Insulin Resistance In 3T3-L1 Adipocytes Research Articles

Reversal of dexamethasone induced insulin resistance in 3T3L1 adipocytes by 3β-taraxerol of Mangifera indica

AimThe present study investigates the efficacy of Mangifera indica ethyl acetate extract (MIEE) and its bioactive compound, 3β-taraxerol in the reversal of dexamethasone (DEX) induced insulin resistance in 3T3L1 adipocytes. Main methodsMIEE and 3β-taraxerol were evaluated for their ability to restore impaired glucose uptake and, expression of molecular markers in the insulin signaling pathway induced by DEX in 3T3L1 adipocytes using 2-deoxy-d-[1-3H] glucose uptake assay and ELISA. Key findingsAn insulin resistant model has been developed using a glucocorticoid, DEX on 3T3L1 adipocytes. Insulin resistant condition was observed at 24h of DEX induction wherein a maximum degree of resistance of about 50% was measured based on inhibition of glucose uptake, which was confirmed using cytotoxicity analysis. The developed model of insulin resistance was studied in comparison to positive control rosiglitazone. DEX induced inhibition of glucose uptake and the expression of insulin signaling markers GLUT4 and PI3K were found to be restored by 3β-taraxerol and MIEE, thus delineating its mechanism of action in the reversal of insulin resistance. Significance3β-Taraxerol effectively restored DEX induced desensitization via restoration of PI3K and GLUT4 expression. To conclude, since 3β-taraxerol exhibits significant effect in reversing insulin resistance it can be further investigated as an insulin resistance reversal agent.

Endogenous CSE/H2S system mediates TNF‐α‐induced insulin resistance in 3T3‐L1 adipocytes

Tumour necrosis factor-α (TNF- α)is a major contributor to the pathogenesis of insulin resistance associated with obesity and type 2 diabetes. It has been found that endogenous hydrogen sulfide (H2 S) contributes to the pathogenesis of diabetes. We have hypothesized that TNF-α-induced insulin resistance is involved in endogenous H2 S generation. The aim of the present study is to investigate the role of endogenous H2 S in TNF-α-induced insulin resistance by studying 3T3-L1 adipocytes. We found that treatment of 3T3-L1 adipocytes with TNF-α leads to deficiency in insulin-stimulated glucose consumption and uptake and increase in endogenous H2 S generation. We show that cystathionine γ-lyase (CSE) is catalysed in 3T3-L1 adipocytes to generate H2 S and that CSE expression and activity are upregulated by TNF-α treatment. Inhibited CSE by its potent inhibitors significantly attenuates TNF-α-induced insulin resistance in 3T3-L1 adipocytes, whereas H2 S treatment of 3T3-L1 adipocytes impairs insulin-stimulated glucose consumption and uptake. These data indicate that endogenous CSE/H2 S system contributes to TNF-α-caused insulin resistance in 3T3-L1 adipocytes. Our findings suggest that modulation of CSE/H2 S system is a potential therapeutic avenue for insulin resistance.

MiRNA-21 Reverses High Glucose and High Insulin Induced Insulin Resistance in 3T3-L1 Adipocytes through Targeting Phosphatase and Tensin Homologue

Our previous study showed there was a change of microRNA (miRNA) expression profile, and miR-21 was significantly down regulated in insulin-resistant adipocytes (IR-adipocytes). Phosphatase and tensin homologs deleted on chromosome 10 (PTEN), a negative regulator of the phosphatidylinositol 3-kinase (PI3K)/AKT pathway, was identified to be a target gene of miR-21, which suggested miR-21 might be associated with insulin resistance (IR) or diabetes. However, it is not known whether miR-21 play any role in the development of IR in 3T3-L1 adipocytes. Normal adipocytes and adipocytes transfected with pre-miR-21(pmiR-21) or negative control (pNeg) were treated with high glucose and high insulin for 24 h, insulin-stimulated glucose uptake was determined by 2-Deoxyglucose transport assay, miR-21 expression level was measured by using quantitative real-time RT-PCR (qRT-PCR). The protein expression levels of PTEN, Akt, phospho-Akt (Ser473), IRβ, GSK3β, phospho-GSK3β (Ser9) and GLUT4 were detected by western blotting assay. We further confirmed that miR-21 was down regulated in IR-adipocytes by qRT-PCR. Over-expression of miR-21 significantly increased insulin-induced glucose uptake and decreased PTEN protein expression, while it had no significant effect on PTEN mRNA expression in IR-adipocytes. Moreover, over-expressing miR-21 significantly increased insulin-induced phosphorylation of AKT (Ser473), GSK3β (Ser9) and the translocation of glucose transporter 4 (GLUT4) in IR-adipocytes. In this study, our data demonstrate that miR-21 reverses high glucose and high insulin induced IR in 3T3-L1 adipocytes, possibly through modulating the PTEN-AKT pathway, and miR-21 may be a new therapeutic target for metabolic diseases such as T2DM and obesity.

