scholarly journals Intermittent Hypoxia Composite Abnormal Glucose Metabolism-Mediated Atherosclerosis In Vitro and In Vivo: The Role of SREBP-1

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Linqin Ma ◽  
Jingchun Zhang ◽  
Yu Qiao ◽  
Xinli Sun ◽  
Ting Mao ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Glucose Metabolism ◽  
Intermittent Hypoxia ◽  
Control Group ◽  
Abnormal Glucose Metabolism ◽  
Mrna And Protein Expression ◽  
Oil Red O Staining ◽  
Oil Red O

Objective. The aim of this study was to establish a 3T3-L1 adipocyte model and ApoE−/− mouse model of intermittent hypoxia (IH) composite abnormal glucose metabolism (AGM) in vitro and in vivo and explore their synergistic damage effect leading to atherosclerosis (AS) and the influence of SREBP-1 signaling molecule-related mechanisms. Methods. Mature 3T3-L1 adipocytes were cultured with complete culture medium containing DEX 1×106 mol/L for 96 h to establish an AGM-3T3-L1 adipocyte model. Then, AGM-3T3-L1 adipocytes were treated with IH for 0 cycles, 2 cycles, 4 cycles, 8 cycles, 16 cycles, and 32 cycles and sustained hypoxia (SH). ApoE−/− mice were treated with high-fat diet and injection of STZ solution to establish an AGM-ApoE−/− mouse model. A total of 16 AGM-ApoE−/− mice were randomly and averagely divided into the normoxic control group (NC) and model group (CIH). AGM-ApoE−/− mice of the CIH group were treated with IH, which meant that the oxygen concentration fell to 10±0.5% in the first 90 seconds of one cycle and then increased to 21±0.5% in the later 90 seconds, continuous for eight hours per day (09 : 00-17 : 00) with a total of eight weeks. Eight C57BL/6J mice were used as the blank control group (Con) which was fed with conventional diet. qPCR and Western blotting were used to detect the expression level of SREBP-1c, FAS, and IRS-1. Oil Red O staining was used to compare the plaque of the aorta among each mouse group. Results. As a result, within 32 cycles of IH, mRNA and protein expression levels of SREBP-1c and FAS in AGM-3T3-L1 adipocytes were elevated with the increase in IH cycles; the mRNA expression of IRS-1 was decreased after IH 32 cycles and lower than that of the SH group. For the study in vivo, Oil Red O staining showed a more obvious AS aortic plaque in the CIH group. After CIH treatment of 4 w and 8 w, fasting blood glucose (FBG) of the NC group and CIH group was higher than that of the Con group, and the insulin level of the CIH group was higher than that of the Con group after IH treatment of 8 w. The expressions of the IRS-1 mRNA level in the aorta, skeletal muscle, and liver of the CIH group were lower than those in the Con group. The mRNA and protein expression of SREBP-1c and its downstream molecule FAS in the aorta, skeletal muscle, and liver significantly enhanced in the CIH group in contrast with those in the Con group. Conclusion. The CIH composite AGM could promote the progress of AS, which might be related to the modulation of the expression of SREBP-1-related molecular pathways.

scholarly journals Insulin action in growth hormone-deficient and age-matched control rats: effect of growth hormone treatment

1999 ◽  
Vol 160 (1) ◽  
pp. 127-135 ◽  
Author(s):  
◽  
JL Laustsen ◽  
BS Hansen ◽  
EA Richter
Keyword(s):  
Skeletal Muscle ◽  
Growth Hormone ◽  
Glucose Metabolism ◽  
Recombinant Human Growth Hormone ◽  
Normal Weight ◽  
Lower Amount ◽  
Control Group ◽  
Muscle Fibre Size ◽  
Igf I

