C/EBPα deficiency in podocytes aggravates podocyte senescence and kidney injury in aging mice.

Kidney aging leads to an increased incidence of end-stage renal disease (ESRD) in the elderly, and aging is a complex biological process controlled by signaling pathways and transcription factors. Podocyte senescence plays a central role in injury resulting from kidney aging. Here, we demonstrated the critical role of C/EBPα in podocyte senescence and kidney aging by generating a genetically modified mouse model of chronological aging in which C/EBPα was selectively deleted in podocytes and by overexpressing C/EBPα in cultured podocytes, in which premature senescence was induced by treatment with adriamycin. Moreover, we illuminated the mechanisms by which podocyte senescence causes tubular impairment by stimulating HK-2 cells with bovine serum albumin (BSA) and chloroquine. Our findings suggest that C/EBPα knockout in podocytes aggravates podocyte senescence through the AMPK/mTOR pathway, leading to glomerulosclerosis, and that subsequent albuminuria exacerbates the epithelial-mesenchymal transdifferentiation of senescent tubular cells by suppressing autophagy. These observations highlight the importance of C/EBPα as a new potential target in kidney aging.

Promoting differentiation and lipid metabolism are the primary effects for DINP exposure on 3T3-L1 preadipocytes.

Diisononyl phthalate (DINP) is a high-molecular-weight phthalate, and has been recently introduced as di-(2-ethyl hexyl) phthalate (DEHP) substitute and commonly used in a large variety of plastic items. The fat tissue is an important target for DINP exposure, however, very little is understood about its toxicity and mechanism(s) in adipocyte cells. Therefore, the present work aimed to investigate the role of DINP in adipogenesis using 3T3-L1 preadipocytes. DINP exposure for 10 days extensively induced adipogenesis in 3T3-L1 preadipocytes to adipocytes as assessed by lipid accumulation and gene expression of adipogenic markers. The RT-qPCR results showed that DINP could upregulate the expression of peroxisome proliferator-activated receptor-gamma (PPARγ), CCAAT/enhancer-binding protein alpha (C/EBPα) and C/EBPß, while the expression of sterol regulatory element binding transcription factor 1 (SREBF1) and C/EBPδ was not affected. The DINP-induced adipogenesis could be inhibited by using the selective PPARγ antagonist GW9662. The RNA-seq analysis was used to study the systemic toxicities of DINP on preadipocytes. A total of 1181 differently expressed genes (DEGs) (640 genes were up-regulated, 541 genes were down-regulated) were detected in 3T3-L1 preadipocytes under 50 µM DINP. The GO enrichment showed the GO term of "fat cell differentiation" was the most significantly affected metabolic functions, and the KEGG pathway enrichment showed the PPAR pathway was the top affected pathway. The interactive pathway (iPath) analysis showed that the changed metabolic pathways were focus on the lipid metabolism.

Vitamin Е activates expression of С/EBP alpha transcription factor and G-CSF receptor in leukemic K562 cells.

BACKGROUND: Chronic myeloid leukemia (CML) is a clonal hematopoietic stem cell disorder associated with the activity of BCR-ABL fusion oncogene. Tyrosine kinase inhibitors are the current treatment of CML, but secondary mutations finally contribute to therapy resistance and blast crisis of the disease. The search for the novel compounds for the effective control of CML is now in the spotlight. The progression of CML to blast crisis is correlated with down-modulation of C/EBP alpha. Therefore, C/EBP alpha may be considered as a putative target in differentiation therapies in myeloid leukemias. The aim of the study was to assess the potential of vitamin E as the possible inducer of C/EBP alpha expression in BCR-ABL-positive CML K562 cells. MATERIALS AND METHODS: RNA extracted from K562 cells cultured with valproic acid or vitamin E was converted to cDNA, RT-PCR reactions were carried out using HotStarTaq DNA polymerase with primers for C/EBP alpha and granulocyte colony-stimulating factor receptor (G-CSFR). RESULTS: We have not found detectable expression of C/EBP alpha in K562 cells. Upon 48-h culture with vitamin E at a dose of 100 µM, K562 cells expressed both C/EBP alpha and G-CSFR. CONCLUSION: Vitamin E restored the expression of C/EBP alpha mRNA in chronic myelogenous leukemia K562 cells. In this setting, G-CSFR expression in vitamin E treated K562 cells seems to suggest the activation to granulocytic differentiation. It should be further elucidated whether such effects of vitamin E on C/EBP alpha transcription factor are direct or mediated indirectly due to antioxidant properties of vitamin E.