α-Lipoic acid ameliorates impaired glucose uptake in LYRM1 overexpressing 3T3-L1 adipocytes through the IRS-1/Akt signaling pathway

Overexpression of the Homo sapiens LYR motif containing 1 (LYRM1) causes mitochondrial dysfunction and induces insulin resistance in 3T3-L1 adipocytes. α-Lipoic acid (α-LA), a dithiol compound with antioxidant properties, improves glucose transport and utilization in 3T3-L1 adipocytes. The aim of this study was to investigate the direct effects of α-LA on reactive oxygen species (ROS) production and insulin sensitivity in LYRM1 overexpressing 3T3-L1 adipocytes and to explore the underlying mechanism. Pretreatment with α-LA significantly increased both basal and insulin-stimulated glucose uptake and insulin-stimulated GLUT4 translocation, while intracellular ROS levels in LYRM1 overexpressing 3T3-L1 adipocytes were decreased. These changes were accompanied by a marked upregulation in expression of insulin-stimulated tyrosine phosphorylation of IRS-1 and serine phosphorylation of Akt following treatment with α-LA. These results indicated that α-LA protects 3T3-L1 adipocytes from LYRM1-induced insulin resistance partially via its capacity to restore mitochondrial function and/or increase phosphorylation of IRS-1 and Akt.

Green tea catechins ameliorate adipose insulin resistance by improving oxidative stress

Epidemiological data have suggested that drinking green tea is negatively associated with diabetes, and adipose oxidative stress may have a central role in causing insulin resistance, according to recent findings. The aim of this work is to elucidate a new mechanism for green tea's anti-insulin resistance effect. We used obese KK-ay mice, high-fat diet-induced obese rats, and induced insulin resistant 3T3-L1 adipocytes as models. Insulin sensitivity and adipose reactive oxidative species (ROS) levels were detected in animals and adipocytes. The oxidative stress assay and glucose uptake ability assay were performed, and the effects of EGCG on insulin signals were detected. Green tea catechins (GTCs) significantly decreased glucose levels and increased glucose tolerance in animals. GTCs reduced ROS content in both models of animal and adipocytes. EGCG attenuated dexamethasone and TNF-α promoted ROS generation and increased glucose uptake ability. EGCG also decreased JNK phosphorylation and promoted GLUT-4 translocation. EGCG and GTCs could improve adipose insulin resistance, and exact this effect on their ROS scavenging functions.

Improvement of insulin resistance by Cyanidin 3-glucoside, anthocyanin from black beans through the up-regulation of GLUT4 gene expression

Black beans have been suggested to have a protective effect against obesity. Cyanidin 3-glucoside (Cy-3-G) belongs to the flavonoid class of molecules and is a member of the anthocyanin family that is present in black beans. Some studies indicated that Cy-3-G from black beans may be beneficial for the improvement of obesity and type Ⅱ diabetes. It has been reported that Cyanidin 3-glucoside ameliorates insulin sensitivity due by down-regulating the retinol binding protein 4 expression in diabetic mice [1]. Accumulation of neutral lipids, triglyceride (TG) in particular, is highly related to the development of insulin resistance and its consequences, such as type Ⅱ diabetes. Lipid droplet size regulation is central in the regulation of metabolism and in adipocytokines secretion such as TNF-α and adiponectin [2,3]. However, little is currently known about how Cy-3-G influences adipocyte differentiation and insulin resistance in 3T3-L1 adipocytes. The purpose of the present study was to determine the effects of Cy-3-G on GLUT4 gene expression to improve insulin resistance in 3T3-L1 adipocytes.

