scholarly journals ZBTB16 Overexpression Enhances White Adipogenesis and Induces Brown-Like Adipocyte Formation of Bovine White Intramuscular Preadipocytes

2018 ◽  
Vol 48 (6) ◽  
pp. 2528-2538 ◽  
Author(s):  
Shengjuan Wei ◽  
Mengmeng Zhang ◽  
Yueying Zheng ◽  
Peishi Yan
Keyword(s):  
Cell Model ◽  
Cell Counting ◽  
Brown Adipocyte ◽  
Mrna Levels ◽  
Mtdna Copy Number ◽  
Gpdh Activity ◽  
Real Time Quantitative Pcr ◽  
Protein Levels ◽  
Bovine Adipocytes

Background/Aims: Our study aims to characterize functions of ZBTB16 gene in the process of intramuscular fat (IMF) deposition and metabolism of bovine, thereby providing insights into mechanisms for the use of ZBTB16 in fat management. Methods: Primary preadipocytes derived from bovine IMF tissue were isolated and used as the in vitro cell model. An adenovirus Ad-ZBTB16 was transfected into bovine preadipocytes to overexpress the ZBTB16 gene. By using real-time quantitative PCR (RT-qPCR), western blotting, Oil Red-O staining, glycerol-3-phosphate dehydrogenase (GPDH) activity assay, and cell counting kit-8 (CCK-8) test, adipogenic and proliferative signals in adipocytes were monitored to investigate effects of ZBTB16 on adipogenesis of bovine preadipocytes. Results: After transfection, mRNA and protein levels of ZBTB16 gene were significantly increased. Enhanced ZBTB16 significantly promoted preadipocyte differentiation, as evidenced by accelerated lipid accumulation, enhanced GPDH activity, consistently increased mRNA expressions of adipogenic key transcription factors PPARγ, C/EBPα, FABP4, and ADIPOQ, and markedly increased protein expressions of PPARγ and FABP4. No difference was observed concerning proliferation of preadipocytes after treatment with Ad-ZBTB16. Furthermore, relative mRNA levels of brown adipocyte selective genes (PRDM16, UCP1, Cidea, Cox8b, and PGC-1α) and beige adipocyte selective genes (CD137, TMEM26, and Tbx1) as well as UCP1 protein expression were significantly increased by Ad-ZBTB16. Meanwhile, Ad-ZBTB16 treatment remarkably induced mitochondrial biogenesis and increased relative mitochondrial DNA (mtDNA) copy number in bovine adipocytes. Conclusion: These results suggest that ZBTB16 overexpression can promote white adipogenesis and induce brown-like adipocyte formation for bovine white intramuscular preadipocytes.

scholarly journals Cyclocarya paliurus Polysaccharide Inhibits Glioma Cell U251 Proliferation, Migration, and Invasion and Promotes Apoptosis via the GSK3β/β-Catenin Signaling Pathway

2020 ◽  
Vol 2020 ◽  
pp. 1-9 ◽  
Author(s):  
Xiaolong Du ◽  
Kai Guo ◽  
Yongqian Ma ◽  
Jianyong Chang
Keyword(s):  
Protein Expression ◽  
Signaling Pathway ◽  
Underlying Mechanism ◽  
Cell Counting ◽  
Migration And Invasion ◽  
Mrna Levels ◽  
Cyclocarya Paliurus ◽  
Protein Levels ◽  
U251 Cells

Objective. To investigate the effects of Cyclocarya paliurus polysaccharide (CPP) on the proliferation, migration, invasion, and apoptosis of human glioma U251 cells and further explore the underlying mechanism. Methods. U251 cells were cultured in vitro and treated with various concentrations (25, 50, 75, 100, 125, and 150 μmol/L) of CPP for 24, 48, and 72 h. Cell counting kit-8 was used to detect the activity of cell proliferation. Wound-healing assay, Transwell assay, and flow cytometry were used to measure the effects of CPP on the migration, invasion, and apoptosis of U251 cells, respectively. Western blotting was used to determine the protein expression involved in the GSK3β/β-catenin signaling pathway and its downstream genes related to proliferation, migration, invasion, and apoptosis including Cyr61, CCND1, Vimentin, and Slug. Meanwhile, qRT-PCR was used to detect the mRNA levels of Cyr61, CCND1, Vimentin, and Slug. Results. We found that CPP not only could inhibit the proliferation, migration, and invasion of U251 cells but also promote its apoptosis in vitro. Besides, CPP could significantly inhibit the phosphorylation and decrease the protein levels of GSK3 β at ser9 site (p<0.05), and thus increasing the phosphorylation of β-Catenin at ser33/37 site (p<0.05), resulting in β-Catenin degradation. In addition, we also found that CPP could downregulate the mRNA (p<0.05) and protein expression (p<0.05) of downstream genes of GSK3 β/β-catenin signaling pathway including Cyr61, CCND1, Vimentin, and Slug, which are related to proliferation, migration, invasion, and apoptosis. Conclusion. CPP could inhibit the expression of GSK3β, promote the degradation of β-catenin, and downregulate the levels of GSK3β/β-catenin downstream genes including Cyr61, CCND1, Vimentin, and Slug, which regulate the proliferation, migration, invasion, and apoptosis of glioma cells.

