MIB-1 labeling indices in benign, aggressive, and malignant meningiomas: A study of 90 tumors

Predicting tumor behavior in meningiomas based on histology alone has been problematic. This study retrospectively compares histology and MIB-1 (cell proliferation marker) labeling indices (LI) in benign, aggressive, and malignant meningiomas. Six histological features, including mitoses, necrosis, loss of pattern, hypervascularity/ hemosiderin deposition, prominent nucleoli, and nuclear pleomorphism, were compared in 90 meningiomas (Fisher's exact test). Tumors with two or more of the above features were designated as aggressive meningiomas. Malignant meningiomas were characterized by brain invasion or metastasis. The MIB-1 Us (% positive tumor cell nuclei) were compared between the three groups (Kruskal-Wallis test, Wilcoxon two-sample test). Of the benign meningiomas (n = 37; mean age, 54 years), 41% had one of the six histological features, with nuclear pleomorphism (n = 10) being the most frequent. The aggressive tumors (n = 29; mean age, 61 years) were characterized by nuclear pleomorphism (n = 28), mitoses (n = 20), necrosis (n = 16), loss of pattern (n = 16), prominent nucleoli (n = 6), and hypervascularity/hemosiderin deposition (n = 5). Malignant tumors (n = 24; mean age, 59 years) were characterized by nuclear pleomorphism (n = 22), mitoses (n = 21), loss of pattern (n = 21), necrosis (n = 21), nucleoli (n = 17), and hypervascularity/hemosiderin deposition (n = 3). Significant differences were found between the aggressive and malignant groups with regard to loss of pattern, necrosis, and nucleoli (P = .0043, .011, and .00029, respectively). Mean MIB-1 LIs for the benign, aggressive, and malignant groups were 1.0% (range, 0 to 5.5%), 5.5% (range, 0.1 to 32.5%), and 12.0% (range, 0.3 to 32.5%), respectively. Differences in the mean MIB-1 LI between groups were statistically significant, with P values of <.0001 (benign v aggressive) and .0012 (aggressive v malignant). Mean MIB-1 LIs for recurrent versus nonrecurrent tumors were 7.1% (range, 0 to 32.5%) versus 3.8% (range, 0 to 20.9%) (P = .32). The mean MIB-1 LI for patients who were alive with or without tumor was 6.2% (range, 0 to 32.5%) versus a mean MIB-1 LI of 14.2% (range, 2.8% to 32.5%) for patients who died of or with tumor (P = .0013). In conclusion, (1) There is a statistically significant difference in the increasing MIB-1 LI means between benign, aggressive, and malignant meningiomas and between patients who were alive versus those who died; (2) there is some overlap in MIB-1 LI ranges between groups, which warrants caution in interpreting an individual MIB-1 LI in a given tumor.     

Transthyretin in endocrine pancreatic tumors.

The occurrence of transthyretin (TTR) in 25 endocrine pancreatic tumors was investigated by immunohistochemical methods using both polyclonal and monoclonal antibodies. All malignant insulinomas were strongly TTR immunoreactive, more so than their benign counterparts, which in some cases were TTR negative. All glucagonomas and nonfunctioning tumors were TTR immunoreactive, whereas gastrinomas and VIPomas were TTR negative. TTR, chromogranin A, and the argyrophil reaction (Grimelius' silver technique) had similar distributions among the cells in many, but not all, tumors. Coexistence of TTR with glucagon, insulin, or pancreatic polypeptide in tumor cells was demonstrated. TTR was also quantitated in preoperative serum samples by electroimmuno assay in some cases. Although one patient with a glucagonoma had a markedly increased serum TTR level, five other patients with endocrine tumors, including two patients with glucagonoma, had TTR levels in serum that were within or below the reference range.     