Moringa concanensis Nimmo ameliorates hyperglycemia in 3T3-L1 adipocytes by upregulating PPAR-γ, C/EBP-α via Akt signaling pathway and STZ-induced diabetic rats.

Moringa concanensis Nimmo is a medicinal plant for treating various human illnesses including menstrual pain, high blood pressure, jaundice, inflammation, pain, fever, sore eyes, and cholesterol in Indian folk medicine. Despite its versatility, its antihyperglycemic mechanism of action (in vitro and in vivo) remains unclear. Therefore, in this study we developed the possible antihyperglycemic mechanism of action in 3T3-L1 cells by evaluating mRNA and protein expression, which are associated with adipogenesis and lipogenesis (insulin sensitizer). Also, the antihyperglycemic activity of the ethanolic extract of M. concanensis Nimmo leaves (EEMCN) was evaluated on glucose, insulin, biochemical, and lipid profile in experimental diabetic rat models induced with streptozotocin (STZ). Results showed that EEMCN leaves enhanced lipid accumulation in 3T3-L1 cells, as assessed by Oil Red O staining, and upregulated gene expression level of PPAR-γ, C/EBP-α, t-SREBP, FAS, Glut-4, adipogenin, DAG, and LPL through Akt signaling in 3T3-L1 cells. Also, EEMCN treatment increased body weight and insulin level and lowered blood glucose, HbA1c, amylase, and lipid profile level in STZ-induced diabetic rats. In conclusion, EEMCN possesses in vivo antidiabetic potential, having such efficacy through a mechanism of action that involves antihyperglycemic, hypoglycemic, and potential insulin sensitizer (PPAR-γ, C/EBP-α/Akt over expression) action.

miR-26b Promotes 3T3-L1 Adipocyte Differentiation Through Targeting PTEN.

microRNAs (miRNAs) play important roles in adipogenesis that is closely linked to obesity and energy homeostasis. Thus far, only a few miRNAs have been identified to regulate adipocyte development, arousing interest in the detailed function of miRNAs during adipogenesis. In this study, we found that the miR-26b expression showed an increasing trend during 3T3-L1 cells differentiation. To investigate the role of miR-26b in adipogenesis, the synthetic miR-26b agomirs and antagomirs were used to perform overexpression and knockdown experiment, respectively. Our data revealed that overexpression of miR-26b significantly accelerated the mRNA expression of the adipogenic markers, peroxisome proliferator-activated receptor gamma (PPARγ), fatty acid synthase (FAS), CCAAT/enhancer binding protein alpha (C/EBPα), and lipoprotein lipase, and the protein level of PPARγ and FAS. miR-26b overexpression also resulted in a significant increase in lipid accumulation. In contrast, inhibition of miR-26b expression decreased differentiation of 3T3-L1 cells. By target gene prediction and luciferase reporter assay, we demonstrated that miR-26b may directly bind to the 3' UTR of phosphatase and tensin homolog (PTEN). Taken together, these results demonstrate that miR-26b might participate in regulating adipogenic differentiation in 3T3-L1 cells by inhibiting the PTEN expression, further highlighting the importance of miRNA in adipogenesis.

miR-128 promoted adipogenic differentiation and inhibited osteogenic differentiation of human mesenchymal stem cells by suppression of VEGF pathway.

CONTEXT: MicroRNA-128 (miR-128), a brain-enriched microRNA, has been reported to participate in the regulation of cell differentiation, but its potential roles in adipogenic and osteogenic differentiation of human mesenchymal stem cells (hMSCs) have not been addressed. OBJECTIVE: The study was conducted to investigate the effects and mechanism of miR-128 on adipogenic and osteogenic differentiation of hMSCs. MATERIALS AND METHODS: Morphology of hMSCs, lipid droplets and calcium nodules were observed and photographed by LSM microscopy. Expression of CCAAT/enhancer binding protein-α (C/EBPα), peroxisome proliferator-activated receptor-γ (PPARγ), miR-128, vascular endothelial growth factor (VEGF), osteocalcin (OCN) and Runt-related transcription factor 2 (RUNX2) was determined by RNA preparation and reverse transcription polymerase chain reaction (RT-PCR), protein expression of VEGF was analyzed by Western blot. RESULTS: It was suggested that miR-128 expression showed a 4.56-fold induction by adipogenic treatment and a 58.8% reduction by osteogenic treatment. Over-expression of miR-128, promoted adipogenic differentiation while inhibited osteogenic differentiation. In contrast, adipocyte formation was inhibited and osteogenesis was enhanced in cells slicing miR-128. Furthermore, over-expression of miR-128 down-regulated VEGF expression in adipogenically and osteogenically differentiated cells. We further identified VEGF as a key regulator in miR-128-induced adipogenic and osteogenic differentiation. Following knockdown of VEGF, the effects of over-expression of miR-128 on adipogenic and osteogenic differentiation of hMSCs were limited. CONCLUSION: It was indicated that miR-128 could regulate adipogenic and osteogenic differentiation of hMSCs significantly through the suppression of VEGF pathway.