scholarly journals Metformin’s Effects on Apoptosis of Esophageal Carcinoma Cells and Normal Esophageal Epithelial Cells: An In Vitro Comparative Study

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Jianwei Peng ◽  
Xubin Jing ◽  
Jialing Wu ◽  
Danmian Hong ◽  
Xi Hu ◽  
...  
Keyword(s):  
Esophageal Carcinoma ◽  
Cell Line ◽  
Molecular Mechanisms ◽  
Carcinoma Cells ◽  
Cell Counting ◽  
Epithelial Cell Line ◽  
Dependent Manner ◽  
Real Time Quantitative Pcr ◽  
Protein Levels

The effect of metformin on human esophageal normal and carcinoma cells remains poorly understood. We aim to investigate the different antiproliferation effects and underlying distinct molecular mechanisms between these two types of cells. Human esophageal squamous cell carcinoma cell line, EC109, and normal esophageal epithelial cell line, HEEC, were used in the experiment. The cell survival rate was determined by cell counting kit-8 (CCK-8). Cell apoptosis was analyzed by flow cytometry. The mRNA and protein levels of signal transducer and activator of transcription 3 (Stat3) were detected by real-time quantitative PCR and western blot. Interleukin-6 (IL-6) was added to activate Stat3. The level of intracellular reactive oxygen species (ROS) was assessed by a DCFH-DA fluorescent probe. Metformin had more significant inhibitory effects on cell proliferation in EC109 cells than HEECs. Metformin induced apoptosis of EC109 cells in a dose-dependent manner instead of HEECs. The expression of Stat3 in both mRNA and protein levels was higher in EC109 cells than HEECs. Further study revealed that metformin may attenuate the phosphorylation of the Stat3 and the Bcl-2 expression, which was restored by IL-6 partly in EC109 cells but not HEECs. On the contrary, metformin increased the level of ROS in both the cell lines, but this intracellular ROS variation had no effect on apoptosis. Metformin has different functional roles on the apoptosis in esophageal carcinoma cells and normal esophageal cells. Therefore, the Stat3/Bcl-2 pathway-mediated apoptosis underlies the cell-type-specific drug sensitivity, suggesting metformin possesses a therapeutic activity and selectivity on esophageal cancer.

scholarly journals TRIM27 interacts with Iκbα to promote the growth of human renal cancer cells through regulating the NF-κB pathway

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Chengwu Xiao ◽  
Wei Zhang ◽  
Meimian Hua ◽  
Huan Chen ◽  
Bin Yang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Prognostic Indicator ◽  
The Cancer Genome Atlas ◽  
Mrna Levels ◽  
Loss Of Function ◽  
Protein Levels ◽  
Kaplan Meier ◽  
Cancer Genome Atlas