Expression of E-cadherin,b-catenin,and Ki-67 in goblet cell carcinoids of the appendix: an immunohistochemical study with clinical correlation

Goblet cell carcinoid (GCC) of the appendix is a rare entity, of which both the histogenesis and biologic behavior remain controversial, and prognostic tools and therapeutic strategies for this unusual tumor have yet to be defined. The aim of this study was to analyze expression of E-cadherin and β-catenin in GCCs of the appendix with long-term follow-up data as related to the expression of Ki-67 proliferation marker to provide a rationale for treatment guidelines. We analyzed the expression of E-cadherin, β-catenin, and Ki-67 in 11 GCCs of the appendix and control groups of typical carcinoids of the large intestine (n=29), well to moderately differentiated adenocarcinomas of the colon (n=10), poorly differentiated adenocarcinomas of the colon (n=12), and normal appendiceal tissues (n=10). There was no significant difference between the GCCs and normal appendiceal tissues regarding the expression of E-cadherin or β-catenin (p=0.297 and 0.103, respectively). The percentage of positive GCC cells ranged between 0.52 and 10.35% (4.27 ± 0.80), and only one case had a score > 10%. Metastatic tumor spread and death were found in high MIB-1 labeling index (LI) cases of GCC (>3%). Our findings suggest that the behavior of the majority of GCCs might be indolent and different from adenocarcinomas because of the preserved expression of E-cadherin and β-catenin and relatively low MIB-1 LI. However, some of these tumors act aggressively and MIB-1 LI might be a good parameter to determine the therapeutic procedure.     

Expression of cyclin A and topoisomerase IIalpha of oligodendrogliomas is correlated with tumour grade,MIB-1 labelling index and survival

AIMS: This study was designed to investigate immunoexpression of cyclin A and D1, and topoisomerase IIalpha in oligodendrogliomas and to evaluate the correlation with MIB-1 (Ki67), tumour grade, and survival of the patients. METHODS AND RESULTS: Forty cases of oligodendrogliomas (20 high- and 20 low-grade) were studied immunohistochemically with the above-mentioned monoclonal antibodies. RESULTS: Normal brain tissues included in tumour sections did not express any of cyclin A, MIB-1 and topoisomerase IIalpha except cyclin D1, which was shown in perineuronal and interfascicular normal oligodendroglial cells. In low-grade and high-grade oligodendrogliomas, the mean cyclin A labelling index (LI) was 1.18 +/- 0.98% versus 4.65 +/- 1.99%, respectively; the mean topoisomerase IIalpha LI was 1.32 +/- 1.04% versus 6.63 +/- 4.31%, respectively; and the mean MIB-1 LI was 1.69 +/- 1.55% versus 9.46 +/- 4.66%, respectively. Interestingly, cyclin D1 was not expressed in any oligodendrogliomas. Both cyclin A and topoisomerase IIalpha LI showed a significant positive correlation with MIB-1 LI and histological grade of oligodendrogliomas (P < 0.01) and an inverse correlation with overall survival (P < 0.01). Univariate analysis showed that cyclin A and topoisomerase IIalpha LIs with a cut-off point at 3% were a significant prognostic factor (P: cyclin A = 0.0040, topoisomerase IIalpha = 0.0033). CONCLUSION: Cyclin A and topoisomerase IIalpha expression are closely correlated with anaplastic oligodendrogliomas and worse clinical outcomes. Cyclin D1 seems not to be involved in the tumorigenesis of oligodendrogliomas.     

Apoptotic activity and bcl-2 immunoreactivity in meningiomas. Association with grade and outcome

We retrospectively evaluated 90 meningiomas for bcl-2 expression, apoptosis counts (per 10 high-power fields [HPF]), MIB-1 labeling indices (LI), and mitosis counts (per 10 HPF). Characteristics were as follows: 37 low-grade (benign) meningiomas: mean apoptosis count, 1.2; MIB-1 LI, 1.0; mitosis count, 0.1; and bcl-2 positivity, 40%; 29 atypical meningiomas: apoptosis count, 3.3; MIB-1 LI, 5.5; mitosis count, 2.2; and bcl-2 positivity, 62%; 24 malignant meningiomas: apoptosis count, 6.5; MIB-1 LI, 12.0; mitosis count, 6.0; and bcl-2 positivity, 67%. By univariate analysis, MIB-1 LI, apoptosis and mitosis counts, and tumor grade were associated significantly with death due to tumor; by multivariate analysis, only mitosis count was independently associated with death due to tumor. We compared similar data for 27 patients with nonrecurrent tumors and 32 patients with recurrent meningiomas. Histologic sections from the initially resected tumor and from the most recent recurrence were reviewed. Only the apoptosis count was significantly higher by univariate analysis in the initial resection specimens from tumors that ultimately recurred vs nonrecurrent tumors. Expression of bcl-2, MIB-1 LI, and mitosis count did not correlate with recurrence. By multivariate analysis, only extent of surgical resection was associated significantly with tumor recurrence. Although bcl-2 immunostaining was not associated statistically with outcome, bcl-2 positivity was more common in atypical and malignant meningiomas than in low-grade tumors.     

