Prognostic Significance of p27, Ki-67, and Topoisomerase lla Expression in Clinically Nonfunctioning Pancreatic Endocrine Tumors

Nonfunctioning islet cell tumors or pancreatic endocrine tumors are the most common type of malignant islet cell tumor. Although previously detected usually at an advanced stage because of mass effect, the early detection rate of small localized disease has been increasing. To date it has been difficult to predict the clinical behavior in localized regional nonfunctioning tumors. To investigate potential markers predicting malignancy and poor prognosis in nonfunctioning pancreatic endocrine tumors, we analyzed the expression of Ki-67, topoisomerase IIα (Topollα), and p27, as well as a variety of clinicopathologic parameters in 76 cases of nonfunctioning islet cell tumors (23 benign cases and 53 malignant cases). Ki-67, Topollα, and p27 labeling indices were significantly different between benign and malignant tumors. Expression of Ki-67, Topollα, and p27 were associated with survival in patients with a malignant tumor in a univariate setting. However, only p27 and Topollα were jointly associated with survival in multivariate analysis. Immunohistochemical staining for p27, Topollα, and Ki-67 can be helpful in the diagnosis of nonfunctioning pancreatic endocrine tumor. Analysis of p27 and Topollα may also have potential utility as prognostic factors for malignant tumors.     

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。     

Effect of prolonged physical exercise on the fibrinolytic system

Summary The effect of a test marathon race on plasma fibrinolytic activity (FA) was studied in 16 endurance athletes before, immediately after, 3 h, and 31 h after the run. Tissue plasminogen activator (t-PA) activity increased about 31-fold immediately after the run. Similar increases were found in t-PA antigen concentration. Plasminogen activator inhibitor (PAI) was not detectable immediately after the race and was significantly decreased 3 h (P < 0.05) and 31 h (P < 0.01) later. B_15–42 peptide increased by 0.63 pmol · ml^–1 (P<0.001),d-dimer by 68.3 ng · ml^–1 (P< 0.05). Euglobulin lysis time (ELT) was reduced from 109 to 18 min (P<0.001). The increased t-PA activity and t-PA antigen concentration disappeared in the course of the first 3 h after exertion. ELT also reached its pre-exercise levels at this time. Thirty-one hours after the race ELT and t-PA antigen levels were slightly but significantly reduced (P<0.05), whereas B_15–42 peptide remained increased (P<0.05). t-PA activity was unchanged compared with pre-exercise values. It seems that the exercise-induced FA is mainly caused by the marked increase of t-PA antigen and t-PA activity.     

Evaluation of proliferation parameters in in vivo bromodeoxyuridine labelled lung cancers

In a series of 44 bronchial biopsies from patients suspected of having endobronchial lung carcinoma, the validity of proliferating cell nuclear antigen (PCNA) and Ki67 antigen as proliferative indicators was evaluated in ethanol fixed, paraffin embedded tissue. The percentages of cells positive for these markers were compared to the in vivo bromodeoxyuridine (BrdU) labelling index. A good correlation was found between PCNA immunoreactivity and BrdU labelling index, while Ki67-antigen expression showed a significant relation with BrdU labelling index and with PCNA expression. All three parameters showed a trend towards similar values for the individual cases. Based on the fact that Ki67 antigen is expressed in all cycling cells, whereas replicon-associated PCNA and BrdU only reflect the S-phase fraction, the differences between Ki67-antigen scores on the one hand and BrdU and PCNA scores on the other were smaller than expected. In order to determine the degree of concordance between immunohistochemically and flow cytometrically detected proliferation variables, BrdU incorporation was measured using both methods in duplicate bronchial specimens. Discrepancies in labelling indices were observed predominantly in DNA diploid samples, with consistently lower values in the flow cytometrically analysed specimens. In tumour specimens with an aneuploid DNA content, flow cytometric determination of proliferative activity yielded results similar to those obtained by tissue section examination. We conclude that the scores for PCNA and Ki67 antigen, immunohistochemically detected in ethanol fixed, paraffin embedded tissue reflect functional proliferative activity.     

