Induction of proliferation in adult human β-cells is challenging. It can be accomplished by introduction of cell cycle molecules such as cyclin-dependent kinase 6 (cdk6) and cyclin D₁, but their continuous overexpression raises oncogenic concerns. We attempted to mimic normal, transient, perinatal human β-cell proliferation by delivering these molecules in a regulated and reversible manner. Adult cadaveric islets were transduced with doxycycline (Dox)-inducible adenoviruses expressing cdk6 or cyclin D₁. End points were cdk6/cyclin D₁ expression and human β-cell proliferation, survival, and function. Increasing doses of Dox led to marked dose- and time-related increases in cdk6 and cyclin D₁, accompanied by a 20-fold increase in β-cell proliferation. Notably, Dox withdrawal resulted in a reversal of both cdk6 and cyclin D₁ expression as well as β-cell proliferation. Re-exposure to Dox reinduced both cdk/cyclin expression and proliferation. β-Cell function and survival were not adversely affected. The adenoviral tetracycline (tet)-on system has not been used previously to drive human β-cell proliferation. Human β-cells can be induced to proliferate or arrest in a regulated, reversible manner, temporally and quantitatively mimicking the transient perinatal physiological proliferation that occurs in human β-cells.