The isolated effect of growth hormone on carbohydrate metabolism in rat skeletal muscle was studied in growth hormone-deficient dwarf rats (dw/dw) treated with either recombinant human growth hormone or saline for 10 days. In addition, age-matched heterozygous (DW/dw) (normal weight and plasma IGF-I) control rats were treated with saline. Growth hormone increased weight gain from 0.1+/-0.1 (s.e.m) to 3.6+/-0.1 g/day and plasma IGF-I concentration from 364+/-23 to 451+/-32 ng/ml. Glucose metabolism in skeletal muscle perfused with basal, submaximal and maximal concentrations (0, 600 and 60 000 pmol/l respectively) of insulin was not changed by growth hormone. No change could be detected in the total number of glucose transporters (GLUT1 and GLUT4) in the skeletal muscles, except from a lower amount of GLUT4 in the soleus muscle in the heterozygous control group. However, at submaximal insulin concentrations, skeletal muscle glucose uptake and transport were significantly lower in the heterozygous control group compared with the growth hormone-deficient group. This could indicate either a direct long-term effect of growth hormone or more likely a secondary effect attributable to the difference in body weight (205.2+/-3.1 vs 361. 6+/-5.9 g for dwarf rats and heterozygous controls respectively), and thereby muscle fibre size, between the groups probably resulting in lower average interstitial insulin and glucose concentrations at a given plasma concentration in the heterozygous rats. It is concluded that restoration of subnormal growth hormone concentrations for 10 days has no effect on insulin-stimulated glucose metabolism in skeletal muscle in vitro.

scholarly journals Mechanism of KLF4 Protection against Acute Liver Injury via Inhibition of Apelin Signaling

2019 ◽  
Vol 2019 ◽  
pp. 1-10 ◽  
Author(s):  
Weitao Ji ◽  
Hongyun Shi ◽  
Hailin Shen ◽  
Jing Kong ◽  
Jiayi Song ◽  
...  
Keyword(s):  
Liver Injury ◽  
Signaling Pathway ◽  
Acute Liver Injury ◽  
Control Group ◽  
Necrosis Factor Alpha ◽  
Protein Levels ◽  
Klf4 Expression ◽  
Mrna And Protein Expression

Krüppel-like factor 4 (KLF4) is a key transcription factor that regulates genes involved in the proliferation or differentiation in different tissues. Apelin plays roles in cardiovascular functions, metabolic disease, and homeostatic disorder. However, the biological function of apelin in liver disease is still ongoing. In this study, we investigated the mechanism of KLF4-mediated protection against acute liver injury via the inhibition of the apelin signaling pathway. Mice were intraperitoneally injected with carbon tetrachloride (CCl4; 0.2 mL dissolved in 100 mL olive oil, 10 mL/kg) to establish an acute liver injury model. A KLF4 expression plasmid was injected through the tail vein 48 h before CCl4 treatment. In cultured LX-2 cells, pAd-KLF4 or siRNA KLF4 was overexpressed or knockdown, and the mRNA and protein levels of apelin were determined. The results showed that the apelin serum level in the CCl4-injected group was higher than that of control group, and the expression of apelin in the liver tissues was elevated while KLF4 expression was decreased in the CCl4-injected group compared to the KLF4-plasmid-injected group. HE staining revealed serious hepatocellular steatosis in the CCl4-injected mice, and KLF4 alleviated this steatosis in the mice injected with KLF4 plasmid. In vitro experiments showed that tumor necrosis factor-alpha (TNF-α) could downregulate the transcription and translation levels of apelin in LX-2 cells and also upregulate KLF4 mRNA and protein expression. RT-PCR and Western blotting showed that the overexpression of KLF4 markedly decreased basal apelin expression, but knockdown of KLF4 restored apelin expression in TNF-α-treated LX-2 cells. These in vivo and in vitro experiments suggest that KLF4 plays a key role in inhibiting hepatocellular steatosis in acute liver injury, and that its mechanism might be the inhibition of the apelin signaling pathway.

scholarly journals Oroxin B Induces Apoptosis by Down-Regulating MicroRNA-221 Resulting in the Inactivation of the PTEN/PI3K/AKT Pathway in Liver Cancer

Molecules ◽  
2019 ◽  
Vol 24 (23) ◽  
pp. 4384 ◽  
Author(s):  
Nannan Li ◽  
Wenxiao Men ◽  
Yibo Zheng ◽  
Hechen Wang ◽  
Xiansheng Meng
Keyword(s):  
Liver Cancer ◽  
Western Blot ◽  
Microfluidic Chip ◽  
Pten Gene ◽  
Control Group ◽  
Rt Pcr ◽  
Mrna And Protein Expression ◽  
Theoretical Evidence

