膀胱尿路上皮癌MMP-2表达及其与FAK、p53、bcl-2、Ki-67的关系

目的 比较不同分化和浸润程度膀胱尿路上皮癌MMP—2表达及其与FAK、p53、bcl—2和Ki—67的关系。方法 采用免疫组化EnVision法对83例膀胱尿路上皮癌和68例非肿瘤尿路上皮，进行MMP—2、FAK、p53、bcl—2和Ki—67的表达检测。结果 MMP—2在肿瘤组织中的表达显著高于非肿瘤性移行上皮，其表达强度随肿瘤分化程度降低和浸润深度增加而显著增强。分化差和浸润性膀胱癌中FAK和p53表达增强，并与MMP—2表达呈正相关；分化差膀胱癌中Ki—67表达增强而bcl—2表达丢失，Ki—67与MMP—2表达呈正相关。结论 在膀胱尿路上皮癌的进展和分化过程中，肿瘤分化程度越低，肿瘤细胞分泌MMP—2越多，其浸润和转移的能力也越强。FAK、p53、Ki—67和bcl—2不但与肿瘤细胞的生长密切相关，还可能直接或间接地参与了MMP—2的调控。     

Tumor specific activation of the VEGF/KDR angiogenic pathway in a subset of locally advanced squamous cell head and neck carcinomas

Vascular endothelial growth factor (VEGF) and its receptors, Flt-1 and flk-1(KDR), constitute an important angiogenic pathway which, under hypoxic conditions, is up-regulated in many solid tumours. We used the monoclonal antibody 11B5, specific for recognizing VEGF expression and the `VEGF/flk-1(KDR) complex' on tumour endothelium, to assess free VEGF protein expression and VEGF/receptor activated microvessel density (aMVD) in a series of 104 inoperable locally advanced squamous cell carcinomas of the head and neck, treated with chemo-radiotherapy. High VEGF expression in cancer cells was strongly associated with high VEGF/receptor expression in the vasculature. The high VEGF expression and the aMVD were not associated with the standard microvessel density (sMVD), as assessed with the monoclonal antibody anti-CD31 and, were not detected in normal tissue. An increased sMVD, however, was significantly related with the expression thymidine phosphorylase (TP), and also with the nuclear accumulation of the oncoprotein p53, but neither p53 nor TP was associated with VEGF expression by cancer cells or VEGF/receptor complex aMVD. In 35% of cancer cases examined, more than 20% of the microvessels assessed with anti-CD31 also expressed the VEGF/KDR complex. The vasculature of the normal head and neck mucosa did not express the VEGF/KDR complex. There was no association between VEGF expression or VEGF/receptor complex aMVD and response to chemo-radiotherapy or patient's survival. It is concluded that activation of the angiogenic pathway VEGF/flk-1(KDR) is tumor specific in a subgroup of locally advanced squamous cell carcinomas of the head and neck. Selective destruction of this type of vasculature, using immunoconjugates directed against the VEGF/receptor complex, may prove therapeutically useful for patients with a high tumoral VEGF/flk-1(KDR) activated microvessel fraction. This revised version was published online in July 2006 with corrections to the Cover Date.     

Expression of the vascular endothelial growth factor receptor-2/Flk-1 in breast carcinomas: correlation with proliferation

