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The tooth root cementum is a thin, mineralized tissue covering the root dentin that is present primarily as acellular cementum on the cervical root and cellular cementum covering the apical root. While cementum shares many properties in common with bone and dentin, it is a unique mineralized tissue and acellular cementum is critical for attachment of the tooth to the surrounding periodontal ligament （PDL）. Resources for methodologies for hard tissues often overlook cementum and approaches that may be of value for studying this tissue. To address this issue, this report offers detailed methodology, as well as comparisons of several histological and immunohistochemical stains available for imaging the cementum-PDL complex by light microscopy. Notably, the infrequently used Alcian blue stain with nuclear fast red counterstain provided utility in imaging cementum in mouse, porcine and human teeth. While no truly unique extracellular matrix markers have been identified to differentiate cementum from the other hard tissues, immunohistochemistry for detection of bone sialoprotein （BSP）, osteopontin （OPN）, and dentin matrix protein 1 （DMP1） is a reliable approach for studying both acellular and cellular cementum and providing insight into developmental biology of these tissues. Histoloeical and immunohistochemical aooroaches Drovide insight on developmental biology of cementum.

Both genetic and environmental factors are important in the pathogenesis of Parkinson＇s disease. As α-synuclein is a major constituent of Lewy bodies, a pathologic hallmark of Parkinson＇s disease, genetic aspects of α-synuclein is widely studied. However, the influence of dietary factors such as quercetin on α-synuclein was rarely studied. Herein we aimed to study the neuroprotective role of quercetin against various toxins affecting apoptosis, autophagy and aggresome, and the role of quercetin on α-synuclein expression. PC12 cells were pre-treated with quercetin（100, 500, 1,000 μM） and then together with various drugs such as 1-methyl-4-phenylpyridinium（MPP＋; a free radical generator）, 6-hydroxydopamine（6-OHDA; a free radical generator）, ammonium chloride（an autophagy inhibitor）, and nocodazole（an aggresome inhibitor）. Cell viability was determined using a 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltertazolium bromide（MTT） assay. Apoptosis was detected by annexin V-fluorescein isothiocyanate and propidium iodide through the use of fluorescence activated cell sorter. α-Synuclein expression was detected by western blot assay and immunohistochemistry. The role of α-synuclein was further studied by knocking out α-synuclein using RNA interference. Cell viability increased at lower concentrations（100 and 500 μM） of quercetin but decreased at higher concentration（1,000 μM）. Quercetin exerted neuroprotective effect against MPP＋, ammonium chloride and nocodazole at 100 μM. MPP＋ induced apoptosis was decreased by 100 μM quercetin. Quercetin treatment increased α-synuclein expression. However, knocking out α-synuclein exerted no significant effect on cell survival. In conclusion, quercetin is neuroprotective against toxic agents via affecting various mechanisms such as apoptosis, autophagy and aggresome. Because α-synuclein expression is increased by quercetin, the role of quercetin as an environmental factor in Parkinson＇s disease pathogenesis needs further investigation.

Estrogen affects the generation and transmission of neuropathic pain,but the specific regulatory mechanism is still unclear.Activation of the N-methyl-D-aspartate acid receptor 1（NMDAR1） plays an important role in the production and maintenance of hyperalgesia and allodynia.The present study was conducted to determine whether a relationship exists between estrogen and NMDAR1 in peripheral nerve pain.A chronic sciatic nerve constriction injury model of chronic neuropathic pain was established in rats.These rats were then subcutaneously injected with 17β-estradiol,the NMDAR1 antagonist D（-）-2-amino-5-phosphonopentanoic acid（AP-5）,or both once daily for 15 days.Compared with injured drug na？ve rats,rats with chronic sciatic nerve injury that were administered estradiol showed a lower paw withdrawal mechanical threshold and a shorter paw withdrawal thermal latency,indicating increased sensitivity to mechanical and thermal pain.Estrogen administration was also associated with increased expression of NMDAR1 immunoreactivity（as assessed by immunohistochemistry） and protein（as determined by western blot assay） in spinal dorsal root ganglia.This 17β-estradiol-induced increase in NMDAR1 expression was blocked by co-administration with AP-5,whereas AP-5 alone did not affect NMDAR1 expression.These results suggest that 17β-estradiol administration significantly reduced mechanical and thermal pain thresholds in rats with chronic constriction of the sciatic nerve,and that the mechanism for this increased sensitivity may be related to the upregulation of NMDAR1 expression in dorsal root ganglia.

