Estrogen receptors and cathepsin D in human thyroid tissue

BACKGROUND: To investigate the significance of estrogen receptors (ER) in the pathogenesis of thyroid dysplasia, the authors analyzed, by analogy with breast cancers, ER and three estrogen-regulated proteins: progesterone receptor (PR), cathepsin D, and pS2 protein, in cytosols of 42 human thyroid tissues. METHODS: ER and PR were measured by an immunoenzymatic assay and cathepsin D and pS2 by an immunoradiometric assay. Tissue specimens included 7 normal tissues, 6 benign nodules, 8 toxic adenomas, 7 from patients with Graves disease, and 14 carcinomas. RESULTS: ER was present at very low concentrations, with no statistical difference between neoplastic and nonneoplastic tissues. The mean levels of cathepsin D, expressed as pmol/mg protein minus thyroglobulin, were higher in the 14 carcinomas (P = 0.0003), the 7 specimens from patients with Graves disease (P = 0.006), and the 8 toxic adenomas (P = 0.04) than in the 7 normal thyroid tissues. A significant difference also was observed between the carcinomas (P = 0.003) and six benign nodules. Compared to TNM parameters, cathepsin D concentrations correlated with tumor size: higher cathepsin D levels were found in pT4 than in pT2 and pT3 carcinomas. All the tissues tested were negative for PR and pS2 protein. CONCLUSIONS: The results clearly indicate a significant difference between neoplastic and normal thyroid tissue in terms of the amount of cathepsin D, but not that of ER. This suggests that cathepsin D probably is not regulated by estrogen but simply is a marker of protease activity during invasion by thyroid carcinomas.     

VCAM (IGSF) adhesion molecule expression in breast carcinomas detected by automated and quantitative immunocytochemical assays

Expression of vascular cell adhesion molecules (VCAM) in tumors is associated with endothelial cell activation and may facilitate adherence of carcinomatous cells to the vessel wall, promoting bloodborne metastases. Expression of VCAM was investigated in 202 breast carcinomas using automated (Ventana System) and quantitative (SAMBA image analyzer) immunoperoxidase staining of frozen sections. Positive VCAM immunoreactivity was observed in 83 tumors (41%) (mean immunostained surface, 12.4%; SD, 10.5). The mean area of immunostaining was correlated with clinical and pathologic prognostic indicators and with the immunohistochemical expression in tissue sections of various indicators of cell proliferation, metastatic potential, and drug resistance or sensitivity, evaluated according to the same method. There was no correlation of VCAM immunoreactivity with tumor size, type, or grade or with nodal status. Also, no significant correlation was observed between VCAM and MIB1/Ki67, p53, Bcl-2, E cadherin, CD44v, cathepsin D, CD31, P-gp, ER, PR, or pS2. However, VCAM immunoreactivity was significantly correlated with ELAM and VLA2 (P = .001) and VLAs (P = .008) expression. The results suggest that VCAM expression in breast carcinoma tissue sections is likely not a prognostic indicator. Its practical clinical relevance, if any, must be established by correlation with patients' outcomes and tumor sensitivity to drugs.     

The extent of proliferative and apoptotic activity in intraductal and invasive ductal breast carcinomas detected by Ki-67 labeling and terminal deoxynucleotidyl transferase-mediated digoxigenin-11-dUTP nick end labeling