Overexpression of CCAAT Enhancer-Binding Protein α Inhibits the Growth of K562 Cells via the Foxo3a-Bim Pathway.

We aimed to investigate the role of CCAAT enhancer-binding protein α (C/EBPα) in the pathogenesis of chronic myeloid leukemia (CML) and the mechanism underlying its effect. Bone marrow specimens from 50 patients with CML and peripheral blood specimens from 20 healthy individuals were collected. K562 cells were treated with imatinib. Subsequently, a stable cell line, K562-C/EBPα, was constructed. Cell proliferation was assayed with cell counting kit-8, and mRNA levels of C/EBPα, forkhead transcription factor FKHRL1 (Foxo3a) and Bim were detected by semiquantitative PCR. The correlation of C/EBPα and BCR-ABL was assessed by Spearman's correlation analysis. The results showed that C/EBPα mRNA levels were significantly reduced in CML patients compared with healthy subjects (p < 0.001) and were negatively correlated with BCR-ABL1 (r = -0.5046, p < 0.01). Additionally, imatinib enhanced the expression of C/EBPα in K562 cells compared with untreated cells (p < 0.05). Overexpression of C/EBPα significantly decreased cell proliferation and upregulated the expressions of the apoptosis-related genes Foxo3a (p < 0.01) and Bim (p < 0.05) in K562 cells. In conclusion, C/EBPα expression was decreased in patients with CML. Imatinib enhances the expression of C/EBPα in K562 cells, and the overexpression of C/EBPα inhibits cell proliferation and increases apoptosis via the Foxo3a-Bim pathway.

A three-dimensional collagen-sponge-based culture system coated with simplified recombinant fibronectin improves the function of a hepatocyte cell line.

Hepatocytes are widely used in pharmaceutical drug discovery tests, but their hepatic functions decrease rapidly during in vitro culture. Many culture systems have been devised to address this problem. We here report that a three-dimensional (3D) collagen-based scaffold coated with simplified recombinant fibronectin (FN) enhanced the function of a hepatocyte cell line. The developed culture system uses a honeycomb collagen sponge coated with collagen-binding domain (CBD)-cell attachment site (CAS), a chimeric protein comprising the CBD and CAS of FN. The function of HepG2 cells grown on honeycomb collagen sponge coated with CBD-CAS was investigated by determining the messenger RNA (mRNA) expression levels of several genes. The mRNA expression level of albumin increased 3.25 times in cells grown on CBD-CAS-coated honeycomb collagen sponge for 3 days; the expression level of CCAAT/enhancer binding protein (C/EBPα) increased 40-fold after 1 d and up to 150-fold after 3 d. These results suggested that CBD-CAS-coated honeycomb collagen sponge could improve the functions of hepatocytes by inducing C/EBPα expression. The activation of cytochrome P450 (CYP) enzymes in HepG2 cells grown on CBD-CAS-coated honeycomb collagen sponge was measured at the mRNA level and was found to increase between two and six times compared to cells grown without the CBD-CAS coating, showing that this culture system induced CYP gene expression and thus may be useful in drug metabolism assays.

GPR39 activates proliferation and differentiation of porcine intramuscular preadipocytes through targeting the PI3K/AKT cell signaling pathway.