Proprotein-Processing Endoprotease Furin and its Substrate Parathyroid Hormone-Related Protein Are Coexpressed in Insulinoma Cells

Parathyroid hormone-related protein (PTHrP) is frequently produced in pancreatic endocrine tumors. PTHrP is synthesized as the precursor pro-PTHrP and undergoes a series of posttranslational processing reactions, among which cleavage of a 12 amino acid sequence from its precursor is crucial for the biological activation of PTHrP. This cleavage is catalyzed by furin, a proprotein-processing endoprotease that cleaves the consensus sequence-Arg-X-(Lys/Arg)-Arg ↓ X-. We previously reported that furin is highly expressed in rat pancreatic islets during the perinatal stage and that the expression of furin in pancreatic β cells induces faster cell growth. From this, we postulated that furin may be co-expressed with PTHrP in insulinomas. We immunostained insulin, PTHrP, and furin in 21 human pancreatic endocrine tumors: 10 insulinomas, 5 VIPomas, 4 gastrinomas, and 2 somatostatinomas. Of these 21 endocrine tumors, furin was positively stained in all 10 insulinomas. Likewise, PTHrP was detected in the same insulinomas. We found one VIPoma and one gastrinoma contained a few insulin-positive cells scatteringly, which were also positive for furin and PTHrP. But other non-insulinoma endocrine tumors did not display furin and PTHrP positivity. We conclude that furin and its substrate pro-PTHrP are co-expressed specifically in insulinomas.     

Cyclin D1 and MIB-1 immunohistochemistry in pilocytic astrocytomas: A study of 48 cases

Pilocytic astrocytoma is an infrequently encountered, generally low-grade neoplasm. No study has extensively looked at both cyclin Dl and MIB-1 labeling indices in pilocytic astrocytoma and their relation to clinical outcome. This study retrospectively examines the clinicopathologic features of 48 patients with pilocytic astrocytoma including MIB-1 (cell proliferation marker) and cyclin Dl (protein that regulates progression from G1 to S phase of the cell cycle) immunohistochemistry. Of 48 patients (27 females and 21 males; mean age, 12.7 years; age range, 2 to 57 years), 26 initially underwent gross total resection; 17, subtotal resection; four, biopsy alone; in one patient, the extent of tumor resection was unknown. Histological features observed included Rosenthal fibers (83.3%), granular bodies (75%), vascular sclerosis (56.2%), vascular proliferation (56.2%), prominent nuclear pleomorphism (14.6%), necrosis (10.4%), and identifiable mitotic figures (2.1%). MIB-1 labeling indices (n = 45) (positive staining tumor nuclei per 1,000 nuclei evaluated) ranged from 0 to 3.5% (mean, 0.6%); seven tumors had a labeling index greater than 1.0%. Cyclin D1 labeling indices (n = 45) ranged from 0 to 0.8% (mean, 0.1%). Most tumors (N = 29, 66.7%) had no immunostaining. At last known follow-up, 27 patients were alive with no evidence of disease (mean, 49.2 months), 17 patients were alive with evidence of disease (mean, 36.8 months), three died with tumor at 2, 22, and 156 months, and one patient was lost to follow-up. Eight patients had at least one tumor recurrence requiring additional surgery; seven of these patients had an initial subtotal resection. In summary, MIB-1 labeling indices were generally low (mean, 0.6%) and are reflective of the slow growth of the tumors. Cyclin D1 immunostaining does not appear to be significantly increased in pilocytic astrocytoma. Adverse outcome in patients with pilocytic astrocytoma may be related to extent of surgical resection and does not seem to correlate with histology, MIB-1 labeling indices, or cyclin D1 immunoreactivity.     

MIB-1 and DNA topoisomerase II alpha could be helpful for predicting long-term survival of patients with glioblastoma

Approximately 10% of patients with glioblastoma survive more than 2 years after diagnosis. Distinguishing these patients from those who died within 2 years of diagnosis is clinically significant. We studied the MIB-1 labeling index (LI) and DNA topoisomerase II alpha LI of glioblastomas from 34 patients who lived for more than 2 years after diagnosis and of glioblastomas from 34 age- and sex-matched control patients who died within 2 years of diagnosis. The means of MIB-1 and topoisomerase II alpha LIs of the group with a better outcome were lower. With 35 as the cutoff point for the MIB-1 LI and 26 as the cutoff point for the topoisomerase II alpha LI, both MIB-1 and topoisomerase II alpha LIs were related significantly to survival. Our study showed that both MIB-1 and topoisomerase II alpha could help predict long-term survival of patients with glioblastomas. Multivariate analyses revealed that MIB-1 was a better prognostic marker than topoisomerase II alpha.     