Vascular endothelial growth factor (VEGF) and its receptor Flk-1/KDR play an important role in vascular permeability and tumor angiogenesis. Prompted by the hypothesis that VEGF/Flk-1 system may have regulatory roles in breast carcinogenesis, we investigated the expression of Flk-1 in 141 invasive breast carcinomas in correlation with clinical and immunohistochemical prognostic parameters, including proliferation indices like Ki-67 and Topoisomerase IIalpha (Topo-IIalpha). The immunohistochemical avidin-biotin-peroxidase method was performed on paraffin sections for the detection of Flk-1, p53, Bcl-2, c-erbB-2, Ki-67, Topo-IIalpha, ER, and PR. Flk-1 was detected in 91 of 141 (64.5%) of invasive breast carcinomas showing a widespread cytoplasmic expression in most of the neoplastic cells. Flk-1 expression was correlated with the menopausal status (P = 0.051) of the patient and the nuclear grade of the invasive breast carcinoma (P = 0.003), but demonstrated no correlation with histologic grade, stage, and patient survival. It is interesting that Flk-1 expression demonstrated a significant correlation with 2 well-established proliferation indices, Ki-67 (P = 0.037) and topo-IIalpha (P = 0.009), whereas there was no correlation with the expression of ER, PR, p53, Bcl-2, and c-erbB-2. Moreover, Flk-1 expression showed an inverse correlation with TIMP-1 mRNA localization in intratumoral stromal cells (P = 0.013). In conclusion, the significant correlation of Flk-1 expression in invasive breast carcinomas with proliferation indices like Ki-67 and topo-IIalpha suggests that VEGF may exert a growth factor activity on mammary cancer cells through its receptor Flk-1. On the other hand, the inverse correlation of Flk-1 with TIMP-1 mRNA in intratumoral stromal cells supports the notion that TIMP-1 may have an inhibitory role on angiogenesis.     

Survivin nuclear labeling index: a superior biomarker in superficial urothelial carcinoma of human urinary bladder.

The caspase family proteases are key proapoptotic proteins while the inhibitor of apoptosis proteins (IAP) prevent apoptosis by antagonizing the caspases or other key proapoptotic proteins. Limited studies of IAPs suggested their deregulation contributed to urothelial neoplasia. However, the expression status and biologic or prognostic significance of the caspase and IAP family proteins in urothelial neoplasms is not clear. In the present study, we first systematically evaluated the expression profile of the major apoptosis regulators, including caspases (CASP3, 6, 7, 8, 9, 10, and 14), IAPs (survivin/BIRC5, CIAP1, CIAP2, XIAP, and LIVIN), APAF1, SMAC, and BCL2, as well as proliferation markers Ki67 and PHH3, in Ta/T1 human urinary bladder urothelial carcinomas and normal urothelium samples by immunohistochemistry. The analysis showed that survivin/BIRC5 nuclear labeling index (BIRC5-N), but not cytoplasmic staining, was the only apoptotic marker which correlated significantly with tumor grade, stage, and patient outcome. We further analyzed the prognostic value of BIRC5-N in 101 Ta/T1 urinary bladder urothelial carcinomas by univariate analysis, which showed that BIRC5-N as well as the more classical prognosticators (stage, grade, and Ki67 index) were of prognostic significance. However, multivariate analysis by Cox proportional hazard regression demonstrated BIRC5-N was a stronger prognosticator than tumor grade, stage, and Ki67 labeling index. BIRC5-N index of 8% or more predicted unfavorable disease-specific survival (relative risk (RR)=6.6, 95% confidence interval=1.6-26.7, P=0.0080) as well as progression-free survival (RR=4.4, 95% confidence interval=1.3-14.6, P=0.0151). We conclude that BIRC5-N is a superior biologic and prognostic marker for Ta/T1 urothelial carcinomas of urinary bladder.     

Proliferation markers and DNA content analysis in urinary bladder TaT1 urothelial cell carcinomas: identification of subgroups with low and high stage progression risks

