亚麻籽木酚素预防乳腺癌的作用及机制研究

目的:探讨亚麻籽木酚素预防乳腺癌的作用及机制。方法:分别采用基础饲料(BD组)和添加亚麻籽木酚素的饲料(FS组)喂养大鼠两周后,通过二甲基苯蒽灌胃复制乳腺癌模型大鼠,连续观察并测定两组大鼠瘤体的体积和重量,采用免疫组织化学技术分别检测和比较两组大鼠乳腺癌组织中PCNA、Her-2、Ki-67的表达。结果:BD组大鼠乳腺癌的发生率及瘤体重量和体积均较FS组更高(P0.05)。FS组乳腺癌组织中Ki-67、Her-2、PCNA阳性表达率均较BD组显著降低(P0.010)。结论:亚麻籽木酚素可发挥预防乳腺癌的效应,该效用可能与其降低Ki-67、Her-2、PCNA的表达有关。     

p16和bcl-2在宫颈上皮瘤样病变及宫颈癌中的表达

目的探讨p16和bcl-2表达产物在宫颈上皮瘤样病变及宫颈癌中表达的意义。方法对正常宫颈鳞状上皮、宫颈上皮内瘤变(CIN)和宫颈癌组织共69例,采用免疫组织化学EliVision法,对宫颈癌变过程中p16和bcl-2蛋白进行研究,将结果进行统计分析。结果p16蛋白在CIN中的表达高于正常宫颈上皮(P<0.01),在宫颈癌中的表达也高于正常宫颈上皮(P<0.01),且高于CIN中的表达(P<0.05);bcl-2蛋白在CIN中的表达高于正常宫颈上皮(P<0.01),在宫颈癌中的表达也高于正常宫颈上皮(P<0.01),但与CIN中表达无差异。p16和bcl-2两种蛋白在CIN和宫颈癌中的表达无明显差异。结论p16和bcl-2蛋白的表达与宫颈癌的发生有关,提示这两种蛋白有可能作为高危人群早期筛查的一种免疫组化指标。     

高危型人乳头瘤病毒及p16和Ki-67联合检测在宫颈鳞状上皮内病变诊断中的价值分析

目的探讨p16和Ki-67在不同宫颈鳞状上皮内病变中的表达情况及不同宫颈鳞状上皮内病变与高危型人乳头瘤病毒(high-risk human papilloma virus, HR-HPV)感染的关系。方法应用免疫组织化学技术检测138例宫颈病变(32例慢性宫颈炎、56例低级别鳞状上皮内病变、50例高级别鳞状上皮内病变)活检组织的p16和Ki-67表达水平,同时回顾性分析活检时HR-HPV的感染情况。结果 HR-HPV感染率、p16和Ki-67阳性表达率和表达水平随着宫颈病变级别增加而逐渐增高;HR-HPV感染率与p16和Ki-67表达水平均呈正相关。结论 HR-HPV的检测可用作临床宫颈病变筛查的常规检查,HRHPV及p16和Ki-67联合检测可降低宫颈病变的漏诊率,可用于指导宫颈病变的分级,提高宫颈活检诊断的准确率。     

乳腺癌细胞肿瘤标记物表达及其与临床病理特征相关性研究

目的 探讨肿瘤标记物14-3-3σ、Akt 和p27Kip1 蛋白在乳腺癌中的表达及其与临床病理特征及Her-2 的相关性.方法 选取43 份乳腺癌石蜡标本和10 份正常乳腺组织标本,采用免疫组织化学技术(SABC)检测组织中14-3-3σ、Akt 和p27Kip1 蛋白的表达,结合临床资料和随访资料,进行回顾性研究,...     

肺癌组织中PCNA、p63和p53蛋白的表达及临床意义

目的:检测蛋白增殖细胞核抗原(PCNA)、p63和p53在肺癌组织中的表达情况,以探讨三者在肺癌的发生、发展中的生物学作用和临床意义。方法:选取195例肺癌组织(其中57例有癌旁组织),应用组织芯片技术和免疫组织化学方法观察三种蛋白的表达情况,并研究三者之间及其与临床病理参数的关系。结果:PCNA、p63和p53蛋白在肺癌组织中的阳性表达率分别为96.41%、38.46%及58.46%,但三者在癌旁组织中均无表达,差异有统计学意义(均P0.05);在肺癌组织中,PCNA、p63和p53蛋白的表达情况均与组织分型有关(P0.05),且PCNA、p53蛋白表达与分化程度有关(P0.05),分化越差,表达越高;p53表达与PCNA表达呈正相关(r=0.352,P=0.043),p63与p53、PCNA的表达不相关(P0.05)。结论:肺癌组织中PCNA、p63和p53蛋白的表达升高,三者均在肺癌的发生、发展中发挥着重要作用,并且临床可通过检测三者的蛋白水平,作为鉴别肺鳞状细胞癌与其他类型癌的重要参考指标,为病理诊断提供依据。     