Immunoreactivity and expression of amylin in gastroenteropancreatic endocrine tumors.

Amylin was isolated from human insulinomas, but there has been only preliminary data regarding whether this peptide can also be detected in other types of gastroenteropancreatic endocrine tumors. In the present study, immunohistochemical staining of 87 gastroenteropancreatic endocrine tumors demonstrated amylin immunoreactivity in 21.8% of the neoplasmas. Thirteen of 15 insulinomas, three of 21 gastrinomas, two of 29 nonfunctioning tumors, and one of 18 carcinoids were amylin-immunoreactive. Seventeen of the 19 amylin-immunoreactive tumors were primarily located in the pancreas, but two tumors were found in the intestine. Measurements of amylin messenger RNA expression in a few tumors revealed amylin synthesis in these tumors. Amylin immunoreactivity did not correlate with invasion and metastasis. However, the rate of curative resections was significantly higher in amylin-immunoreactive tumors. These results demonstrate for the first time that amylin immunoreactivity is not restricted to insulinomas and can also occur rarely in endocrine tumors of the intestine.     

Proliferative index in astrocytic tumours

The accurate grading of astrocytic tumours is of prime importance because it is critical to the patient management and survival/outcome. Although internationally accepted WHO grading system of CNS tumours is based on histological features of H&E stained sections, yet there are cases where differentiation between grade II and grade III is difficult particularly when the biopsy is small. Proliferative index derived from MIB-1 immunostaining has been found to be useful in the distinction between various grades of malignancy. Formalin-fixed paraffin-embedded surgical specimens from 90 cases of astrocytic tumours, 30 each of low-grade astrocytoma (grade II), anaplastic astrocytoma (grade III), and glioblastoma multiforme (grade IV), were immunostained by standard indirect immunoperoxidase technique using MIB-1 monoclonal antibody. MIB-1 labeling index (MIB-1 LI) was calculated. The mean MIB-1 LI values of astrocytomas, anaplastic astrocytomas and glioblastomas were 1.75 +/- 1.5%, 8.74 +/- 6.2%, and 20.54 +/- 12.2% respectively and there was statistically significant difference between grade II and III (Unpaired "t" test, T value 5.907, p value < 0.001) and grade III and grade IV (T value 4.734, p value < 0.001). The statistical analysis also revealed that the mean MIB-1 LI increased with histological grade of malignancy (One way ANOVA test, p value < 0.001). This investigation further reinforces and corroborates the findings that MIB-1 LI is useful tool in assigning grading to the astrocytic tumours and hence in treatment modalities and should be used routinely.     

Correlation between histological grade, MIB-1, p53, and recurrence in 69 completely resected primary intracranial meningiomas with a 6 year mean follow-up

Sixty-nine intracranial, totally excised meningiomas were immunostained for MIB-1 and p53 protein expression. According to the 1993 WHO criteria, revised by Perry et al., the 69 meningiomas were classified into: grade I = 54 benign meningiomas, grade II = 10 atypical meningiomas, grade III = 5 malignant meningiomas. The patients were followed until death or for an average of 6.7 years. The 69 meningiomas were divided into two groups, according to the absence (n = 42) or presence (n = 27) of recurrences. In the last group we included 3 patients who died of meningioma recurrence. According to the percentage of MIB-1 positively stained cells, meningiomas were divided into three groups: <1% (n = 36), 1-10% (n = 28), >10% (n = 5). We found the MIB-1 labeling index (LI) <1% in 33 grade I (61%) and in 3 grade II (30%) meningiomas. On the other hand, 7 grade II (70%) and all grade III (100%) meningiomas presented a MIB-1 LI >1%. Correlation between histological grade and MIB-1 LI was statistically significant (p = 0.0006). The correlation between MIB-1 LI and follow-up was also highly significant (p < 0.001): the majority of meningiomas which did not recur (32/42 equal to 76%) were characterized by a low (<1%) MIB-1 LI. In the recurrence group MIB-1 LI was significantly higher than in the disease-free patients' group. Moreover, MIB-1 appeared to be a prognostic parameter not strongly related to the histological grade. In fact, it was significantly higher in recurrent histologically benign meningiomas, as compared with benign meningiomas without recurrence (p = 0.0006). Positive p53 protein expression (>1%) was shown in 26/45 meningiomas (57%), with an LI of 1-10% in 18 (40%) and an LI of >10% in 8 (17%) meningiomas. Although the p53 LI tended to be higher in atypical and malignant meningiomas, no significant correlation was found between the p53 expression and the recurrence (p = 0.05). The authors conclude that quantitative MIB-1 labeling is a useful technique in the routine diagnostic assessment of meningiomas, and helpful in obtaining more information about prognosis and thereby in planning the most suitable treatment.     