Enhanced polymer one-step staining (EPOS) for proliferating cell nuclear antigen (PCNA) and Ki-67 antigen: Application to intra-operative frozen diagnosis

Enhanced polymer one-step staining (EPOS) is a novel, highly sensitive one-step immunostaining method. This simple and rapid technlque was applied to intra-operattve frozen diagnosis. The markers of choice were proliferating cell nuclear anmen (PCNA) and Ki-67 antigen. These cell prollferation markers were both identifiable in fresh frozen see tions of the human tonsil In approximately 7 min. The suitable staining sequences are as follows. Frozen sections prepared using 3-aminopropyitimethoxysilane-cpated glass slides are immediately fixed, without air drying, for 15s in a mixture of 50% formalin and 50% methanol for PCNA, and in 10% formalln for Ki-67 antigen. After a brief rinse in phosphate-buffered saline (PSS), sections are incubated with the EPOS antibody for 3 min, followed by PBS rinse for 1 min. The peroxidase activity is visualized in diaminobenzidine-H₂O₂ solution containing 10mmol/L imidazole for 2 min. After a light rinse in tap water, the nuclei are briefly counterstained with 5% methyl green. When necessary, endogenous peroxi-dase blockage in 1% periodic acid solution for 1 min is added before the EPOS antibody incubation. This procedure is applicable to frozen sections of gastric cancers, malignant lymphomas, and brain, liver and peritoneal lesions in which differential diagnosis between benignancy and malignancy was required.     

Significance of aberrant (cytoplasmic/nuclear) expression of beta-catenin in pancreatoblastoma

This study concerns the significance of aberrant (nuclear/cytoplasmic) expression of beta-catenin in pancreatoblastoma (PBL). On immunohistochemistry, all seven PBLs examined showed nuclear/cytoplasmic expression of beta-catenin, predominantly in the squamoid corpuscles (SCs). In areas with acinar/ductular differentiation, few tumour cells displayed nuclear/cytoplasmic expression of beta-catenin and more than half of the tumour cells showed membranous expression. Two out of five (40%) tumours examined showed missense mutations in codons 33 and 37 of exon 3 of the beta-catenin gene. No mutation of the adenomatous polyposis coli (APC) gene was detected in two of the remaining three tumours. Amplifiable DNA for APC analysis was not obtained from the one other tumour. Immunoreactivity for cyclin D1, one of the nuclear targets of beta-catenin, was found predominantly in the SCs of the seven tumours. In contrast, the Ki-67 labelling index was 2-4% (median 3%) in the SCs and 8-18% (median 12%) in the other areas, indicating a negative correlation with nuclear cyclin D1 reactivity. These results imply that in PBLs, nuclear/cytoplasmic accumulation of beta-catenin and overexpression of its target gene cyclin D1 are not associated with the induction of tumour cell proliferation. Nuclear/cytoplasmic accumulation of beta-catenin may be related to the morphogenesis of the SCs that are considered most characteristic for PBL.     

p53 and Ki-67 immunoreactivity and nuclear morphometry of 'carcinoma-in-adenoma' and adenoma of the gall-bladder

Twenty lntramucosal tumors of ‘carclnomaln-adenoma’ and 43 ademas (39 pylorlc gland type, 4 Intestinal type) of the gall-bladder were studied to establish more precise histo-logical criteria of carcinoma or adenoma In cases of ‘carcinoma In pyforic gland type adenoma’, to compare carcinoma In adenoma with pure, that Is, without adenomatous components, carcinoma, and to confirm the benign nature of spin-dle cell fd in the adenomas. Ki-67 and p53 immunostaining and nuclear morphomety were used. Eight pure intramucessl cancers were used as controls. The formalin-fixed, paraffln+mbedded sections were stained with p53 and Ki-67 antibodies. Splndle cell foci were observed only In the adenoma area of the pyloric gland type, wlth a frequency of 23% In 39 adenomas, and of 45% in 20 tumors of carclnoma-lrradenoma. Ki-67 staining was negative in 129 of 130 spin-die cell foci examlned, regardless of their size, and positive in only one focus (550 pm in size, Ki-67 Index 0.2%). All of the spindle cell foci were negative for p53 stain. The Ki-67 positive index was 36.6 ± 5.6% In the 8 pure carcinomas, and 12.5 ± 1.9% in the cancer areas of 16 tumors with carcinoma-in-adenoma, while it was 7.9 ± 1.7% in the adenoma areas of 16 tumors with carcinoma-in-adenoma and 4.9 ± 0.5% in the 32 pure pyloric gland adenomas. The p53-protein over-expression was found in seven of eight pure intramucosal cancers, and in one of 16 cancer components of carclnoma-in-adenoma. However, it was not found in any of 16 adenoma components of carcinoma-in-adenoma, and 35 adenomas. Cells of the cancer tissue of carcinoma-In-adenoma showed a significantly larger nuclear area and a larger nuclear minor axis than those of the pyloric gland type adenomas, as well as other architectural and cytologic abnormalities differing from the features of adenomas. These results suggest that clustered spindle cells do not indmte a malignant transformation of adenoma cells and that carcinomas in carcinoma-in-adenoma are dtfferent from pylorlc gland type adenomas In terms of morphology and proliferative activity. Moreover, the results of the present study indicate that carcinomas In carcinoma-ln-adenoma are lower In malignancy than pure carcinomas, and that their genetic abnormaltty may differ from that of pure carcinomas.     