BACKGROUND: The orphan G protein-coupled receptor (GPR) 39 was originally identified as the receptor of obestatin. In this study, the effects and mechanisms of GPR39 on cell proliferation and differentiation were investigated in cultured porcine intramuscular preadipocytes. METHODS: Morphology of preadipocytes and accumulated lipid droplets within cells were identified by an inverted microscope. After transfected with constructed pCMV-GPR39 plasmid, cell proliferation was measured by using methyl thiazolyl tetrazolium method, mRNA expression of GPR39, CCAAT/enhancer binding protein-α (C/EBPα), peroxisome proliferator-activated receptor-γ (PPARγ), Caspase-9 and adipocyte determination and differentiation factor-1 (ADD1) was determined by RNA preparation and reverse transcription polymerase chain reaction, protein expression of phosphoinositide-3 kinase (PI3K), 3-phosphoinositide-dependent protein kinase 1, phosphorylated glycogen synthase kinase 3 (pGSK3), total Akt and phosphorylated Akt (pAkt) was analyzed by Western blot. RESULTS: It found that GPR39 mRNA and protein were expressed in porcine intramuscular preadipocytes and its expression was significantly up-regulated after treatment with Zn(2+) whose function is found to be mediated by GPR39. Furthermore, over-expression of GPR39 further promoted the optical density value of cells, enhanced mRNA expression of PPARγ, C/EBPα and ADD1, and inhibited mRNA expression of Caspase-9. Protein expression of pGSK3 and pAkt was also increased by GPR39 stimulation. In addition, GPR39-induced proliferation and differentiation of porcine intramuscular preadipocytes was partially blocked by the Akt inhibitor (PDTC) and the PI3K inhibitor (LY294002). CONCLUSION: It indicated that GPR39 was a transducer of Zn(2+), and enhanced proliferation and differentiation of porcine intramuscular preadipocytes through activation of the PI3K/Akt signaling pathway.

Restoration of CCAAT enhancer binding protein α P42 induces myeloid differentiation and overcomes all-trans retinoic acid resistance in human acute promyelocytic leukemia NB4-R1 cells.

All-trans retinoic acid (ATRA) is one of the first line agents in differentiation therapy for acute promyelocytic leukemia (APL). However, drug resistance is a major problem influencing the efficacy of ATRA. Identification of mechanisms of ATRA resistance are urgenly needed. In the present study, we found that expression of C/EBPα, an important transcription factor for myeloid differentiation, was significantly suppressed in ATRA resistant APL cell line NB4-R1 compared with ATRA sensitive NB4 cells. Moreover, two forms of C/EBPα were unequally suppressed in NB4-R1 cells. Suppression of the full-length form P42 was more pronounced than the truncated form P30. Inhibition of PI3K/Akt/mTOR pathway was also observed in NB4-R1 cells. Moreover, C/EBPα expression was reduced by PI3K inhibitor LY294002 and mTOR inhibitor RAD001 in NB4 cells, suggesting that inactivation of the PI3K/Akt/mTOR pathway was responsible for C/EBPα suppression in APL cells. We restored C/EBPα P42 and P30 by lentivirus vectors in NB4-R1 cells, respectively, and found C/EBPα P42, but not P30, could increase CD11b, CD14, G-CSFR and GM-CSFR expression, which indicated the occurrence of myeloid differentiation. Further upregulating of CD11b expression and differential morphological changes were found in NB4-R1 cells with restored C/EBPα P42 after ATRA treatment. However, CD11b expression and differential morphological changes could not be induced by ATRA in NB4-R1 cells infected with P30 expressing or control vector. Thus, we inferred that ATRA sensitivity of NB4-R1 cells was enhanced by restoration of C/EBPα P42. In addition, we used histone deacetylase inhibitor trichostatin (TSA) to restore C/EBPα expression in NB4-R1 cells. Similar enhancement of myeloid differentiation and cell growth arrest were detected. Together, the present study demonstrated that suppression of C/EBPα P42 induced by PI3K/Akt/mTOR inhibition impaired the differentiation and ATRA sensitivity of APL cells. Restoring C/EBPα P42 is an attractive approach for differentiation therapy in ATRA resistant APL.

Resetting the transcription factor network reverses terminal chronic hepatic failure.

