【摘要】：目的 對(duì)比Brn-3a與Thy-1.1標(biāo)記大鼠視網(wǎng)膜神經(jīng)節(jié)細(xì)胞(RGCs)的特點(diǎn)及計(jì)數(shù)差異，評(píng)估Brn-3a是否可作為一種可靠的內(nèi)源性標(biāo)記物，標(biāo)記并計(jì)數(shù)大鼠RGCs。 方法 20只SD大鼠，隨機(jī)選取大鼠一眼為T(mén)hy-1.1組，另一眼作為Brn-3a組,過(guò)量麻醉處死，行3點(diǎn)鐘位縫線標(biāo)記,立即摘除眼球，浸泡于4%多聚甲醛的固定液中。48h后沿角鞏膜緣切除眼前節(jié)，剝離晶狀體，余下部分逐節(jié)脫水、透明、石蠟包埋，，于視乳頭顳側(cè)旁開(kāi)1mm，做4μm切片，烘干備用，做視網(wǎng)膜石蠟切片，常規(guī)蘇木素-伊紅染色鏡下觀察RGCs的形態(tài)，運(yùn)用免疫組織化學(xué)法檢測(cè)Brn-3a與Thy-1.1的表達(dá)情況，每只眼球取1張切片，在每張待測(cè)視網(wǎng)膜切片上，以視網(wǎng)膜后極部中點(diǎn)為起點(diǎn)，至近角膜緣視網(wǎng)膜止點(diǎn)處，將視網(wǎng)膜分為三等份，即中央?yún)^(qū)、中間區(qū)及周邊區(qū)，每個(gè)區(qū)域視網(wǎng)膜隨機(jī)取5個(gè)高倍視野，分別計(jì)數(shù)Thy-1.1、Brn-3a陽(yáng)性細(xì)胞均數(shù)。 結(jié)果 光鏡下正常大鼠的視網(wǎng)膜從內(nèi)向外依次為神經(jīng)節(jié)細(xì)胞層(GCL)、內(nèi)核層(INL)、外核層(ONL)。GCL的細(xì)胞呈單層排列，較為整齊，胞核清楚數(shù)目相對(duì)較少，INL和ONL呈多層排列，且細(xì)胞排列緊密，數(shù)目相對(duì)較多。兩種標(biāo)記物染色都僅僅存在于視網(wǎng)膜的GCL，但在大鼠RGCs中分布呈明顯差異：Brn-3a主要在胞核中著色，Thy-1.1主要在胞漿中著色；運(yùn)用此兩種標(biāo)記物標(biāo)記計(jì)數(shù)RGCs,中央?yún)^(qū)Thy-1.1組(n=20)和Brn-3a組(n=20)分別是29.75±2.02（個(gè)/高倍視野）和29.46±1.25（個(gè)/高倍視野），中間區(qū)Thy-1.1組(n=20)和Brn-3a組(n=20)分別是24.72±1.10（個(gè)/高倍視野）和24.43±1.66（個(gè)/高倍視野），周邊區(qū)Thy-1.1組(n=20)和Brn-3a組(n=20)分別是21.79±1.25（個(gè)/高倍視野）和21.84±1.37（個(gè)/高倍視野）,各個(gè)相同區(qū)域內(nèi)Thy-1.1和Brn-3a陽(yáng)性細(xì)胞計(jì)數(shù)比較，差異均無(wú)統(tǒng)計(jì)學(xué)意義（P0.05)。Thy-1.1組中央?yún)^(qū)、中間區(qū)和周邊區(qū)兩兩比較，差異均有統(tǒng)計(jì)學(xué)意義（P 0.05），Brn-3a組中央?yún)^(qū)、中間區(qū)和周邊區(qū)兩兩比較，差異均有統(tǒng)計(jì)學(xué)意義（P 0.05）。 結(jié)論 在大鼠視網(wǎng)膜中Brn-3a陽(yáng)性細(xì)胞僅存在于視網(wǎng)膜的GCL，主要在胞核中著色。與Thy-1.1標(biāo)記大鼠RGCs比較, Brn-3a標(biāo)記的數(shù)量基本相當(dāng)。Brn-3a可以作為一種比較可靠的內(nèi)源性標(biāo)記物，標(biāo)記并計(jì)數(shù)大鼠RGCs。
[Abstract]:Objective to compare the characteristics and counts of (RGCs) labeled by Brn-3a and Thy-1.1 in rat retinal ganglion cells, and to evaluate whether Brn-3a can be used as a reliable endogenous marker for labeling and counting rat RGCs.. Methods Twenty SD rats were randomly selected as Thy-1.1 group and Brn-3a group respectively. The rats were killed under excessive anesthesia and were labeled with suture at 3 o'clock, and the eyeball was removed immediately. 48 hours later, the anterior segment of the eye was removed along the border of the cornea and sclera, and the lens was stripped. The rest was dehydrated, transparent, and paraffin-embedded, and opened at the side of the temporal side of the optic papilla for 1 mm, then sliced 4 渭 m and dried for reserve. The morphology of RGCs was observed under routine hematoxylin-eosin staining. The expression of Brn-3a and Thy-1.1 was detected by immunohistochemical method. One slice was taken from each eye and on each retinal section to be tested. Taking the middle point of the posterior pole of retina as the starting point, to the end point of the retina near the limbus of cornea, the retina is divided into three parts, namely, the central area, the intermediate area and the peripheral area. The retina of each region is randomly taken out of 5 high magnification visual fields, and the Thy-1.1, is counted respectively. Mean number of Brn-3a positive cells. Results under light microscope, the retina of normal rats was composed of ganglion cell layer (GCL), inner nuclear layer (INL), outer nuclear layer (ONL) in turn from inside to outside. The cells of GCL were arranged in a monolayer, and the number of clear nuclei was relatively small. INL and ONL were arranged in multiple layers, and the cells were arranged closely, and the number of cells was relatively large. The staining of the two markers only existed in the retina of GCL, but the distribution of the two markers in the rat RGCs was significantly different: Brn-3a was mainly stained in the nucleus and Thy-1.1 was mainly in the cytoplasm; These two markers were used to count the Thy- 1.1 group (n = 20) and the Brn-3a group (n = 20) in the central area of RGCs, which were 29.75 鹵2.02 (/ HVA) and 29.46 鹵1.25 (HPF), respectively. In the middle region, Thy- 1.1 (n = 20) and Brn-3a (n = 20) were 24.72 鹵1.10 (/ high visual field) and 24.43 鹵1.66 (/ high power visual field), respectively. In peripheral area, Thy- 1.1 group (n = 20) and Brn-3a group (n = 20) were 21.79 鹵1.25 and 21.84 鹵1.37, respectively. The number of Thy-1.1 and Brn-3a positive cells in the same area was significantly higher than that in control group (P < 0.05), and the number of positive cells in the same area was significantly higher than that in control group (P < 0.05). There was no statistically significant difference (P0.05) in the central, middle and peripheral regions of the Brn-3a group (P0.05), and there was a significant difference in the central, intermediate and peripheral regions of the THEY group (P0.05), and there was no significant difference in the central area, middle area and peripheral area between the two groups (P0.05). The difference was statistically significant (P 0.05). Conclusion in rat retina, Brn-3a-positive cells are mainly stained in the nucleus of the retina, but only in the retina. Compared with Thy-1.1 labeled rat RGCs, the number of Brn-3a labeled rats was about the same. Brn-3a could be used as a reliable endogenous marker to mark and count rat RGCs..
【學(xué)位授予單位】：福建醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R774.1

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