AIMS: To evaluate whether in situ biomarkers Ki67, mitotic activity index (MAI), p53, mean area of the 10 largest nuclei (MNA10), and whole genome DNA ploidy by flow and image cytometry (FCM and ICM, respectively) have independent prognostic value in urinary bladder urothelial cell carcinomas (UCs). METHODS: Ki67 and p53 immunoquantitation was performed in TaT1 consensus diagnosis UCs. MAI and MNA10 were also determined. Single cell suspensions were stained (DAPI for FCM; Feulgen for ICM). There was enough material for all measurements in 171 cases. Kaplan-Meier curves and multivariate survival analysis (Cox) were used to assess the prognostic value of all features (including classic clinicopathological risk factors, such as stage, grade, multicentricity, carcinoma in situ). RESULTS: Thirteen (7.6%) patients progressed. Of the classic factors, grade was strongly prognostic in univariate analysis, as were all the biomarkers. In multivariate analysis, the strongest independent combinations for progression were MNA10 (threshold (T) = 170.0 micro m(2)) plus MAI (T = 30), or MNA10 (T = 170.0 micro m(2)) plus Ki67(T = 25.0%). p53 (T = 35.2%) plus Ki67 (T = 25.0%) also predicted progression well, with high hazard ratios, but p53 measurements were not as reproducible as the other features. The prognostic value of the quantitative biomarkers exceeded that of the classic risk factors and DNA ploidy. The sensitivity, specificity, positive, and negative predictive values of MNA10/MAI or MNA10/Ki67 at the thresholds mentioned were 100%, 79%, 57%, and 100%, respectively. These feature combinations were also strongest prognostically in the high risk treatment subgroup. CONCLUSIONS: The combined biomarkers MNA10/Ki67 or MNA10/MAI are more accurate and reproducible predictors of stage progression in TaT1 UCs than classic prognostic risk factors and DNA ploidy.     

Prognostic impact of histological grade and vascular invasion compared with tumour cell proliferation in endometrial carcinoma of endometrioid type

AIMS: The relative impact of different prognostic factors is important for endometrial carcinoma patients. The aim of our study was to examine the combined value of histological grade [International Federation of Gynaecology and Obstetrics (FIGO)] and vascular invasion in comparison with tumour cell proliferation assessed by mitotic count and Ki67. The recently proposed binary architectural grade was also evaluated, in addition to age, depth of myometrial infiltration and FIGO stage in our population-based series of 237 endometrioid carcinomas. METHODS AND RESULTS: The tumours were studied for several histological features, including FIGO grade, binary grade, vascular invasion, mitotic count, myometrial invasion and expression of Ki67. FIGO grade was significantly associated with all investigated histological features, including Ki67 expression. Vascular invasion was significantly more frequent in FIGO grade 3 tumours, and was associated with a diffusely infiltrative growth pattern, solid growth, necrosis and deep myometrial invasion. All variables showed a highly significant relationship with patient survival in univariate analysis. In multivariate models, FIGO grade, vascular invasion, and proliferation assessed by Ki67 expression all had independent prognostic influence in this population-based study. Comparing tumour cell proliferation (Ki67) with vascular invasion as a marker of metastatic spread, the latter had a stronger survival impact. CONCLUSIONS: Vascular invasion and tumour cell proliferation measured by Ki67 both had independent prognostic influence, and should be considered to identify aggressive tumours of the endometrioid subtype.     

Expression of cyclin D1, Ki-67 and PCNA in non-small cell lung cancer: prognostic significance and comparison with p53 and bcl-2

Uncontrolled cell proliferation is the hallmark of malignant tumours. Thus, the proliferative potential of tumour cells is an important prognostic factor. However, evaluation of the prognostic significance of the expression of proteins involved in regulation of cell proliferation remains controversial. In the present study, expression of Ki-67, PCNA and cyclin D1 was estimated in a group of 89 surgically resected non-small cell lung carcinomas using immunohistochemistry. The results were compared with expression of bcl-2 and p53 and with clinicopathological parameters including patients' survival. Ki-67 and PCNA were found to be moderately and highly expressed in 39% and 44% of the tumours, respectively. There was a strong correlation between Ki67 and PCNA expression. Forty five of 88 tumours (51%) showed overexpression of cyclin D1. Surprisingly, cyclin D1 was mainly localized in the cytoplasm and only a small group of tumours (9/88, 10%) showed nuclear staining as well. Bcl-2 and p53 expression was observed in 69% and 30% of the tumours, respectively. All these markers were found to be independent of clinicopathological parameters, except for Ki-67 and bcl-2 expression, which was associated with squamous cell carcinomas. It is concluded that none of the markers that were studied can be used as an independent prognostic factor, whereas the following combinations of markers may have favourable prognostic value: p53 positivity and low Ki-67 expression, p53 positivity and lack of cyclin D1 expression, bcl-2 positivity and low Ki-67 expression, and lack of cyclin D1 expression and low Ki-67 expression.     