The cause of organ failure is enigmatic for many degenerative diseases, including end-stage liver disease. Here, using a CCl4-induced rat model of irreversible and fatal hepatic failure, which also exhibits terminal changes in the extracellular matrix, we demonstrated that chronic injury stably reprograms the critical balance of transcription factors and that diseased and dedifferentiated cells can be returned to normal function by re-expression of critical transcription factors, a process similar to the type of reprogramming that induces somatic cells to become pluripotent or to change their cell lineage. Forced re-expression of the transcription factor HNF4α induced expression of the other hepatocyte-expressed transcription factors; restored functionality in terminally diseased hepatocytes isolated from CCl4-treated rats; and rapidly reversed fatal liver failure in CCl4-treated animals by restoring diseased hepatocytes rather than replacing them with new hepatocytes or stem cells. Together, the results of our study indicate that disruption of the transcription factor network and cellular dedifferentiation likely mediate terminal liver failure and suggest reinstatement of this network has therapeutic potential for correcting organ failure without cell replacement.

The role of C/EBP-α expression in human liver and liver fibrosis and its relationship with autophagy.

AIM: To investigate the expression of CCAAT enhancer binding protein-α (C/EBP-α) in normal human liver and liver fibrosis and its probable association with autophagy. METHODS: Double label immunohistochemistry was used to detect the location of C/EBP-α in hepatocytes and hepatic stellate cells (HSCs). The expression of C/EBP-α, Atg5, and Atg6 was also evaluated by immunohistochemistry in paraffin sections of human liver. HSC-T6 cells were treated with rapamycin and 3-methyladenine (3MA) to induce or inhibit autophagy, and the expression of C/EBP-α protein was detected by Western blotting. RESULTS: Double label immunohistochemistry showed that C/EBP-α was predominantly located in hepatocytes and that its expression was significantly decreased in fibrosis compared with normal liver. Atg5 expression was increased in fibrosis but was located primarily in liver septa and peri-vascular areas, which was consistent with the distribution of HSCs. In contrast, Atg6 was not expressed in normal or fibrotic liver. Treatment of HSC-T6 cells in culture with rapamycin or 3MA decreased or increased C/EBP-α expression, respectively, as shown by Western blotting. CONCLUSION: C/EBP-α was primarily expressed in hepatocytes in normal liver, but its expression decreased significantly in liver fibrosis. Autophagy might play a role in liver fibrosis through its association with C/EBP-α, but this hypothesis warrants further investigation.

miR-199a-5p inhibits monocyte/macrophage differentiation by targeting the activin A type 1B receptor gene and finally reducing C/EBPα expression.

miRNAs are short, noncoding RNAs that regulate expression of target genes at post-transcriptional levels and function in many important cellular processes, including differentiation, proliferation, etc. In this study, we observed down-regulation of miR-199a-5p during monocyte/macrophage differentiation of HL-60 and THP-1 cells, as well as human CD34(+) HSPCs. This down-regulation of miR-199a-5p resulted from the up-regulation of PU.1 that was demonstrated to regulate transcription of the miR-199a-2 gene negatively. Overexpression of miR-199a-5p by miR-199a-5p mimic transfection or lentivirus-mediated gene transfer significantly inhibited monocyte/macrophage differentiation of the cell lines or HSPCs. The mRNA encoding an ACVR1B was identified as a direct target of miR-199a-5p. Gradually increased ACVR1B expression level was detected during monocyte/macrophage differentiation of the leukemic cell lines and HSPCs, and knockdown of ACVR1B resulted in inhibition of monocyte/macrophage differentiation of HL-60 and THP-1 cells, which suggested that ACVR1B functions as a positive regulator of monocyte/macrophage differentiation. We demonstrated that miR-199a-5p overexpression or ACVR1B knockdown promoted proliferation of THP-1 cells through increasing phosphorylation of Rb. We also demonstrated that the down-regulation of ACVR1B reduced p-Smad2/3, which resulted in decreased expression of C/EBPα, a key regulator of monocyte/macrophage differentiation, and finally, inhibited monocyte/macrophage differentiation.

IL-32α down-regulates ß2 integrin (CD18) expression by suppressing PU.1 expression in myeloid cells.

