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Adipocyte renewal from preadipocytes has been shown to occur throughout life and to contribute to obesity, yet very little is known about the molecular circuits that control preadipocyte expansion. The soluble form of the preadipocyte factor (also known as pref-1) delta-like 1 homolog (DLK1(S)) is known to inhibit adipogenic differentiation; however, the impact of DLK1 isoforms on preadipocyte proliferation remains to be determined. We generated preadipocytes with different levels of DLK1 and examined differentially affected gene pathways, which were functionally tested in vitro and confirmed in vivo. Here, we demonstrate for the first time that only membrane-bound DLK1 (DLK1(M)) exhibits a substantial repression effect on preadipocyte proliferation. Thus, by independently manipulating DLK1 isoform levels, we established that DLK1(M) inhibits G1-to-S-phase cell cycle progression and thereby strongly inhibits preadipocyte proliferation in vitro. Adult DLK1-null mice exhibit higher fat amounts than wild-type controls, and our in vivo analysis demonstrates that this may be explained by a marked increase in preadipocyte replication. Together, these data imply a major dual inhibitory function of DLK1 on adipogenesis, which places DLK1 as a master regulator of preadipocyte homeostasis, suggesting that DLK1 manipulation may open new avenues in obesity treatment. [ABSTRACT FROM AUTHOR]

An inducible nitric oxide (NO) synthase isoform (iNOS) is specifically induced in the beta-cells of interleukin (IL)-1 beta-exposed rat islets, suggesting a role for NO in the pathogenesis of type I diabetes. The aim of this study was to clone and characterize iNOS cDNA from cytokine-exposed islets. Neither NO production nor iNOS transcription could be detected in rat islets or in rat insulinoma RIN-5AH beta-cells cultured in the absence of cytokines. Addition of IL-1 beta alone or in combination with tumor necrosis factor-alpha induced a concentration- and time-dependent expression of the iNOS gene and associated NO production (measured as nitrite) from both islets and RIN cells. iNOS transcripts were cloned by reverse transcriptase-polymerase chain reaction from the cytokine-exposed rat islets and RIN cells, and DNA sequence analysis revealed a near 100% identity to the recently published iNOS cDNA cloned from cytokine-exposed rat hepatocytes and smooth muscle cells. Recombinant rat islet iNOS was transiently and stably expressed in human kidney 293 fibroblasts, and the high enzymatic activity was inhibited by addition of the L-arginine analogs, N omega-nitro-L-arginine methyl ester and aminoguanidine. Two-dimensional gel electrophoresis revealed the recombinant iNOS as a series of spots with the expected molecular mass of 131 kDa and pI values in the range of 6.8 to 7.0. In conclusion, the IL-1 beta-induced iNOS cloned and expressed from rat islets and RIN cells is encoded by the same transcript as the iNOS induced in other cell types.