BACKGROUND: The balance among cell proliferation, cell differentiation, and cell death determines the cell number in a population as well as the size or even the stage of a tumor. Thus, to improve our understanding of the pathogenesis of neoplasms, it is important to investigate the regulation of both cell proliferation and cell death. METHODS: This study examined the occurrence of apoptosis and proliferative capacity in 46 breast carcinomas: 20 intraductal carcinomas (ductal carcinomas in situ [DCIS]) and 26 infiltrative ductal carcinomas (IDC). Terminal deoxynucleotidyl transferase-mediated digoxigenin-11-dUTP nick end labeling (TUNEL) and immunostaining with the Ki-67 antibody were used in the examination. A ladder of DNA fragments induced by apoptosis was demonstrated by means of DNA agarose gel electrophoresis in 10 of the available TUNEL positive and negative samples. RESULTS: The results were correlated with p53, bcl-2, estrogen receptor (ER), and progesterone receptor (PR) protein expression, which would suggest association with apoptosis by immunohistochemistry. The apoptosis and proliferation of each cancer were expressed as the number of tumor cells undergoing apoptosis and proliferation per 1000 tumor cells. The extent of apoptosis was more frequently observed in DCIS than in IDC (21.9+/-6.8 vs. 4.0+/-0.9, P < 0.001), and the proliferation activity was significantly higher in IDC than in DCIS (16.8+/-6.5 vs. 3.5+/-0.8, P < 0.006). Apoptosis associated with MIB-1 positive cells and TUNEL labeling was significantly higher in IDC than in DCIS (3.26 vs. 0.42, P=0.001). In DCIS, apoptosis was correlated with p53 (r=0.663, P=0.005), and p53 had a reverse correlation with bcl-2 (r=0.620, P= 0.018). Moreover, bcl-2 expression was associated with ER (P=0.028) and PR (P= 0.005) expression in both DCIS and IDC. CONCLUSIONS: The results of this study show that a higher degree of apoptosis and lower proliferation activity in intraductal carcinoma result in a steady-state, self-renewing condition in which net growth of the tumor is rare. The results also indicate that apoptosis was altered by the expression of p53, bcl-2, ER, and PR.     

A new whole-body exposure chamber for human skin and lung challenge experiments--the generation of wheat flour aerosols

The expression of BCL-2 protein was evaluated immunohistochemically in 23 intracystic papillary carcinomas (IPCs) of the breast. Twenty-two patients were female and one male, aged 49-90 years (median 72). Twenty-one cases had a benign behaviour, while two cases developed local recurrence. Of the 23 tumours, 19 (82%) were immunoreactive for BCL-2, the majority of positive carcinomas showing intense cytoplasmic staining of more than 50% neoplastic cells. The intensity of BCL-2 expression was significantly correlated with prognostic markers such as estrogen receptor (ER) positivity (p = 0.001), cathepsin D (CD) reactivity in the neoplastic cells (p = 0.001) and low growth fraction, evaluated by proliferating cell nuclear antigen (PCNA) immunostaining (p = 0.008). An inverse relationship was also found between BCL-2 and p53 protein (p = 0.001). Three cases of high grade (G3) IPC expressed p53, high PCNA index, and CD (the latter only in the stromal cells), but no immunostaining for BCL-2 and ER. Thus, absence of BCL-2 expression in high grade IPC was associated with ER-negative, rapidly proliferating and p53-positive immunophenotype. All high grade tumours showed invasion of the cystic wall. Local recurrence developed in one of these. The authors conclude that BCL-2 immunoreactivity in IPC is related with tumour grade and with a range of molecular markers of favourable prognosis such as ER positive status, CD expression in the neoplastic cells, and low PCNA index. These findings are consistent with the indolent clinical course and the very favourable prognosis of IPC of the breast.     

A phase II trial of weekly infusional 5-fluorouracil in combination with low-dose leucovorin in patients with advanced colorectal cancer

We examined 59 breast cancers for p53 and bcl-2 protein expression by immunohistochemistry. The results were correlated with Ki-67 immunostaining. p53-negativity was noted in 40 cases and the remaining 19 tumours were p53-positive. Thirty-six tumours showed strong expression of bcl-2 and in 23 no staining for this protein was observed. We found statistically significant reverse correlation between expression of p53 and bcl-2 in majority of carcinomas: 31 cases were bcl-2 positive and p53-negative, and 14 tumours were bcl-2-negative and p53-positive. Six carcinomas showed no nuclear staining for Ki-67 and in the remaining 53 the percent of cancer cells positive for Ki-67 ranged from 1 to 60 (mean: 14.6). In these 53 cases we found that bcl-2-positive tumours were characterized by lower proliferation than bcl-2-negative tumours, the mean value of Ki-67 immunostaining being 10.7% and 23.0%, respectively. p53-negative tumours showed lower proliferation than p53-positive tumours: mean Ki-67 index was 10.2% and 23.9%, respectively. We conclude that immunohistochemically detected p53 and bcl-2 proteins show a significant inverse relationship in majority of breast carcinomas and their expression correlates with tumour proliferation (Ki-67 immunostaining).     