Ki67 immunohistochemistry: a valuable marker in prognostication but with a risk of misclassification: proliferation subgroups formed based on Ki67 immunoreactivity and standardized mitotic index

AIMS : Counting mitotic figures is considered to be a reliable prognosticator, but evaluation of Ki67 immunohistochemistry has become more popular in evaluating proliferation. Our previous studies suggested an occasional discrepancy between mitotic figures and Ki67 fraction. The aim of this study was to investigate this more closely and also to study the associations between bcl-2 and p53 expression and proliferation. METHODS AND RESULTS: Two hundred and sixty-five infiltrating breast carcinomas were immunostained for Ki67, p53 and bcl-2. The standardized mitotic index (SMI) was determined. Four proliferation groups were based on Ki67 positivity fraction and SMI at optimal cut-off points. Cox's multivariate model was used to test the power of the prognosticators. SMI and nodal status were the most powerful individual prognosticators. Ki67 was an independent prognosticator if nodal status, tumour size, age and histological grade were included in the analysis but not if analysed with SMI. The group with low SMI and low Ki67 fraction had the best prognosis. Groups with high SMI had the poorest prognosis. The group with low SMI and high Ki67 fraction had a favourable prognosis. Bcl-2 negativity and p53 positivity correlated with proliferation. CONCLUSIONS: We have found a 'wrong positive' Ki67 group with favourable prognosis. SMI cannot be replaced by Ki67 because of the danger of misclassification of some patients.     

Analysis of Vascular Endothelial Growth Factor (VEGF) and a receptor subtype (KDR/flk-1) in the liver of rats exposed to riddelliine: a potential role in the development of hemangiosarcoma

Riddelliine alters hepatocellular and endothelial cell kinetics and function including stimulating an increase in hepatocytic vascular endothelial growth factor (VEGF) in the absence of increased serological levels of VEGF (Nyska etal. 2002). The objective of this study was to further assess hepatic VEGF and KDR/flk-1 synthesis and expression by hepatic cells under riddelliine treatment conditions. Forty-two male F344/N rats were dosed by gavage with riddelliine (0, 1.0, and 2.5 mg/kg/day) for 6 weeks. Seven animals/group were sacrificed after 8 consecutive daily doses; remaining rats were terminated after 30 daily doses, excluding weekends. Hepatic tissues were evaluated by immunohistochemistry and in situ hybridization. The results showed that VEGF mRNA expression was observed in control and treated animals; however, qualitative differences were noted. Treated animals exhibited VEGF mRNA in clustered, focal hepatocytes and bile duct epithelium, whereas VEGF mRNA in hepatocytes from vehicle control rats was distributed evenly across all hepatocytes. Results evaluating the distribution of the VEGF cognate receptor, KDR/flk-1 showed that randomly distributed, rare sinusoidal endothelium, including those demonstrating karyomegaly and cytomegaly expressed KDR/flk-1. Phosphorylation of KDR/flk-1 at pTyr996 and pTyr1054/1059, but not pTyr951, was also detected, evidence that endothelial cell KDR/flk-1 was activated. These results suggest that both hepatocytes and endothelial cells are targets of riddelliine-induced injury. We speculate that damage to both populations of cells may lead to dysregulated VEGF synthesis by hepatocytes and activation of KDR/flk-1 by endothelium leading to the induction of sustained endothelial cell proliferation, culminating in the development of hepatic hemangiosarcoma.     

