. At the same time, the frayed elastic fibers that were seen with the electron microscope immediately after PPE treatment were replaced by continuous bands of elastin that resembled those in control cultures. Restoration of NaOH resistance did not require a net increase in total cell layer elastin, suggesting that relatively little new tropoelastin incorporation into the cell layer was required for this type of repair. These results suggested a salvage repair mechanism for proteolytically damaged elastin.

Animals↗
PubMed1988

Congo red binding of elastin in aortic smooth muscle cell cultures.

These studies demonstrate that the strong binding capacity of elastin for Congo red can be used to advantage in aortic smooth muscle cell cultures. A fibrous elastin network fluoresces when Congo red is added. Congo red does not alter accumulation of elastin or of total protein, even when the cells are grown in the presence of the dye for long periods of time, indicating that it is not toxic. Porcine pancreatic elastase was used to solubilize elastin in these cultures, to determine the molar ratio of Congo red to elastin, thus making it possible to estimate the amount of elastin solubilized when the cultures are injured. Congo red binding to elastin will be useful in studying elastin accumulation and/or degradation in vitro and in vivo.

Animals↗
PubMed1988

Tissue-specific posttranslational modification of rat apoE. Synthesis of sialated apoE forms by neonatal rat aortic smooth muscle cells.

We have studied the synthesis, modification, and secretion of rat apoE in primary cultures of neonatal aortic smooth muscle cells and adult rat hepatocytes. The cultures were pulsed with [35S]methionine and the intracellular and secreted apoE were immunoprecipitated and analyzed by two-dimensional isoelectric focusing/polyacrylamide gel electrophoresis and autoradiography. A short pulse (10 min) showed the presence of a major unmodified apoE form in both cultures. This form comigrated on two-dimensional gels with the major rat plasma apoE isoprotein. A longer pulse (15-120 min) resulted in the progressive appearance of intracellularly modified apoE isoproteins in both cultures. The apoE secreted by aortic smooth muscle cells consisted exclusively of sialated apoE isoproteins that were sensitive to neuraminidase treatment. In contrast, the apoE secreted by primary cultures of adult rat hepatocytes, organ cultures of neonatal rat liver, as well as rat plasma apoE, contained several minor modified isoproteins. Nascent apoE secreted by the aortic smooth muscle cell cultures floats in the density range of 1.09 to 1.186 g/ml. We conclude that aortic smooth muscle cells can synthesize and secrete sialated apoE isoproteins associated with nascent lipoproteins floating in the high density lipoprotein region.

Animals↗
PubMed1986

Phospholipid methylation in rabbit aorta.

The formation of phosphatidylcholine by successive methylations of phosphatidylethanolamine using S-adenosylmethionine as the methyl donor was studied in homogenates of rabbit aorta. Addition of phosphatidylmonomethylethanolamine and phosphatidyldimethylethanolamine, but not phosphatidylethanolamine, stimulated methyltransferase activity and this activity was further stimulated when the phospholipids were dispersed in taurocholate prior to addition to the assay system. No incorporation of radiolabel into sphingomyelin or lysolecithin was detected indicating minimal metabolism of newly formed phosphatidylcholine. The majority of methyltransferase activity was detected in the high-speed pellet of the aortic homogenate; however, since activity was also detected in the high-speed supernatant, the low-speed supernatant preparation was used as the source of enzyme. Methyltransferase activity was characterized in cultured arterial smooth muscle cells using methionine as the radiolabeled precursor. The major product formed was phosphatidylcholine. No difference in enzyme activity was seen as a function of the length of time that cells were in culture or anatomic location of the aortic explant used as a source of cells. Treatment of the cells with cycloheximide did not affect methyltransferase activity. The ability of catecholamine agonists and vasoactive peptides to influence methyltransferase activity was investigated both in the cell-free preparation and in cultured cells. These compounds did not appear to alter methyltransferase activity in rabbit aortic smooth muscle cells.

Animals↗
PubMed1986

Purification and properties of Clostridium difficile cytotoxin B.

Toxin B, a potent cytotoxin produced by Clostridium difficile, was purified to homogeneity from 6-day broth cultures of a toxigenic isolate. Cytotoxin was purified approximately 4000-fold by sequential ammonium sulfate precipitation, DEAE-Sepharose chromatography, and high performance liquid chromatography on a Mono Q anion-exchange column. The molecular weight of reduced purified toxin was 50,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, compared to 150,000 for unreduced toxin. Dose-response studies indicated that subpicogram concentrations of purified toxin caused rounding of approximately 20,000 IMR-90 fibroblasts. The phenomenon of cell rounding caused by toxin B was correlated with the ratio of globular to filamentous actin in fibroblasts as measured by two techniques. The toxin caused a significant increase in the ratio of globular to filamentous actin which was nearly completed prior to the onset of rounding. We conclude that cell rounding of fibroblasts exposed to toxin B is related to an increase in the ratio of globular to filamentous actin which is produced by small numbers of toxin molecules/cell.