Red wine, white wine, liquor, beer, and risk for coronary artery disease hospitalization

A series for Nm23-protein immunodetection was investigated in human breast carcinomas. Frozen sections were processed by automated immunoperoxidase procedure, and immunoprecipitates in positive tumors were quantified by processing digitized microscopic images. Nm23 immunohistochemical expression in tumors was correlated with clinicopathological data and with intra-tumoral proteins also detected by automated and quantitative immunohistochemistry. A positive Nm23 immunoreaction was observed in 58% of tumors, within cell cytoplasm. Nm23 expression was independent of the patient's age, and of tumor size, type and grade, but an inverse relationship was observed between Nm23 expression and axillary-lymph-node metastasis. An inverse relationship was also observed between Nm23 and P-53, CD-31, cathepsin D, tenascin and P-gp immunohistochemical expressions. But Nm23 expression was independent of c-erb-B product, growth fraction (MIB1/Ki67), and immunohistochemical expression of hormone receptors/P-S2. The results suggest that the anti-metastatic nm23 gene may partly act upon the regulation of tumor-cell proliferation (correlation with P-53) and may have some effects on epithelial-cell/stroma interactions (regulation of extracellular-matrix protease and of angiogenesis) independently of hormone sensitivity.     

The prevalence of bcl-2, p53, and Ki-67 immunoreactivity in transitional cell bladder carcinomas and their clinicopathologic correlates

The aim of this study was to investigate the expression of bcl-2, p53 oncoproteins, and Ki-67 antigen in a series of transitional cell bladder carcinomas and its relation to the traditional prognostic indicators and patient's survival. One hundred six cases with transitional cell carcinoma (TCC) were examined for detection of bcl-2, p53 proteins, and Ki-67 antigen (MIB1 antibody). Bcl-2 immunohistochemical positivity was observed in 52% of TCCs and in 57% of low-grade and 44% of high-grade TCCs. Bcl-2 was also detected in normal urothelium and dysplastic lesions with basal cell expression, and negative staining was observed in carcinomas in situ. Tumor stage showed a significant inverse correlation with overall bcl-2 positivity. The loss of bcl-2 protein expression in higher-stage TCCs was statistically significant (Pt = .01). p53 protein was overexpressed in 50% of TCCs and more frequently in invasive and in carcinomas in situ than in superficial TCCs (Pt = .03). In contrast, detection of p53 was not observed in normal and dysplastic urothelium. p53 positivity was related to the degree of differentiation and to the stage of the disease (Pf = .01 and Pt = .03, respectively). Concerning Ki-67 antigen, its expression was found in 57.5% of TCCs. There was a strong overall correlation of Ki-67 with tumor stage (Pt = .002) and grade (Pf = .002). Univariate statistical analysis showed that the expression of p53 and Ki-67 was significantly correlated to poor prognosis (P = .02, P = .02, respectively). On multivariate analysis, none of these markers but only stage and grade were significantly correlated to prognosis (P = .02, P = .02, respectively). These findings suggest that overexpression of bcl-2 protein may be an early event in tumorigenesis. Tumors with loss of bcl-2 positivity and overexpression of p53 and Ki-67 had an unfavorable prognosis; however, in multivariate analysis, they had no independent prognostic value.     

Expression of osteonectin mRNA in human breast tumours is inversely correlated with oestrogen receptor content

Osteonectin is a secreted glycoprotein which is detected in a number of normal and neoplastic human tissues in vivo. It is an extracellular matrix (ECM)-associated protein which is postulated to regulate cell migration, adhesion, proliferation and matrix mineralisation and previous reports suggest that it may be modulated by steroid hormones in target tissues. The aim of this study was to measure osteonectin mRNA and protein expression in breast tumour biopsies and compare these with oestrogen (ER) and progesterone receptor (PR) levels in the same tumours. An inverse correlation was seen between osteonectin mRNA expression and ER level. Samples with low ER protein expression had a mean osteonectin mRNA level which was almost 4-fold greater than the mean level of expression observed in tumours containing high concentrations of ER protein. This inverse correlation was statistically significant. Despite the strong inverse relationship between osteonectin mRNA levels and tumour ER content, no correlation was seen when osteonectin protein concentration was measured in tumour cytosols on immunoblots and compared to ER and PR levels in the same tumours. However, since it is a secreted protein, osteonectin protein expression may not reflect cellular osteonectin levels in breast tumours. In summary, these data suggest that ER-mediated suppression of osteonectin gene expression may contribute to the less aggressive characteristics associated with receptor-positive tumours and that loss of ER expression may lead to over-expression of osteonectin and contribute to a poorer differentiated, more invasive phenotype.     