大鼠心包组织存在C-kit、Sca-1和Nanog阳性细胞

目的观察大鼠心包组织中C-kit,Sca-1,Nanog阳性细胞的表达及分布,为治疗心肌损伤寻找新的干细胞来源。方法取大鼠新鲜心包,利用石蜡切片和铺片技术,HE和Masson染色,观察其组织结构;利用免疫组织化学染色技术,观察C-kit、Scal-1、Nanog阳性细胞在心包组织中的表达;利用免疫荧光双标技术观察C-kit、Scal-1、Nanog之间的共表达情况;利用免疫印迹法检测心包组织中C-kit、Scal-1、Nanog的表达。结果大鼠心包组织中存在大量散在的小而圆或椭圆形细胞,广泛分布于成纤维细胞之间和小血管周围。免疫组织化学染色显示,这些细胞分别阳性表达C-kit、Sca-1、Nanog。免疫荧光双标染色显示,部分细胞共表达C-kit和scal-1两种表面抗原,部分细胞共表达scal-1和Nanog表面抗原,而C-kit与Nanog共表达于血管弹力膜。结论大鼠心包组织中存在C-kit、Scal-1和Nanog阳性细胞,一种细胞可表达两种表面标记。     

胰腺导管单克隆上皮样干细胞的蛋白表达特征

该文研究1例源于成年大鼠的胰腺导管单克隆上皮样干细胞系的蛋白表达特征、染色体核型及致瘤性。RPMI-1640有血清扩增培养源于成年大鼠的胰腺导管单克隆上皮样干细胞至单层,分别采用流式细胞术或免疫荧光反应检测干细胞蛋白水平的表达特征。常规法制备染色体标本,采用Adobe Photoshop CS6软件进行核型分析。将干细胞移植在裸鼠体内,观察其致瘤性。结果显示,该源于成年大鼠的胰腺导管单克隆上皮样干细胞在蛋白水平表达CK19、Neuro D2、Oct4、PCNA及Nanog,不表达Pdx1、Pax4、Pax6、Maf A、Ptf1a、Ngn3、nestin、Sox2、CD34、CD45、insulin、glucagon、ghrelin、somatostatin及secretin。细胞是正常的二倍体细胞(2n=42),无致瘤性。这表明,该源于成年大鼠的胰腺导管单克隆上皮样干细胞系共表达CK19、Neuro D2、Oct4、PCNA及Nanog蛋白。     

口腔鳞状细胞癌中hMSH2、PCNA和p53的表达及意义

目的:探讨hMSH2、PCNA和p53在OSCC中的表达及可能存在的临床意义.方法:运用免疫组织化学S-P法对56例OSCC组织中hMSH2、PCNA和p53的表达进行检测.结果:(1)3种基因产物在OSCC中的阳性率均高于正常口腔黏膜,中-低分化癌的阳性率高于高分化癌,有淋巴结转移的阳性率高于无淋巴结转移.(2)hMSH2与PCNA、p53,p53与PCNA的表达均呈正相关.(3)5年生存率与hMSH2蛋白表达和淋巴结转移有关.结论:hMSH2、PCNA和p53的异常表达及其相互之间的调节可能与OSCC的发生发展有关.     

