P物质受体在大鼠纹状体边缘区内的表达

我们以前的工作观察到纹状体边缘区内有密集的P物质纤维及终末分布,本用原位杂交和免疫组织化学方法研究了大鼠纹状体边缘区内P物质受体（SPR）的表达及分布,原位杂交结果发现P物质mRNA阳性杂交信号在纹状体内的分布不均匀,尾壳核内只有少量中等大小的阳性胞体,苍白球内只有少量较大的阳性胸体,而在尾壳核和苍白球之间的边缘区部位则可见许多中等大小的梭形阳性神经元胞体,并呈现密集的带关分布。免疫组织化学结果观察到P物质阳性神经元胞体在纹状体内的分布与原位杂交结果一致。推测大鼠纹体边缘区内可以合成P物质受体,具有接受和整合P物质神经递质的功能,推测边缘区内SPR神经元可能对SP递质的接受、调节有重要作用。     

大鼠视网膜中HAP1的免疫组织化学定位及不同光照条件对HAP1表达的影响

目的探讨亨廷顿蛋白相关蛋白1(huntingtin associated protein 1, HAP1)是否存在于视网膜内及是否与视觉有关.方法对正常大鼠眼球壁用ABC法进行免疫组织化学染色,观察HAP1在视网膜中的定位;用半定量免疫印迹方法(Western blotting)检测不同光照条件对大鼠视网膜中HAP1表达的影响.结果 HAP1较广泛地分布在大鼠视网膜各层,但以内核层及外核层中免疫反应较强,阳性反应产物主要定位在节细胞层和内核层/外核层中部分细胞胞体内;其余各层中,HAP1免疫反应较弱,阳性产物呈弥散分布,未见明显的阳性胞体.在连续处于黑暗环境中72小时大鼠视网膜中,HAP1表达较常规光照动物明显减少,而连续光照72h大鼠视网膜内HAP1表达无明显变化.结论 HAP1在视网膜中的存在及不同光照条件对视网膜HAP1表达的影响表明,HAP1可能与视觉活动有关.     

大鼠颌下腺促性腺激素释放激素及其mRNA的免疫组织化学与原位杂交 …

用免疫组织化学及原位杂交法,研究了促性腺激素释放激素及其ｍＲＮＡ在大鼠颌下腺的分布。结果显示,大鼠颌下腺的浆液腺泡的上皮细胞,各级导和的上皮细胞及副交感神经节细胞均呈促性腺激素释和激素免疫反应阳性,阳生反应物质分布在胞质,胞核呈阴性反应。     

自分泌运动因子受体在大鼠胃肠的定位

目的 研究自分泌运动因子受体在大鼠胃肠的表达及分布特点。方法应用免疫组织化学方法染色。结果在大鼠胃壁内有自分泌运动因子受体的表达,免疫反应产物主要定位于胃底腺,阳性细胞胞体大,胞核呈阴性,小肠和大肠组织均呈阴性反应。结论正常大鼠胃底腺有自分泌运动因子受体的表达,提示可能参与胃腺的多种生理活动。     

大鼠颌下腺及其离体培养细胞脱氢表雄酮（DHEA）的免疫组织化学定位

用免疫组织化学ABC法,研究了颌下腺及无血清培养的颌下腺上皮细胞DHEA的定位,结果显示,大鼠颌下腺的浆液性腺泡的上皮细胞及各级导管上皮细胞均呈DHEA免疫反应阳性,无血清培养腺上皮细胞也呈DHEA免疫反应阳性,阳性物质分布于胞质,胞核呈阴性反应,此结果提示：大鼠颌下腺能自身合成DHEA,DHEA对消化功能可能具有重要的调节作用。     

大鼠颌下腺促性腺激素释放激素及其mRNA的免疫组织化学与原位杂交研究

用免疫组织化学及原位杂交法,研究了促性腺激素释放激素及其ｍＲＮＡ在大鼠颌下腺的分布。结果显示,大鼠颌下腺的浆液性腺泡的上皮细胞,各级导管的上皮细胞及副交感神经节细胞均呈促性腺激素释放激素免疫反应阳性,阳性反应物质分布在胞质,胞核呈阴性反应。颌下腺的浆液性腺泡上皮细胞,各级导管上皮细胞同样被检测到很强的促性腺激素释放激素ｍＲＮＡ杂交信号。以上结果提示,大鼠颌下腺能自身合成促性腺激素释放激素,促性腺激素释放激素对消化功能可能有重要调节作用。     

