高糖对人晶状体上皮细胞iNOS表达机制及丹酚酸B作用的研究.docx

高糖对人晶状体上皮细胞iNOS表达机制及丹酚酸B作用的研究
摘要：
本研究旨在探讨高糖条件下人晶状体上皮细胞iNOS表达机制及丹酚酸B的作用。通过使用人晶状体上皮细胞系进行实验，使用不同浓度的葡萄糖处理晶状体上皮细胞，检测iNOSmRNA和蛋白质表达水平，同时通过细胞计数和MTT测定细胞增殖情况。结果表明，高浓度葡萄糖处理可显著增加人晶状体上皮细胞iNOSmRNA和蛋白质表达水平，抑制细胞增殖。丹酚酸B预处理可逆转高糖诱导的iNOSmRNA和蛋白质高表达，并且显著促进细胞增殖。进一步的研究表明，丹酚酸B可通过抑制NF-κB和MAPKs通路的活化及其下游的相关分子表达来逆转高糖诱导的iNOS表达。
本研究结果表明，高糖条件下的iNOS表达主要通过NF-κB和MAPKs通路的活化调节，并且丹酚酸B可以拮抗高糖对iNOS表达的影响，其机制可能涉及到NF-κB和MAPKs通路的抑制。这一研究结果有望为晶状体疾病的发病机制及治疗提供新的思路和方法。
关键词：高糖，iNOS，丹酚酸B，NF-κB，MAPKs
Introduction
Thelensepithelialcells(LECs)existasasinglecelllayersituatedattheperipheryofthelensandplayanimportantroleinmaintainingocularhomeostasis.However,ocularcomplications,suchasdiabeticcataracts,canarisewhenthesecellsareexposedtochronichyperglycemia,leadingtoexcessiveoxidativestressandinflammation.Nitricoxide(NO)isakeymediatorofinflammation,andelevatedlevelsofinduciblenitricoxidesynthase(iNOS),whichproducesNOfromarginine,havebeenreportedindiabeticcataracts.Inaddition,previousstudieshaveshownthatiNOSoverexpressioninLECscancauseoxidationofproteinsandlipids,whichmaycontributetocataractformation.Therefore,itisimportanttoinvestigatetheunderlyingmechanismofiNOSexpressioninLECsunderhighglucoseconditions.
Danshensu(DSS)isanaturallyoccurringphenolicacidderivativewidelyfoundintherootsofSalviamiltiorrhiza,whichhasawiderangeofpharmacologicaleffects,includinganti-inflammatoryandanti-oxidativeproperties.StudieshaveshownthatDSScaninhibitinflammationandoxidativestressinvariouscelltypes.However,whetherDSScaninhibitiNOSexpressioninLECsunderhighglucoseconditionsremainsunclear.
Materialsandmethods
Cellculture:Ahumanlensepithelialcelllinewasculturedinhighglucosemedium(25mM)ornormalglucosemedium(5mM).
Cellproliferationassay:CellproliferationwasassessedusinganMTTassay.
Westernblotanalysis:TotalproteinwasextractedfromthecellsandsubjectedtoWesternblotanalysistodetermineiNOSproteinexpression.
Real-timePCR:TotalRNAwasextractedfromthecellsandsubjectedtoreal-timePCRtodetermineiNOSmRNAexpression.
NF-κBandMAPKspathwayassay:TheexpressionofNF-κBan