高危型人乳头瘤病毒感染后TopoⅡα和p16INK4A蛋白的表达及诊断价值

目的 探讨高危型人乳头瘤病毒(HR-HPV)感染后TopoⅡα和p16INK4A在宫颈组织中的表达及其对癌前病变的诊断价值.方法 选取在温州医学院附属二院行宫颈病变诊治的患者135例,包括各级宫颈上皮内瘤变(CIN)和宫颈鳞癌(SCC)的患者.随机选取同期就诊的慢性宫颈炎患者30例作为对照.运用第二代杂交捕获法(HC2)检测高危型人乳头瘤病毒(HR-HPV)的表达.活体提取组织,运用免疫组织化学法(SP法)检测宫颈组织中Topo Ⅱα和p16INK4A蛋白表达.结果 在HR-HPV阳性患者的宫颈组织中Topo Ⅱα和p16INK4A表达率高于HR-HPV阴性患者,差异有统计学意义.Topo Ⅱα和p16INK4A在正常宫颈上皮组织、各级CIN和SCC中的表达率逐渐增加,差异有统计学意义.p16INK4A用于筛查宫颈癌前病变(包括SCC)时阳性预测值99.3％.Topo Ⅱα用于筛查宫颈严重病变(CINⅢ和SCC)时阳性预测值67.9％.结论 HR-HPV感染后Topo Ⅱα和p16INK4A蛋白表达明显增高,并且Topo Ⅱα和p16INK4A蛋白表达率随着宫颈癌前病变的严重程度而上调,参与宫颈上皮的异常增殖和恶性转变.p16INK4A免疫组化检测提高了宫颈癌前病变筛查的特异性,Topo Ⅱα免疫组化检测提高了宫颈严重病变筛查的特异性,当Topo Ⅱα蛋白表达阳性,宫颈组织可能已发生不可逆性病变.     

人乳头状瘤病毒感染与宫颈癌患者临床病理特征和Ki-67、PCNA的关系

目的:研究人乳头状瘤病毒(HPV)感染与宫颈癌患者临床病理特征和Ki-67、细胞增殖抗原(PCNA)的相关性,从而为临床宫颈癌的诊治提供参考依据。方法:选取2016年3月～2018年6月于我院接受手术治疗的宫颈病变患者130例为研究对象。其中宫颈癌患者30例记为宫颈癌组,宫颈上皮内瘤变患者68例记为宫颈上皮内瘤变组,慢性宫颈炎患者32例记为对照组。采用免疫组织化学法检测各组宫颈组织中HPV感染、Ki-67以及PCNA阳性表达情况,并分析HPV与宫颈癌患者临床病理特征的关系及其与Ki-67、PCNA的相关性。结果:宫颈癌组、宫颈上皮内瘤变组患者HPV、Ki-67以及PCNA阳性率均高于对照组,宫颈癌组高于宫颈上皮内瘤变组(均P0.05)。临床分期Ⅲ～Ⅳ期以及淋巴结转移宫颈癌患者HPV感染率均明显高于临床分期Ⅰ～Ⅱ期与无淋巴结转移患者(均P0.05)。经Spearman相关性分析可得:宫颈癌患者HPV感染与Ki-67、PCNA表达均呈正相关关系(均P0.05)。结论:宫颈癌患者存在明显的HPV感染,且HPV感染与宫颈癌患者临床分期、淋巴结转移、Ki-67、PCNA表达存在一定相关性,临床可通过对HPV、Ki-67、PCNA进行联合检测,从而有助于宫颈癌的早期诊断。     

Expression of Caveolin-1 in tongue squamous cell carcinoma by quantum dots

Quantum dots (QDs) are a new class of fluorescent probes to detect biomarker expression. The role of caveolin-1 (Cav-1) in tongue squamous cell carcinoma (TSCC) is still unknown. This study aimed to investigate the expression profile of Cav-1 in carcinogenesis and development of TSCC by QDs immunofluorescence histochemistry (QDs-IHC) and discuss the relationship between the Cav-1 expression and the clinicopathological outcomes. QDs-IHC was used to detect Cav-1 expression in tissue microarrays including normal tongue mucosa (NTM; n=10), hyperplastic tongue mucosa (HTM; n=10), tongue pre-cancer lesions (TPL; n=15) and primary tongue squamous cell carcinoma (PTSCC; n=61). Correlations between the Cav-1 expression and clinicopathologic variables were evaluated statistically. Cells positive for Cav-1 were clearly detected and bright images were obtained in a fine, granular pattern at the cell membrane and cytoplasm using QDs-IHC. The rate of Cav-1 immunoreactivity increased progressively from NTM (0%), HTM (0%), TPL (36%) to PTSCC (74%). When compared with each other, there was statistical significance among PTSCC, TPL and NTM as well as among PTSCC, TPL and HTM. Moreover, Cav-1 expression level in PTSCC was correlated positively with clinical stage and histologic grade. QDs-IHC could accurately detect protein location in tongue mucosa. An increased expression of Cav-1 in the stepwise carcinogenesis from NTM, HTM, TPL to PTSCC suggested that Cav-1 might be an oncogene in the development of tongue squamous cell carcinoma.Key words: tongue squamous cell carcinoma, caveolin-1, quantum dots.     