Electroacupuncture at Shangjuxu （ST37） and Tianshu （ST25） can improve visceral hypersensitivity in rats. Colorectal distension was used to establish a rat model of chronic visceral hypersensitivity. Immunohistochemistry was used to detect P2X2 and P2X3 receptor expression in dorsal root ganglia from rats with chronic visceral hypersensitivity. Results demonstrated that abdominal withdrawal reflex scores obviously increased following establishment of the model, indicating visceral hypersensitivity. Simultaneously, P2X2 and P2X3 receptor expression increased in dorsal root ganglia. After bilateral electroacupuncture at Shangjuxu and Tianshu, abdominal withdrawal reflex scores and P2X2 and P2X3 receptor expression decreased in rats with visceral hypersensitivity. These results indicated that electroacupuncture treatment improved visceral hypersensitivity in rats with irritable bowel syndrome by reducing P2X2 and P2X3 receptor expression in dorsal root ganglia.

Glypican-3 （GPC3） is a promising tumor marker for hepatocellular carcinoma （HCC） diagnosis with high sensitivity and specificity. The aim of this study was to establish an immunohistochemical detection method for GPC3 using the 7D 11 monoclonal antibody （7D11 mAb） and evaluate its application for HCC diagnosis. The feasibility of the 7D11 mAb was evaluated by immunohistochem- istry performed on adjacent normal liver and intrahepatic cholangiocarcinoma （ICC） samples, Furthermore, the serum GPC3 levels were evaluated in 40 HCC patients, 7 ICC patients and 50 healthy donors. The results showed that GPC3 was expressed in 85% of HCC tissues （34/40）, but was undetectable in ICC tissues and adjacent normal tissues.GPC3 was significantly increased in the serum of HCC patients （17/40, 42.5%） but was undetectable in the serum of ICC patients （0/7, 0%） and healthy donors（0/50, 0%）. This prospective study evaluated the clinical usefulness of 7D11 mAb for GPC3 detection in HCC patients. In conclusion, the use of 7D 11 mAb might be good for GPC3 large-scale applications for clinical diagnosis of HCC.

Autoimmune activities have been implicated in the pathogenesis of hypertension.High levels of autoantibodies against the second extracellular loop of α1-adrenoceptor（α1-AR autoantibody,α1-AA） are found in patients with hypertension,and α1-AA could exert a α1-AR agonist-like vasoconstrictive effect.However,whether the vasoconstrictive effect of α1-AA is enhanced in hypertension is unknown.Using aortic rings of spontaneously hypertensive rats（SHR） and normotensive Wistar-Kyoto（WKY） rats,we observed the vasoconstrictive responses to α1-AA with phenylephrine（α1-AR agonist） as a positive control drug.Aortic nitrotyrosine levels were also measured by ELISA and immunohistochemistry.The results showed that the aortic constrictive responses to α1-AA and phenylephrine（both 1 nmol L-1-10 μmol L-1） were greater in SHR than in WKY rats.Endothelial denudation or L-NAME（a non-selective NOS inhibitor）（100 μmol L-1） increased α1-AA- or phenylephrine-induced vasoconstrictions both in SHR and WKY.However,selective iNOS inhibitor 1400W（10 μmol L-1） enhanced the α1-AA-induced aortic constriction in WKY,but not in SHR.The aortic nitrotyrosine level was significantly higher in SHR than WKY,as shown by both ELISA and immunohistochemistry.These results indicate that the vasoconstrictive response to α1-AA is enhanced in SHR,and this altered responsiveness is due to endothelial dysfunction and decreased NO bioavailability.The study suggests an important role of α1-AR autoimmunity in the pathogenesis and management of hypertension especially in those harboring high α1-AA levels.