Heparin-Binding EGF-Like Growth Factor (HB-EGF) Mediates 5-HT-Induced Insulin Resistance Through Activation of EGF Receptor-ERK1/2-mTOR Pathway

Although an inverse correlation between insulin sensitivity and the level of Gq/11-coupled receptor agonists, such as endothelin-1, thrombin, and 5-hydroxytryptamine (5-HT), has been reported, its precise mechanism remains unclear. In this report, we provide evidence that 5-HT induced production of heparin-binding epidermal growth factor-like growth factor (HB-EGF) and caused insulin resistance in 3T3-L1 adipocytes, primary adipocytes, and C2C12 myotubes. In 3T3-L1 adipocytes, 5-HT stimulated HB-EGF production by promoting metalloproteinase-dependent shedding of transmembrane protein pro-HB-EGF. HB-EGF then bound and tyrosine-phosphorylated EGF receptors, which activated the mammalian target of rapamycin pathway through ERK1/2 phosphorylation. Mammalian target of rapamycin activation caused serine phosphorylation of insulin receptor substrate-1, which attenuated insulin-stimulated tyrosine phosphorylation of insulin receptor substrate-1 and glucose uptake. Pharmacological inhibition of either Gq/11-coupled receptors or metalloproteinases, as well as either inhibition or knockdown of HB-EGF or Gαq/11, restored insulin signal transduction impaired by 5-HT. Inhibition of metalloproteinase activity also abolished HB-EGF production and subsequent EGF receptor activation by other Gq/11-coupled receptor agonists known to cause insulin resistance, such as endothelin-1 and thrombin. These results suggest that transactivation of the EGF receptor through HB-EGF processing plays a pivotal role in 5-HT-induced insulin resistance.

Pioglitazone upregulates adiponectin receptor 2 in 3T3-L1 adipocytes

AimsPioglitazone, a full peroxisome proliferator-activated receptor (PPAR)-γ agonist, improves insulin sensitivity by increasing circulating adiponectin levels. However, the molecular mechanisms by which pioglitazone induces insulin sensitization are not fully understood. In this study, we investigated whether pioglitazone improves insulin resistance via upregulation of either 2 distinct receptors for adiponectin (AdipoR1 or AdipoR2) expression in 3T3-L1 adipocytes. Main methodsGlucose uptake was evaluated by 2-[3H] deoxy-glucose uptake assay in 3T3-L1 adipocytes with pioglitazone treatment. AdipoR1 and AdipoR2 mRNA expressions were analyzed by qRT–PCR. Key findingsWe first confirmed that pioglitazone significantly increased insulin-induced 2-deoxyglucose (2-DOG) uptake in 3T3-L1 adipocytes. Next, we investigated the mRNA expression and regulation of AdipoR1 and AdipoR2 after treatment with pioglitazone. Interestingly, pioglitazone significantly induced AdipoR2 expression but it did not affect AdipoR1 expression. In addition, adenovirus-mediated PPARγ expression significantly enhanced the effects of pioglitazone on insulin-stimulated 2-DOG uptake and AdipoR2 expression in 3T3-L1 adipocytes. These data suggest that pioglitazone enhances adiponectin's autocrine and paracrine actions in 3T3-L1 adipocytes via upregulation of PPARγ-mediated AdipoR2 expression. Furthermore, we found that pioglitazone significantly increased AMP-activated protein kinase (AMPK) phosphorylation in insulin-stimulated 3T3-L1 adipocytes, but it did not lead to the phosphorylation of IRS-1, Akt, or protein kinase Cλ/ζ. SignificanceOur results suggest that pioglitazone increases insulin sensitivity, at least partly, by PPARγ-AdipoR2-mediated AMPK phosphorylation in 3T3-L1 adipocytes. In conclusion, the upregulation of AdipoR2 expression may be one of the mechanisms by which pioglitazone improves insulin resistance in 3T3-L1 adipocytes.