This study aims to investigate the anticancer effect of Oroxin B (OB) both in vitro and in vivo, and the molecular mechanism involved in microRNA-221 and the PI3K/Akt/PTEN pathway through modulation of apoptosis in Hepatocellular carcinoma (HCC). DEN-induced rats and HepG2 cells based on the microfluidic chip were employed, while the mRNA and protein expression of microRNA-221, PI3K, p-Akt and PTEN were evaluated by RT-PCR and Western blot analysis. Based on Microfluidic Chip and DENinduced rat model, OB effectively exerts anti-liver cancer effect both in vitro and in vivo, and the expression of miR-221 in OB treated groups was significantly lower than that in the control group (** p < 0.01). The RT-PCR and Western blot results suggested the PI3K mRNA and protein in OB treated groups were both lower than those in control group and indicated the overexpression of PTEN. Therefore, OB effectively exerts anticancer effects by positively regulating the PTEN gene and then inactivating the PI3K/Akt signaling pathway through down-regulating the expression of the microRNA-221, thereby inducing apoptosis of liver cancer cells. This study offers a theoretical evidence for further development and clinical guidance of OB as an anti-tumor agent.

scholarly journals Improved Tetanic Force and Mitochondrial Calcium Homeostasis by Astaxanthin Treatment in Mouse Skeletal Muscle

Antioxidants ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 98 ◽  
Author(s):  
Sztretye ◽  
Singlár ◽  
Szabó ◽  
Angyal ◽  
Balogh ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Calcium Homeostasis ◽  
Inhibitory Effect ◽  
Calcium Transients ◽  
Muscle Performance ◽  
Control Group ◽  
Mitochondrial Calcium ◽  
Tetanic Force

Background: Astaxanthin (AX) a marine carotenoid is a powerful natural antioxidant which protects against oxidative stress and improves muscle performance. Retinol and its derivatives were described to affect lipid and energy metabolism. Up to date, the effects of AX and retinol on excitation-contraction coupling (ECC) in skeletal muscle are poorly described. Methods: 18 C57Bl6 mice were divided into two groups: Control and AX supplemented in rodent chow for 4 weeks (AstaReal A1010). In vivo and in vitro force and intracellular calcium homeostasis was studied. In some experiments acute treatment with retinol was employed. Results: The voltage activation of calcium transients (V50) were investigated in single flexor digitorum brevis isolated fibers under patch clamp and no significant changes were found following AX supplementation. Retinol shifted V50 towards more positive values and decreased the peak F/F0 of the calcium transients. The amplitude of tetani in the extensor digitorum longus was significantly higher in AX than in control group. Lastly, the mitochondrial calcium uptake was found to be less prominent in AX. Conclusion: AX supplementation increases in vitro tetanic force without affecting ECC and exerts a protecting effect on the mitochondria. Retinol treatment has an inhibitory effect on ECC in skeletal muscle.

scholarly journals Therapeutic potential of small extracellular vesicles derived from lipoma tissue in adipose tissue regeneration—an in vitro and in vivo study

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Pengyu Hong ◽  
Xiaoyang Xu ◽  
Xin Hu ◽  
Hao Yang ◽  
Yue Wu ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Extracellular Vesicles ◽  
Tissue Regeneration ◽  
Lipid Droplets ◽  
Adipogenic Differentiation ◽  
Rt Pcr ◽  
Oil Red O Staining ◽  
Oil Red O