Pediatric intracranial ependymomas: prognostic relevance of histological, immunohistochemical, and flow cytometric factors.

The correlation between the histological features and clinical outcome remains poor in pediatric intracranial ependymomas. We performed a retrospective study of a group of 31 patients (diagnosed from 1985 to 1995) to assess prognostic implications of the current grading system, of histological and immunohistochemical features, and of ploidy status estimated by flow cytometry. Immunoexpression of a broad spectrum of antigens was evaluated, including MIB-1, topoisomerase-IIalpha, cyclin D1, glial and epithelial proteins (GFAP, EMA, cytokeratins), molecules involved in controlling apoptosis (bcl-2, caspase-3/CPP32), and p53 oncoprotein. Univariate and multivariate statistical analyses were performed to evaluate the influence of each variable on both the progression free survival (PFS) and the overall survival (OS) with at least 7-year follow up. Although we showed a significant correlation between histological grade and prognosis, the current grading system failed in predicting outcome in nearly one third of individual cases. Problems with interpathologist reproducibility were also demonstrated. The extent of surgical resection was the only clinical factor that was associated with survival. Both the PFS and the OS were significantly decreased for the following pathological variables: increased cellularity (>300 nuclei per HPF), mitotic activity of >7 per 10 HPF, increased MIB-1 labeling index (LI), topoisomerase-IIalpha LI, S-phase fraction, and p53 and bcl-2 positivity. Increased cyclin D1 LI was demonstrated to have only a marginally significant impact on PFS. A flow chart modeling was further performed to formulate a scheme for discriminating of prognostic subgroups. Based on that, p53 immunopositivity and/or MIB-1 LI of >5% (after subtotal resection) or MIB-1 LI of >15% (after complete resection) were the strongest indicators of the tumor's aggressive behavior and of a poor prognosis of the disease. Foci of hypercellularity should be specifically looked for in ependymomas for assessing the immunohistochemical studies.     

Immunocytochemical Localization of Glucose Transporter-2 (GLUT-2) in Pancreatic Islets and Islet Cell Tumors

Glucose is a major metabolic fuel in mammals and is transported into organs and cells by a facilitated diffusion which involves binding of glucose to glucose transporters (GLUTs). Among several GLUTs so far indentified, GLUT-2 is specifically localized immunocytochemically in beta-islet cells. Using immunocytochemical staining, normal pancreases and 27 cases of islet cell tumors, including insulinomas, gastrinomas, glucagonomas, pancreatic polypeptide-omas (PPomas), and a nonfunctioning islet cells tumor, were systematically stained for four different pancreatic hormones, gastrin, and GLUT-2. GLUT-2 staining in beta-islet cells was more diffuse than that of insulin immunostaining, and corresponded with the positive staining in the lateral segments of beta-cell plasma membrane, that faced adjacent beta-cells. Glucagon, somatostatin (SRIF) and PP cells stained weakly for GLUT-2, weaker than that of beta-cells. Some nonbeta cells, especially extra-islet PP cells were not stained for GLUT-2. Among islet cell tumors, insulinomas stained less strongly for GLUT-2 than normal beta-cells from the adjacent normal pancreas. Gastrinomas, glucagonomas, and PPomas stained weaker than insulinomas. Even nonfunctioning islet cell tumors were weakly stained for GLUT-2. The positive staining for GLUT-2 observed for islets cells and all islet tumors is consistent with the notion that all pancreatic islet cells and islet cell tumors utilize glucose as a major fuel, requiring transporter-facilitated diffusion of glucose into the cells of normal organ and their tumors.     

