Protocollagen Proline Hydroxylase Activity in the Skin of Normal Human Subjects and of Patients with Scleroderma

Uitto, Jouni; Halme, Jouko; Hannuksela, Matti; Peltokallio, P; Kivirikko, Kari I.

doi:10.3109/00365516909077656

Cited by 99 publications

(27 citation statements)

References 27 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Selected steps in collagen synthesis have been analyzed in scleroderma. Proline hydroxylase levels have been variably elevated in skin biopsy material from scleroderma patients, as well as skin tissue from several other connective tissue syndromes (21,22). The incorporation of labeled proline, estimated either as overall incorporation or as hydroxyproline counts, has been increased in about 50% of the observations made (23).…”

Section: Resultsmentioning

confidence: 81%

Increased Collagen Synthesis by Scleroderma Skin Fibroblasts In Vitro A POSSIBLE DEFECT IN THE REGULATION OR ACTIVATION OF THE SCLERODERMA FIBROBLAST

Ec¹

1974

J. Clin. Invest.

528

177

View full text Add to dashboard Cite

A B S T R A C T Cultures of dividing skin fibroblasts from normal and sclerodermatous human skin have permitted estimations of soluble collagen concentration, net collagen accumulation, cell-doubling times, and the comparison of morphologic and ultrastructural characteristics. In vitro, the scleroderma fibroblast produces more soluble collagen, synthesizes collagen more rapidly, and fourfold more of its protein synthetic activity is directed to collagen production than in the normal skin fibroblast. Cell-doubling times and morphologic and ultrastructural observations of cells in culture have not provided clues to the nature of the biologic defect in the regulation or activation of collagen synthesis by the scleroderma fibroblast.

show abstract

Section: Resultsmentioning

confidence: 81%

Increased Collagen Synthesis by Scleroderma Skin Fibroblasts In Vitro A POSSIBLE DEFECT IN THE REGULATION OR ACTIVATION OF THE SCLERODERMA FIBROBLAST

Ec¹

1974

J. Clin. Invest.

528

177

View full text Add to dashboard Cite

show abstract

“…human prolyl hydroxylase was isolated as a pure protein. Foetal tissues were used as starting material for this, as it was earlier reported that the enzyme activity is markedly higher in foetal than in adult human tissues [29]. The human enzyme was found to be markedly similar to the chick enzyme [12,13] with respect to its molecular weight, subunit structure, amino acid composition, and specific activity.…”

Section: Discussionmentioning

confidence: 99%

“…However, several modifications were necessary to obtain optimal results with the human tissues. Although most of the prolyl hydroxylase activity can be extracted from 12-day-old chick embryos with 0.01 M potassium chloride [6], the use of a higher salt concentration [29] and the addition of Triton X-100 [24,30,31] were found to be necessary for optimal extraction of the enzyme from human foetal tissues. The ammonium sulphate fractionation step [32] was also slightly modified to increase the recovery of enzymic activity.…”

Section: Purification Of Human Prolyl Hydroxylusementioning

confidence: 99%

Human Prolyl Hydroxylase. Purification, Partial Characterization and Preparation of Antiserum to the Enzyme

1975

Self Cite

View full text Add to dashboard Cite

Prolyl hydroxylase was purified from human foetal skin and from a mixture of human foetal tissues by the affinity chromatography procedure using poly(L-proline). The enzyme from both sources was pure, when examined by polyacrylamide gel electrophoresis, as a native protein or in the presence of sodium dodecylsulphate, and enzyme activity recovery varied from 38% to 70% with seven enzyme preparations. The enzyme synthesized from 61.0 pmol to 82.7 pmol hydroxyproline mg protein-' h-' at 37 "C with a saturating concentration of (Pro-Pro-Gly), as substrate. The molecular weight of the enzyme was identical with that of the chick prolyl hydroxylase when studied by gel filtration, and the molecular weights of the subunits of the enzyme were about 61 000 and 64000 as determined by sodium dodecylsulphate -polyacrylamide gel electrophoresis. The amino acid composition of the human enzyme was very similar to that of the chick prolyl hydroxylase.Antisera to human and chick prolyl hydroxylases were prepared in rabbits. A single precipitin line was seen between the antiserum to human prolyl hydroxylase and the human enzyme in double immunodiffusion, and no cross-reactivity was detected between the human and chick enzymes by this technique. However, a distinct cross-reactivity was observed between the human and chick enzymes in inhibition experiments.

show abstract

“…Early evidence demonstrated an increased rate of collagen synthesis in the skin ofpatients with scleroderma (5). This conclusion was based on the observations that the activity of prolyl hydroxylase, a key enzyme in collagen biosynthesis, was increased in skin biopsy specimens taken from patients with active disease (6,7). Furthermore, the incorporation of radioactive proline into cutaneous collagen and the synthesis of radioactive hydroxyproline were also increased under tissue culture conditions (8,9).…”

Section: Introductionmentioning

confidence: 99%

Scleroderma

Uitto¹,

Bauer²,

Eisen³

1979

J. Clin. Invest.

223

View full text Add to dashboard Cite

A B S T R A C T To assess potential abnormalities in collagen metabolism in systemic scleroderma, skin fibroblast lines from patients with this disease were established and compared to control cell lines derived from healthy subjects. For studies on the biosynthesis of procollagen, the cells were incubated with [14C]proline in a medium supplemented with ascorbic acid and ,8-aminopropionitrile, and the synthesis of nondialyzable [14C]hydroxyproline, in relation to DNA or cell protein, was taken as an index of procollagen formation. Five of eight scleroderma fibroblast cell lines demonstrated procollagen biosynthesis rates significantly higher than the controls, and the mean rate of procollagen synthesis by scleroderma fibroblasts was about twice that of the control cells. Control experiments demonstrated that the specific activity of the intracellular free proline was not different in scleroderma and control fibroblasts, and the mean population doubling times of the scleroderma and the control fibroblast cell lines were the same. The relative synthesis of the genetically distinct procollagens was examined by isolating type I and type III procollagens from the cell culture medium using DEAE-cellulose chromatography. The ratios of type I/III procollagens in scleroderma cell lines did not differ from the controls. The helical stability of the collagenous portion of type I and type III procollagens, estimated by the resistance of 14C-collagen to limited proteolytic digestion with pepsin under nondenaturing conditions, was the same in both A preliminary report of part of this work has been presented at

show abstract

Protocollagen Proline Hydroxylase Activity in the Skin of Normal Human Subjects and of Patients with Scleroderma

Cited by 99 publications

References 27 publications

Increased Collagen Synthesis by Scleroderma Skin Fibroblasts In Vitro A POSSIBLE DEFECT IN THE REGULATION OR ACTIVATION OF THE SCLERODERMA FIBROBLAST

Increased Collagen Synthesis by Scleroderma Skin Fibroblasts In Vitro A POSSIBLE DEFECT IN THE REGULATION OR ACTIVATION OF THE SCLERODERMA FIBROBLAST

Human Prolyl Hydroxylase. Purification, Partial Characterization and Preparation of Antiserum to the Enzyme

Scleroderma

Contact Info

Product

Resources

About