海北高寒草甸土壤有机碳同位素组成及C3/C4碳源的变化

通过对高寒嵩草草甸土壤剖面不同深度(0～5cm,5～15cm,15～25cm,25～35cm,35～50cm,50～65cm)有机碳稳定性碳同位素的测定发现,土壤有机碳稳定性同位素(δ^13C)随土壤深度的增加而变大。表层土壤(0～5cm,定义为现代土壤)的δ^13C值最小,基本上接近现代植被的碳同位素特征。在土层5～10cm深度以下(粗略地定为古土壤),土壤有机碳稳定性同位素骤然上升,与表层土壤的同位素特征明显不同。考虑到影响土壤碳同位素的诸多因素,通过稳定性碳同位素的质量平衡模型计算,得出初步结果：来自C4(或CAM)植物的碳源随土壤深度的增加而增大。进一步推测,该地区植被可能经历由C4植物占优势的群落向C3植物占优势的群溶演化的过程。在这个过程中,大气碳同位素的变化和土壤有机质的形成过程(有机质淋溶过程)等也会引起土壤碳同位素的升高,因此质量平衡模型可能会过多地估算C4组分,而低估C3组分。     

Expression and function of CCAAT/enhancer binding proteinbeta (C/EBPbeta) LAP and LIP isoforms in mouse mammary gland, tumors and cultured mammary epithelial cells

CCAAT/Enhancer binding proteins (C/EBPs) play important roles in the regulation of cell growth and differentiation. This study investigated the expression and function of C/EBPbeta isoforms in the mouse mammary gland, mammary tumors, and a nontransformed mouse mammary epithelial cell line (HC11). C/EBPbeta mRNA levels are 2-5-fold higher in mouse mammary tumors derived from MMTV/c-neu transgenic mice compared with lactating and involuting mouse mammary gland. The "full-length" 38 kd C/EBPbeta LAP ("Liver-enriched Activator Protein") isoform is the predominant C/EBPbeta protein isoform in mammary tumor whole cell lysates, however, the truncated 20 kd C/EBPbeta LIP ("Liver-enriched Inhibitory Protein") isoform is also present at detectable levels (mean LAP:LIP ratio 5.3:1). The mammary tumor C/EBPbeta LAP:LIP ratio decreases 70% (from 5.3:1 to 1.6:1) when lysate preparation is switched from a rapid whole cell lysis protocol to a multistep nuclear/cytoplasmic fractionation protocol. In contrast to mammary tumors, only the C/EBPbeta LAP isoform is detectable in the mammary gland whole cell and nuclear lysates; the truncated "LIP" isoform is undetectable regardless of isolation protocol. Ectopic over expression of C/EBPbeta LIP or C/EBPbeta LAP did not alter HC11 growth rates. However, C/EBPbeta LIP over expressing HC11 cells (LAP:LIP ratio of approximately 1:1) exhibited a consistent 2-4 h delay in G(0)/S phase transition. C/EBPbeta LIP overexpressing HC11 cells did not express beta-casein mRNA (mammary epithelial cell differentiation marker) in response to lactogenic hormones. This defect in beta-casein expression was not corrected by carrying out the differentiation protocol in the presence of an artificial extracellular matrix. These results demonstrate that the "full-length" C/EBPbeta LAP isoform is the predominant C/EBPbeta protein isoform expressed in mouse mammary gland in vivo and mouse mammary epithelial cell cultures in vitro. C/EBPbeta LIP detected in mammary tumor lysates may result from in vivo production or ex vivo isolation-induced proteolysis of C/EBPbeta LAP. Ectopic overexpression of C/EBPbeta LIP (LAP:LIP ratio of approximately 1:1) inhibits mammary epithelial cell differentiation (beta-casein expression).     