Abstract Background The tripartite motif (TRIM) family proteins exhibit oncogenic roles in various cancers. The roles of TRIM27, a member of the TRIM super family, in renal cell carcinoma (RCC) remained unexplored. In the current study, we aimed to investigate the clinical impact and roles of TRIM27 in the development of RCC. Methods The mRNA levels of TRIM27 and Kaplan–Meier survival of RCC were analyzed from The Cancer Genome Atlas database. Real-time PCR and Western blotting were used to measure the mRNA and protein levels of TRIM27 both in vivo and in vitro. siRNA and TRIM27 were exogenously overexpressed in RCC cell lines to manipulate TRIM27 expression. Results We discovered that TRIM27 was elevated in RCC patients, and the expression of TRIM27 was closely correlated with poor prognosis. The loss of function and gain of function results illustrated that TRIM27 promotes cell proliferation and inhibits apoptosis in RCC cell lines. Furthermore, TRIM27 expression was positively associated with NF-κB expression in patients with RCC. Blocking the activity of NF-κB attenuated the TRIM27-mediated enhancement of proliferation and inhibition of apoptosis. TRIM27 directly interacted with Iκbα, an inhibitor of NF-κB, to promote its ubiquitination, and the inhibitory effects of TRIM27 on Iκbα led to NF-κB activation. Conclusions Our results suggest that TRIM27 exhibits an oncogenic role in RCC by regulating NF-κB signaling. TRIM27 serves as a specific prognostic indicator for RCC, and strategies targeting the suppression of TRIM27 function may shed light on future therapeutic approaches.

YAP1 Overexpression Is Associated with Kidney Dysfunction in Lupus Nephritis

Pathobiology ◽  
2021 ◽  
pp. 1-12
Author(s):  
Ying Xie ◽  
Yuanyuan Ruan ◽  
Huimei Zou ◽  
Yixin Wang ◽  
Xin Wu ◽  
...  
Keyword(s):  
Lupus Nephritis ◽  
Epithelial Mesenchymal Transition ◽  
Kidney Tissue ◽  
Mouse Kidney ◽  
Experimental Comparison ◽  
Control Group ◽  
Mrna Levels ◽  
Mesenchymal Transition ◽  
Protein Levels

<b><i>Objective:</i></b> The goal of the present study was to determine the expression of yes-associated protein 1 (YAP1) in renal tissues of mice with lupus nephritis (LN) and elucidate its role in the progression of renal fibrosis. <b><i>Methods:</i></b> C57BL/6 mice and MRL/lpr mice were selected for experimental comparison. Mouse kidney tissues were removed and sectioned for hematoxylin and eosin staining, Masson’s trichome staining, Sirius staining, and immunohistochemistry. The mRNA and protein levels of YAP1 in mouse kidney tissues were detected, and the correlation between YAP1 and fibronectin (FN) mRNA levels was analyzed. Mouse renal epithelial cells were used for in vitro experiments. After transfection and stimulation, the cells were divided into 4 groups, namely the C57BL/6 serum group (group 1), the MRL/lpr serum group (group 2), the MRL/lpr serum + siRNA-negative control group (group 3), and the MRL/lpr serum + siRNA-YAP1 group (group 4). Epithelial-mesenchymal transition (EMT) markers in each group were detected by Western blotting and immunofluorescence staining. Serum creatinine, blood urea nitrogen, and urinary protein levels were detected and assessed for their correlation with YAP1 mRNA levels by Spearman’s analysis. <b><i>Results:</i></b> Compared to C57BL/6 mice, MRL/lpr mice exhibited obvious changes in fibrosis in renal tissues. In addition, YAP1 expression was significantly higher in the renal tissues of MRL/lpr mice than in those of C57BL/6 mice, and YAP1 mRNA levels were positively correlated with those of FN. YAP1 silencing in lupus serum-stimulated cells could effectively relieve serum-induced EMT. Finally, we observed that YAP1 mRNA levels in mouse kidney tissue were significantly and positively correlated with the degree of renal function injury. <b><i>Conclusion:</i></b> YAP1 expression in the kidney tissues of LN mice was higher than that observed in normal mice, indicating that YAP1 may play an important role in the occurrence and development of LN.

scholarly journals Maternal obesity during pregnancy leads to adipose tissue ER stress in mice via miR-126-mediated reduction in Lunapark

Diabetologia ◽  
2021 ◽  
Author(s):  
Juliana de Almeida-Faria ◽  
Daniella E. Duque-Guimarães ◽  
Thomas P. Ong ◽  
Lucas C. Pantaleão ◽  
Asha A. Carpenter ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Glucose Tolerance ◽  
Er Stress ◽  
Maternal Obesity ◽  
Luciferase Assay ◽  
Whole Body ◽  
Mrna Levels ◽  
Protein Levels ◽  
Novel Targets