胰岛素对牛胸主动脉内皮细胞血管内皮生长因子及其受体表达的影响

目的:评价胰岛素对培养的牛胸主动脉内皮细胞血管内皮生长因子(VEGF)及其受体表达的影响。方法: 取新生的小牛胸主动脉,做血管内皮细胞原代及传代培养,取4-6代培养细胞分组,应用不同浓度的胰岛素(30 mU/L、300 mU/L、3 000 mU/L)干预培养过程,48 h后应用免疫组化法测定内皮细胞VEGF及其受体(flt-1、flk-1/KDR)的表达水平。结果: 低浓度胰岛素组(30 mU/L、300 mU/L)内皮细胞VEGF表达明显高于不用胰岛素组(P<0.01);高浓度组(3 000 mU/L)内皮细胞VEGF表达明显低于不用胰岛素组(P＜0.05);各组内皮细胞VEGF受体(flt-1及flk-1/KDR)的表达无明显差异(P＞0.05)。结论: 低浓度胰岛素促进小牛主动脉血管内皮细胞VEGF表达;高浓度胰岛素可抑制血管内皮细胞VEGF表达;胰岛素对小牛主动脉血管内皮细胞VEGF受体(flt-1、flk-1/KDR)的表达无直接影响。     

Expression of p21 WAF1/CIP1 protein in colorectal cancers: study of its relation to p53 mutation and Ki67 antigen expression

Mutations of the p53 gene are the most common genetic alteration in malignant human tumors. A cyclin-dependent kinase inhibitor, p21WAF1/CIP1, is thought to be an important mediator of p53-induced cell cycle arrest. Although numerous studies have reported p53 expression and mutation in colorectal cancer few of them have correlated p53 expression with that of its downstream effector p21 and with the proliferation index as measured by expression of the Ki67 nuclear antigen. We studied p53, p21 and Ki67 expression by immunohistochemistry and molecular biology in 35 colorectal carcinomas. We compared these findings with each other and with clinical factors. Sixty three percent of tumors expressed p53 whereas seventy one percent expressed p21WAF1/CIP1. In adenocarcinomas, p21 staining was heterogeneous: p21-reactive cells were seen in the most differentiated areas. There was no correlation between p21WAF1/CIP1 and p53 expression, p53 mutation, Ki67 expression or clinical factors such as sex or location of the tumor. On the other hand, there was a statistical relationship between p21 expression and survival: our results indicated an association between high p21 expression and lower stages p21WAF1/CIP1 appears to be induced independently of p53 in these tumors and may be associated with differentiation rather than proliferation.     

Micropapillary lung adenocarcinoma: a distinctive histologic subtype with prognostic significance. Case series

The aims of the present work were to evaluate the prognostic significance of the micropapillary pattern of lung adenocarcinoma and determine whether there are differences in the behavior of this type of tumor according to its immunohistochemical profile. A series of 191 consecutively resected pulmonary adenocarcinomas were divided into those with (n = 62) and those without (n = 129) micropapillary components. The disease was stage I in 38 and 54 patients, respectively. The 5-year survival rates of patients with and without micropapillary components were 54% and 77%, respectively (log rank P = .03). In multivariate survival analysis, the micropapillary component proved to be an independent prognostic factor (hazard ratio, 3.2). Five autopsy cases were used to investigate the immunohistochemical profile. The percentages of cases positive for various markers were 56.7 for p53, 94 for Ki67, 85.1 for c-myc, 2.9 for Bcl-2, 35.8 for epidermal growth factor receptor, 43.3 for cyclin D1, and 46.3 for Bax. The prognostic value was evaluated according to the expression of the different markers in micropapillary carcinomas in stage I. In univariate analysis, only cyclin D1 expression and Bax expression were associated with significantly worse survival (log rank P = .03 and P = .02, respectively). We conclude that it is important to recognize the micropapillary growth pattern in lung adenocarcinoma. Moreover, cyclin D1 and Bax seem to be markers of a poor prognosis.     

Evaluation of DNA topoisomerase IIalpha expression provides independent prognostic information in non-Hodgkin's lymphomas