Immunohistochemical expression of p63, p53 and MIB-1 in urinary bladder carcinoma. A tissue microarray study of 158 cases

P63 is a member of the p53 family, which plays a role in the differentiation of urothelium and is supposed to play a role in urothelial carcinogenesis. P53 and MIB-1 are recognised in many studies as predictive markers of progression, but few studies in the literature have examined p63. The aims of our study were to explore the expression of p63 in bladder carcinomas and to compare this expression to p53 and MIB-1, as well as to stage and grade. Tissue microarrays were performed on 158 urothelial carcinomas (56 pTa, 45 pT1 and 57≥pT2). Immunohistochemical studies were performed with p63, p53 and MIB-1 antibodies. In our study we observed that p63 immunostaining is present in all cell layers in papillary urothelial neoplasm of low malignant potential (PUNLMP), but partially lost in non-invasive papillary urothelial carcinoma low grade (NILGC) and in pT1/≥pT2 bladder cancers. P53 and MIB-1 displayed lower expression in PUNLMP/NILGC vs non-invasive papillary urothelial carcinoma high grade (NIHGC)/pT1, but there was no correlation between the expression of p63, p53 and MIB-1. Our study demonstrates that p63 expression distinguishes between PUNLMP/NILGC and NIHGC/pT1 (p=4.10⁵). A statistical difference disserving pTa and pT1/≥pT2 with a statistical significance (p<10⁻⁶) could also be observed. P63 should be considered as an additional biomarker that might help pathologists to classify their patients.     

Histopathologic features and MIB-1 labeling indices in recurrent and nonrecurrent meningiomas.

BACKGROUND: Predicting the behavior of meningiomas based on histopathologic features alone has remained problematic. DESIGN: This study retrospectively compared several histopathologic features and MIB-1 labeling indices (LIs) in recurrent meningiomas with those of nonrecurrent meningiomas. Six histopathologic features, including mitoses, necrosis, loss of architectural pattern, hypervascularity/hemosiderin deposition, prominent nucleoli, and nuclear pleomorphism, were compared between 32 recurrent and 27 nonrecurrent meningiomas using Fisher exact tests. MIB-1 LIs (% positive tumor cell nuclei) were compared using the Wilcoxon rank sum test. RESULTS: The patients in the recurrent group included 26 women (mean age, 55 years), who developed 1 to 5 recurrences. Time intervals to the first recurrence ranged from 5 to 183 months (mean, 55 months). The nonrecurrent group included 21 women (mean age, 56 years), with follow-up ranging from 88 to 124 months (mean, 109 months). Of the histopathologic features evaluated, statistically significant differences between the recurrent and nonrecurrent groups were found only with respect to prominent nucleoli (P =.024) and nuclear pleomorphism (P <.001), both of which were more common in the recurrent group. In the recurrent group, 9 tumors were considered malignant (defined by brain invasion or metastasis) versus 2 of the nonrecurrent meningiomas. Nineteen percent of nonrecurrent tumors versus 41% of recurrent tumors had 2 or more of the 6 histopathologic features. MIB-1 LIs in the nonrecurrent group ranged from 0 to 8.3 (mean, 1.5) and were generally lower than those in the recurrent group (range, 0-32.5; mean, 5.4); no statistical difference was identified between these groups. No statistically significant difference with regard to histology or MIB-1 LIs was noted between the initially excised recurrent tumor and the most recently resected recurrence. CONCLUSIONS: Of the histopathologic features examined, only prominent nucleoli and nuclear pleomorphism were found to be statistically more common in recurrent than nonrecurrent meningiomas. The mean MIB-1 LI was higher in the recurrent than in the nonrecurrent group, although there was no statistical difference between means and there was clear overlap with regard to MIB-1 LI ranges.     