Abstract Aims/hypothesis Levels of the microRNA (miRNA) miR-126-3p are programmed cell-autonomously in visceral adipose tissue of adult offspring born to obese female C57BL/6J mice. The spectrum of miR-126-3p targets and thus the consequences of its dysregulation for adipocyte metabolism are unknown. Therefore, the aim of the current study was to identify novel targets of miR-126-3p in vitro and then establish the outcomes of their dysregulation on adipocyte metabolism in vivo using a well-established maternal obesity mouse model. Methods miR-126-3p overexpression in 3T3-L1 pre-adipocytes followed by pulsed stable isotope labelling by amino acids in culture (pSILAC) was performed to identify novel targets of the miRNA. Well-established bioinformatics algorithms and luciferase assays were then employed to confirm those that were direct targets of miR-126-3p. Selected knockdown experiments were performed in vitro to define the consequences of target dysregulation. Quantitative real-time PCR, immunoblotting, histology, euglycaemic–hyperinsulinaemic clamps and glucose tolerance tests were performed to determine the phenotypic and functional outcomes of maternal programmed miR-126-3p levels in offspring adipose tissue. Results The proteomic approach confirmed the identity of known targets of miR-126-3p (including IRS-1) and identified Lunapark, an endoplasmic reticulum (ER) protein, as a novel one. We confirmed by luciferase assay that Lunapark was a direct target of miR-126-3p. Overexpression of miR-126-3p in vitro led to a reduction in Lunapark protein levels and increased Perk (also known as Eif2ak3) mRNA levels and small interference-RNA mediated knockdown of Lunapark led to increased Xbp1, spliced Xbp1, Chop (also known as Ddit3) and Perk mRNA levels and an ER stress transcriptional response in 3T3-L1 pre-adipocytes. Consistent with the results found in vitro, increased miR-126-3p expression in adipose tissue from adult mouse offspring born to obese dams was accompanied by decreased Lunapark and IRS-1 protein levels and increased markers of ER stress. At the whole-body level the animals displayed glucose intolerance. Conclusions/interpretation Concurrently targeting IRS-1 and Lunapark, a nutritionally programmed increase in miR-126-3p causes adipose tissue insulin resistance and an ER stress response, both of which may contribute to impaired glucose tolerance. These findings provide a novel mechanism by which obesity during pregnancy leads to increased risk of type 2 diabetes in the offspring and therefore identify miR-126-3p as a potential therapeutic target. Graphical abstract

scholarly journals The role of RXFP2 in mediating androgen-induced inguinoscrotal testis descent in LH receptor knockout mice

Reproduction ◽  
2010 ◽  
Vol 139 (4) ◽  
pp. 759-769 ◽  
Author(s):  
F P Yuan ◽  
X Li ◽  
J Lin ◽  
C Schwabe ◽  
E E Büllesbach ◽  
...  
Keyword(s):  
Mesenchymal Cell ◽  
Postnatal Period ◽  
Testosterone Replacement Therapy ◽  
Developmental Defect ◽  
Mrna Levels ◽  
Lh Receptor ◽  
Protein Levels ◽  
Testis Descent

LH receptor knockout (LhrKO) male mice exhibit a bilateral cryptorchidism resulting from a developmental defect in the gubernaculum during the inguinoscrotal phase of testis descent, which is corrected by testosterone replacement therapy (TRT).In vivoandin vitroexperiments were conducted to investigate the roles of the androgen receptor (AR) and RXFP2 signals in regulation of gubernacular development inLhrKO animals. This study demonstrated that AR and RXFP2 proteins were expressed in the gubernaculum during the entire postnatal period. TRT normalized gubernacular RXFP2 protein levels inLhrKO mice. Organ and primary cell cultures of gubernacula showed that 5α-dihydrotestosterone (DHT) upregulated the expression ofRxfp2which was abolished by the addition of an AR antagonist, flutamide. A single s.c. testosterone injection also led to a significant increase inRxfp2mRNA levels in a time-dependent fashion inLhrKO animals. DHT, natural and synthetic insulin-like peptide 3 (INSL3), or relaxin alone did not affect proliferation of gubernacular mesenchymal cells, while co-treatments of DHT with either INSL3 or relaxin resulted in an increase in cell proliferation, and they also enhanced the mesenchymal cell differentiation toward the myogenic pathway, which included a decrease in a mesenchymal cell marker, CD44 and the expression of troponin. These effects were attenuated by the addition of flutamide, siRNA-mediatedRxfp2knockdown, or by an INSL3 antagonist. Co-administration of an INSL3 antagonist curtailed TRT-induced inguinoscrotal testis descent inLhrKO mice. Our findings indicate that the RXFP2 signaling pathway plays an important role in mediating androgen action to stimulate gubernaculum development during inguinoscrotal testis descent.

