Skp2与p27蛋白的表达与肺癌的关系

目的 探讨S期蛋白激酶相关蛋白(S-phase kinase-associated protein 2,Skp2)和p27蛋白在肺癌中表达及临床意义.方法 应用免疫组织化学S-P法检测Skp2和p27在60例肺癌(鳞癌25例.腺癌20例,小细胞肺癌15例)和15例经手术证实、病理检测肺良性病变(炎性假瘤、局灶性肺脓肿、包裹性肺霉菌病、错构瘤和支气管腺瘤)正常细胞中的表达.结果 Skp2仅在肺癌组织中表达,且在吸烟者、鳞癌和低分化患者中过表达,而与患者年龄、性别、肿瘤位置和病理分期无关.p27在正常支气管上皮中均有表达,在肺癌中表达降低,但其表达仅与淋巴结转移和病理分期有关.Skp2阳性表达患者中.p27表达降低.结论 Skp2与p27蛋白的表达与肺癌的发生、发展有一定关系,在肺癌中二者存在某种相互作用机制.     

MAP-2 Expression in the Human Adenohypophysis and in Pituitary Adenomas. An Immunohistochemical Study

MAP-2, a well characterized member of the microtubule associated protein (MAP) family, binds to and stabilizes microtubules and is involved in cell proliferation as well as neuronal differentiation. The aim of the present work was to study MAP-2 expression in human adenohypophyses and pituitary adenomas. To our knowledge, data regarding MAP-2 expression in human pituitaries has not been reported to date. For immunohistochemistry, the streptavidin-biotin-peroxidase complex method was used. Nine non-tumorous adenohypophyses and 77 adenomas (GH-, PRL-, ACTH-, TSH-, FSH/LH- and/or alpha subunit- producing or immunonegative tumors) were investigated. The results show that MAP-2 is expressed in the cytoplasm of non-tumorous adenohypophysial cells as well as of various pituitary adenoma types. No significant correlation was found between MAP-2 expression and gender, patient age, mitotic activity, MIB-1 labelling indices, hormone immunoprofile, and endocrine status, ie. hormonal activity or lack thereof. Thus MAP-2 expression cannot be used to estimate cell proliferation rate, growth potential, endocrine activity or biologic behaviour of an adenoma. Immunopositivity appeared to be stronger in the cytoplasm of adenoma cells than in that of non-tumorous adenohypophysial cells, implying that the adenoma cells contain larger quantities of MAP-2. It can be concluded that the functional activity of MAP-2 is not associated with the manufacture of any specific adenohypophysial hormone(s) and is not limited to one specific cell type.     

Skp2-RNAi suppresses proliferation and migration of gallbladder carcinoma cells by enhancing p27 expression

AIM:To explore the role of S-phase kinase-associated protein-2(Skp2)in gallbladder carcinoma and to identify whether depletion of Skp2 by Skp2-RNAi could attenuate proliferation and migration of gallbladder carcinoma.METHODS:Skp2-RNAi was transduced into cells of the gallbladder carcinoma cell line GBC-SD,using a lentiviral vector.The effect of Skp2-RNAi on the proliferation,migration,invasion and cell cycle of GBC-SD cells was studied using in vitro assays for cell proliferation,colony formation,wound healing and cell cycle.The expression of Skp2 and p27 was detected by real-time polymerase chain reaction and Western immunoblotting.The effect of Skp2-RNAi on the proliferation of GBC-SD cells in vivo was investigated by tumorigenicity experiments in nude mice.RESULTS:Lentivirus-mediated RNAi reduced the expression of Skp2 in cultured cells.The expression of the p27 protein increased along with the down-regulation of Skp2,although no significant difference was found in p27 mRNA expression.Flow cytometry revealed that Skp2-RNAi transfection significantly increased the proportion of cells in the S phase and significantly decreased the proportion of cells in the G 2 /M phase.No significant difference in the frequency of cells in the G0/G1 phase was observed.The results from the cell proliferation,colony formation and wound healing assays revealed that Skp2-RNAi transfection markedly inhibited the proliferation and migration of GBC-SD cells in vitro.Additionally,tumorigenicity experiments showed that suppression of Skp2 significantly decreased the weights of the tumors(0.56 ± 0.11 and 0.55 ± 0.07 g in the control and Scr-RNAi groups vs 0.37 ± 0.09 and 0.35 ± 0.08 g in the Skp2-RNAi-L and Skp2-RNAi-H groups).CONCLUSION:The expression of Skp2 in GBC-SD cells was inhibited following Skp2-RNAi transfection.Silencing of the Skp2 gene inhibited proliferation,migration and invasiveness of GBC-SD cells by mechanisms dependent on enhanced expression of the p27 protein.     