Myeloid-specific CD18 associates with CD11 and plays a critical role in leukocyte adhesion to the endothelium. In this study, we observed that CD18 expression was decreased by IL-32α in THP-1 and K562 cells upon PMA stimulation, and investigated the mechanism by which IL-32α down-regulated CD18 expression. We found that IL-32α suppressed the expression of PU.1, a major transcription factor for CD18. Because we previously demonstrated that IL-32α mediated STAT3 S727 phosphorylation by PKCε, and STAT3 regulates PU.1 expression, we performed time-course analyses of STAT3 S727 phosphorylation and found that IL-32α induces prolonged phosphorylation of STAT3 S727 until 72h after PMA stimulation. The expression pattern of C/EBPα, another transcriptional regulator of PU.1, was not affected by IL-32α. In addition, we showed that STAT3 binding to the PU.1 promoter was suppressed by IL-32α. Thus, we examined the relatedness among these factors and found that IL-32α-mediated STAT3 S727 phosphorylation induced C/EBPα association. When STAT3 was mutated at S727 to proline (S727P), the mutant STAT3 S727P did not interact with C/EBPα. We further demonstrated that only the intact STAT3 interacted with the basic leucine zipper region of C/EBPα. The PU.1 promoter was activated by co-expression of STAT3 and IL-32α upon PMA stimulation. However, the promoter activity was inhibited with STAT3 and C/EBPα co-expression. Therefore, our data suggest that IL-32α-mediated STAT3 S727 phosphorylation induced C/EBPα association, which inhibited PU.1 expression, and then resulted in the down-regulation of CD18 expression.

Estrogen suppresses adipogenesis by inhibiting S100A16 expression.

The aim of this study is to determine the effects of E2 on metabolic syndrome and the molecular mechanisms involving S100A16. Ovariectomized (OVX) rat models and mouse embryonic fibroblasts cell models were used. E2 loss in OVX rats induced body weight gain and central abdominal fat accumulation, which were ameliorated by E2 treatment under chow and high-fat diet (HFD) conditions. E2 decreased the expression of the adipocyte marker genes PPARγ, aP2, C/EBPα, and S100A16. E2 inhibited adipogenesis. Overexpression of S100A16 reversed the E2-induced adipogenesis effect. A luciferase assay showed that E2 inhibited the expression of S100A16. E2 treatment decreased body weight gain and central abdominal fat accumulation under both chow and HFD conditions. Also, E2 suppressed adipogenesis by inhibiting S100A16 expression.

Obestatin changes proliferation, differentiation and apoptosis of porcine preadipocytes.

Obestatin, originally identified and purified from rat stomach extracts, was reported to bind to orphan G protein-coupled receptor, GPR39, and inhibit appetite and gastric motility. This study was conducted to investigate the effects of porcine obestatin on proliferation, differentiation and apoptosis of porcine preadipocytes isolated from subcutaneous fat of piglets. At indicated times of culture, morphology of preadipocytes and accumulated lipid droplets within the cells were identified by invert microscope. After treating with obestatin (0, 0.1, 1, 10 and 100nM), cell proliferation was measured by MTT method and protein expression of CCAAT/enhancer binding protein-α (C/EBPα), peroxisome proliferator-activated receptor-γ (PPARγ), Caspase-7 and Caspase-9 was determined by Western Blot, mRNA expression of GPR39 and Caspase-3 was analyzed by RT-PCR, and the activity of Caspase-3 was measured by spectrophotometric method. The results showed that obestatin had no effect on GPR39 expression, while promotes the optical density (OD) value of cells, enhanced protein expression of PPARγ and C/EBPa, decreased mRNA expression and activity of Caspase-3, and inhibited protein expression of Caspase-7 and Caspase-9 in a dose-dependent manner. These results suggested that obestatin enhances proliferation and differentiation of preadipocytes promoting PPARγ and C/EBPa expression, and inhibiting preadipocyte apoptosis by decreasing expression of Caspase-3, Caspase-7 and Caspase-9.

Hyperglycemia diverts dividing osteoblastic precursor cells to an adipogenic pathway and induces synthesis of a hyaluronan matrix that is adhesive for monocytes.

Isolated rat bone marrow stromal cells cultured in osteogenic medium in which the normal 5.6 mm glucose is changed to hyperglycemic 25.6 mm glucose greatly increase lipid formation between 21-31 days of culture that is associated with decreased biomineralization, up-regulate expression of cyclin D3 and two adipogenic markers (CCAAT/enhancer binding protein α and peroxisome proliferator-activated receptor γ) within 5 days of culture, increase neutral and polar lipid synthesis within 5 days of culture, and form a monocyte-adhesive hyaluronan matrix through an endoplasmic reticulum stress-induced autophagic mechanism. Evidence is also provided that, by 4 weeks after diabetes onset in the streptozotocin-induced diabetic rat model, there is a large loss of trabecular bone mineral density without apparent proportional changes in underlying collagen matrices, a large accumulation of a hyaluronan matrix within the trabecular bone marrow, and adipocytes and macrophages embedded in this hyaluronan matrix. These results support the hypothesis that hyperglycemia in bone marrow diverts dividing osteoblastic precursor cells (bone marrow stromal cells) to a metabolically stressed adipogenic pathway that induces synthesis of a hyaluronan matrix that recruits inflammatory cells and establishes a chronic inflammatory process that demineralizes trabecular cancellous bone.