AIMS: In view of the dual role that DNA topoisomerase IIa (TopoIIa) plays as a cell proliferation marker and as a possible indicator of chemosensitivity, we investigated its expression in non-Hodgkin's lymphomas (NHL) in relation to conventional clinicopathological parameters, cell proliferation (as defined by Ki67 immunoreactivity), response to therapy and patient outcome. METHODS and RESULTS: Formalin-fixed paraffin-embedded tissues from 153 patients with NHL were immunohistochemically stained for TopoIIalpha. Patients were followed up until death (n = 63) or for an average of 68 months (median 64 months, n = 90). The percentage of TopoIIalpha positive cells (TopoIIalpha LI) increased with grade (P < 0.001), extranodal location (P = 0.05) and Ki67 LI (P = 0.01, r = 0.673). In most cases (58%), Ki67 LI exceeded TopoIIalpha LI (TopoIIalpha/Ki67 < 1), especially within the indolent group (P < 0.001). TopoIIalphaLI, Ki67LI and TopoIIalpha/Ki67 ratio were all adversely related to overall survival in univariate analysis, though their significance was not maintained after adjustment for grade. In multivariate analysis high TopoIIalpha/Ki67 ratio and high TopoIIalpha LI independently predicted shortened overall and post-relapse survival, respectively. Most importantly, low TopoIIalpha/Ki67 ratio was the only independent predictor of diminished disease-free survival. However, there was no relationship between TopoIIalpha expression and response. CONCLUSIONS: Our results suggest that evaluation of TopoIIalpha expression and TopoIIalpha/Ki67 ratio as cell proliferation markers provides independent prognostic information in relation to post-relapse and overall survival. Furthermore, TopoIIalpha/Ki67 ratio appears to play a key role in the identification of patients prone to early relapse.     

Microsatellite abnormalities and somatic down-regulation of mismatch repair characterize nodular-trabecular muscle-invasive urothelial carcinoma of the bladder

AIMS: To correlate histological infiltration patterns with genetic and mismatch repair (MMR) profiles in muscle-invasive bladder urothelial carcinomas (UroC). METHODS AND RESULTS: Infiltration patterns were assessed in the deep compartment of muscle-invasive UroC (nodular-trabecular, 45 cases; infiltrative, 27 cases). Tumour compartment (superficial and deep to muscularis mucosa) analysis included: microsatellite pattern of TP53, RB1, WT1 and NF1 by polymerase chain reaction/denaturing gradient gel electrophoresis; mitotic, Ki67, in situ end labelling (ISEL) indices and DNA ploidy. MMR was assessed by MLH1 and MSH2 sequencing and immunohistochemistry in UroC with two or more abnormal microsatellite loci. Statistical differences were tested using anova and Fisher's exact tests. Infiltrative UroC showed lower Ki67 index 14.94 +/- 4.28, ISEL index 14.1 +/- 10.0 and shorter median survival (20 months) than nodular-trabecular UroC (Ki67 index 20.65 +/- 4.94, ISEL 20.2 +/- 22.7, 37-month survival, respectively). The genetic profile was significantly different for RB1 (P = 0.0003) and NF1 (P = 0.0023) only, being more frequently abnormal in nodular-trabecular UroC. A significant decrease in MLH1 or MSH2 protein expression with no gene mutations was identified in UroC with microsatellite abnormalities and a nodular-trabecular growth pattern. CONCLUSIONS: Somatic down-regulation of MMR proteins in nodular-trabecular muscle-invasive UroC results in RB1/NF1 microsatellite abnormalities, correlating with higher cellular turnover and longer survival.     

Prognostic and biological significance of cytokeratin 19 in pancreatic endocrine tumours

AIM: The prognosis of well-differentiated pancreatic endocrine tumours (PETs) is difficult to establish on a histological basis. The expression of cytokeratin (CK19) has recently been proposed as an indicator of unfavourable outcome. However, this finding still needs to be verified and to be compared with more frequently used prognosticators such as proliferative indices, vascular and/or perineural invasion. The aim of this study was to evaluate the prognostic value of CK19 expression in PETs. METHODS AND RESULTS: One hundred and forty-five PETs were studied using two different anti-CK19 monoclonal antibodies (BA17 and RCK108). The results were statistically compared with proliferation markers, vascular and perineural invasion and the presence of metastases. On univariate analysis, CK19 immunoreactivity correlated with prognosis only when it was detected with the RCK108 antibody and only in the whole group of PETs and in insulinomas. Conversely, it did not predict survival in non-functioning neoplasms. Ki67 index, mitotic count, vascular and perineural invasion were all statistically correlated with prognosis. On multivariate analysis, only the Ki67 index and metastases were independent prognosticators. CONCLUSIONS: CK19 expression correlates with patient survival only when detected with the RCK108 antibody and mainly in insulinomas. Ki67 index and metastases represent the only two independent predictors of survival.     

