Reduced matrix metalloproteinase and collagen transcription mediated by the TGF‐β/Smad pathway in passaged normal human dermal fibroblasts

BACKGROUND: Transforming growth factor-beta (TGF-beta) is a major regulator of extracellular matrix (ECM) events, particularly collagen production. AIM: We explored whether the expression of matrix metalloproteinases (MMPs) and collagen are transcriptionally regulated by the TGF-beta and Smad signaling pathways, and the roles played by NF-kappaB and mitogen-activated protein kinase (MAPK) signaling in normal, aged, human dermal fibroblasts. METHODS: We quantified mRNA and protein expression using real-time PCR and immunoblotting of proteins from cells in passage 5-15. RESULTS: The levels of mRNAs encoding TGF-beta1, TGF-beta3, and TGF-beta receptor type I (TGFbeta RI) decreased with increasing passage number. The levels of mRNAs encoding TGF-beta2, TGFbeta RII, and TGFbeta RIII increased to passage 10 but decreased by passage 15. The levels of mRNAs encoding Smad-2, -3, -4, and -7 decreased with increasing passage number. The level of mRNA encoding MMP-1 increased with increasing passage number, and the levels of mRNAs encoding MMP-2, TIMP-1, and TIMP-2 increased to passage 10 but decreased by passage 15. The levels of mRNAs encoding collagen types I and II decreased with increasing passage number. At the protein level, NF-kappaB, IkappaBalpha, p38, ERK, Akt, and JNK became increasingly phosphorylated at higher passage numbers. CONCLUSION: Our results suggest that reductions in the expression levels of MMPs and collagen types I and III in aging human dermal fibroblasts reflect reduced expression of TGF-beta/Smad and TGF-beta receptors, thus compromising the TGF-beta receptor-binding capacity of fibroblasts; the NF-kappaB and Akt-JNK/MAPK signaling pathways may play active roles in this process.