Background The principal function of skin is to provide a barrier to water loss and percutaneous penetration of exogenous substances. Any compromise of the skin homeostasis can lead to dryness, itchiness, or even pathogenic conditions. This study aimed to explore the relationship among psychological stress, skin homeostasis, sleep quality, and emotion. MethodsThis prospective observational study was conducted using a random sample of medical students from Shanghai Medical College, Fudan University. A questionnaire including the validated Chinese version of Pittsburgh Sleep Quality Index (PSQI), Beck Anxiety Inventory (BAI), and Beck Depression Inventory-II (BDI) was distributed. The skin barrier function was measured by the transepidermal water loss (TEWL), hydration of skin. Dermoscopic images of the nose and infraorbital skin were obtained. We compared the skin homeostasis, sleep quality, and emotional state of medical students at different time points of the semester and explored the correlation between the variables. Results As the semester progressed, the sleep quality and severity of anxiety of medical students got significantly worse, and they were closely related to the impaired skin barrier function. Both irregular schedule and stress can aggravate dark circles. Conclusions Mental stress can do harm to skin barrier through poor sleep and severe anxiety. The amount of sebum was positively correlated with the severity of anxiety while blackheads are mainly influenced by season change. Dermoscopy has a unique role in assessing blackheads and dark circles.