| Diabetes is an endocrine disease.It has been confirmed by the long-term research that the genetic and environmental factors are related to the disease which seriously affects human health.In synthetic drug clinical treatment of diabetes,there are many problems such as adverse reactions.In comparison,natural medicine has the advantages of high safety,convenient use and long service time.Sulforaphane(SFN)is a natural monomer extracted from plants.Studies have shown that it has a good effect in relieving the symptoms of insulin resistance(IR)in patients with type 2 diabetes,but its mechanism remains to be explored.Our experiments demonstrated that sulforaphane can improve the insulin resistance in IR-Hep G2 cells,and the mechanism may be related to the inhibition of the expression of protein,Lipin1.Lipin1 mainly distributed in the liver,and the improving of the drug's targeting to the liver can also play a better role.In addition,in the experiment,we found that sulforaphane in high doses has certain toxicity,and the limiting of the use of the drugs thus affecting the pharmacological effects.As a drug carrier,liposomes have some special advantages such as targeting,reducing toxicity and side effects of drugs.Therefore,this study combines the sulforaphane and liposomes treated as a new dosage to develop a sulforaphane liposomes,for the purpose of improving drug's targeting to the liver and solving the dosage limits of drugs.In this way,the efficiency can be enhanced and toxicity can be reduced,which will be beneficial to the clinical treatment in the future.This topic is mainly divided into two parts:1.The study on the effect of sulforaphane on improving insulin resistance of Hep G2 and the study of its mechanismThe insulin resistance model(IR-Hep G2)of Hep G2 cells was established by palmitic acid(PA).By detecting the levels of glucose consumption(GC),the changes of intracellular diacylglycerol(DAG)content as well as the expression of cytochrome P4503A4(CYP3A4)and Lipin1 protein in the supernatant of medium before and after using drugs,we tried to reveal the mechanism that associated with the potential hypoglycemic effect in the sulforaphane on IR-Hep G2 cells.By analyzing the glucose consumption,we found that the glucose consumption levels in the model group were significantly lower than that of the control group,indicating that the IR-Hep G2 model was successfully established.Compared with the model group,sulforaphane with the doses of low,medium and high could increase the consumption of glucose in supernatant,especially the consumption of high group.The results of the diacylglycerol test kit showed that low,medium and high doses of sulforaphane could reduce the diacylglycerol content in IR-Hep G2 cells when compared with the model group.The western blot experiment showed that sulforaphane could decrease the expression of Lipin1 and CYP3A4.Suggesting that sulforaphane can improve the insulin resistance in IR-Hep G2 cells,and its mechanism may be related to the inhibition of the expression of Lipin1 which resulting in reduced diacylglycerol content.2.Preparation and quality evaluation of sulforaphane liposomesThrough the orthogonal design,the sulforaphane liposomes were prepared by reverse evaporation method and their quality was evaluated.Using the orthogonal experiment method,the most suitable preparation formula for the preparation of sulforaphane liposomes was selected by using the encapsulation efficiency as the evaluation index.The morphological,particle size,electronegativity,stability and other properties of liposomes were also investigated.The suitable method of tissue sample was selected,and the method of in vivo analysis of sulforaphane was established.Mice were divided into two groups according to the experimental design: one for sulforaphane and the other for sulforaphane liposomes.After administration,the tissues was analyzed and the drug was analyzed in vivo.After treatment,the drug was analyzed in vivo,and then the tissue distribution of sulforaphane liposomes was studied preliminarily.The optimum extraction conditions were as this: the ratio of organic phase and aqueous phase was 1:3,the temperature was 30?,the hydration time was 5min,and the phospholipid cholesterol molar ratio was 6: 1,according to this prescription,the encapsulated rate of liposomes was up to 51.11%.The obtained sulforaphane liposomes were spherical or near spherical monolayer liposomes under the result coming from the transmission electron microscopy.The particles were uniform in size and good in flow ability.The average particle size was(176.733 ± 5.443)nm and the PDI was(0.197 ± 0.032),which indicated that the distribution of liposomes was not wide and the size was uniform.The average zeta potential was(21.13±0.17)mv,indicating that the liposomes had some stability.The results showed that at a dose of 4?for 10 days,a small amount of liposomes appeared to precipitate.After one month,the liposomes partially precipitated,and the encapsulation efficiency also showed a decreasing trend,and the overall liposome system was stable.The HPLC method showed that the distribution of sulforaphane liposomes in liver and spleen tissue in mice was more than that of the free drug groups.The results of cell proliferation test showed that the toxicity of sulforaphane liposomes in Hep G2 cells was lower than sulforaphane of the same dosage,indicating that it can enhance the efficiency as well as reduce the toxicity of sulforaphane. |
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