Apelin stimulates glucose uptake through the PI3K/Akt pathway and improves insulin resistance in 3T3-L1 adipocytes

Apelin, a cytokine mainly secreted by adipocytes, is closely related with insulin resistance. The underlying molecular mechanisms of how apelin affects insulin resistance, however, are poorly understood. This study aimed to investigate the effect of apelin on glucose metabolism and insulin resistance in 3T3-L1 adipocytes. After 10 ng/ml TNF-α treatment for 24 h, insulin-stimulated glucose uptake was reduced by 47% in 3T3-L1 adipocytes. Apelin treatment improved glucose uptake in a time- and dose-dependent manner. Treatment of 1,000 nM apelin for 60 min maximally augmented glucose uptake in insulin-resistant 3T3-L1 adipocytes. Furthermore, apelin pre-incubation also increased adipocytes' insulin-stimulated glucose uptake, and PI3K/Akt pathway were involved in these effects. In addition, immunocytochemistry staining and western blotting analysis indicated that apelin could increase glucose transporter 4 translocation from the cytoplasm to the plasma membrane. Apelin also increased the anti-inflammatory adipokine adiponectin mRNA expression while reducing that of pro-inflammatory adipokine interleukin-6 in insulin-resistant 3T3-L1 adipocytes. These results suggest that apelin stimulates glucose uptake through the PI3K/Akt pathway, promotes GLUT4 translocation from the cytoplasm to the plasma membrane, and modulates inflammatory responses in insulin-resistant 3T3-L1 adipocytes.

Andrographolide attenuates tumor necrosis factor-alpha-induced insulin resistance in 3T3-L1 adipocytes

Andrographolide (AG), the primary bioactive component of Andrographils paniculate Nees, has showed an anti-diabetic effect. However, the molecular mechanism has not been clarified. In this study, we demonstrated that AG increased glucose uptake in 3T3-L1 cells in a time- and dosedependent manner. The activation of insulin signaling by AG was initiated from phosphotyrosine of IRS-1 and further passed on through phosphatidylinositol 3-kinase (PI3K) and the downstream signaling cascades. Moreover importantly, pretreatment cells with AG suppressed the TNF-α induced activation of NF-κB signaling pathway and its downstream inflammatory factors expression, therefore ameliorating insulin resistance. In conclusion, AG can improve insulin sensitivity through inhibition of NF-κB pathway. These findings are helpful in understanding the anti-diabetic properties of AG and can be of interest for the therapeutic application of AG in glucose controlling.

E0012 Apelin stimulates glucose uptake through PI3KAkt pathway in insulin resistant 3T3L1 adipocytes

BackgroundApelin, a cytokine mainly secreted by adipocytes, shows beneficial effect on insulin resistance. However, the molecular mechanism underlying is still poorly understood. This study was to investigate the mechanisms of...

Inhibition of O-GlcNAcase Using a Potent and Cell-Permeable Inhibitor Does Not Induce Insulin Resistance in 3T3-L1 Adipocytes

SummaryTo probe increased O-GlcNAc levels as an independent mechanism governing insulin resistance in 3T3-L1 adipocytes, a new class of O-GlcNAcase (OGA) inhibitor was studied. 6-Acetamido-6-deoxy-castanospermine (6-Ac-Cas) is a potent inhibitor of OGA. The structure of 6-Ac-Cas bound in the active site of an OGA homolog reveals structural features contributing to its potency. Treatment of 3T3-L1 adipocytes with 6-Ac-Cas increases O-GlcNAc levels in a dose-dependent manner. These increases in O-GlcNAc levels do not induce insulin resistance functionally, measured using a 2-deoxyglucose (2-DOG) uptake assay, or at the molecular level, determined by evaluating levels of phosphorylated IRS-1 and Akt. These results, and others described, provide a structural blueprint for improved inhibitors and collectively suggest that increased O-GlcNAc levels, brought about by inhibition of OGA, does not by itself cause insulin resistance in 3T3-L1 adipocytes.