MIB-1 and PCNA immunostaining as a diagnostic adjunct to cervical Pap smear

The present study was done to determine the role of MIB-1 (Molecular Immunology Borstel) and proliferating cell nuclear antigen (PCNA) proliferative index as a diagnostic adjunct to cervical Papanicolaou (Pap) smear for the identification of ascending grades of cervical intraepithelial neoplasia (CIN) developing into cancer in the human uterine cervix. A total of 49 adequate Pap smears with consensus diagnosis were destained for immunocytochemical staining (MIB-1 and PC10). Staining was done by streptavidin-biotin method after antigen retrieval. MIB-1 and PC10 labeling index (LI) were calculated in each case and divided into three groups, i.e., <10%, 10-20%, and >20%, respectively. Statistical analysis was done by using the SPSS 10.0 package. The comparisons were made using analysis of variance (ANOVA) and independent sample t-test. Bivariate and Pearson's correlation coefficient were used to obtain correlations between different groups. Out of 49 cases, 40 cases (81.6%) showed positive immunostaining with MIB-1 and PCNA. Proliferative LI of MIB-1 and PCNA increased with the ascending grades of CIN lesions to carcinoma. The highest proliferative index (mean +/- SD) for PCNA and MIB-1 were observed for the carcinoma group (PCNA LI, 39.200 +/- 1.6865; MIB-1LI, 35.300 +/- 1.8886). A significant positive correlation between ascending grades of squamous intraepithelial lesion (SIL) and labeling indices of markers (r = 0.87 for MIB-1 and r = 0.88 for PCNA) suggests that MIB-1/PCNA proliferative markers can be used as an adjunct to cytomorphological interpretation of conventional cervical Pap smear.     

Correlation of Ki-67 and p53 with the new World Health Organization/International Society of Urological Pathology Classification System for Urothelial Neoplasia

OBJECTIVE: The present study examines p53 and Ki-67 staining patterns of the diagnostic entities included within the new World Health Organization/International Society of Urological Pathology (WHO/ISUP) classification of urothelial neoplasms. DESIGN: We retrospectively studied 151 bladder biopsies from 81 patients with the following neoplasms: normal urothelium (n = 34 biopsies); low-grade intraurothelial neoplasia (LGIUN; n = 19); high-grade intraurothelial neoplasia (HGIUN; n = 20); papillary hyperplasia (n = 4); papilloma (n = 3); papillary neoplasm of low malignant potential (LMP; n = 12); low-grade papillary carcinoma (n = 28); and high-grade papillary carcinoma (n = 31). Sections were labeled immunohistochemically with antibodies to p53 and Ki-67 (MIB-1). Two hundred cells from each lesion were visually counted, and the percentage of positive cells was tabulated without knowledge of the WHO/ISUP diagnosis. RESULTS: In flat lesions, p53 positivity was of limited diagnostic utility; the marker was present in 6 of 34 benign biopsies, 6 of 19 LGIUNs, and 10 of 20 HGIUNs. In one case in which HGIUN was present elsewhere in the bladder, 29% of the benign urothelial cells were p53 positive. In papillary lesions, p53 positivity was not seen in 4 of 4 cases of papillary hyperplasia, 3 of 3 papillomas, and 8 of 12 LMP tumors. In contrast, p53 was detected in 18 of 28 low-grade and 26 of 31 high-grade papillary urothelial carcinomas. A p53 labeling index (LI) greater than 30% was only seen in HGIUNs and high-grade papillary carcinomas. In flat lesions, an increased Ki-67 LI separated out benign urothelium (mean LI, 0.62%) from dysplasia (mean LI, 3.3%) and HGIUN (mean LI, 11.6%). In papillary lesions, Ki-67 positivity was as follows: papillary hyperplasia (mean LI, 1.1%); papilloma (mean LI, 4.3%); LMP tumors (mean LI, 2.5%), low-grade papillary carcinoma (mean LI, 7.3%); and high-grade carcinoma (mean LI, 15.7%). A Ki-67 LI greater than 10% was seen only in low- and high-grade papillary carcinomas, HGIUN, and single cases of LGIUN and papillary neoplasm of LMP. CONCLUSIONS: An increased proliferative index as demonstrated by immunohistochemical staining for Ki-67 (MIB-1) is most often seen in papillary carcinoma and HGIUN. Marked p53 positivity is also characteristic of carcinoma but may be seen in benign-appearing urothelium, suggesting a "field effect" with occult molecular aberration.     

