| Encephalitozoonosis is an obligate intracellular,eukaryotic microsporidiosis and a world widely epidemic zoonosis.Brain is one of the target organs attacked by Encephalitozoon cunicul(E.cuniculi)i.Main pathological lesions caused by E.cuniculi are non-suppurative encephalitis and cerebral granuloma.Up to now,relationship between cerebral granuloma by E.cuniculi and ERK/PLCγ signal transduction pathway remain unclear.In order to research the formation and characteristics of cerebral granuloma by E.cuniculi,and effect of granulomatous formation through signal transduction pathway,this experiment was performed.30 brains of sick rabbits and 10 brains of control rabbits were used in this study.The brains were treated by frozen section and paraffin section.The sections were stained by HE,special staining,immunohistochemical staining and TUNEL.The occurrence,formation and classify were observed by microscopy.The expression of GFAP is tested by immunohistochemistry or Western-blot.To research critical proteins and enzymes within ERK/PLCγ,such as NGF,MAPK,ERK,PLCγ and PKC,the paraffin sections and cerebral proteins were tested by immunohistochemistry or Western-blot.The changes of ERK/PLCγ signal transduction pathway were researched while cerebral granuloma was formed by E.cuniculi.The results showed that E.cuniculi could intrude into vascular epithelial cell and cause the injury of blood vessel,could be devoure by mononuclear cell and enter into the brain later on.There were more microglial cells invasion and lots of astrocytes proliferation surrounding parasite colony and infiltrated macrophages in focal despaired brain tissues.The E.cuniculi granuloma was formed in this way by continuous development.The granuloma had three types that werecellular,proliferative and necrotic granuloma.Lots of blue E.cuniculi in epithelioid cell and necrotic tissues of granulomas were checked out by modified gram stain.A lot of red E.cuniculi in cytoplasm of epithelioid cells were detected by methyl green pyronine stain.The main cells forming cerebral granuloma were epithelioid cell,microglial cell,astrocytes,vascular epithelium and lymphocyte.In those cells the astrocytes were the most important cells.Plenty of reactrive-astrocyte proliferations showed around the periphery granuloma and vascular cuffing by anti-GFAP staining.The foot processes of proliferative astrocytes formed thicker barrier membranes around blood vessels,under cerebral pia meter and in basilar part of ependymocytes.GFAP abundance in sick rabbit’s brain tissues was higher than control rabbits by western-blot and was in line with the results of anti-GFAP immunohistochemical staining.It was proved that the activity of NGF,MAPK4,ERK,PLCγ and PKC were increased obviously in astrocytes,vascular epithelium and fibroblast fibrocyte by immunohistochemical staining.The above several signal pathway protein abundance of sick rabbit’s brain tissue was much higher than control rabbit’s.It is proved that the ERK/PLCγ signal pathway was activated in process of forming cerebral granuolma by E.cuniculi.Activated-ERK could fornated the dipolymer in cytoplasm via the strong cascade reaction,then entered into the nucleus,actived the transcription factor,induced the expression of c-Fos,c-Jun,c-Myc and Egr1,lead to the translation of tubulin,such as GFAP,and promoted the proliferation of cells.Activated-PLCγ could hydrolysed IP2,produced IP3 and DAG,in turn actived PKC,phosphorylated transcription factors,such as Elk-1,NF-κB,AP-1,Jun,Fos and so on,and facilitated the genetic expression.In conclusion,E.cuniculi could entered into brain tissue by destroyed blood brain barrier or followed with infiltration of mononuclear cells.Three kinds of granulomatousmorphological changes were actually different development stage in a pathological process.All kinds of barrier membrane formed by reactive astrocyte were able to protect brain tissue against more injury.ERK/PLCγ signal pathway activited by the brain injury of E.cuniculi could induced the proliferation of astrocytes,vascular epithelium and fibroblast fibrocyte.These proliferous new cells and transudatory inflammatory cells gathered in the cerebral tissues and leaded to the formation of granuloma. |