scholarly journals The Role of the FOXO1/β2-AR/p-NF-κB p65 Pathway in the Development of Endometrial Stromal Cells in Pregnant Mice under Restraint Stress

2021 ◽  
Vol 22 (3) ◽  
pp. 1478
Author(s):  
Jiayin Lu ◽  
Yaoxing Chen ◽  
Zixu Wang ◽  
Jing Cao ◽  
Yulan Dong
Keyword(s):  
Oxidative Stress ◽  
Stromal Cells ◽  
Restraint Stress ◽  
Target Genes ◽  
Mrna Levels ◽  
Endometrial Stromal Cells ◽  
Protein Levels ◽  
Pregnant Mice

Restraint stress causes various maternal diseases during pregnancy. β2-Adrenergic receptor (β2-AR) and Forkhead transcription factor class O 1 (FOXO1) are critical factors not only in stress, but also in reproduction. However, the role of FOXO1 in restraint stress, causing changes in the β2-AR pathway in pregnant mice, has been unclear. The aim of this research was to investigate the β2-AR pathway of restraint stress and its impact on the oxidative stress of the maternal uterus. In the study, maternal mice were treated with restraint stress by being restrained in a transparent and ventilated device before sacrifice on Pregnancy Day 5 (P5), Pregnancy Day 10 (P10), Pregnancy Day 15 (P15), and Pregnancy Day 20 (P20) as well as on Non-Pregnancy Day 5 (NP5). Restraint stress augmented blood corticosterone (CORT), norepinephrine (NE), and blood glucose levels, while oestradiol (E2) levels decreased. Moreover, restraint stress increased the mRNA levels of the FOXO family, β2-AR, and even the protein levels of FOXO1 and β2-AR in the uterus and ovaries. Furthermore, restraint stress increased uterine oxidative stress level. In vitro, the protein levels of FOXO1 were also obviously increased when β2-AR was activated in endometrial stromal cells (ESCs). In addition, phosphorylated-nuclear factor kappa-B p65 (p-NF-κB p65) and its target genes decreased significantly when FOXO1 was inhibited. Overall, it can be said that the β2-AR/FOXO1/p-NF-κB p65 pathway was activated when pregnant mice were under restraint stress. This study provides a scientific basis for the origin of psychological stress in pregnant women.

scholarly journals A Theacrine-Based Supplement Increases Cellular NAD+ Levels and Affects Biomarkers Related to Sirtuin Activity in C2C12 Muscle Cells In Vitro

Nutrients ◽  
2020 ◽  
Vol 12 (12) ◽  
pp. 3727
Author(s):  
Petey W. Mumford ◽  
Shelby C. Osburn ◽  
Carlton D. Fox ◽  
Joshua S. Godwin ◽  
Michael D. Roberts
Keyword(s):  
Cell Line ◽  
Mitochondrial Biogenesis ◽  
C2c12 Myoblast ◽  
Mrna Levels ◽  
Nicotinamide Phosphoribosyltransferase ◽  
Protein Levels ◽  
Myoblast Cell Line ◽  
Rodent Cell ◽  
Myoblast Cell

There is evidence in rodents to suggest that theacrine-based supplements modulate tissue sirtuin activity as well as other biological processes associated with aging. Herein, we examined if a theacrine-based supplement (termed NAD3) altered sirtuin activity in vitro while also affecting markers of mitochondrial biogenesis. The murine C2C12 myoblast cell line was used for experimentation. Following 7 days of differentiation, myotubes were treated with 0.45 mg/mL of NAD3 (containing ~2 mM theacrine) for 3 and 24 h (n = 6 treatment wells per time point). Relative to control (CTL)-treated cells, NAD3 treatments increased (p < 0.05) Sirt1 mRNA levels at 3 h, as well as global sirtuin activity at 3 and 24 h. Follow-up experiments comparing 24 h NAD3 or CTL treatments indicated that NAD3 increased nicotinamide phosphoribosyltransferase (NAMPT) and SIRT1 protein levels (p < 0.05). Cellular nicotinamide adenine dinucleotide (NAD+) levels were also elevated nearly two-fold after 24 h of NAD3 versus CTL treatments (p < 0.001). Markers of mitochondrial biogenesis were minimally affected. Although these data are limited to select biomarkers in vitro, these preliminary findings suggest that a theacrine-based supplement can modulate select biomarkers related to NAD+ biogenesis and sirtuin activity. However, these changes did not drive increases in mitochondrial biogenesis. While promising, these data are limited to a rodent cell line and human muscle biopsy studies are needed to validate and elucidate the significance of these findings.