Aquaporin-1在大鼠睾丸输出小管的免疫组织化学定位研究

目的研究正常大鼠睾丸输出小管上皮细胞上Aquaporin．1(AQP-1)的定位分布以期了解其在水重吸收上的作用机制。方法对正常wistar大鼠睾丸输出小管进行常规免疫组织化学方法染色观察。结果在睾丸输出小管非纤毛上皮细胞的刷状缘及基侧部AQP-1阳性表达强烈,核上区的胞内体的质膜上也有阳性表达;纤毛上皮细胞的纤毛亦呈阳性反应。结论Aquaporin-1可能与睾丸输出小管非纤毛上皮细胞水重吸收功能有密切关系。     

大鼠胃卵泡刺激素及其受体的免疫荧光定位及原位杂交

目的探讨大鼠胃组织中是否表达卵泡刺激素（FSH）及其受体（FSHR）,为进一步研究FSH对消化系统的功能调节提供理论依据。方法选取SD大鼠20只,雌雄不拘,经腹腔注射戊巴比妥钠麻醉成功后,用4％多聚甲醛先快后慢灌流固定2h,开腹取胃组织置于300g／L蔗糖液中直至组织沉底,恒冷箱切片机切成厚度为6／zm的组织切片用于单标记免疫荧光定位研究。另一部分胃组织放入4％多聚甲醛室温固定6—8h,按石蜡包埋程序包埋后制成4pm石蜡切片用于原位杂交研究。结果在大鼠胃底腺的壁细胞和主细胞呈FSH和FSHR免疫反应阳性,阳性物质分布于细胞质,细胞核呈阴性反应;上述细胞同样含有FSH和FSHRmRNA杂交信号,信号物质亦分布于细胞质内,细胞核呈阴性反应。结论FSH及其受体定位于大鼠壁细胞和主细胞,同时大鼠壁细胞和主细胞又能产生FSH及其受体,说明FSH对壁细胞和主细胞功能作用可能是通过旁分泌或自分泌的作用来实现的。     

地高辛标记寡核苷酸探针检测大鼠胰岛素mRNA

本实验采用地高辛标记前胰岛素寡核苷酸探针原位杂交组织化学技术,研究了Ｗｉｓｔａｒ大鼠胰腺组织胰岛素基因的表达。实验用Ｗｉｓｔａｒ大鼠５只,胰腺经４％多聚甲醛灌注固定,并在同一固定液中后固定２４ｈ,常规石蜡包埋切片。结果表明,经前胰岛素寡核苷酸探针杂交的胰腺切片中,胰岛Ｂ细胞呈蓝色。杂交反应物位于Ｂ细胞的细胞质和部分核仁中,胞核无色。胰岛其它细胞及胰腺外分泌部腺泡细胞无杂交反应物。对照切片中均无阳性信号出现。结果表明地高辛标记寡核苷酸探针不仅具备非同位素标记探针的优点而且能精确检测特异性ｍＲＮＡ的表达。本研究方法操作便利,可靠精确,重复性好。     

生长休止蛋白7在大鼠小脑皮质中的免疫细胞化学研究

目的探讨生长休止特定蛋白7(Gas7)在大鼠小脑中的表达定位。方法应用Gas7抗血清,对大鼠小脑组织切片进行免疫组织化学染色。结果在小脑皮质分子层可见大量的Gas7阳性神经纤维;蒲氏细胞层中,Gas7主要表达在神经元胞膜和部分胞质处;颗粒层中可见Gas7阳性神经纤维。结论Gas7主要在小脑神经元的定位特征可能与Gas7促进神经元和神经突起发育的调节功能有关。     

大白鼠颌下腺促性腺激素释放激素受体(GnRHR)mRNA的RT-PCR和原位杂交的研究

本实验采用RT－PCR法探讨大白鼠颌下腺是否存在GnRH受体mRNA,并用原位杂交法对其细胞定位进行了研究。结果显示RT－PCR可扩增出大白鼠颌下腺GnRH受体mRNA的特异性片段,其碱基数与设计的一致,原位杂交发现颌下腺浆液性腺泡上皮细胞、颗粒曲管、排泄管及分泌管上皮细胞内有GnRH受体mRNA的杂交信号,信号物质分布于胸质内,胞核阴性。上述结果表明大白鼠颌下腺能合成GnRH受体,颌下腺产生的GnRH可作用于颌上腺的靶细胞,参与颌下腺生理功能的调节。     

Immunohistochemical and in situ hybridization studies of gonadotropin releasing hormone (GnRH) and its receptor in rat digestive tract

