Objective: To evaluate the effects of long-term wear and discontinuation of the orthokeratology lenses (Orth-K) on the biological parameters of eyeballs in children with myopia.Methods: In this prospective study, a total of 308 subjects with myopia were randomized to receive Orth-K (n = 154) or single vision spectacles (SVS) (n = 154) for 12 months followed by a 1-month withdrawal period. The axial length (AL), the central corneal thickness (CCT), the anterior chamber depth (ACD) and the central lens thickness (CLT) were assessed at the baseline, 6 months, 12 months, and 13 months (1-month after lens withdrawal).Results: A total of 279 subjects completed the 13-month follow-up (142 in Orth-K group and 137 in SVS group). No statistical difference was noted in AL, CCT, ACD and CLT between the two groups at the baseline (all p > 0.05). However, compared with the baseline, the AL from the two groups became elongated 12 months after wearing Orth-K or SVS. The increase of AL in Orth-K group was 0.22 t 0.11 mm, significantly smaller than 0.35 t 0.08 mm in SVS group (p < 0.05). In addition, CCT in Orth-K group was 544.26 t 11.69 mu m at 12 months, significantly thinner than 550.49 t 12.13 mu m in SVS group (p < 0.05). Interestingly, the change in CCT between the baseline and 1-month after withdrawal of the lens was not statistically different in either group (all p > 0.05). Furthermore, at 12-months, CLT in Orth-K group was 3.35 t 0.21 mm, significantly thicker than 3.31 t 0.15 mm at baseline and thicker than 3.30 t 0.05 mm in SVS group at 12 months (all p < 0.05). Lastly, ACD was not statistically different between Orth-K and SVS groups at any time point (p > 0.05).Conclusion: Orthokeratology lenses can effectively retard axial elongation, reversibly reduce CCT, increase CLT in myopic children, but have no obvious effect on ACD, indicating that Orth-K may significantly retard myopia without noticeable myopia rebound after interruption of Orth-K.