Enhanced long-chain fatty acid uptake contributes to overaccumulation of triglyceride in hyperinsulinemic insulin-resistant 3T3-L1 adipocytes

The precise pathogenesis of obesity remains controversial. In obesity, diminished adipose glucose utilization suggests that some other substrates may be responsible for the adipose triglyceride (TG) overaccumulation. Here we attempted to evaluate if long-chain fatty acid (LCFA) flux was modulated by a physiologically relevant condition of hyperinsulinemia in 3T3-L1 adipocytes and if the altered LCFA influx might eventually contribute to the TG overaccumulation in obesity. The effects of prolonged insulin exposure to adipocytes on basal, insulin-stimulated LCFA uptake as well as intracellular LCFA metabolism were measured. Prolonged insulin exposure was found to induce insulin resistance (IR) yet enhance basal and insulin-stimulated LCFA uptake in normoglycemic condition, and the addition of high glucose exacerbated these abnormalities of both glucose and LCFA influx. Along with the enhanced LCFA uptake was an increase in the rates of intracellular LCFA deposition and incorporation into TG; but a decrease was found in basal and insulin-suppressive LCFA oxidation, as well as in isoproterenol-induced fatty acid efflux. Inhibition of either phosphatidylinositol 3-kinase or mitogen-activated protein kinase (MAPK) pathway did not prevent the induction of IR, whereas the enhanced basal and insulin-stimulated LCFA uptake was abrogated by inhibition of MAPK pathway. In hyperinsulinemic insulin-resistant 3T3-L1 adipocytes, basal and insulin-stimulated LCFA uptake tends to increase via a MAPK-dependent mechanism. The increment of LCFA influx predominantly accounts for TG overaccumulation, but not for mitochondrial oxidation, and is prone to retain within adipocytes. These findings may interpret the plausible mechanism of pathogenesis for obesity in hyperinsulinemia-associated IR.

Bis(α-furancarboxylato)oxovanadium(IV) prevents and improves dexamethasone-induced insulin resistance in 3T3-L1 adipocytes

Abstract Previous studies showed that bis(α-furancarboxylato)oxovanadium(IV) (BFOV), an orally active antidiabetic organic vanadium complex, could improve insulin resistance in animals with type 2 diabetes. The present study has been carried out to evaluate the effects of BFOV on insulin-resistant glucose metabolism using dexamethasone-treated 3T3-L1 adipocytes as an in-vitro model of insulin resistance. The results showed that BFOV, similar to vanadyl sulfate and rosiglitazone, caused a concentration-dependent increase in glucose consumption by insulin-resistant adipocytes. Moreover, BFOV enhanced the action of insulin and completely prevented the development of insulin resistance induced by dexamethasone, leading to glucose consumption equal to that by normal cells. In addition, dexamethasone reduced the mRNA expression of insulin receptor substrate 1 (IRS-1) and glucose transporter 4 (GLUT4) in 3T3-L1 adipocytes, while BFOV normalized the expression of IRS-1 and GLUT4. These findings suggest that BFOV prevents and improves dexamethasone-induced insulin resistance in 3T3-L1 adipocytes by enhancing expression of IRS-1 and GLUT4 mRNA.

β-Carotene accumulation in 3T3-L1 adipocytes inhibits the elevation of reactive oxygen species and the suppression of genes related to insulin sensitivity induced by tumor necrosis factor-α

Objective β-Carotene is an abundant carotenoid with potent antioxidative activities and accumulates in adipose tissue. However, its physiologic functions are poorly understood. In this study, we examined whether accumulation of β-carotene for 4 d in insulin-resistant 3T3-L1 adipocytes alters the expression of genes related to insulin sensitivity. Methods The 3T3-L1 adipocytes were treated with/without 10 or 20 μM β-carotene during differentiation for 4 d. The cells treated with 10 μM β-carotene for 4 d were subsequently incubated with/without 5 ng/mL of tumor necrosis factor-α for 48 h in the medium without β-carotene. The mRNA levels of genes in the cells and adiponectin protein levels in the medium were determined by real-time reverse transcription–polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Reactive oxygen species levels in the cells were assessed by oxidation of 2′,7′-dichlorodihydrofluorescein diacetate. Result β-Carotene treatment at a concentration of 20 μM, but not 10 μM, in 3T3-L1 adipocytes during differentiation for 4 d enhanced the expression of genes related to insulin sensitivity, including adiponectin, adipocyte lipid-binding protein, glucose transporter-4, peroxisome proliferator-activated receptor-γ2, and adiponectin protein in the medium. Tumor necrosis factor-α treatment repressed the expression of these genes and adiponectin protein in the medium and induced reactive oxygen species levels. In contrast, cells that accumulated β-carotene at a concentration of 10 μM did not show these alterations. Conclusion The accumulation of the β-carotene in 3T3-L1 adipocytes restores the expression of genes related to insulin sensitivity and reactive oxygen species levels in insulin-resistant adipocytes.