scholarly journals ATF6 is required for efficient rhodopsin clearance and retinal homeostasis in the P23H rho retinitis pigmentosa mouse model

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Eun-Jin Lee ◽  
Priscilla Chan ◽  
Leon Chea ◽  
Kyle Kim ◽  
Randal J. Kaufman ◽  
...  
Keyword(s):  
Retinitis Pigmentosa ◽  
Retinal Degeneration ◽  
Mrna Levels ◽  
Photoreceptor Layer ◽  
Protein Levels ◽  
Unfolded Protein ◽  
Activating Transcription Factor ◽  
The Unfolded Protein Response ◽  
Protein Response

AbstractRetinitis Pigmentosa (RP) is a blinding disease that arises from loss of rods and subsequently cones. The P23H rhodopsin knock-in (P23H-KI) mouse develops retinal degeneration that mirrors RP phenotype in patients carrying the orthologous variant. Previously, we found that the P23H rhodopsin protein was degraded in P23H-KI retinas, and the Unfolded Protein Response (UPR) promoted P23H rhodopsin degradation in heterologous cells in vitro. Here, we investigated the role of a UPR regulator gene, activating transcription factor 6 (Atf6), in rhodopsin protein homeostasis in heterozygous P23H rhodopsin (Rho+/P23H) mice. Significantly increased rhodopsin protein levels were found in Atf6−/−Rho+/P23H retinas compared to Atf6+/−Rho+/P23H retinas at early ages (~ P12), while rhodopsin mRNA levels were not different. The IRE1 pathway of the UPR was hyper-activated in young Atf6−/−Rho+/P23H retinas, and photoreceptor layer thickness was unchanged at this early age in Rho+/P23H mice lacking Atf6. By contrast, older Atf6−/−Rho+/P23H mice developed significantly increased retinal degeneration in comparison to Atf6+/−Rho+/P23H mice in all retinal layers, accompanied by reduced rhodopsin protein levels. Our findings demonstrate that Atf6 is required for efficient clearance of rhodopsin protein in rod photoreceptors expressing P23H rhodopsin, and that loss of Atf6 ultimately accelerates retinal degeneration in P23H-KI mice.

Secreted-Osteopontin Contributes to Brown Adipogenesis In Vitro via a CD44-Dependent Pathway

2019 ◽  
Vol 51 (11) ◽  
pp. 741-748
Author(s):  
Mengxi Wang ◽  
Yaoyao Guo ◽  
Yumeng Zhou ◽  
Wanwan Yuan ◽  
Huixia Li ◽  
...  
Keyword(s):  
Lipid Droplets ◽  
Biological Activities ◽  
Tumor Formation ◽  
Brown Adipocyte ◽  
Brown Adipocytes ◽  
Protein Levels ◽  
Infected Cells ◽  
Oil Red O Staining ◽  
Oil Red O

AbstractOsteopontin (OPN), a secreted glycoprotein, is involved in various pathophysiological processes including immune response, inflammation, tumor formation, and metabolism. OPN exists in 2 forms, secreted-OPN (sOPN) and intracellular-OPN (iOPN). While they might have different biological activities, it remains largely unknown whether sOPN and iOPN induce the differentiation of brown adipocytes. To test this possibility, 3T3-L1 cells were induced by DMI induction with or without recombinant human OPN (rhOPN, 10, 50, 100, 200 μM), respectively. Meanwhile, another batch of 3T3-L1 cells were infected with Ad-GFP-ap2-OPN and followed by DMI differentiation. Subsequently, the infected cells were treated with either anti-CD44 antibody or immunoglobulin G (Ig G). Accumulation of lipid droplets was visualized by Oil red O staining and protein levels were assayed by western blotting analysis. The results showed that sOPN and not rhOPN, notably increased the accumulation of lipid droplets and the expression of brown adipocyte-related genes. Moreover, neutralization of CD44 partially abrogated the effects induced by sOPN. These data demonstrate that sOPN and not rhOPN has the capacity to induce the differentiation of white preadipocytes into brown adipocytes through a CD44-dependent mechanism. The findings might provide a potential target for sOPN to combat obesity.

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