Senescent dermal fibroblasts negatively influence fibroblast extracellular matrix-related gene expression partly via secretion of complement factor D.

Tomonobu Ezure,Satoshi Amano,Misato Sugahara

doi:10.1002/biof.1512

Abstract

Aging is associated with a decrease of extracellular matrix and an increase of senescent cells in the dermal layer. Here, to examine whether and how senescent cells are involved in aging‐related deterioration of the dermal layer, we cocultured dermal young fibroblasts (low‐passage number) with senescent cells (high‐passage number) in Transwells, in which the two cell types are separated by a semipermeable membrane. Young fibroblasts in coculture showed decreased collagen type I alpha 1 chain and elastin gene expression, and increased matrix metalloproteinase 1 (MMP1) gene expression. To identify causative factors, we compared gene expression of young and senescent cells and selected candidate secretory factors whose expression was increased by ≥2.5 in senescent fibroblasts. Then, we used siRNAs to knock down each of the 11 candidate genes in senescent fibroblasts in the coculture system. Knockdown of complement factor D (CFD) in senescent fibroblasts significantly reduced the increase of MMP1 in the cocultured young fibroblasts. In monocultures, treatment of young fibroblasts with CFD resulted in increased MMP1 gene expression, while knockdown of CFD in senescent fibroblasts decreased MMP1 gene expression. In addition, production of CFD was increased in culture medium of untreated senescent fibroblasts. Furthermore, CFD gene and protein expression were increased in the dermal layer of skin specimens from aged subjects (>70 years old), compared to young subjects (<20 years old). Overall, these results suggest that senescent cells negatively influence matrix production and promote degradation of nearby fibroblasts in the dermal layer, in part through secretion of CFD.

Highlights

Cellular senescence is associated with reduced cell proliferation, and has a critical influence on the response to cellular damage, inhibiting the progression of aberrant cells.[1,2] Further, wileyonlinelibrary.com/journal/biofBioFactors. 2019;45:556–562
Herefore, in order to clarify the role of secretory factors released from senescent fibroblasts in the dermal layer, we investigated the effects of senescent dermal fibroblasts on young dermal fibroblasts by using a coculture system in which the two types of cells are separated by a semipermeable membrane
CFDdirected siRNA almost completely blocked complement factor D (CFD) mRNA expression in the senescent fibroblasts (Figure 2c). This knockdown of CFD in senescent fibroblasts significantly reduced the increment of matrix metalloproteinase 1 (MMP1) gene expression in cocultured young fibroblasts by about 50% (Figure 2d), while there was no significant change in collagen type I alpha 1 chain (COL1A1) or elastin gene expression in the young fibroblasts

Summary

| INTRODUCTION

Senescent cells contribute to age-associated tissue disorders, such as impaired tissue homeostasis and tumorigenesis, because of the impairment of tissue regeneration.[3,4]. When old mice were surgically linked to young mice so that their blood mutually circulated, some tissues of the young mice developed agingassociated phenotypes, such as delay of regeneration after injury. This effect was mediated by complement factor C1q.9. Senescent cells have been reported to increase with aging in the dermal layer.[16]. It is still not clear whether and how senescent cells and SASP contribute to skin aging. We aimed to identify the factor(s) mediating these effects

| EXPERIMENTAL PROCEDURES

| RESULTS

Findings

| DISCUSSION