Molecular pathology of non-invasive urothelial carcinomas (part I)

An international consultation on the diagnosis of non-invasive urothelial neoplasms was held in Ancona, Italy in May 2001. Besides histology and problems of classification, one group of experts (Committee no. 3) discussed the molecular pathology and cytometry of non-invasive urothelial carcinomas. In the following first part, special immunohistochemical and molecular markers for stratifications in bladder cancer were discussed including different cytokeratins (clone 34betaE12, CK 20), cell proliferation markers (Ki67/MIB-1, PCNA, AgNOR, DNA-cytometry), tumor suppressor genes and oncogenes (p53, p21, erb-B2, bcl-2), different receptor expressions of epidermal growth factor and vascular endothelial growth factor and others. These molecular markers were analyzed in diagnosis of urothelial carcinomas, recurrences, progression and response to treatment.     

Membranous and cytoplasmic staining of Ki67 is associated with HER2 and ER status in invasive breast carcinoma

Aims:  Membranous and cytoplasmic Ki67 immunoreactivity has recently been observed in a number of histopathological entities, but frequency of occurrence and relationship to prognosis in more common cancers have not been described. The aim was to describe the pattern and frequency of membranous/cytoplasmic Ki67 in a cohort of invasive breast carcinomas, and their associations with grade, HER2 amplification and oestrogen receptor (ER) expression.
Methods and results:  Three hundred and twenty-two cases of invasive ductal carcinoma were assessed for histological grade, Ki67 (MIB-1 clone) proliferation index and pattern of immunoreactivity, ER expression by immunohistochemistry, and HER2 amplification status by fluorescence in situ hybridization. Overall, 26/322 (8%) breast carcinomas showed membranous/cytoplasmic Ki67, and expression was significantly associated with grade 3, HER2-amplified and ER− tumours. Membranous/cytoplasmic Ki67 was not, however, an independent prognostic factor on multivariate analysis.
Conclusions:  Membranous/cytoplasmic Ki67 identifies a group of breast carcinomas that may be important to consider separately in prognostic and predictive studies. The mechanism of subcellular Ki67 relocalization remains elusive and further studies are required to establish both the cause and effect of this unusual pattern of Ki67 immunoreactivity.     

Enhanced cell kinetics, p53 accumulation and high p21WAF1 expression in chronic cholecystitis: comparison with background mucosa of gallbladder carcinomas

AIMS: Since neoplasia resulting from chronic inflammation has recently attracted increasing attention, we have investigated surgically removed gallbladders to examine the relationship between chronic cholecystitis and carcinogenesis. METHODS AND RESULTS: The mucosa of 108 cholecystectomy specimens without gallbladder cancer and 54 surgically resected gallbladder carcinomas were classified into three groups according to the degree of lymphocytic infiltration, and assessed immunohistochemically for Ki67, p53, p21WAF1 and apoptosis. In gallbladder mucosa without carcinoma, all four parameters tended to increase with the inflammation score (IS). Significantly positive correlations were revealed between Ki67 and p53, Ki67 and p21WAF1, and p53 and p21WAF1. However, in gallbladder carcinoma cases, values of p53 and p21WAF1 for background mucosa were elevated as compared to the mucosa of cholecystitis with low IS, but there was no correlation between their expression and IS, except for Ki67. CONCLUSIONS: Severe chronic cholecystitis is associated with acceleration of epithelial cell turnover, damaged cells being eliminated by apoptosis. The background mucosa of gallbladder carcinomas showed similar cell proliferative activity (Ki67) to that in cholecystitis, with no parallel changes of p53 and p21WAF1 expression, suggesting the possibility of unknown cofactors causing genomic damage.     