Skp2与C-myc在食管癌中的表达及其意义

目的探讨Skp2、C-myc蛋白在食管癌发生、恶性增殖及转移扩散中的作用及两者间的相互关系。方法运用免疫组织化学EnVision法检测skp2、C-myc蛋白在97例食管癌、30例食管癌旁组织标本中的表达情况。结果Skp2与C-myc蛋白的阳性表达率在癌旁组织组（O％、0％）与食管癌组（48．45％、70．10％）、淋巴结转移阳性组（63．41％、92．68％）与淋巴结转移阴性组（37．50％、53．57％）、高分化组（29．63％、44．44％）与中分化组（53．33％、77．78％）、低分化组（60．00％、84．00％）之间差异有统计学意义（P〈0．01）,而与食管癌的肿瘤大小、患者、性别年龄和临床分期无关（P〉0．05）。两种蛋白表达之间存在交互作用。结论skp2和C-myc蛋白的表达与食管癌的分化程度及淋巴结的转移情况有关,分化程度低者或伴随有淋巴结转移的患者C-myc与skp2蛋白表达增高,且两者的表达存在交互作用。     

Rb/Cdk2/Cdk4 triple mutant mice elicit an alternative mechanism for regulation of the G1/S transition

The G₁/S-phase transition is a well-toned switch in the mammalian cell cycle. Cdk2, Cdk4, and the rate-limiting tumor suppressor retinoblastoma protein (Rb) have been studied in separate animal models, but interactions between the kinases and Rb in vivo have yet to be investigated. To further dissect the regulation of the G₁ to S-phase progression, we generated Cdk2^−/−Cdk4^−/−Rb^−/− (TKO) mutant mice. TKO mice died at midgestation with major defects in the circulatory systems and displayed combined phenotypes of Rb^−/− and Cdk2^−/−Cdk4^−/− mutants. However, TKO mouse embryonic fibroblasts were not only resistant to senescence and became immortal but displayed enhanced S-phase entry and proliferation rates similar to wild type. These effects were more remarkable in hypoxic compared with normoxic conditions. Interestingly, depletion of the pocket proteins by HPV-E7 or p107/p130 shRNA in the absence of Cdk2/Cdk4 elicited a mechanism for the G₁/S regulation with increased levels of p27^Kip1 binding to Cdk1/cyclin E complexes. Our work indicates that the G₁/S transition can be controlled in different ways depending on the situation, resembling a regulatory network.     

糖尿病大鼠肾小球周期素激酶抑制剂p27的变化及意义

目的 :探讨糖尿病肾小球周期素激酶抑制剂p2 7水平的变化及p2 7在糖尿病 (DM)肾小球细胞 (特别是系膜细胞MC)肥大中的作用。　 方法 :蛋白印迹 (Western杂交 )方法测定肾小球裂解液p2 7蛋白水平 ,RT PCR方法测定肾小球p2 7mRNA ,ELISA方法测定肾小球裂解液细胞外基质 (ECM)蛋白 (Ⅳ型胶原及纤维连接蛋白 )和尿白蛋白。　 结果 :随着DM病程的延长 ,DM大鼠肾小球p2 7水平和肾小球ECM蛋白水平逐渐增高 ,2 4h尿白蛋白排泄量逐渐增加 ,同时DM大鼠肾重 /体重呈增高趋势。DM(2 8天 )大鼠与正常大鼠间肾小球p2 7mRNA无差别。不同病程糖尿病大鼠间全血血糖浓度无明显差异。　 结论 :DM大鼠肾小球p2 7水平增高 ,p2 7水平增高可能在DM大鼠肾小球细胞肥大 (特别是MC肥大 )中起重要作用。     

曲古菌素A对血管平滑肌细胞p27kip1表达的调控机制研究

目的探讨组蛋白去乙酰化酶抑制剂曲古菌素A（Trichostatin A，TSA）对血管平滑肌细胞（VSMC）的p27kip1表达的影响和调控机制。方法半定量逆转录聚合酶链反应（RT—PCR）检测p27kip1的mRNA水平，蛋白印迹测定p27kip1和S-phase kinase—associated protein-2（skp2）蛋白表达，荧光分光光度法测定20S蛋白酶体活性。结果100ng／ml TSA不影响VSMC中p27kip1的mRNA水平。100ng／ml TSA显著抑制血清诱导的p27kip1蛋白下调，并延长p27kip1蛋白的半衰期。100ng／mlTSA抑制血清诱导的skp2表达上调，且skp2表达与相应时点p27kip1蛋白呈负相关。100ng／ml TSA对20S蛋白酶体活性物影响。结论TSA对VSMC的p27kip1表达调控不是在转录水平上，而是通过翻译后机制抑制血清诱导VSMC的p27kip1蛋白降解，其机制可能与TSA抑制泛素连接酶亚单侍skp2表达有关．     

