| Objective:Spontaneous intraventricular hemorrhage (SIVH) is a common clinicalemergency that the blood enters the ventricular system with the rupture ofintracranial vessels caused by non-traumatic factors. Its' incidence is from20%to60%of spontaneous intracerebral hemorrhage and Its' mortality isfrom14%to83.3%. Its' clinical features depend on the extent ofintraventricular hemorrhage. Hydrocephalus and cerebral vasospasm are themost familiar complications of intraventricular hemorrhage. At present,researches of intraventricular hemorrhage concentrate on the etiology ofpost-hemorrhagic hydrocephalus and there are few studies on the nervoustissue harm and the process of damage is unclear. For this reason, weestablished a rat model of IVH, observed the changes of subventricular zone atdifferent times after IVH specially and we tried to explore the impact andmechanism of changeable expression of Nestin and CD133during the processof nervous tissue damage.Methods:1.55Sprague Dawley male rats(300g±10g) were divided into3groupsrandomly:the IVH group(n=25),the saline control group(n=25),and thenomal control group(n=5).2. The rat mode of IVH consulted the reports of K.R.Lodhia, etc and madesome improvements.The IVH group was made by infusing autologous wholetail-artery blood into the lateral ventricle(160μl).The saline control group wasmade by injecting160ul saline to the lateral ventricle. The normal controlgroup was made by the same treatment except for the ventricular injection. Allthe rats were fed in the same environment.3. The rats of experimental groups and control groups were examined according to the neurological severity score standard and weighted, then therats were sacrificed at five time points of1,2,3,7,14days. After fixed in4％paraform for24h, The brain tissues were coronally sliced from the point0.40mm posterior to the bregma for observing the hematoma in lateralventricle and measuring the volume of lateral ventricle (the width of lateralventricle at the coronal plane at least1.0mm is the standard ofhydrocephalus),Then the brain tissue samples were embedded in paraffin andmade into5μm thick serial sections. The paraffin sections were then stained byHE And specificity immunohistochemistry was applied to the brain tissuesamples for observation. Then, at high magnification, we counted the numberof Nestin and CD133expression cells in each group at different time points.4. Statistical analysis: Use SPSS15.0statistical software to conductstatistical analysis of the experimental data. T test was used to comparemeasurement data between groups. P <0.05indicated that there werestatistically significant differences.Results:1．General description: the IVH group's weight loss after the operationat each time point were respectively5.75±1.50g,10.75±2.99g,21.25±6.29g,45.00±5.77g,71.25±2.50g, the rats of saline control group's weight lossduring the3days after operation were respectively4.50±0.58g,6.00±1.83g,8.63±1.11g, the following time points rats restored, the added weightwere respectively4.50±0.58g,6.00±1.83g,8.63±1.11g. The IVH group dr-opped more than ratsin the saline control group in the former2days and3days time points. there were significant differences between the IVH gr-oup and the saline control group(P＜0.05).The weight of normal controlgroup increased52.75±3.20g.The neurological severity score of the rats was operated at each time p-oint, the IVH group's scores were respectively11.8±0.8,15.2±0.8,16.6±0.5,16.0±0.7,15.4±0.5, the saline control group's scores were respectively13.0±1.0,17.6±0.5,18.0±0.7,18.6±0.5,18.8±0.4, there was noabnormality in thenormal control group. There were significant differrences in the neurologi- cal severity score between the IVH group andthe saline control group at1day time point and14days time point (P＜0.05).2. Dilatation of ventricle: the width of lateral ventricle of the points1,3,7,14days in the IVH group was:1.24±0.11mm,1.43±0.08mm,1.28±0.08mm,1.61±0.05mm; the data of the saline control group at each timepoint was1.09±0.09mm,1.04±0.05mm,1.10±0.03mm,1.06±0.05mm; therewere significant differences between the IVH group and the saline controlgroup (P＜0.05). The width of lateral ventricle of normal control groupwas0.94±0.04mm, there was no changes in every time point.3. Immunohistochemical staining results: the cell rates of positive exp-ression of nestin in subventricular zone at1,2,3,7,14days after intravent-ricular hemorrhage were respectively24.71±4.76%,15.27±4.04%,15.69±2.13%,15.99±8.45%,16.77±1.42%, at the same points with the IVHgroup, the cell rates of positive expression of Nestin in subventricular z-one of the saline control group were respectively7.33±2.66%,7.79±1.52%,8.09±2.18%,8.62±7.50%,8.56±6.05%; the cell rates of positive exp-ression of CD133in subventricular zone at1,2,3,7,14days after intravent-ricular hemorrhage were respectively11.49±3.78%,7.16±2.72%,8.53±4.30%,8.64±1.35%,12.34±2.67%; at the same points with the IVH group,the cell rates of positive expression of CD133in subventricular zoneof the saline control group were respectively4.11±1.13%,4.08±0.57%,3.97±3.55%,3.82±0.77%,3.40±1.08%; Using T test to compare the me-ans of Nestin-positive cell rates and CD133-positive cell rates of thetwo groups at the same time points, we observed that, there were signific-ant differences in cell rates between the IVH group and the saline cont-rol group(P＜0.05).4. Immunofluoresence staining of Nestin-positive cells: We usedimmunofluorescence technique to stain the paraffin sections. Under afluorescence microscope, we observed the expression of nestin, there weremore positive expression of nestin in subventricular zone, at each time point,there were more Nestin-positive cells in the IVH group and the saline control group. At the3days time point, there were significant differences of theexpression of nestin in bilateral subventricular zone of the same brain tissuesample, however, the other time points suggested the opposite.Conclusions:1. The rat IVH model in the experiment prepared by fractionated infusingunanticoagulation autologous whole tail-artery blood in the lateral cerebralventricle was stable and it can preferably imitate the natural process andclinical changes of intraventricular hemorrhage.2. The expression of Nestin in the subventricular zone was correlated with thetime variation, On1day time point, The expression of Nestin increased rapidly;on2days time point the expression of Nestin decreased slightly, on3daystime point, the expression increased more than the former time point; on7days time point, the expression increased further up; on14days time point, theexpression continued increasing, but the level is lower than the level of1daytime point. The expression of Nestin in the saline control group increasedslightly.3. The expression of CD133was correlated with the time variation too, on the1day time point, the expression reached peak; on the2days time point, theexpression decreased; on the3days time point, the expression increasedcomparing with the2days time point; the expression increased by small rangeon the7days time point; on the14days time point, the expression increasedto higher level than the7days time point. The expression of the control groupwas stable.4. The volume of the ventricle on the1day time point dilated obviously on theright side; on the3days time point, bilateral ventricles dilated; on the7daystime point, bilateral ventricles dilated further more; on the14days time point,bilateral ventricles dilated comparing with the former time points, but still onthe slightly dilated state.5. The infusing of blood into the ventricle can activated the potentialdifferentiated cells in the subventricular zone. |
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