GnRH(LH-RH) is first discovered in the hypothalamus and found to have a role in regulation of reproduction. With the study on it deepening, GnRH was demonnstrated that it also exists in a number of organs beyond the hypothalamus and acts on extrapituitary organs. To study whether digestive tract synthesizes GnRH and its receptor and, if it does, by what cells. In the experiment, the locallizations of GnRH and its receptors in rat digestive tract were studied using immunohistochemistry and in situ hybridization. The parietal cells of gastric gland, the villous and glandular epithelium in small and large intestine and parasympathetic ganglion cells of myenteric plexus showed GnRH immunoreactivity; GnRH mRNA hybridization signal was detected. The epithelium of gastric pit and the cells above in digestive tract showed GnRH receptor immunoreactivity; GnRH receptor mRNA hybridization signal was detected. The immunoreactive and signal materials distributed in cytoplasm of all positive cells, with nuclei being immunonegative and with no hybridization signal. These results suggested that the digestive tract can produce GnRH and express GnRH receptor; GnRH may also be a gastrointestinal hormone.     

Distribution, cloning and sequencing of GnRH, its receptor, and effects of gastric acid secretion of GnRH analogue in gastric parietal cells of rats

Our objective was to study the distribution of gonadotropin-releasing hormone (GnRH) and its receptor, cloning and sequencing of GnRH and its receptor gene in cultured gastric parietal cells of rats. The distribution of GnRH and its receptor mRNA were investigated through immunocytochemical ABC methods and in situ hybridization methods in cultured gastric parietal cells of rats. After isolation of the total RNA from the parietal cells, RT-PCR was conducted to obtain GnRH and its receptor cDNA. Then, the products of PCR was purified, digested by the restriction enzyme of Hind III and EcoR I, and DNA fragments of interests were cloned into pUC19 vector. The products of PCR were analyzed by sequencing with Sanger's method after identified by PCR and digestion of restriction enzyme. Gastric parietal cells showed GnRH and its receptor immunoreactivity; positive material was located in cytoplasm other than in nuclei. GnRH and its receptor mRNA hybridized signals were also detected in cytoplasm with negative nuclei. The specific amplified band of GnRH and its receptor sequences were detected through Agarose gel electrophoresis, and GnRH gene sequence is identical to that of GnRH which has been reported in rat hypothalamus and GnRH receptor sequence is identical to that of the pituitary of rat. GnRH analogue (Alarelin) could inhibit the gastric acid secretion both by direct actions on parietal cells and by inhibiting vagous function. Our data suggest that GnRH could be produced by gastric parietal cells of rats and may modulate physiological function of gastric parietal cells of rats through autocrinal and paracrinal way.     

Expression and regulation of Fas and Fas ligand on thyrocytes and infiltrating cells during induction and resolution of granulomatous experimental autoimmune thyroiditis.

Granulomatous experimental autoimmune thyroiditis (G-EAT) is induced by mouse thyroglobulin-sensitized spleen cells activated in vitro with mouse thyroglobulin, anti-IL-2R, and IL-12. G-EAT lesions reach maximal severity 19-21 days after cell transfer, and lesions almost completely resolve by day 35. Depletion of CD8+ cells delays resolution and reduces Fas ligand (FasL) mRNA expression in thyroids. This study was undertaken to analyze Fas and FasL protein expression in the thyroid during induction and resolution of G-EAT and to determine whether CD8+ cells might regulate Fas or FasL expression in the thyroid. Fas and FasL expression was analyzed by immunohistochemical staining or in situ hybridization in thyroids of mice with or without depletion of CD8+ cells. Fas and FasL proteins were not detectable in normal thyroids, but expression of both proteins increased during development of G-EAT. Fas was expressed primarily by inflammatory cells; some enlarged thyrocytes were also Fas+. Thyrocytes had intense FasL immunoreactvity, and many CD8+ cells were also FasL positive. Depletion of CD8+ cells resulted in decreased FasL expression by thyrocytes and inflammatory cells, but had no effect on Fas expression. TUNEL assay detected many apoptotic inflammatory cells in proximity to thyrocytes. CD8-depleted thyroids had ongoing inflammation with fewer apoptotic infiltrating cells at day 35. Administration of a neutralizing anti-FasL mAb had no apparent effects on development of G-EAT, but anti-FasL was as effective as anti-CD8 in preventing G-EAT resolution. These results suggested that CD8+ T cells and thyrocytes may kill inflammatory cells through the Fas pathway, contributing to G-EAT resolution.     

Enzymic markers of thyroid C cells in some rodents.