Occurrence of asthma and chronic bronchitis among female hairdressers. A questionnaire study

AIMS: We examined the relationship between apoptosis and three different major stages of human breast carcinoma: intraductal carcinoma (DCIS), infiltrating duct carcinoma (IDC) and metastatic carcinoma in lymph nodes. We also determined the correlation between apoptosis and oestrogen receptor (ER), progesterone receptor (PR) and p53. METHODS AND RESULTS: The study investigates the extent of apoptosis in 63 breast carcinomas by in-situ end-labelling, in formalin-fixed, paraffin-processed tissue sections. The 63 breast carcinomas, included 22 DCISs, 26 IDCs, three infiltrating lobular carcinomas (ILC) and 12 metastatic lymph nodes. The apoptotic labelling index was higher in DCIS than IDC and metastatic carcinoma (P < 0.001, P < 0.007, respectively). By immunohistochemistry, we also analysed p53, ER and PR. Apoptosis correlated significantly with p53 (r = 0.748, P = 0.0004) in IDC. Also, ER correlated significantly with PR (r = 0.629, P = 0.00001). No apparent correlation was found between the apoptosis and ER or PR. CONCLUSION: Our data suggest that not only does apoptosis differ between intraductal carcinoma and infiltrating carcinoma but also it might be regulated by altered p53 expression.     

Bcl-2 expression and its association with cell kinetics in human gastric carcinomas and intestinal metaplasia

The bcl-2 protooncogene was initially discovered at the t(14;18) chromosomal breakpoint in follicular lymphomas. It has been demonstrated that bcl-2 protein (Bcl-2) expression blocks apoptosis and plays an important role in cell development and maturation. In the present study, Bcl-2 expression was immunohistochemically examined in 103 cases of gastric carcinoma, as well as 64 cases of non-carcinous gastric mucosa, and its correlation with apoptosis, cell proliferation and p53 immunoreactivity was investigated. Bcl-2 was detected in 18.0% of differentiated-type gastric carcinomas (9 of 50) and 7.5% of the undifferentiated type (4 of 53). In adjacent intestinal metaplastic gastric epithelium, the incidence of Bcl-2 positivity in the incomplete type (21/23, 91.3%) was significantly higher than in the complete type (23/41, 56.1%) (P < 0.04). Double immunostaining for Bcl-2 and Ki-67 clearly revealed the majority of Bcl-2-positive cancer cells to be in a nonproliferating state, although some cancer cells expressed both proteins together. Statistical assessment demonstrated that the average Ki-67 labeling index and apoptotic labeling index in Bcl-2-positive foci were significantly lower than in Bcl-2-negative foci (P < 0.0001, P < 0.0003). In addition, a significant dissociation between Bcl-2 and p53 immunoreactivity was found in cancer tissues. These results indicate that aberrant Bcl-2 expression in gastric carcinomas possibly originates from intestinal metaplastic epithelium, and suggest a possible role in tumor development and growth.     

In vivo stimulation by tamoxifen of cathepsin D RNA level in breast cancer

We have previously shown that 3 weeks of treatment with tamoxifen, of patients with primary breast carcinomas, increased cytosolic cathepsin D protein in oestrogen receptor (ER) positive tumours [Maudelonde et al., Cancer 1989, 63, 1265-1270]. In order to investigate the mechanism of this increase and to eliminate a transient flare-up effect, we semi-quantified cathepsin D RNA levels by in situ hybridisation in 32 breast carcinomas from patients treated with tamoxifen for 3 weeks prior to surgery and in 35 breast cancer patients receiving no tamoxifen. We found that tamoxifen increased cathepsin D RNA level regardless of the ER status of the tumours. In ER positive tumours, tamoxifen increased the cathepsin D RNA level to the same extent as cytosolic cathepsin D protein but not in ER negative tumours. The induction of cathepsin D RNA by tamoxifen in ER positive tumours was probably due to its agonist activity, also observed in vitro in breast cancer cell lines. These results suggest that the cathepsin D gene is inducible by oestrogens in ER positive breast cancer as it is in breast cancer cell lines.     