Skp2在人类消化系统肿瘤中的研究进展

Skp2是F-box蛋白家族中的一员,对正常细胞周期起调控作用,主要通过泛素蛋白酶体途径,对细胞周期素依赖性蛋白激酶抑制物中的p27的降解调控.研究发现,Skp2与多种肿瘤的发生密切相关.此文就Skp2在消化系统肿瘤中的研究进展作一综述.     

胰腺癌组织中SKP2和P27蛋白表达及其相互关系

目的:研究胰腺导管癌和慢性胰腺炎组织中(S期激酶相关蛋白-2)SKP2和P27表达,探讨其临床病理意义及两者在胰腺导管癌中表达的相互关系．方法:51例胰腺导管癌和10例慢性胰腺炎手术切除标本常规制作石蜡包埋切片,SKP2和P27染色方法为SP免疫组化法．结果:51例胰腺导管癌SKP2阳性表达28例(54．9％)和P27阳性表达25例(49．0％),10例慢性胰腺炎SKP2阳性2例(20．0％) 和P27阳性9例(90．0％),两者之间均存在显著差异(P<0．05), 高分化腺癌(7／20,35．0％)和未转移(5／16,31.2％)病例SKP2表达阳性率明显低于低分化腺癌(14／19,73．7％)和转移(23／35, 65．7％)病例,均存在显著差异(P<0．05);高分化腺癌(13／20, 65．0％)和未转移(12／16,75．0％)病例P27表达阳性率明显高于低分化腺癌(6／19,31．5％)和转移(13／35,37．1％)病例,有显著或高度显著差异(P<0．05或P<0．01)．SKP2和P27在胰腺导管癌中表达呈密切相关(x2=14．33,P<0．01)．结论:SKP2和P27表达是反映胰腺导管癌发生,发展及预后的重要生物学标记物,SKP2阳性表达或P27阴性表达者恶性度高、易发生转移及预后不良,且两者表达存在相互调控作用．     

Overexpression of Stimulatory G Protein Alpha-subunit Is a Hallmark of Most Human Somatotrophic Pituitary Tumours and Is Associated with Resistance to GH-releasing Hormone

The heterotrimeric Gs protein–adenylyl cyclase (AC) cascade plays a pivotal role in controlling hormone secretion by endocrine glands. Consequently, deficiency of the alpha-subunit of Gs leads to endocrine hypofunction and hypoplasia in the affected cells whereas AC hyperactivity results from activating point mutations within the Gs-alpha gene. The latter, termed gsp oncogenes, are found primarily in a subset of growth hormone (GH) -secreting human pituitary tumours (somatotrophinomas) and are thus associated with excessive GH secretion. We present here evidence that another type of defect in human somatotrophinomas may be overexpression of the Gs-alpha subunit. Immunohistochemistry using an antibody against recombinant human Gs-alpha revealed high levels of expression in 25 of 39 somatotrophinomas but weak staining in normal human pituitary cells. These results were confirmed by Western blot analysis. Additionally, cholera toxin-mediated ADP-ribosylation in the presence of 32P-labelled N+ resulted in an autoradiographic signal intensity which correlated directly with magnitude of immunostaining and amount of antigen shown by Western blot analysis, providing evidence for overexpression of functionally active subunit. Finally, reconstitution assays were applied and directly demonstrated the increased activity of overexpressed Gs-alpha. In vivo, the effect of Gs-alpha on AC activity may be partially counterregulated by high levels of inhibitory G protein that also occurred in these tumours. In culture, GH-releasing hormone (GHRH) had markedly reduced effects on GH secretion by somatotrophinomas exhibiting Gs-alpha overexpression, whereas powerful stimulation occurred in weakly staining tumours. In contrast to these observations with Gs-alpha, immunostaining for the phospholipase C-coupled G11-alpha subunit was relatively weak in all somatotrophinomas studied and synthetic GH-releasing peptide, which acts via a specific G11-coupled receptor, led to powerful and consistent stimulation of GH secretion by different tumours. These results indicate that Gs-alpha overexpression is associated with dysfunction in hormone secretion by some somatotrophinomas.     