Abstract Objective To explore the adipogenic effects of the small extracellular vesicles derived from the lipoma tissues (sEV-LT), and to find a new cell-free therapeutic approach for adipose tissue regeneration. Methods Adipose tissue-derived stem cells (ADSCs) and small extracellular vesicles derived from the adipose tissues (sEV-AT) were isolated from human adipose tissue, while sEV-LT were isolated from human lipomatous tissue. ADSCs were characterized by using flow cytometric analysis and adipogenic and osteogenic differentiation assays. sEV was identified by electron microscopy, nanoparticle tracking, and western blotting. ADSCs were treated with sEV-LT and sEV-AT, respectively. Fluorescence confocal microscopy was used to investigate whether sEV-LT and sEV-AT could be taken by ADSCs. The proliferation and migration abilities and adipogenic differentiation assay of ADSCs were evaluated by CCK-8 assays, scratch test, and oil red O staining test, and the expression levels of adipogenic-related genes C/EBP-δ, PPARγ2, and Adiponectin in ADSCs were assessed by real-time quantitative PCR (RT-PCR). The sEV-LT and sEV-AT transplantation tubes were implanted subcutaneously in SD rats, and the neotissues were qualitatively and histologically evaluated at 2, 4, 8, and 12 weeks after transplantation. Hematoxylin and eosin (H&E) staining was subsequently used to observe and compare the adipogenesis and angiogenesis in neotissues, while immunohistochemistry was used to examine the expression and the distribution of C/EBP-α, PPARγ, Adiponectin, and CD31 at the 4th week. Results The in vitro experiments showed that both sEV-LT and sEV-AT could be taken up by ADSCs via endocytosis. The scratch experiment and CCK-8 experiment showed that the migration area and proliferation number of ADSCs in sEV-LT group and sEV-AT group were significantly higher than those in the non-sEV group (p < 0.05). Compared with sEV-AT group, sEV-LT group had larger migration area and proliferation number of ADSCs (p < 0.05). Oil red O staining and RT-PCR experiments showed that, compared with the non-sEVs group, the lipid droplets and the mRNA expression levels of adipogenesis-related genes PPARγ2 and Adiponectin of ADSCs in sEV-LT group and sEV-AT group were significantly upregulated (p < 0.05); however, there was no statistical significance in the expression level of C/EBP-δ (p > 0.05). In addition, no significant difference in the amount of lipid droplets and adipogenesis-related genes between the sEV-LT groups and sEV-AT was seen (p > 0.05). At 2, 4, 8, and 12 weeks, the adipocyte area and the number of capillaries in neotissues in the sEV-LT groups and sEV-AT groups were significantly increased compared with the Matrigel group (p < 0.05); however, there was no dramatic difference between sEV-LT groups and sEV-AT groups (p > 0.05). At the 4th week, neotissues in the sEV-LT groups and sEV-AT groups all showed upregulated expression of C/EBP-α, PPARγ, Adiponectin, and CD31 protein, while neotissues in the Matrigel group only showed positive expression of CD31 protein. Conclusions This study demonstrated that sEV-LT exerted promotion effects on adipose tissue regeneration by accelerating the proliferation, migration, and adipogenic differentiation of ADSCs in vitro and recruiting adipocytes and promoting angiogenesis in vivo. The sEV-LT could serve as an alternative cell-free therapeutic strategy for generating adipose tissue, thus providing a promising application prospect in tissue engineering.

scholarly journals ROS/PI3K/Akt and Wnt/β-Catenin Signalings Activate HIF-1α-Induced Metabolic Reprogramming to Impart 5-Fluorouracil Resistance in Colorectal Cancer

2021 ◽  
Author(s):  
Shuohui Dong ◽  
Shuo Liang ◽  
Zhiqiang Cheng ◽  
Xiang Zhang ◽  
Li Luo ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Glucose Metabolism ◽  
Metabolic Reprogramming ◽  
Cell Models ◽  
Primary Tumors ◽  
Pharmacological Inhibition ◽  
Abnormal Glucose Metabolism ◽  
Potential Biomarker