Expression of protooncogene bcl-2 in thyroid tumors

OBJECTIVE: To determine the expression of protooncogene bcl-2 in thyroid tumors and its relationship to the development and prognosis of the tumor. METHODS: 124 cases of thyroid tissues (41 thyroid carcinomas, 53 thyroid adenomas, 20 thyroid tissues adjacent to cancer and 10 normal thyroid tissues) were immunohistochemically stained for bcl-2, by using bcl-2 protein monoclonal antibody. The positive-staining rates in different thyroid tissues were compared statistically. RESULTS: Bcl-2 immunoreactivity was found in thyroid carcinomas (43.9%), thyroid adenomas (22.6%), and thyroid tissues adjacent to cancer (15.0%). The positive-staining rate in thyroid carcinomas was higher than that in thyroid adenomas (P = 0.0439) and that in thyroid tissues adjacent to cancer (P = 0.0430). In thyroid carcinoma, the higher positive-staining rates were found in the cases of undifferentiated carcinoma and follicular carcinoma, as well as in the cases of positive lymph nodes or at tumor stage II and IV. CONCLUSION: The results suggest that the over-expression of bcl-2, a possible prognostic marker of thyroid cancer, may be related to the development of thyroid tumor.     

Role of Mycoplasma pneumoniae in acute respiratory-tract infections in Saudi paediatric patients

The expression of RB and bcl-2 proteins was investigated in 135 thyroid carcinomas (130 papillary carcinomas and five follicular carcinomas) by the immunohistochemical technique with RB and bcl-2 monoclonal antibodies. The results were as follows: 1) RB protein nuclear staining was shown in 117 of the 135 thyroid carcinomas (87%). Bcl-2 protein staining was shown in 82 of the carcinomas (61%). 2) There was a higher incidence of recurrence in RB-negative cases than in RB-positive cases (chi 2 test p = 0.001). But there is no significant relationship between the incidence of recurrence and the results of bcl-2 staining (chi 2 test p = 0.287). 3) The expression of RB protein was related to age and clinical N and M categories. Bcl-2 protein expression was related to clinical T categories. 4) The 10-year overall survival rate for RB-positive patients was higher than that for negative patients (long-rank test p < 0.0001). But the survival rate was not associated with bcl-2 expression (log-rank test p = 0.3). Our results suggest that assessment of RB protein expression by the immunohistochemical technique may be useful to determine the possibility of recurrence and to predict the outcome of thyroid carcinomas. We think that assessment of RB protein expression by the immunohistochemical technique can be applied as a more sensitive prognostic factor.     

bcl-2 automated and quantitative immunocytochemical assays in breast carcinomas: correlation with 10-year follow-up

PURPOSE: bcl-2 protein is detectable in human cancers and may be involved in the response to antineoplastic drugs or endocrine therapy in breast carcinomas. In a previous study, we had developed optimal technical conditions for bcl-2 immunodetection. The aim of the present report was to determine the prognostic significance of bcl-2 expression in breast carinomas by the use of a similar immunocytochemical procedure. METHODS: bcl-2 immunocytochemical assays were performed on frozen sections by automated immunoperoxidase technique (Ventana) and computer-assisted analysis of digitized colored microscopic images (SAMBA) in a series of 170 breast carcinomas. The results of automated quantitative immunocytochemical assays were correlated with patient follow-up (120 months). RESULTS: Intense bcl-2 immunocytochemical expression in tumors (cutpoint, 15%) significantly correlated with longer disease-free survival and longer recurrence-free survival in the entire cohort of patients (P = .028 and P = .035, respectively) and also in node-negative subgroups of patients (P = .028 and P = .01; Kaplan-Meier long-rank test; NCSS 6.0.1 software). But bcl-2 immunostained surfaces (cutpoint, 15%) did not correlate with overall survival. In multivariate analysis (proportional hazards regression, Cox model), bcl-2 prognostic significance in terms of disease-free survival was only independent of the tumor size and grade and histoprognostic index (Nottingham prognostic index [NPI]). CONCLUSION: bcl-2 immunohistochemical expression is a significant indicator of favorable outcome only in terms of disease-free and local recurrence-free survival. However, bcl-2 expression in tumors is an independent weakly prognostic indicator in breast carcinomas. bcl-2 immunodetection assessed in optimal technical conditions (frozen samples, automation, quantitative analysis, scatter diagram cutoffs) may have some limited practical clinical relevance for the management of patients with breast carcinomas.     