Expression of von Hippel-Lindau Protein (VHL-P) in Nontumorous and Adenomatous Human Pituitaries

We studied the presence of von Hippel-Lindau protein (VHL-P) in 7 nontumorous pituitaries and 68 pituitary adenomas by immunocytochemistry using a polyclonal antibody which detects both normal and mutated forms. In nontumorous pituitary VHL-P was variable expressed in the cytoplasm of most adenohypophysial cells. In addition, weak diffuse staining was noted in the posterior lobe. Among the 53 VHL-P immunopositive adenomas (78%), 32 showed only cytoplasmic, 7 only nuclear, and 14 both cytoplasmic and nuclear immunoreactivity. In densely (8 cases) and sparsely (7 cases) granulated somatotroph adenomas nuclear and weak cytoplasmic immunoreactivity were common; all 4 sparsely granulated lactotroph adenomas had only moderate cytoplasmic immunostaining; all 7 functioning corticotroph adenomas presented intense cytoplasmic immunoreactivity and 4/7 showing nuclear immunostaining as well; all 3 silent subtype 1 adenomas were negative; 3/4 silent subtype 2 and 4/7 silent subtype 3 adenomas exhibited only cytoplasmic immunoreactivity; thyrotroph adenomas showed variable patterns from negative in 1/7, to only cytoplasmic in 2/7, nuclear alone in 2/7, or both in 2/7. Only weak, focal cytoplasmic immunoreactivity was noted in gonadotroph adenomas of female (2/4) and male 1/5 type. Lastly, all 4 null cell and 8 oncocytic adenomas showed moderate to intense cytoplasmic immunoreactivity. It can be concluded that the majority of pituitary adenomas express the VHL-P with variable distributions and intensity in different tumor types. The frequent localization of VHL-P in the nuclei of somatotroph adenomas, the least vascularized tumor type, suggests a possible inhibitory role of VHL-P in pituitary angiogenesis.     

Correlation of Bcl-2 and Bax with Apoptosis in Human Pituitary Adenomas

Bcl-2 oncogene and Bax gene play an important role in regulating apoptosis. In the present study, the expression of bcl-2 and bax was investigated and correlated with apoptosis in a series of 81 pituitary adenomas. Bcl-2 and bax proteins were localized by immunohistochemistry and the histoscore (HSC) was assessed by multiplying the immunohistostaining grade (1 to 4) by the staining intensity grade (1 to 3). According to bcl-2/bax HSC the tumors were separated in group A when > or = 1 and group B when < 1. The apoptotic labeling index (ALI) was accessed by the in situ end-labeling (ISEL) technique. Bcl-2 protein was equally detected in functioning and nonfunctioning adenomas with statistically significant higher HSC in nonfunctioning tumors (P < 0.03). Bax protein was immunopositive in the substantial majority of adenomas with significantly higher HSC in functioning as compared to nonfunctioning adenomas (P < 0.0009). The ALI was significantly higher in functioning adenomas as compared to nonfunctioning adenomas (P < 0.04). In addition, ALI was significantly higher in group B than in group A (P < 0.004) and it was correlated with bax HSC (P < 0.004). Finally, the group B of bcl-2/bax significantly predominated in nonfunctioning tumors (P < 0.0009) and in microadenomas (P = 0.05), as compared with functioning adenomas and macroadenomas respectively. In conclusion, our findings suggest that bcl-2 and bax molecules play a role in the regulation of apoptotic mechanisms in pituitary adenomas.     

SKP2在大肠癌组织中的表达及预后意义

目的:探讨SKP2在大肠癌中的表达和预后作用．方法:采用SP免疫组化法检测68例大肠癌手术切除组织标本中SKP2和P27的表达,用Kaplan-Meier和Cox回归分析法进行生存分析．结果:在68例大肠癌组织中,SKP2和P27的阳性表达率分别为41．2％(n=28)和52．9％(n= 36)．SKP2的表达与组织分级显著相关(X2= 14．073,P=0．001)．SKP2表达与年龄、性别和AJCC分期无关(P>0．05)．SKP2和P27负相关(r=-0．528,P=0．0001)．SKP2高表达组的总生存期较SKP2低表达组短(31．5±4．0 mo vs 54．5±2．1 mo,P<0．01)．多因素Cox回归分析表明,SKP2表达是大肠癌的独立预后因素(RR= 6．227．P=0．033)．结论:SKP2表达可以作为大肠癌患者预后的指标．     

Serial MR Intensity Changes of the Posterior Pituitary in Patients with Diabetes Insipidus After Transsphenoidal Surgery for Pituitary Adenomas: Report of Two Cases

