Objective: To investigate the effect of overnight orthokeratology (OOK) on the ocular surface and changes  in meibomian gland morphology in children and adolescents with myopia. Methods: In this prospective  cohort study, choosing myopia children from the Second Affiliated Hospital of Zhejiang Chinese Medical University, sixty myopic patients (60 eyes) with OOK were selected as the OOK group, and 60 patients (60  eyes) with eye glasses were selected as the glasses group. The following tests were performed before OOK  and eyeglass wear and after 1, 3, 6 and 12 months of wear. Tests included ocular surface disease index  (OSDI) questionnaire, noninvasive tear break-up time (NIBUT), tear meniscus height (TMH), corneal  epithelial staining (CSF), and meiboscore using noncontact meibography. The effect of OOK wear on the  ocular surface and changes that occurred were evaluated and summarized by comparing the differences  between these two groups. Results: There were no significant differences in the OSDI score or tear  meniscus height between these two groups. The NIBUT of the OOK group showed a significant decrease  after 6 and 12 months of lens wear (both P<0.05). There were no statistically significant differences at other  time points. Corneal fluorescence staining in the OOK group increased after wearing the lenses (χ2 =19.511,  P=0.013), but there were no statistically significant differences at the other time points. The score of the  meibomian gland orifice in the OOK group was slightly increased before wearing the lenses and after 1, 3  and 6 months, but the differences were not statistically significant. The final 12 months were significantly  higher than before wearing eye glasses and 1 month after wearing eye glasses, and the difference was  statistically significant (P<0.05). Conclusions: It is relatively safe to wear OOK lenses overnight. Shortterm wear of OOK will not cause discomfort and it will not reduce tear meniscus height or the amount of  tear secretion in the short term, however, it reduces the stability of the tear film and increases the corneal  fluorescence staining score. OOK lens wear affected meibomian gland function. In particular, the upper  meibomian gland changed first.