Abstract Background: Acquired resistance of 5-fluorouracil (5-FU) remains a clinical challenge in colorectal cancer (CRC), and efforts to develop targeted agents to reduce resistance have not yielded success. Metabolic reprogramming is a key cancer hallmark and confers several tumor phenotypes including chemoresistance. Glucose metabolic reprogramming events of 5-FU resistance in CRC has not been evaluated, and whether abnormal glucose metabolism could impart 5-FU resistance in CRC is also poorly defined.Methods: We generated three acquired 5-FU resistance CRC cell line models, and the detailed assessment of glucose metabolism was performed by in vitro and in vivo experiments, including glucose and lactate utilization, the RNA and protein expressions of glucose metabolism‐related enzymes, the changes of intermediate metabolites of glucose metabolite pool and so on. We detected the protein levels of hypoxia inducible factor 1α (HIF-1α) in primary tumors and circulating tumor cells of CRC patients by immunohistochemistry and immunofluorescence. Stably HIF1A knockdown in cell models were established with a lentiviral system. The influence of both HIF1A gene knockdown and pharmacological inhibition on 5-FU resistance in CRC was detected in cell models in vivo and in vitro.Results: Here we describe the condition of abnormal glucose metabolism in 5-FU-resistant CRC in detail, and we demonstrate that the enhanced glycolysis and pentose phosphate pathway in CRC are associated with increased HIF-1α expression. We also show that HIF-1α-induced glucose metabolic reprogramming imparts 5-FU resistance in CRC. HIF-1α showed enhanced expression in 5-FU-resistant CRC cells and clinical specimens, and increased HIF-1α levels were associated with failure of fluorouracil analog-based chemotherapy in CRC patients and poor survival. Upregulation of HIF-1α in 5-FU-resistant CRC occurs through non-oxygen-dependent mechanisms of reactive oxygen species-mediated activation of PI3K/Akt signaling, and aberrant activation of β-catenin in the nucleus. Both HIF-1α gene knock-down and pharmacological inhibition restored the sensitivity of CRC to 5-FU, indicating the potential efficacy of strategies targeting HIF-1α as an upstream glycolytic pathway regulator. Conclusions: Our results indicate HIF-1α is a potential biomarker for 5-FU-resistant CRC, and targeting HIF-1a in combination with 5-FU may represent an effective therapeutic strategy in 5-FU-resistant CRC.

scholarly journals Stevioside fromStevia rebaudianaBertoni Increases Insulin Sensitivity in 3T3-L1 Adipocytes

2013 ◽  
Vol 2013 ◽  
pp. 1-8 ◽  
Author(s):  
Nabilatul Hani Mohd-Radzman ◽  
Wan Iryani Wan Ismail ◽  
Siti Safura Jaapar ◽  
Zainah Adam ◽  
Aishah Adam
Keyword(s):  
Insulin Sensitivity ◽  
Cell Viability ◽  
Glucose Uptake ◽  
Insulin Signalling ◽  
Stevia Rebaudiana ◽  
Control Group ◽  
Insulin Signalling Pathway ◽  
Oil Red O Staining ◽  
Oil Red O

Stevioside fromStevia rebaudianahas been reported to exert antihyperglycemic effects in both rat and human subjects. There have been few studies on these effectsin vitro. In this paper, radioactive glucose uptake assay was implemented in order to assess improvements in insulin sensitivity in 3T3-L1 cells by elevation of glucose uptake following treatment with stevioside. Oil Red-O staining and MTT assay were utilized to confirm adipocyte differentiation and cell viability, respectively. Findings from this research showed a significant increase in absorbance values in mature adipocytes following Oil Red-O staining, confirming the differentiation process. Stevioside was noncytotoxic to 3T3-L1 cells as cell viability was reduced by a maximum of 17%, making it impossible to determine its IC50. Stevioside increased glucose uptake activities by 2.1 times (p<0.001) in normal conditions and up to 4.4 times (p<0.001) in insulin-resistant states. At times, this increase was higher than that seen in positive control group treated with rosiglitazone maleate, an antidiabetic agent. Expressions of pY20 and p-IRS1 which were measured via Western blot were improved by stevioside treatment. In conclusion, stevioside has direct effects on 3T3-L1 insulin sensitivity via increase in glucose uptake and enhanced expression of proteins involved in insulin-signalling pathway.

scholarly journals Protein kinase N2 regulates AMP kinase signaling and insulin responsiveness of glucose metabolism in skeletal muscle

2017 ◽  
Vol 313 (4) ◽  
pp. E483-E491 ◽  
Author(s):  
Maxwell A. Ruby ◽  
Isabelle Riedl ◽  
Julie Massart ◽  
Marcus Åhlin ◽  
Juleen R. Zierath
Keyword(s):  
Skeletal Muscle ◽  
Protein Kinase ◽  
Glucose Metabolism ◽  
Glucose Uptake ◽  
Muscle Metabolism ◽  
Whole Body ◽  
Skeletal Muscle Metabolism ◽  
Ampk Signaling

