Experimental Study Of The Endogenous NPCs Proliferation,Migration, Differentiation In The Brain Of Amyotrophic Lateral Sclerosis's Disease-Like Transgenic (G93A SOD1) Mice

Objective:To observe the proliferation,migration and differentiation of neural precursors(NPCs)in brain-related anatomical regions of normal adult wild-type mice and SOD1-G93 A transgenic mice with amyotrophic lateral sclerosis(ALS)during different stages,which may provide some experimental evidences for the further regulation of proliferation,migration,differentiation of NPCs and a potential way for the treatment of ALS.Methods:We construct the animal models of G93A-SOD1 C57BL/6J transgenic mice which expression were identified by the PCR.The G93A-SOD1 transgenic mice were divided into three different groups: pre-onset stage(60-70d),onset stage(90-100d)and progression stage(120-130d).To mark the proliferated newborn cells,we injected the wild-type and G93A-SOD1 C57BL/6J transgenic mice with intraperitoneal injection of Brdu(5-bromodeoxyuridine).Immunohistochemical and double immunofluorescence techniques were employed in this experiment.The anti-nestin and vimentin,NeuN,GFAP,Olig antibodies were used to mark NPCs,neurons,astrocytes,oligodendrocytes respectively.We observed positive cells under the same microscope view of magnification and field,overplaying different stained positive cells by image processing technique to observe whether there was a co-immunoreaction.Then,the number of positive cells in different anatomical regions and different stages of disease was calculated.Finally,we get to make a statistical analysis of the characteristics of proliferation,migration and differentiation of NPCs in brain-related anatomical area in wild-type mice and G93A-SOD1 transgenic mice.Results:1?the vimentin-containing cells(VCCs)were mainly distributed in the ependymal zone,subventricular zone,hippocampus,cerebral cortex,Olfactory bulb of adult wild-type mice brain.2?The VCCs appeared in many nucleis in ventral brainstem of G93A-SOD1 ALS transgenic mice.The number of positive cells increased progressively with the disease progression.The rank order of VCCs amount from groups was progression> onset>pre-onset.3?The increased VCCs in brainstem were almost co-immunologically reactive with GFAP,suggesting that almost all VCCs differentiate into astrocytes.4?The amount of VCCs decreased in the olfactory cortex,cingulate gyrus and motor cortex of G93A-SOD1 ALS transgenic mice compared with the normal control.5?The increased VCCs were few BrdU-positive,suggesting that increased VCCs may have two sources: 1)a few VCCs which co-immunoreactive with BrdU were proliferated newborn cells;2)most positive cells which not co-immunoreactive with BrdU may be late proliferation and differentiation NPCs,this part of VCCs may also be migrate and differentiate from other regions.6.In the pons and medulla oblongata of G93A-SOD1 ALS transgenic mice,all Nestin positive cells were VCCs,but co-immunoreaction between vimentin and nestin was only occupied a small part of them,which indicated that the major VCCs were the late differentiation NPCs/NSCs Conclusions:The NPCs were extensively found in various cerebral regions of normal adult wild-type mice.The mass proliferation of NPCs in various ventral nucleus of the brainstem is involved in the onset of ALS,the small part were early proliferation NPCs,Other were late proliferation and differentiation NPCs,and most of them differentiate into astrocytes.