Remodeling of the dermal extracellular matrix occurs during photoaging. Here, the effect of repetitive UVB irradiation on dermal hyaluronic acid (HA) was examined. C57/BL6 mice were chronically (182 days) irradiated with UVB, and consecutive skin biopsies were collected during the irradiation period and afterward (300 and 400 days of age). UVB caused marked loss of HA from the papillary dermis and down-regulation of HA synthase 1 (HAS1), HAS2, and HAS3 mRNA expression. In contrast, hyaluronidases (HYAL) 1, HYAL2, and HA receptor CD44 were unchanged. Furthermore, transforming growth factor β-1 (TGF-β1) and TGF-β1-receptor II expression were decreased in UVB-irradiated biopsies, and TGF-β1 strongly induced HAS1 and HAS2 expression in cultured dermal fibroblasts. Therefore, TGF-β1 might be one factor involved in UVB-induced down-regulation of HAS enzymes. In addition, total cell number and the percentage of proliferating fibroblasts in the papillary dermis of UVB-irradiated mice were decreased. Down-regulation of HAS2 by lentiviral overexpression of short hairpin RNA in vitro caused inhibition of HA synthesis, DNA synthesis, and migration of dermal fibroblasts. In conclusion, chronic UVB irradiation induces loss of HA from the dermis, thereby contributing to the quiescent phenotype of dermal fibroblasts.