Insulin resistance is central to the development of type 2 diabetes and related metabolic disorders. Because skeletal muscle is responsible for the majority of whole body insulin-stimulated glucose uptake, regulation of glucose metabolism in this tissue is of particular importance. Although Rho GTPases and many of their affecters influence skeletal muscle metabolism, there is a paucity of information on the protein kinase N (PKN) family of serine/threonine protein kinases. We investigated the impact of PKN2 on insulin signaling and glucose metabolism in primary human skeletal muscle cells in vitro and mouse tibialis anterior muscle in vivo. PKN2 knockdown in vitro decreased insulin-stimulated glucose uptake, incorporation into glycogen, and oxidation. PKN2 siRNA increased 5′-adenosine monophosphate-activated protein kinase (AMPK) signaling while stimulating fatty acid oxidation and incorporation into triglycerides and decreasing protein synthesis. At the transcriptional level, PKN2 knockdown increased expression of PGC-1α and SREBP-1c and their target genes. In mature skeletal muscle, in vivo PKN2 knockdown decreased glucose uptake and increased AMPK phosphorylation. Thus, PKN2 alters key signaling pathways and transcriptional networks to regulate glucose and lipid metabolism. Identification of PKN2 as a novel regulator of insulin and AMPK signaling may provide an avenue for manipulation of skeletal muscle metabolism.

Abstract 637: Allograft Inflammatory Factor 1 Promotes Key Macrophage Functions that Limit Plaque Destabilization

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Prameladevi Chinnasamy ◽  
Isabel Casimiro ◽  
Lander E Gorroño ◽  
Nicholas Sibinga
Keyword(s):  
Aortic Root ◽  
Lesion Size ◽  
Inflammatory Factor ◽  
Plaque Stability ◽  
Fibrous Cap ◽  
En Face ◽  
Oil Red O Staining ◽  
Oil Red O

Objective: Allograft inflammatory factor-1 (Aif-1) is best characterized as a cytoplasmic protein involved in actin bundling and phagocytosis. We investigated Aif-1 function in macrophage biology and the pathogenesis of atherosclerosis. Hypothesis: Aif-1-mediated potentiation of NFκB signaling promotes key macrophage functions and supports characteristics of plaque stability. Methods: We generated apoE -/- and aif-1 -/- ; apoE -/- mice (SKO and DKO, respectively), fed them a high fat diet for 18 weeks, and analyzed lesion burden in en face aorta, aortic root, and brachiocephalic arteries (BCA). We used immunofluorescence to assess marker expression and FACS analysis for proliferation and efferocytosis. Results: Lesion burden was not significantly different in aortic roots (DKO vs. SKO; 1.385e+006 ± 2065 vs 1.957e+006 ± 558, total lesion area), BCAs (DKO vs. SKO; 63 vs.65, % of Oil red O staining) and en face aortas (9 vs. 10, % of Oil red O staining). On the other hand, aortic root lesions from DKO mice showed more necrosis (DKO vs. SKO; 0.65 vs 0.4) and decreased fibrous cap thickness (DKO vs. SKO; 0.3 vs. 0.45, p0.05 respectively) normalized to lesion size. We also observed less macrophage staining (DKO vs. SKO; 0.6 vs. 2.2 % area/aortic root) and increased apoptosis (14 vs. 6 %Tunel+ cells), which correlated with decreased lesional P-NFκB p65 (DKO vs. SKO; 4 vs. 7 +nuclei/aortic root) activation. In vitro , aif-1 -/- macrophages showed decreases in oxidative stress-induced survival (40 vs. 80%), phagocytosis (30-60% decrease in zymosan particle uptake) and efferocytosis (40 vs.60 %F4/80;Tamra double +ve cells), and interleukin-6 release (1.2 vs 3 ng/ml in conditioned media). Mechanistically, these increased key macrophage functions correlated with P-NFκB p65 activation, which was impaired in aif-1 -/- macrophages. Conclusion: Our in vivo and in vitro data point to an essential role of Aif-1, via NFκB activation, in support of key macrophage functions, including survival and efferocytosis, that limit necrotic core size and increase atherosclerotic plaque stability.

Sign in / Sign up

Export Citation Format

Share Document