EMS1 gene expression in primary breast cancer: relationship to cyclin D1 and oestrogen receptor expression and patient survival

The EMS1 and CCND1 genes at chromosome 11q13 are amplified in about 15% of primary breast cancers but appear to confer different phenotypes in ER positive and ER negative tumours. Since there are no published data on EMS1 expression in large series of breast cancers we examined the relationship of EMS1 expression with EMS1 gene copy number and expression of mRNAs for cyclin D1 and ER. In a subset of 129 patients, where matched tumour RNA and DNA was available, EMS1 mRNA overexpression was associated predominantly with gene amplification (P = 0.0061), whereas cyclin D1 mRNA overexpression was not (P = 0.3142). In a more extensive series of 351 breast cancers, there was no correlation between cyclin D1 and EMS1 expression in the EMS1 and cyclin D1 overexpressors (P = 0.3503). Although an association between EMS1 mRNA expression and ER positivity was evident (P = 0.0232), when the samples were divided into quartiles of EMS1 or cyclin D1 mRNA expression, the increase in the proportion of ER positive tumours in the ascending EMS1 mRNA quartiles was not statistically significant (P = 0.0951). In marked contrast there was a significant stepwise increase in ER positivity in ascending quartiles of cyclin D1 mRNA (P = 0.030). A potential explanation for this difference was provided by the observation that in ER positive breast cancer cells oestradiol treatment resulted in increased cyclin D1 gene expression but was without effect on EMS1. The relationship between EMS1 expression and clinical outcome was examined in a subset of 234 patients with median follow-up of 74 months. High EMS1 expression was associated with age > 50 years (P = 0.0001), postmenopausal status (P = 0.0008), lymph node negativity (P = 0.019) and an apparent trend for worse prognosis in the ER negative subgroup. These data demonstrate that overexpression of EMS1 mRNA is largely due to EMS1 gene amplification, is independent of cyclin D1 and ER expression and, in contrast to cyclin D1, is not regulated by oestrogen. Independent overexpression of these genes may confer different phenotypes and disease outcomes in breast cancer as has been inferred from recent studies of EMS1 and CCND1 gene amplification.     

Different expression of Bcl-2 protein in gastric adenomas and carcinomas

In order to cast light on the possible role of bcl-2 protein (Bcl-2) expression in gastric tumorigenesis, 33 cases of gastric adenomas and carcinomas originating from the same stomachs were immunohistochemically investigated for Bcl-2 protein (Bcl-2) expression, accumulation of p53 protein and cell proliferation as determined by the Ki-67 labeling index (LI). Bcl-2 expression was detected in 24/33 (72.7%) adenomas and in 6/33 (18.2%) carcinomas, the difference being statistically significant (P = 0.0001). Only 4 of 33 (12.1%) cases exhibited expression in both adenoma and carcinoma lesions in the same stomachs. Immunoreactivity was decreased in areas of cellular and structural atypia in adenoma lesions (P < 0.008), and appeared to be positively linked to the tumor progression and the degree of differentation in carcinomas, although it did not reach statistical significance. Accumulation of p53 protein was rare in the adenomas but was found in 15/33 (45.5%) of carcinoma lesions, with a significant dissociation from Bcl-2 immunoreactivity. No apparent relation between Ki-67 LI and either adenoma grading or carcinoma typing was noted, although average Ki-67 LI of the highest labeling areas in carcinomas was statistically higher than in adenomas (P = 0.0001). These results indicate that the regulation of Bcl-2 expression may differ between gastric adenomas and carcinomas, may be correlated with tumor differentiative features. In addition, p53 accumulation may play an important role in the onset of malignancy.     

