Skin dryness and thickening are hallmarks of systemic sclerosis (SSc) disease. Aquaporins (AQPs) are plasma membrane proteins that transport glycerol and water, resulting in water retention and skin hydration. Expression of AQPs has been evaluated in human normal skin. However, expression of these proteins in SSc dermal Fibroblasts has not yet been reported. The aim of this study was to assess the expression profile of AQPs in dermal Fibroblasts of SSc patients. Fibroblast cells were extracted from SSc and healthy skin biopsies and characterized using fibroblast surface protein antibody. To evaluate the mRNA expression of AQP1, 3, 5, 7, 9, and 10 in dermal Fibroblasts, Real-time PCR was performed using SYBR Green Master mix. Immunoblotting was performed to confirm the results of Real-time PCR. Our data demonstrated that only AQP1, AQP3, and AQP9 were expressed in human skin Fibroblasts. Moreover, the expression of AQP3 mRNA and protein were significantly decreased in SSc dermal Fibroblasts compared with healthy Fibroblasts. AQP3, which involves in skin hydration and wound healing through water and glycerol transmission, is downregulated in SSc Fibroblasts. Based on previous studies and our results, it seems that SSc manifestations like skin dryness, abnormal wound healing, and fibrotic lesions may be related to downregulation of AQP3 in SSc Fibroblasts. Therefore, induction of AQP3 expression can be a potential treatment to relieve SSc skin thickness in the future.