Bone Morphogenic Protein-7의 복막섬유화 억제효과와 기전 = Antifibrotic Effect of BMP-7 in the Peritoneum and the Mechanism

다국어 초록 (Multilingual Abstract)

Purpose: Bone morphogenic protein (BMP)-7, a member of TGF-β1 superfamily, is an endogenous antifibrotic protein highly expressed in normal kidney. It is not known, however, whether human peritoneal mesothelial cells (HPMC) express BMP-7 or if BMP-7 protects against peritoneal fibrosis and by what mechanism. We examined the effect of BMP-7 overexpression in TGF-β1-induced epithelial-mesenchymal transition (EMT) of HPMC and in TGF-β1 signaling in HPMC to elucidate the mechanisms of antifibrotic effect of BMP-7.
Methods: Growth arrested and synchronized HPMC were stimulated with 2 ng/mL of TGF-β1 to induce EMT. HPMC were transiently transfected with adenovirus-mediated human BMP-7 (AdBMP-7) or with GFP (AdGFP). EMT was defined as downregulation of E-cadherin and upregulation of α-smooth muscle actin (SMA).
Results: HPMC constitutively expressed BMP-7 mRNA and protein. BMP-7 mRNA and protein expression were significantly inhibited by 50 mM D-glucose, 2x diluted commercial peritoneal dialysis solution, and 2 ng/ml of TGF-β1. Transfection of AdBMP-7 resulted in 2.5-fold increase in BMP-7 mRNA expression in HPMC. TGF-β1 significantly decreased E-cadherin and increased α-SMA expression in GFP transfected cells. BMP-7 overexpression effectively reversed TGF-β1-induced E- cadherin and α-SMA expression and significantly suppressed TGF-β1-induced phosphorylation of Smad2/3, ERK1/2, JNK, and p38 MAPK in HPMC as compared to GFP transfected cells.
Conclusion: BMP-7 is an endogenous antifibrotic protein and downregulation of BMP-7 in HPMC by high glucose, PD solution, and TGF-β1 may permit the development of peritoneal fibrosis during long-term PD. Our data demonstrate that BMP-7 overexpression reverses TGF-β1-induced EMT of HPMC and consequent peritoneal fibrosis possibly through inhibition of Smad2/3 and MAPK phosphorylation.

국문 초록 (Abstract)

목 적: Bone morphogenic protein (BMP)-7은 TGF-β superfamily에 속하는 내인성 항섬유화 단백질로 신장내에 대량 표현되어 있다. 그러나 사람 복막중피세포 (HPMC)가 BMP-7을 표현하는지, BMP-7이 복막섬유화를 억제하는지 또 그 기전이 무엇인지는 밝혀진바 없다. 본 연구에서는 HPMC에서 BMP-7의 과발현이 TGF-β1에 의한 상피세포-중간엽세포 이행 (EMT)과 TGF-β1 신호전달계에 미치는 영향을 관찰함으로써 BMP-7의 복막섬유화 억제 효과와 그 기전을 검색하였다.
방 법: 배양된 HPMC에 adenovirus vector를 매개로 human BMP-7 (AdBMP-7)과 GFP (AdGFP)를 transfection하였다. EMT는 2 ng/mL의 TGF-β1으로 유발하였고 E-cadherin의 감소와 α-smooth muscle actin (α- SMA)의 증가로 확인하였다. mRNA는 real time PCR로 단백은 Western blot으로 측정하였다.
결 과: HPMC는 기저 상태에서 BMP-7 mRNA와 단백을 표현하였다. 50 mM 포도당과 2배 희석한 상용 복막투석액 그리고 2 ng/mL TGF-β1은 HPMC의 BMP-7 mRNA 와 단백 표현을 의미 있게 억제하였다. AdBMP-7은 AdGFP에 비하여 BMP-7 mRNA 표현을 2.5배 증가시켰다. AdBMP-7은 TGF-β1에 의한 E-cadherin mRNA와 단백 감소 및 α-SMA mRNA와 단백 증가를 회복시키고 Smad2/3, ERK1/2, JNK, P38 MAPK의 인산화를 의미 있게 억제하였다.
결 론: HPMC에서 BMP-7은 내인성 항섬유화 단백으로 고포도당, 상용 복막투석액 그리고 TGF-β1은 모두 BMP- 7 표현을 억제함으로써 장기간의 복막투석 중 복막섬유화의 발생을 허용할 수 있음을 시사한다. BMP-7의 과발현은 TGF-β1에 의한 EMT 발생을 예방하는 동시에 Smad와 MAPK 인산화를 억제하여 Smad와 MAPK 활성억제가 HPMC의 EMT와 복막섬유화 억제에 관여할 수 있음을 시사한다.

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