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In breast cancer, oestrogen regulated genes, such as pS2, may be expressed in well differentiated tumours with a good prognosis. We have examined pS2 mRNA expression in 78 primary, untreated breast cancers and related pS2 expression to disease behaviour and known prognostic factors. pS2 mRNA expression was detected in 25/78 (32%) of cancers and was significantly associated with a moderate/high oestrogen receptor content (P = 0.045, Chi Square test). pS2 mRNA expression was associated with freedom from disease at median 31 months clinical and radiological follow-up (P = 0.015, Fisher's exact test, odds ratio 8.6). Using multiple logistic regression analysis of six potential prognostic factors only pathological axillary node status (P < 0.01) and pS2 mRNA expression (P < 0.05) provided independent prognostic information. Furthermore, pS2 was associated with a good prognosis in the axillary node positive patients where only 1/13 (8%) with pS2 mRNA expression compared with 13/29 (45%) without detectable expression had recurrence of their disease. These data provides strong support for pS2 as a useful independent prognostic factor in primary breast cancer.     

Impact of holmium laser settings and fiber diameter on stone fragmentation and endoscope deflection

To evaluate the relationship between cell proliferation and apoptosis in sporadic colorectal carcinogenesis, immunohistochemistry for proliferation-associated antigen Ki-67 and in situ end labelling for identifying apoptotic bodies were performed on paraffin sections from 59 adenomas and 22 carcinomas. These results were correlated with the expression of the proliferation and apoptosis modulators Bcl-2 and p53. Carcinomas showed increased proliferation and apoptosis compared with adenomas (P<0.0001, P<0.001, respectively). There were positive linear correlations between proliferation and apoptosis in adenomas and carcinomas (P<0.02, P<0.05, respectively). The proliferative rate increased significantly from mild to moderate, and from moderate to severe dysplasia (P<0.002, P<0.001, respectively). Apoptotic rate also increased in this sequence, but the increases did not reach statistical significance (both P>0.05). Expression of Bcl-2 was associated with lower apoptotic rate in adenomas (P<0.025) but not in carcinomas (P>0.25), whereas p53 expression was correlated with higher proliferative rate in both adenomas and carcinomas (P<0.01, P<0.05, respectively). An inverse relationship between Bcl-2 and p53 expression was seen in both adenomas and carcinomas (P<0.05, P<0.005, respectively). These data suggest that the normal balance between proliferation and apoptosis is disturbed in colorectal carcinogenesis, both being increased, but proliferation occurs in excess. Bcl-2 and p53 may each play a role in modulating cell apoptosis or proliferation during the development of colorectal carcinoma.     

Proliferation and DNA fragmentation in meningioma subtypes

Atypical meningioma has been introduced as tumour subtype of intermediate biological behaviour between classical and malignant meningiomas. To substantiate this three-step scale of malignancy, we assessed the proliferative activity reflected by Ki-67 (MIB1) labelling index (LI) in a series of 89 meningiomas, including 15 classical, 29 atypical, 35 anaplastic tumours, and 10 haemangiopericytomas and papillary meningiomas. The possible correlation of proliferation with the frequency of apoptosis and their relations to BCL-2 immunoexpression was investigated in seven classical, 10 atypical and 10 malignant meningiomas. Apoptosis was demonstrated by evaluation of the frequency of apoptotic figures, by the enzymatic technique of in situ tailing (IST) which stains apoptotic DNA fragments, and by DNA preparation and gel electrophoresis demonstrating DNA laddering in frozen tissues of five meningiomas. MIB1 LI revealed a highly significant increase from classical through atypical to anaplastic meningiomas (P < 0.0001); haemangiopericytomas and papillary meningiomas were well within the range of atypical meningiomas. IST indices rose with increasing malignancy and correlated with MIB1 LI (P < 0.0001): they showed a weak inverse correlation with BCL-2 immunoexpression (P = 0.05). BCL-2 expression tended to decrease with malignancy grade and was unrelated to MIB1 LI or frequency of apoptosis. Our data show that (i) apoptosis is a feature of meningiomas, significantly correlated with the malignancy scale. (ii) DNA fragmentation shows significant correlation with proliferation and inversely with BCL-2 expression; (iii) proliferation indices and frequencies of apoptosis/DNA fragmentation within meningioma subgroups corroborate the intermediate biological position of the atypical meningioma between classical and malignant meningiomas.