This is the first report describing magnetic resonance (MR) intensity changes of the posterior pituitary gland in the patients suffering from the classical "triphasic" diabetes insipidus (DI) after transsphenoidal surgery for pituitary adenomas. A 21-year-old female and a 54-year-old female were admitted to our hospital with the diagnosis of Cushing's disease and acromegaly due to pituitary microadenomas, respectively. No evidence of DI was found, and T1-weighted MR images exhibited "bright spot" corresponding to the posterior pituitary in both cases. Both experienced the classical "triphasic" pattern of water metabolism disturbance after successful transsphenoidal resection of pituitary adenomas, that is, polyuria-oliguria-polyuria. The MR signal hyperintensity in posterior pituitary was detected during the first polyuric phase, but the hyperintensity disappeared during the second polyuric phase. In addition, "bright spot" was restored along with the recovery from DI in the chronic phase. These findings of serial MR images supported that the first DI phase of the classical triphasic course of water metabolism disturbance was caused by secretional dysfunction of stored vasopressin from the posterior gland, whereas the second DI phase was due to impairment in the functional integrity producing vasopressin-containing granules after depletion of vasopressin in the oliguric phase.     

Relation of overexpression of S phase kinase-associated protein 2 with reduced expression of p27 and PTEN in human gastric carcinoma

AIM: To investigate the significance of S phase kinase associated protein 2 (Skp2) expression in human gastric carcinoma and the relation between expressions of Skp2, p27 and PTEN. METHODS: Immunohistochemical analysis was performed on 138 gastric carcinoma specimens, their paired adjacent mucosa specimens, 102 paired lymphatic metastatic carcinoma tissue specimens, 30 dysplasia specimens, 30 intestinal metaplasia specimens, 10 chronic superficial gastritis specimens and 5 normal gastric mucosa specimens for Skp2 expression and on 138 gastric carcinoma specimens for p27 and PTEN expression. RESULTS: Skp2 labeling frequency was significantly higher in intestinal metaplasia (12.68±0.86) and adjacent mucosa (19.32±1.22) than in normal gastric mucosa (0.53±0.13) and chronic superficial gastritis (0.47±0.19) (P = 0.000); in dysplasia (16.74±0.82) than in intestinal metaplasia (P = 0.000); in gastric primary carcinoma (31.34±2.17) than in dysplasia and adjacent mucosa (P = 0.000); in metastasis gastric carcinoma in lymph nodes (39.76±2.00) than in primary gastric carcinoma (P = 0.037), respectively. Skp2 labeling frequency was positively associated with differentiation degree (rho = 0.315, P = 0.000), vessel invasion (rho = 0.303, P = 0.000) and lymph node metastasis (rho = 0.254, P = 0.000) of gastric cancer. Expression of Skp2 was negatively associated with p27 (rho = -0.451, P = 0.000) and PTEN (rho = -0.480, P = 0.000) expression in gastric carcinoma. p27 expression was positively associated with PTEN expression in gastric carcinoma (rho = 0.642, P = 0.000). CONCLUSION: Skp2 overexpression may be involved in carcinogenesis and progression of human gastric carcinoma in vivo, possibly via p27 proteolysis. PTEN may regulate the expression of p27 by negatively regulating Skp2 expression.     

吲哚美辛对胃癌SGC 7901细胞增殖及Cyclin D1蛋白表达的影响

目的:观察引哚美辛对胃癌SGC 7901细胞增殖及细胞周期调控蛋白Cyclin D1表达的影响,探讨吲哚美辛抑制细胞增殖的机制.方法:采用MTT法观察吲哚美辛对胃癌细胞SGC 7901增殖的影响,采用流式细胞仪观察细胞周期分布的变化,采用免疫细胞化学方法检测Cyclin D1蛋白的表达.结果:吲哚美辛呈时间、浓度依赖方式抑制胃癌SGC 7901细胞增殖,改变细胞周期的分布,增加G0/G1期细胞的比例,下调Cyclin D1蛋白的表达.结论:吲哚美辛可能通过下调Cyclin D1蛋白表达,影响细胞周期的分布来抑制胃癌SGC7901细胞增殖.     

p27蛋白在肺腺癌中的表达及临床意义

目的 探讨p27蛋白在肺腺癌的表达及临床意义.方法 应用免疫组化技术检测44例肺腺癌患者的p27蛋白表达,分析p27蛋白表达与性别、年龄、肿瘤大小、临床分期、分化程度和淋巴结转移以及预后的关系.结果 p27蛋白水平与肺腺癌的分化程度相关,与患者的年龄、性别、肿瘤大小、淋巴结转移、TMN分期无关.p27蛋白低表达者的平均生存期（18.6个月）明显低于p27蛋白高表达者（34.5个月）.结论 p27蛋白表达可以作为评价肺腺癌恶性程度和预后的一个重要指标.