Actins↗
PubMed1986

Elastase effect on the extracellular matrix of rat aortic smooth muscle cells in culture.

The effect of elastase on the extracellular matrix of neonatal rat aortic smooth muscle cell cultures was monitored both chemically and ultrastructurally. Porcine pancreatic elastase was shown to decrease the elastin content in these cultures. Although chemically no distinction could be made between the elastin remaining in the culture matrix after elastase when compared to that in the nontreated cultures, the elastin was dramatically altered morphologically. The elastin assumed a "mottled" appearance after elastase treatment similar to that seen in vivo in emphysema models. A highly sensitive immunogold staining technique was used to detect elastin at the earliest stages of accumulation. Pulse experiments demonstrated an increase in protein synthesis by the cells 20 hr after elastase exposure. The culture system described here provides a model for probing in vivo elastase effects on elastin-containing tissues.

Amino Acid Sequence↗
PubMed1986

Plasma amino acid levels in cataract.

Fasting plasma amino-acid profiles were determined in 32 subjects with either cataractous lenses (24 subjects) or normal lenses (eight subjects). The findings were compared with established laboratory normal values. The amino-acid levels of all subjects with normal lenses were within the normal range for the subjects' age groups. In contrast, subjects with presenile or senile cataracts had markedly elevated levels of glutamine, with similar elevations of alanine in presenile cataract and of histidine in senile cataract. To the best of our knowledge, this is the first report of such a plasma amino-acid pattern in subjects with presenile and senile cataracts.

Adult↗
PubMed1985

Processing of soluble elastin in cultured neonatal rat smooth muscle cells.

The synthesis and extracellular deposition of elastin by cultured neonatal rat aorta smooth muscle cells has been followed. The addition of beta-aminopropionitrile to the culture medium promotes accumulation of soluble precursors of elastin. Under such conditions, a protein possessing characteristics of a soluble elastin precursor with an apparent molecular weight of 77,000 was detected and partially purified. Pulse-chase studies suggested that this 77-kDa protein undergoes an extracellular, enzymatically catalyzed process to a 71-kDa protein. This 71-kDa protein is strikingly similar to tropoelastins isolated from other tissue systems, in which no evidence for higher molecular weight soluble precursors is at present available. Data presented in this communication suggest that the 77-kDa protein, which we have designated protropoelastin, represents a precursor to the tropoelastin moiety produced in the neonatal rat smooth muscle cell culture.

Aminopropionitrile↗
PubMed1985

Alteration of the extracellular matrix of smooth muscle cells by ascorbate treatment.

The protein composition in the extracellular matrix of cultured neonatal rat aortic smooth muscle cells has been monitored over time in culture. The influence of ascorbate on insoluble elastin and collagen has been described. In the absence of ascorbate, the cells accumulate an insoluble elastin component which can account for as much as 50% of the total protein in the extracellular matrix. In the presence of ascorbate, the amount of insoluble collagen increases, while the insoluble elastin content is significantly less. When ascorbate conditions are varied at different times during the culture, the extracellular matrices are altered with respect to collagen and elastin ratios. The decrease in elastin accumulation in the presence of ascorbate may be explained by an overhydroxylation of tropoelastin. Approximately 1/3 of the prolyl residues in the soluble elastin fractions isolated from cultures grown in the presence of ascorbate are hydroxylated. Since the insoluble elastin accumulated in these cultures contain the unique lysine-derived cross-links in amounts comparable to aortic tissue, this culture system proves ideal for studying the influence of extracellular matrix elastin on cell growth and metabolism.

Amino Acids↗
PubMed1985

Aging effects on the elastin composition in the extracellular matrix of cultured rat aortic smooth muscle cells.

Elastin accumulation in the extracellular matrix of cultured rat aortic smooth muscle cells was monitored as a function of age. The effect of the animal donor age and time in culture in single or consecutive passages on the cells' ability to accumulate total protein as well as elastin was evaluated. Smooth muscle cells were obtained from animals ranging in age from 2 d to 36 mo. Protein accumulation by the cells based on DNA content was similar regardless of which of the above aging parameters was examined. Although there were significant amounts of elastin present in the extracellular matrix of those cells originating from the younger animals (2 d and 6 wk old), little or none was detected in cell cultures derived from the oldest animals. A soluble elastin-like fraction which was isolated from the cultures of the 2-d-old rats seemed to be lacking in the cultures of cells from the 36-mo-old animals. This observation may, in part, explain the absence of insoluble elastin in the matrix of some cultures obtained from older animals. The data strongly suggest that the age of the donor animal from which the cells originate has the greatest influence on in vitro elastin accumulation.

