Growth-arrested 3T3-L1 preadipocytes rapidly express CCAAT/enhancer-binding protein-␤ (C/EBP␤) upon hormonal induction of differentiation. However, the DNA binding activity of C/EBP␤ is not activated until the cells synchronously reenter S phase during the mitotic clonal expansion (MCE) phase of differentiation. In this period, C/EBP␤ is sequentially phosphorylated by MAPK and glycogen synthase kinase-3␤, inducing C/EBP␤ DNA binding activity and transcription of its target genes. Because the DNA binding activity of C/EBP␤ is further enhanced by oxidation in vitro, we investigated how redox state affects C/EBP␤ DNA binding and MCE during adipogenesis. When 3T3-L1 cells were treated with H 2 O 2 and hormonal stimuli, differentiation was accelerated with increased expression of peroxisome proliferator-activated receptor ␥. Interestingly, cell cycle progression (S to G 2 /M phase) was markedly enhanced by H 2 O 2 , whereas antioxidants caused an S phase arrest during the MCE. H 2 O 2 treatment resulted in the early appearance of a punctate pattern observed by immunofluorescent staining of C/EBP␤, which is a hallmark for C/EBP␤ binding to regulatory elements, whereas a short antioxidant treatment rapidly dispersed the centromeric localization of C/EBP␤. Consistently, reactive oxygen species production was increased during 3T3-L1 differentiation. Our results indicate that redox-induced C/EBP␤ DNA binding activity, along with the dual phosphorylation of C/EBP␤, is required for the MCE and terminal differentiation of adipocytes.

Apoptosis is characterized by a series of distinct morphological changes including cell shrinkage, membrane bleebing, nuclear breakdown, and DNA fragmentation, and is essential for the maintenance of tissue homeostasis and the elimination of harmful cells.1,2) An apoptotic death stimulus activates caspases, the major executioners of this process, either directly or via the activation of the mitochondrial death program.3,4) The mitochondria-mediated pathway involves an increase of mitochondrial permeability and the release of cytochrome c into the cytosol. 5,6) Cytochrome c promotes the activation of caspase-3 in the cytosol, resulting in the cleavage of an inactive 32 kDa precursor into a 17 kDa and an 11 kDa subunit, which dimerize to form an active enzyme. 7)During the progression of apoptosis, caspase-3 is one of the strongest candidates for the cleavage of 116 kDa native poly (ADP-ribose) polymerase (PARP) into an 86 kDa and a 36 kDa fragment.8) The cleavage of this PARP protein ultimately results in cellular, morphological, and biochemical alterations of apoptosis. 9)Dong-Chong-Xia-Cao in Chinese, which translates as "winter worm and summer grass," is an entomogenous fungus that colonizes the larvae or pupae of insects. It includes many different genera, such as Cordyceps, Paecilomyces, Torrubiella, and Podonectria. The entomopathogenic mycelia naturally grow in the intestine of pupa or larva in the autumn, and fruiting bodies of the fungus protrude from the body in the following summer.10) The fungal fruiting body and the caterpillar host are chemically similar, have similar pharmacological properties, and are traditionally dried and consumed together.Cordyceps sinensis and Cordyceps militaris are representative insect-born fungi of the genus Cordyceps used for medicinal purposes in eastern Asian countries. The known bioactive compounds of these Cordyceps species include cordycepin, 11) ophicordin, 12) a polysaccharide, 13) galactomannan, 14) and L-tryptophan. 15) The recent scientific evaluation of Cordyceps sinensis has confirmed the efficacy of this extract in the treatment of cardiovascular and endocrine disorders, 16) in immune enhancement, 17) in hepatic protection, 18) in the inhibition of tumor growth, 19) and in relieving fatigue and stress. 20)Cordyceps militaris, together with Paecilomyces japonica, another entomopathogenic fungi, has been listed as a food additive in Korea since 2000,21) and is often used in different types of tonic drinks sold in Korea due to its folkloric activities, which have not been verified scientifically. Although some recent studies have reported its effectiveness in inhibiting angiogenesis, 22) cancer cell growth, 23,24) and inflammation, 25) scientific evidences of the biological activities of Cordyceps militaris are still very limited compared to other frequently used insect-born fungi of the Cordyceps species, such as Cordyceps sinensis. It was previously reported by these authors that the hot water extracts of Cordyceps militaris of silkworm pupa (CMP) or silkwo...

KLF8 (Krüppel-like factor 8) is a zinc-finger transcription factor known to play an essential role in the regulation of the cell cycle, apoptosis, and differentiation. However, its physiological roles and functions in adipogenesis remain unclear. In the present study, we show that KLF8 acts as a key regulator controlling adipocyte differentiation. In 3T3-L1 preadipocytes, we found that KLF8 expression was induced during differentiation, which was followed by expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer-binding protein α (C/EBPα). Adipocyte differentiation was significantly attenuated by the addition of siRNA against KLF8, whereas overexpression of KLF8 resulted in enhanced differentiation. Furthermore, luciferase reporter assays demonstrated that overexpression of KLF8 induced PPARγ2 and C/EBPα promoter activity, suggesting that KLF8 is an upstream regulator of PPARγ and C/EBPα. The KLF8 binding sites were localized by site mutation analysis to −191 region in C/EBPα promoter and −303 region in PPARγ promoter, respectively. Taken together, these data reveal that KLF8 is a key component of the transcription factor network that controls terminal differentiation during adipogenesis.

Taurine-induced changes in the expression profiles of HepG2 cells were assessed using a cDNA microarray technology, and confirmed by real-time reverse transcription-polymerase chain reaction (RT-PCR) analyses. Of 8,298 human genes on the microarray, 128 genes (87 known genes) were up-regulated, and 349 (206 known genes) were down-regulated more than 2.0-fold by taurine. Among the 293 known genes regulated by taurine, a total of 44 genes were involved in signal transduction; 16 genes were up-regulated greater than 2.0-fold, and 28 genes were down-regulated more than 2.0-fold by taurine. The results of RT-PCR analyses for the five genes selected were consistent with our microarray data, although the fold changes in the expression level differed somewhat between the two analytical methods. Among signal transduction-related genes affected by taurine, four genes--mitogen-activated protein kinase (MAPK) kinase kinase 7, p21-activated kinase 4, sprouty homolog 2, and MAPK kinase 1--are implicated in the MAPK signaling pathway. Taurine also regulated the expression of signal transducer and activator of transcription (STAT) 3 gene involved in the Janus kinase-STAT pathway, and diacylglycerol kinase, zeta 104 kDa, the downstream mediator of the protein kinase C transmembrane signaling pathway. In conclusion, gene expression profiling of HepG2 cells treated with taurine provided us with new insights into the novel aspects of taurine as a possible regulator of MAPK signaling cascades and protein kinase C signaling pathways involved in cellular processes such as cell growth, differentiation, and apoptosis.