Analysis of vascular gene expression in arthritic synovium by laser-mediated microdissection.
OBJECTIVE
In rheumatoid arthritis (RA), formation of new blood vessels is necessary to meet the nutritional and oxygen requirements of actively proliferating synovial tissue. The aim of this study was to analyze the specific synovial vascular expression profiles of several angiogenesis-related genes as well as CD82 in RA compared with osteoarthritis (OA), using laser-mediated microdissection (LMM).


METHODS
LMM and subsequent real-time polymerase chain reaction were used in combination with immunohistochemical analysis for area-specific analysis of messenger RNA (mRNA) and protein expression of vascular endothelial growth factor (VEGF), VEGF receptor 1 (VEGFR-1), VEGFR-2, hypoxia-inducible factor 1alpha (HIF-1alpha), HIF-2alpha, platelet-derived growth factor receptor alpha (PDGFRalpha), PDGFRbeta, inhibitor of DNA binding/differentiation 2 (Id2), and CD82 in RA and OA synovial microvasculature and synovial lining.


RESULTS
Expression of Id2 mRNA was significantly lower in RA synovial vessels compared with OA synovial vessels (P=0.0011), whereas expression of VEGFR-1 was significantly higher in RA (P=0.0433). No differences were observed for the other parameters. At the protein level, no statistically significant differences were observed for any parameter, although Id2 levels were 2.5-fold lower in RA (P=0.0952). However, the number of synovial blood vessels and the number of VEGFR-2-expressing blood vessels were significantly higher in RA compared with OA.


CONCLUSIONS
Our results underscore the importance of area-specific gene expression analysis in studying the pathogenesis of RA and support LMM as a robust tool for this purpose. Of note, our results indicate that previously described differences between RA and OA in the expression of angiogenic molecules are attributable to higher total numbers of synovial and vascular cells expressing these molecules in RA rather than higher expression levels in the individual cells.
Hashimoto
Atsushi
A
Department of Medicine and Rheumatology, Justus-Liebig-University of Giessen, Giessen, and University Hospital Regensburg, Germany.
Tarner
Ingo H
IH
Bohle
Rainer M
RM
Gaumann
Andreas
A
Manetti
Mirko
M
Distler
Oliver
O
Steinmeyer
Jürgen
J
Ulfgren
Ann-Kristin
AK
Schulz
Andreas
A
Gay
Steffen
S
Müller-Ladner
Ulf
U
Neumann
Elena
E
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001172:Arthritis, Rheumatoid; D051792:Basic Helix-Loop-Helix Proteins; D005260:Female; D015870:Gene Expression; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism; D051797:Inhibitor of Differentiation Protein 2 / Q000235:genetics / Q000378:metabolism; D051861:Kangai-1 Protein / Q000235:genetics / Q000378:metabolism; D008297:Male; D042282:Microdissection; D008875:Middle Aged; D009389:Neovascularization, Pathologic / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D010003:Osteoarthritis; D012333:RNA, Messenger / Q000378:metabolism; D020796:Receptor, Platelet-Derived Growth Factor alpha / Q000235:genetics / Q000378:metabolism; D020797:Receptor, Platelet-Derived Growth Factor beta / Q000235:genetics / Q000378:metabolism; D013583:Synovial Membrane / Q000098:blood supply* / Q000378:metabolism / Q000473:pathology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics* / Q000378:metabolism; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000235:genetics* / Q000378:metabolism; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics* / Q000378:metabolism; D000099162:Endothelial PAS Domain-Containing Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D051797:Inhibitor of Differentiation Protein 2; D051861:Kangai-1 Protein; D012333:RNA, Messenger; D020796:Receptor, Platelet-Derived Growth Factor alpha; D020797:Receptor, Platelet-Derived Growth Factor beta; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; D040281:Vascular Endothelial Growth Factor Receptor-1; D040301:Vascular Endothelial Growth Factor Receptor-2; D000099162:Endothelial PAS Domain-Containing Protein 1; C497688:CD82 protein, human; C497442:HIF1A protein, human; C497450:ID2 protein, human; C467484:VEGFA protein, human