On-chip detection of protein glycosylation based on energy transfer between nanoparticles

We describe a chip-based method to detect protein glycosylation based on the energy transfer between quantum dots (QDs) and gold nanoparticles (AuNPs). Our assay system relies on modulations in the energy transfer between the nanoparticles on a surface. The photoluminescence (PL) of lectin-coated QDs (energy donor) immobilized on a glass slide is quenched by carbohydrate-coated AuNPs (energy acceptor), and the presence of the glycoprotein causes the increase of the PL of QDs. As a proof-of-concept, Concanavalin A-coated QDs (ConA-QDs) and dextran-coated AuNPs (Dex-AuNPs) were used to detect the mannosylated proteins. As a result, the PL intensity of QDs was found to be linearly correlated with the concentration and the number of glycan moiety of the glycoprotein. We anticipated that our simple assay system will find applications for the analysis of glycoproteins with high selectivity and sensitivity in a high-throughput manner.     

Comparison of quantum dot immunofluorescence histochemistry with conventional immunohistochemistry in detecting Helicobacter pylori infection in paraffin-embedded tissues of gastric biopsy

Quantum dots (QDs) are a new type of fluorescent label, which has been widely used in many biological and biomedical imaging applications. In this study, we used QDs-based immunofluorescence histochemistry (QDs-IHC) and conventional immunohistochemistry (IHC) techniques to perform a retrospective analysis on paraffin-embedded tissues of gastric biopsies in 203 patients (112 of which were HP positive and 91 were negative). The ability of QDs-IHC to detect Helicobacter pylori (HP) in gastric biopsies compared to IHC technology was evaluated. In our study, both methods showed consistent HP morphology and localization. The positive detection rate of HP for QDs-IHC in formalin-fixed and paraffin-embedded (FFPE) tissue was 54.7% (111/203), and the sensitivity and specificity reached 99.11% and 100%, respectively. However the positive detection rate of HP for IHC was 53.7% (109/203), with a sensitivity and specificity of 97.32% and 100%, respectively. Weak positives (1+) were detected in 2 case of QDs-IHC with negative in IHC, and moderate positives (2+) were detected in 3 case of QDs-IHC with weak positives (1+) in IHC. The consistency test showed that the two methods showed good agreement (κ?=?0.980, P?=?0.014), but the sensitivity of QDs-IHC was slightly higher than that of conventional IHC. Our results show that QDs-IHC has strong sensitivity and high specificity. It is superior to conventional IHC in detecting HP infection in FFPE tissues of gastric biopsy, especially in tissues with low HP content.

     

Comparison of quantum dots immunofluorescence histochemistry and conventional immunohistochemistry for the detection of caveolin-1 and PCNA in the lung cancer tissue microarray

Luminescent semiconductor quantum dots (QDs) are a new class of fluorescent label with wide ranges of applications in cell imaging. In this study, we evaluated the capability of QDs immunofluorescence histochemistry (QDs-IHC) for detecting antigens of caveolin-1 and PCNA in the lung cancer tissue microarray (TMA) in comparison with the conventional immunohistochemistry (IHC) technique. Both methods revealed consistent antigen localization and statistically non-significant detection rates of caveolin-1 and PCNA expressions in our study. However, the sensitivity of QDs-IHC was higher than IHC. The positive detection rates of caveolin-1 and PCNA by QDs-IHC were 57% (40/70) and 86% (60/70), respectively, which were higher than the detection rates of 47% (33/70) and 77% (54/70), respectively, by IHC. Moreover, QDs exhibited a much better photostability, a broader excitation spectrum and a longer fluorescence lifetime. We showed here the advantages of QDs-IHC over IHC for the detection of caveolin-1 and PCNA in lung cancer TMA.     

In situ immunocytochemical detection of potyviral proteins in plant cells

A method for in situ protein immunodetection using a peroxidase labeling system is described for detecting functional and structural proteins encoded by potato virus Y (Tunisian isolate) in plant tissues. Such Potyviruses are characterized by the accumulation of inclusion bodies containing viral encoded proteins other than coat protein. These proteins are functional at early stages of infection, making them easy to detect. Data are compared to those obtained by immunofluorescence techniques. Our technique can be used as a preliminary method for rapid detection of virus infection using antibodies directed against functional proteins.     

