| Summary: | 碩士 === 國立臺灣大學 === 微生物與生化學研究所 === 97 === The goal of this study was to investigate whether dietary factors prescreened by NF-kappaB transactivation assay could alleviate the development of insulin resistance and inflammatory state. We employed a NF-kappaB promoted luciferase reporter gene plasmid to transfect RAW264.7 macrophage cell lines for pre-screening of anti-inflammatory activity. Proinflammatory cytokine productions from LPS-stimulated primary peritoneal macrophages and ConA-stimulated primary splenocytes isolated from C57BL/6 mice were also measured. The results showed that drugs, including sodium salicylate and rosiglitazone, and dietary factors, including bitter gourd, GMI, and f-GMI, inhibited NF-kappaB-dependent luciferase activity, TNF-alpha secretion from peritoneal macrophages, and IFN-gamma secretion from splenocytes.
To further investigate the hypoglycemic effect in vivo, in StageⅠ, five groups of 12-week-old C57BL/6 mice were fed with AIN-76 diets supplemented with tested samples, including BGP (bitter gourd powder) and different doses of GMI (G1x, G4x, and G20x). After 5 weeks of feeding, G4x group had significantly lower serum glucose level compared to the control, and after 6 weeks of feeding, G1x group also had lower fasting glucose. In StageⅡ, a 30% high fat diet was applied to increase the body weight of mice. After 8 weeks of high-fat diet feeding, no tested sample had hypoglycemic effects on mice, but BGP group had significantly lower AUC of OGTT and HOMA-IR index, whereas G40x group elevated HOMA-IR index. After 4-week wash out period, we switched the tested samples to f-GMI, and continued with StageⅢ of the experiment. After 4 weeks of f-GMI feeding, no hypoglycemic effect was observed. However, the serum glucose levels of BGP group significantly lowered after 2 weeks into StageⅢ, and the AUC of OGTT and HOMA-IR index were also lowered after 3 weeks of feeding.
To further investigate the hypoglycemic effect of f-GMI, 6-week-old C57BL/6J mice were divided randomly into two groups: the Basal group was fed AIN-76 diet, whereas the HF (high fat) group was fed with a 30% butter diet. After 9 weeks of induction, the HF group was subdivided into the following 7 groups: containing HF, salicylate (0.7% sodium salicylate in diet), Rosi (0.005% rosiglitazone in diet), BGP (5% BGP in diet), G6x (0.4 mg/mouse/day), G12x (0.8 mg/mouse/day), and G18x (1.2 mg/mouse/day). After 5 weeks of feeding, BGP, salicylate, and G12x group were shown to be effective in ameliorating the HF diet-induced hyperglycemia, whereas G18x group led to an increase in serum glucose. BGP and Rosi group also decreased hyperleptinemia, AUC of OGTT, and HOMA-IR index. In the adipose tissue of high fat-fed mice, Rosi, salicylate, G6x, and G12x group decreased MCP-1 and F4/80 mRNA expression, resulting in reduced macrophage infiltration in adipose tissue. Furthermore, Rosi, BGP, salicylate, G6x, and G18x group increased the mRNA levels of M2 macrophage markers, such as IL-10 and arginase, and tended to induce M2 macrophage polarization. In addition, Rosi, BGP, and G18x group had higher TNF-alpha mRNA levels, while Basal and BGP group also upregulated IL-1beta mRNA expression, which suggest that these samples might have proinflammatory effect in the adipose tissue of diet-induced obese mice. Moreover, salicylate, G6x, G12x, and G18x significantly reduced CRP levels in liver. BGP group decreased IL-6 content in liver, but raised MCP-1 levels in liver and serum. BGP also increased CRP concentration in serum. The results showed that bitter gourd might have proinflammatory effect in high-fat fed C57BL/6J mice.
Administering the tested samples prescreened by NF-kappaB transactivation assay to high-fat fed C57BL/6J mice showed lower fasting glucose and anti-inflammatory effect through not only reduced F4/80 and MCP-1 mRNA levels, but also increased the levels of M2 macrophage markers. Moreover, BGP also had anti-inflammatory activity by reducing IL-6 levels in liver, leptin and NEFA levels in serum. In conclusion, dietary factors prescreened by NF-kappaB transactivation assay may alleviate the development of insulin resistance through reducing inflammatory state.
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