Effect of lysyl hydroxylase inhibitor, minoxidil, on ultrastructure and behavior of c

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Effect of lysyl hydroxylase inhibitor, minoxidil, on ultrastructure and behavior of cultured rabbit subconjunctival fibroblasts.


Abstract

BACKGROUND:

Minoxidil is an inhibitor of lysyl hydroxylase, an enzyme involved in collagen production, and decreases collagen production in vitro. We investigated the in vitro effects of minoxidil on behavior such as proliferation and migration of rabbit subconjunctival fibroblasts (SCFs). The ultrastructural effect of the drug on SCFs was also examined.
METHODS:

Proliferation of SCFs and closure of the defect produced in monolayer cultures in the presence or absence of minoxidil was studied. The ultrastructure of SCFs treated with minoxidil was also examined.
RESULTS:

Minoxidil inhibited SCF proliferation and the closure of the defect produced in monolayer cell sheets. Ultrastructural observations revealed extensive areas of irregularly dilated endoplasmic reticulum in cells treated with minoxidil, indicating the accumulation of protein, probably underhydroxylated collagen precursors, in the cisternae of endoplasmic reticulum.
CONCLUSIONS:

The results indicated that minoxidil attenuated cellular activities of SCFs such as proliferation and migration in vitro. The exact mechanism of the inhibitory effects of minoxidil on these cellular activities is unknown. The findings suggest that the drug might help to prevent bleb scarring after glaucoma filtering surgery.

http://www.ncbi.nlm.nih.gov/pubmed/7672621

 

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Proof that Minoxidil inhibit collagen production and fibrosis... again..


Minoxidil exerts different inhibitory effects on gene expression of lysyl hydroxylase 1, 2, and 3: implications for collagen cross-linking and treatment of fibrosis.

Zuurmond AM, van der Slot-Verhoeven AJ, van Dura EA, De Groot J, Bank RA.
Source

TNO Quality of Life, Biomedical Research, Department of Tissue Repair, P.O. Box 2215, 2301 CE Leiden, The Netherlands.

Abstract

Collagen deposits in fibrotic lesions often display elevated levels of hydroxyallysine (pyridinoline) cross-links. The relation between the occurrence of pyridinoline cross-links and the irreversibility of fibrosis suggests that these cross-links contribute to the aberrant accumulation of collagen. Based on its inhibitory effect on lysyl hydroxylase activity minoxidil has been postulated to possess anti-fibrotic properties by limiting the hydroxylysine supply for hydroxyallysine cross-linking. However, to interfere with hydroxyallysine cross-linking specifically lysyl hydroxylation of the collagen telopeptide should be inhibited, a reaction predominantly catalysed by lysyl hydroxylase (LH) 2b. In this study, we demonstrate that minoxidil treatment of cultured fibroblasts reduces LH1>>LH2b>LH3 mRNA levels dose-and time-dependently, but has essentially no effect on the total number of pyridinoline cross-links in the collagen matrix. Still the collagen produced in the presence of minoxidil displays some remarkable features: hydroxylation of triple helical lysine residues is reduced to 50% and lysylpyridinoline cross-linking is increased at the expense of hydroxylysylpyridinoline cross-linking. These observations can be explained by our finding that LH1 mRNA levels are the most sensitive to minoxidil treatment, corroborating that LH1 has a preference for triple helical lysine residues as substrate. In addition, the non-proportional increase in cross-links (20-fold) with respect to the decrease in lysyl hydroxylation state of the triple helix (2-fold) even suggests that LH1 preferentially hydroxylates triple helical lysine residues at the cross-link positions. We conclude that minoxidil is unlikely to serve as an anti-fibroticum, but confers features to the collagen matrix, which provide insight into the substrate specificity of LH1.


Minimum structural requirements for minoxidil inhibition of lysyl hydroxylase in cultured fibroblasts.

Murad S, Walker LC, Tajima S, Pinnell SR.
Source

Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710.

Abstract

The structural features of minoxidil contributing to its inhibitory effect on lysyl hydroxylase in cultured fibroblasts were investigated. Minoxidil analogs in which the pyrimidine ring (two nitrogens) was replaced with a pyridine ring (one nitrogen) or a sym-triazine ring (three nitrogens) suppressed lysyl hydroxylase activity without affecting prolyl hydroxylase activity, as did an analog in which the piperidine ring was replaced with an N,N-diethyl group. By contrast, minoxidil analogs bearing an N-monoalkyl (ethyl or butyl) group in place of the piperidine substituent failed to suppress lysyl hydroxylase activity. The results indicate that nitroxides of pyridine and triazine, in addition to pyrimidine, having an ortho amino group can act as specific inhibitors of lysyl hydroxylase in the cell. The minimum structural requirement for inhibitory activity appears to be an organic moiety containing a tertiary nitrogen para to the nitroxide oxygen, a condition that is best fulfilled by the piperidine ring in minoxidil. Hydroxylation of minoxidil at the 3 or 4 position of the piperidine ring had no impact on its ability to inhibit the post-translational hydroxylation of lysine during collagen biosynthesis. Fibroblasts treated with minoxidil, 3'-hydroxyminoxidil, or 4'-hydroxyminoxidil synthesized a collagen specifically deficient in hydroxylysine by approximately 70%, which completely accounted for the diminished lysyl hydroxylase activity. The percentage of total proteins synthesized as collagen was reduced minimally by minoxidil but not by 3'- or 4'-hydroxyminoxidil. The studies offer a potential means for therapeutic intervention of excessive collagen deposition during fibrosis, using minoxidil or preferably its hydroxy derivatives to limit the supply of hydroxylysine for collagen crosslink formation.

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