Signaling pathways known to have components with mutations in human medulloblastoma include sonic hedgehog, Wnt/beta-catenin and insulin-like growth factor. Microarray analysis was applied to examine the gene expression changes in medulloblastomas of pTet-on/pTRE-SV40Tag transgenic mice. Altogether, 14 112 genes were detectable, including 152 genes with significantly different expression levels. These genes were associated with immunity, the cell cycle, signal transduction, cytoskeleton and metabolism. To further confirm the microarray data, real-time polymerase chain reactions were used to examine the expression changes of genes related to sonic hedgehog, Wnt/beta-catenin and insulin-like growth factor signal pathways. Immunohistochemistry detected insulin receptor substrate-1 in the nuclei of brain tumor tissue cells from pTet-on/pTRE-SV40Tag transgenic mice, suggesting that SV40 large T antigen may activate the insulin-like growth factor signal pathway to promote tumorigenesis.

Animal models of intracerebral hemorrhage were established by injection of autologous blood into the caudate nucleus in rats. Cell apoptosis was measured by flow cytometry and immunohistochemical staining of the p75 neurotrophin receptor. p75 neurotrophin receptor protein was detected by immunohistochemistry. p75 neurotrophin receptor mRNA was examined by quantitative real-time polymerase chain reactions. At 24 hours after modeling, cellular apoptosis occured around hematoma with upregulation of p75 neurotrophin receptor protein and mRNA was observed, which directly correlated to apoptosis. This observation indicated that p75 neurotrophin receptor upregulation was associated with cell apoptosis around hematomas after intracerebral hemorrhage.

背景与目的脆性组氨酸三联体（fragile histidine triad，FHIT）是1996年由Ohta等克隆的候选抑癌基因，文献报道在多种与环境致癌物有关的恶性肿瘤中，常有FHIT的改变。本研究应用免疫组化方法检测FHIT蛋白在细胞芯片中的表达情况，进一步验证细胞芯片，为快速、简便、经济、规范地进行各项细胞学检查提供一种新方法。方法收集天津医科大学总医院、天津市第一中心医院及天津市胸科医院2005年5月-8月有肿瘤细胞的肺癌患者的胸水50例、非肿瘤患者胸水6例，运用自制的细胞芯片检测装置，制备112点细胞芯片。应用免疫组织化学方法检测细胞芯片中FHIT蛋白的表达，采用SPSS11．5统计软件进行数据处理。结果细胞芯片排列整齐，分布均匀，无掉片，符合观察需要。细胞芯片免疫组化染色结果与进行FHIT免疫组化染色的组织芯片中Ⅲ＋Ⅳ期肺癌的检测结果进行比较，差异均无统计学意义（P〉0．05）。结论细胞芯片具有操作简单，信息量大，节约试剂，减少误差，诊断决速的特点，在临床诊断、科学研究和流行病学筛选中具有广泛的应用前景。

目的 研究p63基因在人类非小细胞肺癌及肺良性病变组织中的转录表达水平及其意义。方法 采用RT-PCR(rcversc transcriptase—PCR)法对40例非小细胞肺痛及10例肺良性病变组织中p63基因两种亚型(TAp63及△Np63)转录表达情况进行检测和比较，同时用免疫组化法检测p63蛋白在上述组织中的表达情况。结果 18例肺鳞癌(18／23)、1例细支气管肺泡癌(1／6)及2例肿鳞癌癌旁组织(2／23)中△Np63呈阳性表达，其余肿瘤、癌旁及远癌组织中△Np63表达均为阴性。所有良、恶性肺组织中均未见TAp63表达。免疫组化研究显示p63蛋白在肺鳞癌组织中的阳性表达率和表达强度明显大于腺癌组织及良性病变肺组织(P<0．01)。结论 p63基因主要存肺鳞癌组织中呈阳性表达，在肺鳞癌的发生、发展中可能起到癌基因作用。