Arthritis & Rheumatism, Volume 60,
October 2009 Abstract Supplement
The 2009 ACR/ARHP Annual Scientific Meeting
Philadelphia October 16-21, 2009.
Specific Histone Deacetylases (HDACs) Modulate Fibrosis and Angiogenesis in Systemic Sclerosis (SSc)
Hemmatazad, Hossein, Maurer, Britta, Rodrigues, Hanna Maciejewska, Gay, Renate E., Michel, Beat A., Gay, Steffen, Distler, Oliver
Recently, we have shown that Trichostatin A (TSA) does not only inhibit the activity of HDACs but also modulates significantly the transcriptional expression of two isoforms of HDACs, HDAC3 and 7. While the expression of HDAC7 is almost completely down-regulated by TSA, HDAC3 expression is up-regulated. Furthermore, we demonstrated that silencing of HDAC7 has anti-fibrotic effects in skin fibroblasts from patients with SSc. However, there are reports indicating that silencing of HDAC7 affects angiogenesis in human umbilical vein endothelial cells (HUVECs). Since microvascular damage is characteristic for SSc, we investigate here the role of HDAC3 in fibrosis and in angiogenesis in human uterine microvascular endothelial cells (HUMECs).
Over-expression of HDAC3 in SSc skin fibroblasts and HUMECs was achieved by transfecting a HDAC3-expression vector/plasmid into cells using Amaxa nucleotransfection. An empty vector was used as control. Gene expression patterns of collagen types I and III, connective tissue growth factor (CTGF) and intercellular adhesion molecule-1 (ICAM-1) were analyzed by Sybr Green Real-time PCR. To assess the expression on the protein level, antibodies against collagen types I and III, CTGF and ICAM-1 were used for Western blot. Endothelial capillary sprouting was assessed using tube formation assay.
After over-expression of HDAC3 in skin fibroblasts from patients with SSc, the transcription of collagen types I and III was reduced by 38 ± 16 % and 25 ± 20 % (n=6 each, p<0.05) respectively. These changes could be confirmed on the protein level by Western blot. In addition, the mRNA expression of other pro-fibrotic molecules such as CTGF remained unchanged in cells over-expressing HDAC3, while the expression of ICAM-1 was significantly reduced (50 ± 20 %, n=4, p<0.05). Western blot confirmed the results on the protein level. Over-expression of HDAC3 did not affect the expression levels of HDAC7, suggesting that HDAC3 exerts its anti-fibrotic effects independently of HDAC7. Using the tube formation assay, over-expression of HDAC3 did not inhibit the angiogenesis in HUMECs. However, in HDAC3 over-expressing cells, the accumulative length and number of tubes were increased to 204 ± 46 and 185 ± 52 % (p<0.05), after 18h of seeding the cells on Matrigel, compared to cells transfected with the empty vector.
Based on the fact that the altered angiogenesis is a major factor in the pathogenesis of SSc, we could show here that the over-expression of HDAC3 improves angiogenesis and has anti-fibrotic effects in SSc.
To cite this abstract, please use the following information:
Hemmatazad, Hossein, Maurer, Britta, Rodrigues, Hanna Maciejewska, Gay, Renate E., Michel, Beat A., Gay, Steffen, et al; Specific Histone Deacetylases (HDACs) Modulate Fibrosis and Angiogenesis in Systemic Sclerosis (SSc) [abstract]. Arthritis Rheum 2009;60 Suppl 10 :1813