Progesterone inhibits glucose uptake by affecting diverse steps of insulin signaling in 3T3-L1 adipocytes

Maternal insulin resistance is essential for efficient provision of glucose to the fetus. Although elevation of placental hormones is known to relate to the development of insulin resistance, the precise underlying mechanism of maternal insulin resistance is unknown. Therefore, we examined the molecular mechanisms of progesterone causing insulin resistance in 3T3-L1 adipocytes. Progesterone at 10(-4) M, but not 10(-5) M, reduced the amount of IRS-1. As a result, insulin-induced phosphorylation of IRS-1, the association of IRS-1 with p85alpha, and subsequent phosphorylation of Akt1 and -2 was decreased moderately by 10(-4) M progesterone. Subsequently, insulin-induced translocation of GLUT4 to the plasma membrane evaluated by immunostaining on the plasma membrane sheet by confocal laser microscope was also decreased by 10(-4) M progesterone. In contrast, 2-[(3)H]deoxyglucose (2DG) uptake was markedly inhibited by both 10(-5) and 10(-4) M progesterone in a dose-dependent manner. Surprisingly, 2DG uptake elicited by adenovirus-mediated expression of constitutive-active mutant of PI 3-kinase (myr-p110) and Akt (myr-Akt) was suppressed by progesterone. Interestingly, insulin-induced tyrosine phosphorylation of Cbl and activation of TC10 were inhibited by progesterone at 10(-5) M. These results indicate that progesterone is implicated in insulin resistance during pregnancy by inhibiting the PI 3-kinase pathway at the step of 1) IRS-1 expression and 2) distal to Akt and 3) by suppressing the PI 3-kinase-independent pathway of TC10 activation by affecting Cbl phosphorylation.

Transcription factor AP-2β inhibits expression and secretion of leptin, an insulin-sensitizing hormone, in 3T3-L1 adipocytes

We have previously reported an association between the activator protein-2beta (AP-2beta) transcription factor gene and type 2 diabetes. This gene is preferentially expressed in adipose tissue, and subjects with a disease-susceptible allele of AP-2beta showed stronger AP-2beta expression in adipose tissue than those without the susceptible allele. Furthermore, overexpression of AP-2beta led to lipid accumulation and induced insulin resistance in 3T3-L1 adipocytes. We found that overexpression of AP-2beta in 3T3-L1 adipocytes decreased the promoter activity of leptin, and subsequently decreased both messenger RNA (mRNA) and protein expression and secretion. Furthermore, knockdown of endogenous AP-2beta by RNA-interference increased mRNA and protein expression of leptin. Electrophoretic mobility shift and chromatin immunoprecipitation assays revealed specific binding of AP-2beta to leptin promoter regions in vitro and in vivo. In addition, site-directed mutagenesis of the AP-2-binding site located between position +34 and +42 relative to the transcription start site abolished the inhibitory effect of AP-2beta. Our results clearly showed that AP-2beta directly inhibited insulin-sensitizing hormone leptin expression by binding to its promoter. AP-2beta modulated the expression of leptin through direct interaction with its promoter region.