Enhancement of differentiation induction and upregulation of CCAAT/enhancer-binding proteins and PU.1 in NB4 cells treated with combination of ATRA and valproic acid.

The effects of all-trans retinoic acid (ATRA) and valproic acid (VPA), alone and in combination, on the human acute promyelocytic leukemia (APL) cell line NB4 were investigated in view of differentiation induction and growth inhibition. After 48 h of treatment, not only ATRA but also VPA induced differentiation in NB4 cells, and their combination further augmented the differentiation activity. Furthermore, the upregulation of transcription factors including CCAAT/enhancer-binding proteins (CEBPα, ß, ε) and PU.1, which are known to be critical factors for normal myelopoiesis, granulocytic maturation and being repressed in APL, concurred with the differentiation induction. A significant cell growth inhibition was observed after the treatment with VPA, which was further strengthened by the addition of ATRA. Given the importance of C/EBPs and PU.1 in myeloid development, these results, thus, suggest that restoration of the normal function of the myeloid cell transcriptional machinery is a major molecular mechanism underlying the differentiation induction in NB4. Therefore, these results may provide novel insights into a possible combinational therapeutic approach for APL patients.

Genistein exposure during the early postnatal period favors the development of obesity in female, but not male rats.

Genistein (Gen), the primary isoflavone in soy, has been shown to adversely affect various endocrine-mediated endpoints in rodents and humans. Soy formula intake by human infants has been associated with early age at menarche and decreased female-typical behavior in girls. Adipose deposition and expansion are also hormonally regulated and Gen has been shown to alter these processes. However, little is known about the impact of early-life soy intake on metabolic homeostasis in adulthood. The current study examined the impact of early-life Gen exposure on adulthood body composition (by magnetic resonance imaging) and the molecular signals mediating adipose expansion. From postnatal day (PND) 1 to 22, rat pups were daily orally dosed with 50mg/kg Gen to mimic blood Gen levels in human infants fed soy formula. Female but not male Gen-exposed rats had increased fat/lean mass ratio, fat mass, adipocyte size and number, and decreased muscle fiber perimeter. PND22 Gen-exposed females, but not males, had increased expression of adipogenic factors, including CCAAT/enhancer binding protein alpha (Cebpα), CCAAT/enhancer binding protein beta (Cebpß), and peroxisome proliferator-activated receptor gamma (Pparγ). Furthermore, Wingless-related MMTV integration site 10b (Wnt10b), a critical regulator of adipogenic cell fate determination, was hypermethylated and had decreased expression in adipose of PND22 Gen-exposed females. These data suggest that developmental Gen exposure in rats has gender-specific effects on adiposity that closely parallel the effects of a postweaning high-fat diet and underscore the importance of considering timing of exposure and gender when establishing safety recommendations for early-life dietary Gen intake.

Heat stress stimulates hepcidin mRNA expression and C/EBPα protein expression in aged rodent liver.

Elevations in hepatic iron content occur with aging and physiological stressors, which may promote oxidative injury to the liver. Since dysregulation of the iron regulatory hormone, hepcidin, can cause iron accumulation, our goal was to characterize the regulation of hepcidin in young (6 mo) and old (24 mo) Fischer 344 rats exposed to environmental heat stress. Liver and blood samples were taken in the control condition and after heating. Hepcidin expression did not differ between young and old rats in the control condition, despite higher levels of hepatic iron and IL-6 mRNA in the latter. Following heat stress, pSTAT3 increased in both groups, but C/EBPα and hepcidin mRNA increased only in old rats. Despite this, serum iron decreased in both age groups 2 h after heat stress, suggesting hepcidin-independent hypoferremia in the young rats. The differential regulation of hepcidin between young and old rats after hyperthermia may be due to the enhanced expression of C/EBPα protein in old rats. These data support the concept of "inflammaging" and suggest that repeated exposures to stressors may contribute to the development of anemia in older individuals.