Stimulation of proteoglycan synthesis by glucuronosyltransferase-I gene delivery: a strategy to promote cartilage repair.
Osteoarthritis is a degenerative joint disease characterized by a progressive loss of articular cartilage components, mainly proteoglycans (PGs), leading to destruction of the tissue. We investigate a therapeutic strategy based on stimulation of PG synthesis by gene transfer of the glycosaminoglycan (GAG)-synthesizing enzyme, beta1,3-glucuronosyltransferase-I (GlcAT-I) to promote cartilage repair. We previously reported that IL-1beta down-regulated the expression and activity of GlcAT-I in primary rat chondrocytes. Here, by using antisense oligonucleotides, we demonstrate that GlcAT-I inhibition impaired PG synthesis and deposition in articular cartilage explants, emphasizing the crucial role of this enzyme in PG anabolism. Thus, primary chondrocytes and cartilage explants were engineered by lipid-mediated gene delivery to efficiently overexpress a human GlcAT-I cDNA. Interestingly, GlcAT-I overexpression significantly enhanced GAG synthesis and deposition as evidenced by (35)S-sulfate incorporation, histology, estimation of GAG content, and fluorophore-assisted carbohydrate electrophoresis analysis. Metabolic labeling and Western blot analyses further suggested that GlcAT-I expression led to an increase in the abundance rather than in the length of GAG chains. Importantly, GlcAT-I delivery was able to overcome IL-1beta-induced PG depletion and maintain the anabolic activity of chondrocytes. Moreover, GlcAT-I also restored PG synthesis to a normal level in cartilage explants previously depleted from endogenous PGs by IL-1beta-treatment. In concert, our investigations strongly indicated that GlcAT-I was able to control and reverse articular cartilage defects in terms of PG anabolism and GAG content associated with IL-1beta. This study provides a basis for a gene therapy approach to promote cartilage repair in degenerative joint diseases.
Venkatesan
N
N
Unité Mixte de Recherche 7561, Centre National de la Recherche Scientifique, Université Henri Poincaré-Nancy 1, Faculté de Médecine, B.P. 184, 54505 Vanoeuvre-lès-Nancy, France.
Barré
L
L
Benani
A
A
Netter
P
P
Magdalou
J
J
Fournel-Gigleux
S
S
Ouzzine
M
M
D000818:Animals; D015153:Blotting, Western; D050260:Carbohydrate Metabolism; D002356:Cartilage / Q000378:metabolism / Q000473:pathology*; D002358:Cartilage, Articular / Q000473:pathology; D019902:Chondrocytes / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D015536:Down-Regulation; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D014453:Glucuronosyltransferase / Q000235:genetics*; D006801:Humans; D007150:Immunohistochemistry; D007375:Interleukin-1 / Q000378:metabolism; D050356:Lipid Metabolism; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D010003:Osteoarthritis / Q000628:therapy*; D011509:Proteoglycans / Q000378:metabolism*; D051381:Rats; D013997:Time Factors; D014162:Transfection; D014945:Wound Healing
D018076:DNA, Complementary; D007375:Interleukin-1; D016376:Oligonucleotides, Antisense; D011509:Proteoglycans; C067806:glucuronyltransferase GlcAT-1; D014453:Glucuronosyltransferase