The paper provides comparative data of the localization of histochemical reactions demonstrating the activities of alpha-glycerophosphate and succinate dehydrogenases, acid phosphatase, non-specific esterases and non-specific acetylcholinesterase in the C cells of thyroids of 26 animals belonging to 5 rodent species. The family Muridae is represented by the Wistar albino rat and albino mouse, the family Microtidae by the bank vole Clethrionomys glareolus (Schreber 1780), the field vole Microtus agrestis L. 1761, and the pine vole Pitymys subterraneus De Selys-Longchamps 1825. The observed enzyme activity differences were most conspicuous on comparing the rat and mouse thyroids and in a much less degree the Microtidae thyroids. Among the histochemical reactions tested that for succinate dehydrogenase proved to be least effective as a C cell marker, alpha-glycerophosphate dehydrogenase being better, and acid phosphatase and non-specific esterases the best (not in the rat thyroid). The reaction for non-specific cholinesterase (with some limitations) gave satisfactory results in the C cells of all animal's thyroids. The present paper continues earlier studies [19] on the morphology of the C cells in thyroid glands of the rodents of the families Muridae and Microtidae and aims at supplementing them with histochemical data of enzymic activities. It deals with enzyme reactions that are employed as C cell markers in Mammals other than Rodents.     

Localization of growth hormone receptors in rat and human thyroid cells

Growth hormone (GH) exerts its multiple actions by binding to a specific receptor (GHR) widely distributed in the organism. It is well established that, in acromegaly, the thyroid gland is larger than normal and that GH increases triiodothyronin concentrations and decreases those of tetraiodothyronin (thyroxine). The aim of the present study was to analyze the presence of GHR and its mRNA in rat and human thyroid gland by Western blot, in situ hybridization techniques, and immunohistochemistry. A band of the expected size for GHR was shown in rat and human thyroid by Western blot. GHR immunoreactivity was found in virtually all follicles. The signal was mainly localized in the cytoplasm, although a nuclear positivity was also found. In situ hybridization techniques demonstrated the presence of GHR messenger RNA in the thyroid gland (cytoplasm of the follicular cells). These results provide direct morphological evidence that GHR is localized in the thyroid gland of mammals and opens up the possibility that GH regulates thyroid cell function directly or via local autocrine or paracrine production of insulin-like growth factor I.     

A study on the localization and distribution of GnRH and its receptor in rat submaxillary glands by immunohistochemical,in situ hybridization and RT-PCR

We investigated the rat submaxillary gland for the presence of GnRH and GnRH receptors, the localization and colocalization of GnRH, GnRH receptor and their mRNA, and studied the sequence of GnRH receptor complementary DNA (cDNA) by immunohistochemistry, in situ hybridization and RT-PCR. The results showed that GnRH and GnRH receptor immunoreactive materials were colocalized in the epithelial cells of the serous acinus and glandular duct. The GnRH and GnRH receptor mRNA hybridization signals were detected in the above cells. The sequence obtained from the RT-PCR product was identical to the published cDNA sequence of GnRH receptor in the rat pituitary. The results suggested that the rat submaxillary gland was capable of synthesizing GnRH and GnRH receptors. GnRH may be involved in the functional regulation of the submaxillary gland through autocrine or paracrine activity.     

Studies on expression of FSH and its anti-apoptotic effects on ischemia injury in rat spinal cord

Studies indicated that many tissues could express FSH. New functions of FSH have been recognized beyond reproduction regulation. However, no report has been made about the expression and function of FSH in rat spinal cord. Double-labeled immunofluorescence stain and in situ hybridization were used to study the co-localization of FSH with its receptor and co-localization of FSH with GnRH receptor in rat spinal cord. Spinal cord ischemia injury models were built, TUNEL stain and Fas immunostaining were made to observe the anti-apoptotic effects of FSH to neurons induced by spinal cord ischemia injury. The results found that some neurons and glias of rat spinal cord showed both FSH immunoreactivity and FSH mRNA positive signals; not only FSH and its receptor but also FSH and GnRH receptor co-located in cells of both gray matter and white matter; treatment with certain concentration of FSH before ischemia–reperfusion injury, less TUNEL positive cells and Fas positive cells were found in motor neurons of ventral gray matter in FSH experiment group than that in control group. These suggested that rat spinal cord could express FSH, it is also a target organ of FSH; FSH might exert functions through its receptor by paracrine or autocrine effects; GnRH in spinal cord might regulate FSH positive neurons through GnRH receptor; FSH might inhibit ischemia induced neuron apoptosis by down-regulating Fas expression in spinal cord.