Regulation of Lysyl Oxidase mRNA in Dermal Fibroblasts from Normal Donors and Patients with Inherited Connective Tissue Disorders

Lysyl oxidase (LO) is an extracellular copper-dependent enzyme that catalyzes the initial reaction in the formation of lysine or hydroxylysine-derived crosslinks during collagen biosynthesis. We have isolated a cDNA for human LO from skin fibroblast poly(A+)RNA by PCR using primers based on the recently published sequence of human LO. This cDNA probe detects a major mRNA of 4.2 kh on Northern blots of RNA from normal fibroblasts. The level of LO mRNA was not significantly affected by cell density or by ascorbate treatment. Treatment of skin fibroblasts with hydralazine (50μM), which increases the mRNAs for both the α and the β subunits of prolyl hydroxylase (PH) and the mRNAs for lysyl hydroxylase, also increased LO mRNA by fourfold over a 72-h time course. In contrast, hydralazine dramatically decreased the mRNAs for α1(I) collagen. Administration of minoxidil (500 μM), which specifically decreases LH activity without affecting PH activity or collagen biosynthesis in skin fibroblasts, stimulated the level of LO mRNA. Neither the administration of penicillamine (100 μM), which interferes with collagen cross-linking, nor the administration of β-aminopropionitrile, which is a strong irreversible inhibitor of LO, to fibroblasts significantly changed the levels of LO mRNA over a 72-h time course. However, bleomycin (0.6 μg/ml) significantly decreased the 4.2-kb LO mRNA in contrast to the levels of the α1(I) collagen mRNAs, which were unchanged. No significant change was observed in the steady-state levels of LO mRNAs in fibroblasts isolated from patients with certain connective tissue disorders, including Marfan syndrome, Menkes disease, cutis laxa, and pseudoxanthoma elasticum.