Down-regulated expression of exocytotic proteins in pancreatic islets of diabetic GK rats

Exocytosis is regulated by exocytotic proteins, which are present in insulin-secreting beta-cells and play regulatory roles in insulin secretion. Non-insulin dependent diabetes mellitus (type 2 diabetes) is a disease characterized by impaired insulin secretion and insulin resistance. Exocytotic protein immunoreactivities were studied in pancreatic islets of type 2 diabetic Goto-Kakizaki (GK) rats using immunofluorescence histochemistry. The immunoreactivities for vesicle-associated membrane protein-2 (VAMP-2), synaptotagmin III, cysteine string protein (CSP), mammalian homologue of the unc-18 gene (Munc-18), alpha-soluble N-ethylmaleimide-sensitive attachment protein (alpha-SNAP), N-ethylmaleimide-sensitive factor (NSF) and synaptosomal-associated protein of 25 kDa (SNAP-25) exhibited weaker immunofluorescence intensity in islets of GK rats as compared to control Wistar rats. Insulin immunoreactivity was also decreased in GK rat beta-cells, whereas no detectable alterations in the expression of actin immunoreactivity could be detected. The data suggest that reduced expression of exocytotic proteins and decreased insulin content may contribute to the diabetic syndrome in the GK rat.     

UDP-glucuronate decarboxylase, a key enzyme in proteoglycan synthesis: cloning, characterization, and localization

UDP-glucuronate decarboxylase (UGD) catalyzes the formation of UDP-xylose from UDP-glucuronate. UDP-xylose is then used to initiate glycosaminoglycan biosynthesis on the core protein of proteoglycans. In a yeast two-hybrid screen with the protein kinase Akt (protein kinase B), we detected interactions with a novel sequence, which we cloned and expressed. The expressed protein displayed UGD activity but did not display the activities of homologous nucleotide sugar epimerases or dehydratases. We did not detect phosphorylation of UGD by Akt nor did we detect any influence of Akt on UGD activity. Effects of UGD on Akt kinase activity were also absent. Northern blot and Western blot analyses revealed the presence of UGD in multiple tissues and brain regions. Subcellular studies and histochemistry localized UGD protein to the perinuclear Golgi where xylosylation of proteoglycan core proteins is known to occur.     

Monoclonal antibodies to different antigens of skin epithelium obtained by immunizing mice with streptococcus group A antigens

Monoclonal antibodies (MCA) were obtained by immunization of BALB/c mice with streptococcal group A protein antigens of the cellular wall, or with whole microbial cells. In immunofluorescence test, MCA react with different skin epithelial structures (basal, suprabasal or all the epidermal layers). The majority of MCA belong to autoantibodies. The same MCA revealed no cross-reactions with streptococcal antigens in immunoenzyme and inhibition tests. MCA reacting with epithelial cells are, apparently, obtained as a result of polyclonal activation of the autoreactive clones by streptococcal antigens.     

Molecularly imprinted polymer anchored on the surface of denatured bovine serum albumin modified CdTe quantum dots as fluorescent artificial receptor for recognition of target protein

A new type of molecularly imprinted polymer (MIP)-based fluorescent artificial receptor was developed by anchoring MIP on the surface of denatured bovine serum albumin (dBSA) modified CdTe quantum dots (QDs) using the surface molecular imprinting process. The approach combined the merits of molecular imprinting technology and the fluorescent property of the CdTe QDs. The dBSA was used not only to modify the surface defects of the CdTe QDs, but also as assistant monomer to create effective recognition sites. Three different proteins, namely lysozyme (Lyz), cytochrome c (Cyt) and methylated bovine serum albumin (mBSA), were tested as the template molecules and then the receptors were synthesized by sol-gel reaction (imprinting process). The results of fluorescence and binding experiments demonstrated the recognition performance of the receptors toward the corresponding template. Under optimum conditions, the linear range for Lyz was from 1.4×10(-8) to 8.5×10(-6) M, and the detection limit was 6.8 nM. Moreover, the new artificial receptors were applied to separate and detect Lyz in real samples. This fluorescent artificial receptor may serve as a starting point in the design of highly effective synthetic fluorescent receptor for recognition of target protein.     

石蜡切片行免疫荧光染色后再行PAS或HE染色确定NPR-A蛋白在肾脏的定位

目的 介绍一种新方法来明确NPR-A蛋白在大鼠肾组织的定位.方法 采用肾脏石蜡切片先行NPR-A免疫荧光染色,然后再行PAS或HE染色.结果 NPR-A免疫阳性物在大鼠肾组织主要沉积于皮质的近端小管、外髓的髓袢升支粗段以及内髓集合管,直小血管、肾小球、远曲小管和细段也有一定量的表达,而皮质及外髓集合管仅有少量的表达.结论 研究采用石蜡切片先行免疫荧光染色后再行PAS或HE染色,在不用或少用特异性抗体的情况下,成功的解决了NPR-A蛋白在大鼠肾组织表达的分布位置及细胞定位的难题.