Phospholipase C delta-type consists of three isozymes: bovine PLCdelta2 is a homologue of human/mouse PLCdelta4

To date, 12 phospholipase C (PLC) isozymes have been identified in mammals, and they are divided into five classes, beta-, gamma-, delta-, epsilon-, and zeta-type. PLCdelta-type is reported to be composed of four isozymes, PLCdelta1-delta4. Here we report that a screening for mouse PLCdelta2 from a BAC library with primers that amplify a specific region of bovine PLCdelta2 resulted in isolation of one clone containing the mouse PLCdelta4 gene. Furthermore, a database search revealed that there is only one gene corresponding to PLCdelta2 and PLCdelta4 in the mouse and human genomes, indicating that bovine PLCdelta2 is a homologue of human and mouse PLCdelta4. However, PLCdelta2 Western blot analysis with a widely used commercial anti-PLCdelta2 antibody showed an expression pattern distinct from that of PLCdelta4 in wild-type mice. In addition, an 80-kDa band, which was recognized by antibody against PLCdelta2, was smaller than an 85-kDa band detected by anti-PLCdelta4 antibody, and the 80-kDa band was detectable in lysates of brain, testis, and spleen from PLCdelta4-deficient mice. We also found that immunoprecipitates from brain lysates with this PLCdelta2 antibody contained no PLC activity. From these data, we conclude that bovine PLCdelta2 is a homologue of human and mouse PLCdelta4, and that three isozymes (delta1, delta3, and delta4) exist in the PLCdelta family.     

Wnt/Lrp/beta-catenin signaling suppresses adipogenesis by inhibiting mutual activation of PPARgamma and C/EBPalpha

Wnt/beta-catenin signaling has been implicated in repressing adipogenesis. Several lines of evidence show that the possible mechanism is blockade of PPARgamma induction. However, the precise mechanisms remain to be elucidated. In this study, we demonstrated that Wnt3a conditioned medium suppresses C/EBPbeta/delta-induced adipogenesis of 3T3-L1 cells by inhibiting PPARgamma induction. In addition, the mutual activation of PPARgamma and C/EBPalpha was also repressed in the presence of Wnt3a. To further investigate the role of the canonical Wnt pathway in adipogenesis, we used mouse embryonic fibroblasts (MEFs) isolated from Lrp6-deficient embryos. Contrary to wild-type MEFs, Lrp6-deficient MEFs showed spontaneous adipogenesis and escaped the suppressive effect of exogenous Wnt3a. These findings suggest a critical role of Wnt/Lrp6/beta-catenin signaling in adipogenesis and cell fate decision of mesenchymal stem cells.     

D4F alleviates the C/EBP homologous protein-mediated apoptosis in glycated high-density lipoprotein-treated macrophages by facilitating autophagy

The present study aimed to investigate the role of D4F, an apolipoprotein A-I mimetic peptide, in macrophage apoptosis induced by the glycated high-density lipoprotein (gly-HDL)-induced endoplasmic reticulum (ER) stress C/EBP homologous protein (CHOP) pathway, and unravel the regulatory role of autophagy in this process. Our results revealed that except for suppressing the accumulation of lipids within RAW264.7 macrophages caused by gly-HDL, D4F inhibited gly-HDL-induced decrease in the cell viability and increase in lactate dehydrogenase leakage and cell apoptosis, which were similar to 4-phenylbutyric acid (PBA, an ER stress inhibitor). Besides, similar to PBA, D4F inhibited gly-HDL-induced ER stress response activation evaluated through the decreased PERK and eIF2α phosphorylation, together with reduced ATF6 nuclear translocation as well as the downregulation of GRP78 and CHOP. Interestingly, D4F facilitated gly-HDL-triggered activation of autophagy, measured as elevated levels of beclin-1, LC3-II, and ATG5 expressions in macrophages. Furthermore, the inhibition effect of D4F on gly-HDL-induced ER stress-CHOP-induced apoptosis of macrophages was restrained after beclin-1 siRNA and 3-methyladenine (3-MA, an inhibitor of autophagy) treatments, while this effect was further reinforced after rapamycin (Rapa, an inducer of autophagy) treatment. Furthermore, administering D4F or Rapa to T2DM mice upregulated LC3-II and attenuated CHOP expression, cell apoptosis, and atherosclerotic lesions. However, the opposite results were obtained when 3-MA was administered to these mice. These results support that D4F effectively protects macrophages against gly-HDL-induced ER stress-CHOP-mediated apoptosis by promoting autophagy.