Altered expression of cell cycle regulators in myxofibrosarcoma, with special emphasis on their prognostic implications

Myxofibrosarcoma/myxoid malignant fibrous histiocytoma (MFH) has continued to be considered a distinct entity even after recently published reassessments of pleomorphic sarcomas and MFH. Several cell cycle-regulated proteins have already been screened by immunohistochemistry with the aim of finding the reliable prognostic indicator of soft tissue sarcomas; however, it is still unknown whether their altered expression affects patient survival in myxofibrosarcoma. In this study, we evaluated the expression of p53, MDM2, MIB-1 (Ki-67), p21, p27, p16, cyclin A, cyclin D1, and cyclin E by immunohistochemistry in 45 cases of myxofibrosarcoma. First, we searched for possible clinicopathologic prognostic factors in 61 cases of myxofibrosarcoma for which follow-up data were available. In univariate analysis, large tumor size (> or =5 cm), deeply situated tumor, and high histological grade (grade 2 or 3) significantly decreased survival (log-rank test, P <0.05). Among 43 cases of myxofibrosarcoma for which immunohistochemical findings were available, high MIB-1 labeling index (LI) (cutoffs of 10 and 22.5 on average), high cyclin A LI (cutoffs 10% and 13.8% on average), low p21 LI (cutoffs 10 and 20.7 on average), and reduced abnormal expression of p16 were adverse prognostic factors. In multivariate analysis (Cox proportional hazards model), high mitotic rate (>15/10 high-power fields), p53 immunoreactivity (cutoff 10%), high MIB-1 LI (>22.5), low p21 LI (<20.7), and low p27 LI (<47.8 on average) were independent poor prognostic factors. Our results suggest that reduced expression of p21 could be considered a new parameter to be evaluated, along with classical clinicopathologic prognostic factors, for identifying those at high risk for myxofibrosarcoma.     

Proliferation in Adrenocortical Tumors: Correlation with Clinical Outcome and p53 Status

There appears to be a relationship between mitotic activity and malignant behavior in adrenocortical tumors, and carcinomas with a high mitotic index may have a poorer prognosis. This has been investigated further by quantifying and comparing the Ki-67 index using antibody MIB-1 in a series of 14 adrenocortical adenomas and 40 carcinomas. The levels have been correlated with survival and disease-free survival in carcinomas and with evidence of abnormal p53 expression as detected by immunohistochemistry. Nevertheless, many carcinomas have a low level of proliferation that may reflect varying abnormalities within the regulation of both cell division and apoptosis. Expression of bcl-2 protein, an inhibitor of apoptosis has therefore also been examined. The Ki-67 index in carcinomas was significantly higher than in adenomas, but below 4% there was overlap. There was no significant difference in survival between carcinomas with MIB-1 index <3% and those greater, but the lower group had significantly longer disease-free survival (p=0.02). There was no significant difference between p53 immunopositive and p53 immunonegative carcinomas. No tumor showed immunopositivity for bcl-2 protein. It is concluded that MIB-1 index may contribute additional prognostic information in adrenocortical tumors. Inhibition of apoptosis by bcl-2 does not appear to play a role in tumor growth.     

Cell proliferation indices,morphometry and DNA flow cytometry provide objective criteria for distinguishing low and high grade bladder carcinomas

Argyrophilic nucleolar organizer region (Ag-NOR) analysis, proliferating cell nuclear antigen (PC-NA/PC10) and MIB-1 immunohistochemistry, nuclear morphometry and DNA flow cytometry have been performed on formalin-fixed, paraffin-embedded biopsies from 50 patients with transitional cell carcinoma of the urinary bladder. The mean AgNOR count was 6.01 for the 17 grade 1 (G1), 7.59 for the 21 G2 and 13.33 for the 12 G3 carcinomas (p<0.001). The mean PCNA score was 15.03% for G1, 24.04% for G2 and 40.01% for G3 cases (p<0.001). The mean MIB-1 score was 11.31% for G1, 17.09% for G2 and 34.47% for G3 carcinomas (p<0.001). The mean nuclear area was 35.53 m² for G1, 38.65 m² for G2 and 83.62 m² for G3 cases (p<0.001). Aneuploidy rates were significantly higher (91.7%) in G3 than in G2 (42.9%, p<0.01) or G1 cases (47.1%, p<0.05) but not different for G1 versus G2 cases (p=0.94). While many overlaps of values were seen between G1 and G2 tumours, no overlaps were found between G3 and G1/G2 tumours. Significant differences of values were also found between pTa and invasive tumours (p<0.0001 for AgNOR count and PCNA score; p<0.001 for MIB-1 score and mean nuclear area; p<0.01 for DNA ploidy); however many overlaps were seen. Our findings indicate that the quantitative parameters obtained with different methods are associated with histological grade of bladder urotheliomas and may improve the grading reproducibility. In addition, the absence of overlaps between G3 and G2/G1 carcinomas supports the tendency to classify bladder urotheliomas in only two categories of malignancy.