Amino Acids↗
PubMed1985

A case of pulmonary tuberculosis presenting with a choroidal tuberculoma.

A 17-year-old woman was referred because of a choroidal mass, with marked decrease in vision, in her left eye. Her medical history was negative. However, preliminary examination revealed a positive skin tuberculin test (PPD) and a chest x-ray picture of pulmonary tuberculosis. Bronchoscopy documented the diagnosis. Findings of other systemic investigations were negative. Fluorescein angiographic findings were compatible with the clinical diagnosis of a choroidal tuberculoma. A regimen of antituberculous medications was started, and both the choroidal and the pulmonary lesions gradually subsided. To our knowledge this is the first reported case of nonmiliary tuberculosis that presented solely with a choroidal lesion.

Adolescent↗
PubMed1985

Isolation and characterization of a rat lung fraction enriched in alveolar wall basement membranes.

Separation of lung alveolar basement membranes from interstitial connective tissue protein has proved difficult, and a pure preparation of alveolar wall basement membranes (AWBM) is not available. We have modified a technique employing the detergent Triton X-100 for isolating AWBM from rat lungs by adding a step utilizing human skin collagenase (HSC), a highly purified enzyme obtained from skin fibroblasts that specifically cleaves non-basement membrane collagens. Triton extraction of both lungs yields 15-20 mg of basement membrane-enriched material referred to as crude fraction (CF). Ultrastructural studies show that CF includes both epithelial and endothelial basement membranes that appear similar to their in vivo counterparts and contain heparan sulfate proteoglycans. Extraction of type IV collagen is documented by the appearance of highly glycosylated hydroxylysine. This CF contains minimal amounts of contaminating elastin but significant amounts of interstitial collagens. CF was further purified for biochemical studies by incubation with HSC. HSC solubilized 20% of CF hydroxyproline resulting in a final fraction highly enriched in AWBM. Lung minces incubated in tritiated lysine produced a CF extract rich in newly formed type IV collagen, showing that lung tissue synthesizes AWBM collagen in vitro.

Animals↗
PubMed1984

Effect of varying amounts of ascorbate on collagen, elastin and lysyl oxidase synthesis in aortic smooth muscle cell cultures.

In the presence of ascorbate, there is an increase in collagen synthesis with a concomitant decrease in insoluble elastin and lysyl oxidase activity in cultured rabbit aortic smooth muscle cells. While the addition of 0.5 micrograms ascorbate per ml of medium enhances collagen synthesis and accumulation, detectable insoluble elastin and lysyl oxidase activity remain essentially unchanged. However, at 2 micrograms ascorbate per ml, the integrity of the insoluble elastin is lost and lysyl oxidase activity is decreased. These studies suggest that by modifying the levels of ascorbate in the culture medium one can alter the nature of the extracellular matrix produced by the smooth muscle cells.

Amino Acid Oxidoreductases↗
PubMed1984

Changes in the connective tissue proteins, glycosaminoglycans and calcium in the arteries of the cynomolgus monkey during atherosclerotic induction and regression.

The chemical composition of the aorta, carotid, coronary and cerebral arteries of the cynomolgus monkey was determined during the induction and 'regression' of atherosclerosis. The feeding of a 2% cholesterol and 10% butter diet for 6 months resulted in extensive and severe atherosclerosis involving the aorta, carotid and coronary arteries. The involvement of these vessels was reflected by increases in arterial weight and chemical content of cholesterol, collagen, elastin, glycosaminoglycans (GAGs) and calcium. The cerebral arteries, which showed no atherosclerotic involvement, likewise showed no significant changes in weight and composition. During the 12-month regression period marked changes in the chemical composition of the involved arteries occurred and these included further increases in the collagen, GAG and calcium content of the vessels and decreases in the free and esterified cholesterol content. These changes were consistent with the gross and microscopic findings which revealed that during regression the pre-established lesions had not decreased in size but had become more fibrotic and calcified while the number of foam cells and amount of lipid contained in the lesion had decreased. During induction and regression, much of the cholesterol contained in the involved vessels appeared to be present in a crystalline form as indicated by the appearance of cholesterol clefts in the lesions. Aortic collagen was not altered with respect to amino acid composition and behavior in acrylamide gels throughout the study. However, elastin prepared by hot alkali treatment from diseased vessels, showed minor changes in amino acids during induction and marked changes during regression presumably due to the binding of glycoproteins to the elastin. The GAG composition of the involved arteries did not change during induction, whereas during regression the percent dermatan sulfate increased while the percent of heparan sulfate decreased. The over-all findings are consistent with the concept that the interaction of the connective tissue proteins with the GAGs, lipoproteins and calcium of the artery plays an important role in the development and regression of advanced atherosclerotic disease.

Animals↗