Survey of the human pancreatic beta-cell G1/S proteome reveals a potential therapeutic role for cdk-6 and cyclin D1 in enhancing human beta-cell replication and function in vivo.
OBJECTIVE
To comprehensively inventory the proteins that control the G1/S cell cycle checkpoint in the human islet and compare them with those in the murine islet, to determine whether these might therapeutically enhance human beta-cell replication, to determine whether human beta-cell replication can be demonstrated in an in vivo model, and to enhance human beta-cell function in vivo.


METHODS
Thirty-four G1/S regulatory proteins were examined in human islets. Effects of adenoviruses expressing cdk-6, cdk-4, and cyclin D1 on proliferation in human beta-cells were studied in both in vitro and in vivo models.


RESULTS
Multiple differences between murine and human islets occur, most strikingly the presence of cdk-6 in human beta-cells versus its low abundance in the murine islet. Cdk-6 and cyclin D1 in vitro led to marked activation of retinoblastoma protein phosphorylation and cell cycle progression with no induction of cell death. Human islets transduced with cdk-6 and cyclin D1 were transplanted into diabetic NOD-SCID mice and markedly outperformed native human islets in vivo, maintaining glucose control for the entire 6 weeks of the study.


CONCLUSIONS
The human G1/S proteome is described for the first time. Human islets are unlike their rodent counterparts in that they contain easily measurable cdk-6. Cdk-6 overexpression, alone or in combination with cyclin D1, strikingly stimulates human beta-cell replication, both in vitro as well as in vivo, without inducing cell death or loss of function. Using this model, human beta-cell replication can be induced and studied in vivo.
Fiaschi-Taesch
Nathalie
N
Division of Endocrinology, The University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA. taeschn@dom.pitt.edu
Bigatel
Todd A
TA
Sicari
Brian
B
Takane
Karen K
KK
Salim
Fatima
F
Velazquez-Garcia
Silvia
S
Harb
George
G
Selk
Karen
K
Cozar-Castellano
Irene
I
Stewart
Andrew F
AF
D000818:Animals; D002453:Cell Cycle; D002455:Cell Division; D019938:Cyclin D1 / Q000235:genetics / Q000502:physiology*; D051361:Cyclin-Dependent Kinase 6 / Q000235:genetics / Q000502:physiology*; D017931:DNA Primers; D016193:G1 Phase / Q000235:genetics*; D005786:Gene Expression Regulation; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000166:cytology* / Q000201:enzymology / Q000502:physiology* / Q000637:transplantation; D007700:Kinetics; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D010766:Phosphorylation; D020543:Proteome; D016160:Retinoblastoma Protein / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016196:S Phase / Q000235:genetics*; D013045:Species Specificity
D017931:DNA Primers; D007328:Insulin; D020543:Proteome; D016160:Retinoblastoma Protein; D019938:Cyclin D1; C495911:CDK6 protein, human; D051361:Cyclin-Dependent Kinase 6; D005947:Glucose