Effects of a grapeseed procyanidin extract (GSPE) on insulin resistance

Flavonoids are beneficial compounds against risk factors for metabolic syndrome, but their effects and the mechanisms on glucose homeostasis modulation are not well defined. In the present study, we first checked the efficacy of grapeseed procyanidin extract (GSPE) for stimulating glucose uptake in insulin-resistant 3T3-L1 adipocytes. Results show that when resistance is induced with chronic insulin treatment, GSPE maintain a higher stimulating capacity than insulin. In contrast, when dexamethasone is used as the resistance-inducing agent, GSPE is less effective. Next we evaluated how effective different GSPE treatments are at improving glucose metabolism in hyperinsulinemic animals (fed a cafeteria diet). GSPE reduced plasma insulin levels. The lower dose (25 mg GSPE/kg body weight per day) administered for 30 days improved the HOmeostasis Model Assessment-insulin resistance index. This was accompanied by down-regulation of Pparg2, Glut4 and Irs1 in mesenteric white adipose tissue. Similarly, a chronic GSPE treatment of insulin-resistant 3T3-L1 adipocytes down-regulated the mRNA levels of those adipocyte markers, although cells were still able to respond to the acute stimulation of glucose uptake. In summary, 25 mg/kg body weight per day of GSPE has a positive long-term effect on glucose homeostasis, and GSPE could be targeted at adipose tissue, where it might directly stimulate glucose uptake. This work also highlights the need to carefully consider the bioactive dose, since a higher dose does not necessarily correlate to a greater positive effect.

CHROMIUM PICOLINATE INHIBITS RESISTIN SECRETION IN INSULIN‐RESISTANT 3T3‐L1 ADIPOCYTES VIA ACTIVATION OF AMP‐ACTIVATED PROTEIN KINASE

1. Chromium picolinate (CrPic) has been recommended as an alternative therapeutic regimen for Type 2 diabetes mellitus (T2DM). However, the molecular mechanism underlying the action of CrPic is poorly understood. 2. Using normal and insulin-resistant 3T3-L1 adipocytes, we examined the effects of CrPic on the gene transcription and secretion of adiponectin and resistin. In addition, using immunoblotting, ELISA and real-time reverse transcription-polymerase chain reaction (RT-PCR), we investigated the effects of 10 nmol/L CrPic for 24 h on AMP-activated protein kinase (AMPK) to determine whether this pathway contributed to the regulation of adiponectin and resistin expression and secretion. 3. Chromium picolinate did not modulate the expression of adiponectin and resistin; however, it did significantly inhibit the secretion of resistin, but not adiponectin, by normal and insulin-resistant 3T3-L1 adipocytes in vitro. Furthermore, although CrPic markedly elevated levels of phosphorylated AMPK and acetyl CoA carboxylase in 3T3-L1 adipocytes, it had no effect on the levels of AMPK alpha-1 and alpha-2 mRNA transcripts. Importantly, inhibition of AMPK by 2 h pretreatment of cells with 20 micromol/L compound C completely abolished the CrPic-induced suppression of resistin secretion. 4. In conclusion, the data suggest that CrPic inhibits resistin secretion via activation of AMPK in normal and insulin-resistant 3T3-L1 adipocytes.

Biological effects of THC and a lipophilic cannabis extract on normal and insulin resistant 3T3-L1 adipocytes

Type 2 diabetes, a chronic disease, affects about 150 million people world wide. It is characterized by insulin resistance of peripheral tissues such as liver, skeletal muscle, and fat. Insulin resistance is associated with elevated levels of tumor necrosis factor alpha (TNF-alpha), which in turn inhibits insulin receptor tyrosine kinase autophosphorylation. It has been reported that cannabis is used in the treatment of diabetes. A few reports indicate that smoking cannabis can lower blood glucose in diabetics. Delta(9)-tetrahydrocannabinol (THC) is the primary psychoactive component of cannabis. This study aimed to determine the effect of a lipophilic cannabis extract on adipogenesis, using 3T3-L1 cells, and to measure its effect on insulin sensitivity in insulin resistant adipocytes. Cells were cultured in Dulbecco's modified eagle medium (DMEM) with 10% fetal bovine serum (FBS) and differentiated over a 3 day period for all studies. In the adipogenesis studies, differentiated cells were exposed to the extract in the presence and absence of insulin. Lipid content and glucose uptake was subsequently measured. Insulin-induced glucose uptake increased, while the rate of adipogenesis decreased with increasing THC concentration. Insulin-resistance was induced using TNF-alpha, exposed to the extract and insulin-induced glucose uptake measured. Insulin-induced glucose was increased in these cells after exposure to the extract. Semiquantitative real time polymerase chain reaction (RT-PCR) was performed after ribonucleic acid (RNA) extraction to evaluate the effects of the extract on glucose transporter isotype 4 (GLUT-4), insulin receptor substrate-1 (IRS-1) and IRS-2 gene expression.