Laser microdissection and pressure catapulting (LMPC) in paraffin sections mounted on glass slides. A methodological report

The technique of laser microdissection together with laser pressure catapulting (LMPC) is demonstrated in paraffin sections obtained from surgical specimens of brain tumors mounted on glass slides. A sufficient and precise application of microdissection techniques in tissue on glass slides is worthwhile, since it offers the possibility of a retrospective analysis of archived paraffin sections in histopathology. We could demonstrate a precise dissection of areas in tissues of different thicknesses (4 microm and 20 microm). Areas of tissue mounted directly on glass need to be dissected in a scanning mode in order to remove the total region in form of small tissue fragments row by row. This mode provided a precise microdissection of tissue areas of different sizes and shapes. A successful molecular biological analysis of the microdissected regions could be demonstrated. As an example for such an analysis, differential-PCR for detecting an amplification of the gene for the epidermal growth factor receptor (EGFR) was performed.     

The anaplastic variant of centrocytic lymphoma is marked by frequent rearrangements of the bcl-1 gene and high proliferation indices

Ten cases of classic centrocytic lymphoma as defined in the Kiel classification system were investigated for their immunophenotype, their proliferation activity and by means of molecular diagnostics. The findings were compared to those obtained from a group of nine cases of anaplastic centrocytic lymphoma. Both groups showed virtually identical immunohistochemical characteristics with positivity for CD5 and negativity for CD10 and CD23. In the group of anaplastic centrocytic lymphoma, there were considerably higher proliferation indices as documented by staining for the Ki-67 antigen, up to 80% of the tumour cells being positive. Moreover, the cases of anaplastic centrocytic lymphoma had bcl-1 gene rearrangements in eight out of nine cases compared with three out of 10 cases of classic centrocytic lymphoma. DNA analysis was not able to detect bcl-2 gene rearrangement in any case, pointing to a difference compared with lymphomas of germinal centre origin. The coincidence of anaplastic and sometimes blast-like morphology of the tumour cells, high proliferation index and a rearranged bcl-1 gene in nearly all cases of anaplastic centrocytic lymphoma support their classification as high-grade malignant variants of centrocytic lymphoma and suggest a possible role for the bcl-1 locus not only in the origin but also in the progression of centrocytic lymphomas.     

Telomerase activity and proliferation index in aggressive mature B-cell lymphoma: comparison to germinal center phenotypic markers

Cell proliferation may be evaluated by various methods, including Ki-67 immunohistochemistry and measures of telomerase activity. Both methods would theoretically show comparable increases in a given case. To evaluate the relationship between these 2 markers of proliferation in aggressive mature B-cell lymphomas, 48 cases were studied. The study group included 5 cases of mantle cell lymphoma (MCL); 6 cases of Burkitt's/Burkitt's-like lymphoma (BL); 9 cases of follicular lymphoma, grade 3 (FLC); and 28 cases of diffuse large B-cell lymphoma (DLC). Telomerase activity was measured as total product generated (TPG) units, and TPG results for the aforementioned cases were compared to the TPG results for 10 cases of reactive follicular hyperplasia. An overlap in TPG scores between reactive cases and lymphoma cases was found. Significant differences in both log TPG (P = 0.0443) and Ki-67 (P = 0.0006) were seen in the different lymphoma types. A positive correlation between Ki-67 percentage and TPG score was identified in FLC (r = 0.9281; P = 0.0003), but a poor correlation between these 2 indicators was seen in the other lymphoma types. Cluster analysis identified distinct patterns for MCL, FLC, and BL, but heterogeneous patterns for DLC. Because increases in both Ki-67 proliferation and telomerase activity are reported in normal germinal centers (GCs), these tests were also evaluated for usefulness as markers of a GC cell phenotype. Among the FLC and DLC cases, features of a GC phenotype significantly correlated with increased Ki-67 percentage (P = 0.0152), but not with increased log TPG. An elevated log TPG correlated with CD10 expression, and elevated Ki-67 percentage correlated with both CD10 and BCL-6 expression. TPG level and Ki-67 percentage did not correlate with the presence of t(14;18) or BCL-2 protein expression. Although the proliferation patterns were fairly distinctive for MCL, FLC, and BL, these studies show that markers of cell proliferation do not by themselves,identify distinct subtypes of large cell lymphomas. With the exception of FLC, the tumors exhibited poor correlation between telomerase activity and Ki-67 proliferation index. These tests did show some correlation with expression of GC cell phenotypic markers, however.