The angiogenic pathway "vascular endothelial growth factor/flk-1(KDR)-receptor" in rheumatoid arthritis and osteoarthritis

Active angiogenesis, together with an up-regulation of angiogenic factors, is evident in the synovium of both rheumatoid arthritis (RA) and osteoarthritis (OA). The present study assessed, by immunohistochemistry, the microvessel density in the synovium of these arthritides and in normal controls, in relation to the expression of the angiogenic factors vascular endothelial growth factor (VEGF) and platelet-derived endothelial cell growth factor (PD-ECGF) and the apoptosis-related proteins bcl-2 and p53. More importantly, using the novel 11B5 MAb, the activated "VEGF/flk-1(KDR)-receptor" microvessel density was assessed. VEGF expression in fibroblasts was diffuse in both RA and OA. Diffuse PD-ECGF expression of fibroblasts was noted in all cases of RA, while fibroblast reactivity was focal in the OA material. The standard microvessel density (sMVD), as assessed with the anti-CD31 monoclonal antibody (MAb), was higher in RA (64+/-12) and in OA (65+/-16) than in normal tissues (52+/-8; p=0.008 and 0.0004, respectively). The activated microvessel density (aMVD), assessed with the 11B5 MAb, was significantly higher in RA (29+/-10) than in OA (17+/-4; p<0.0001) and than in normal tissues (14+/-2; p<0.0001). The "activation ratio" (aMVD/sMVD) was statistically higher in RA (0.46+/-0.17) than in OA and normal synovial tissues, the latter two having a similar ratio (0.28+/-0.08 and 0.26+/-0.03, respectively). Cytoplasmic bcl-2 expression was frequent in the synovial cells of OA, but rare in RA. Nuclear p53 protein accumulation was never observed. It is suggested that the angiogenic pathway VEGF/flk-1(KDR) may play an important role in the pathogenesis of RA and OA. Thus, failure of VEGF/flk-1(KDR) activation, in the presence of increased VEGF expression, may indicate a synovium with an impaired capacity to establish a viable vasculature, consistent with the degenerative nature of OA. On the other hand, the activated angiogenesis in RA shows a functional, still pathologically up-regulated VEGF/flk-1(KDR) pathway. Whether restoration of an impaired VEGF/flk-1(KDR) pathway in OA, or inhibition of this in RA, would prove of therapeutic importance requires further investigation.     

Minichromosome maintenance proteins 2 and 5 expression in muscle-invasive urothelial cancer: a multivariate survival study including proliferation markers and cell cycle regulators

Evaluation of cell cycle regulators has gained special interest in the effort to increase the amount of prognostic information in malignant tumors. Minichromosome maintenance proteins (MCMs) drive the formation of prereplicative complexes, which is the first key event during G1 phase. Therefore, altered MCM expression may be a hallmark of cell cycle deregulation, which is supposed to be the most essential mechanism in the development and progression of bladder cancer. Our aim was to investigate the value of MCMs as proliferation markers and prognostic indicators in detrusor muscle-invasive urothelial bladder carcinomas. We analyzed immunohistochemically the expression of MCM-2 and MCM-5 in 65 patients with detrusor muscle-invasive urothelial bladder carcinomas in relation with clinicopathologic parameters, patients' overall and disease-free survival, and the expression of the conventional proliferation index Ki-67 and other cell cycle modulators (p53, pRb, p21(WAF1), and p27(Kip1)). The levels of MCM-2 and MCM-5 were significantly higher in high-grade (P < .0001), advanced-stage (P = .001), and nonpapillary tumors (P < .0001). The expression of MCM-2 and MCM-5 significantly associated with the conventional proliferation index Ki-67 (P = .0001 for each protein). The expression of MCM-2 or MCM-5 positively correlated with p53 labeling index (P = .014 and P = .009, respectively). Also, median p21(WAF1) labeling index was higher in MCM-5 high expressors (P = .028). Finally, both MCM-2 and MCM-5 associated significantly with adverse patients' outcome in both univariate (P = .0072 and P = .0074, respectively) and multivariate (P = .0001) analysis. In conclusion, MCM-2 and MCM-5 appear to be reliable proliferation indexes and useful prognostic markers in patients with muscle-invasive urothelial bladder carcinomas.