Synovial sarcoma. Evaluation of prognosis with emphasis on the study of DNA ploidy and proliferation (PCNA and Ki-67) markers

Controversy still exists regarding the validity of parameters commonly used in the evaluation of prognosis of patients with synovial sarcoma (SS). Forty-nine cases of previously untreated primary SS (23 females and 26 males, ranging in age from 7 to 81, with 31 tumors located in the lower extremity, 8 at the upper extremity and 10 at the trunchus), without regional lymph-node or distant metastases were studied. We investigated the relationship between (flow and image) DNA cytometry, proliferation activity, clinicopathologic parameters, and relapse-free and overall survival of the patients. The prognostic value of gender, age, duration of symptoms, location, compartmentalization, size, adequacy of surgical margins, residual tumor, adjuvant therapy, histologic subtype, extent of necrosis, glandular differentiation, calcification, and extent of hemangiopericytic areas, mitotic rate, amount of mast cells, blood vessel invasion, histologic (UICC and NCI) grades, DNA ploidy, percentage of cells in S and S+G2 phases, PCNA and Ki-67 labeling indices (LI), and TNM (UICC) stage of the tumors, were evaluated by univariate and multivariate (Cox hazard model) analyses. Short duration of symptoms (<12 months), biphasic SS, scarcity of mast cells (<10/10 HPF), high mitotic rate (> or =10/10 HPF), high histologic grade (grade 3), high PCNA-LI (> or =20%), high Ki-67-LI (> or =10%), DNA aneuploidy, and advanced TNM stage (stage III) were features associated with significantly shorter relapse-free and overall 5-year survival rates in the univariate analyses. Scarcity of mast cells, high mitotic rate, or high PCNA-LI were significant predictors of poor survival, in addition to TNM stage in the multivariate analyses. The amount of mast cells was inversely correlated with mitotic rate and PCNA-LI. Scarcity of mast cells, high mitotic rate, or high PCNA-LI are factors associated with poor prognosis, in addition to advanced TNM stage in patients with localized SS.     

Kinetic analysis of glucose metabolism by FDG-PET versus proliferation index of Ki-67 in meningiomas--comparison with gliomas

I compared glucose metabolism by 18F-fluorodeoxy-glucose (FDG)-PET with proliferative potentials determined by using Ki-67 in meningiomas and gliomas. Ki-67 labeling index (LI) as proliferation index was used to assess tumor aggressiveness. In FDG-PET, I measured tumor versus contralateral gray matter ratio (T/N), standardized uptake value (SUV), kinetic rate constants (k1, k2, k3) which were analyzed according to the three compartment FDG model, and kinetic cerebral metabolic rate of glucose (kCMRGl). Significantly elevated FDG uptake in T/N, kCMRGl was found in a high Ki-67 LI (above 2%) group compared to a low Ki-67 LI group in gliomas, but there was not found significant difference between these two groups in meningiomas. Tumor k3 value, an indicator of hexokinase activity, of a high Ki-67 LI group was significantly higher than that of a low Ki-67 LI group. It was found that k3 correlated with Ki-67 LI. The k3 value is useful for estimating the biological aggressiveness of meningiomas.     

Interleukin-6 in multiple myeloma: correlation with disease activity and Ki-67 proliferation index

Interleukin-6 (IL-6) is supposed to be a growth factor in multiple myeloma (MM). Applying a bioassay and a modified ELISA, we measured serum IL-6 values in 64 patients with overt MM, seven patients with smouldering myeloma (SMM), 57 patients with monoclonal gammopathy of undetermined significance (MGUS) and 40 healthy volunteers as controls. IL-6 failed to discriminate between MGUS and MM, stage I, whereas comparison of MM, stage I and stage II/III (p = 0.0143), or comparison of stable disease/remission and progressive disease (p < 0.0001) revealed significant differences. Furthermore, we found significantly higher IL-6 values in overt MM compared to SMM (p = 0.0018). Using a Ki67/CD38 immunohistological double staining method we found a significant correlation between proliferation of bone marrow myeloma cells and serum IL-6 values in 15 patients (p = 0.005). These data demonstrate that IL-6 is a parameter of disease activity in MM and, beside its role in tumor biology, may become a valuable supplement to the established risk factors when selecting patients with unfavourable clinical course for more aggressive treatment modalities.     

Immunohistochemical analysis of progesterone receptor and Ki-67 labeling index in astrocytic tumors

BACKGROUND: Intracranial tumors such as meningiomas express steroid hormone receptors but little is known regarding progesterone receptor (PR) in astrocytic tumors. The authors evaluated expression of PR in 86 astrocytic tumors in relation to tumor proliferative potential. METHODS: Paraffin embedded tumor sections were stained with polyclonal antiprogesterone antibody by the peroxidase-antiperoxidase method and with monoclonal MIB-1-Ki-67 antibody by avidin-biotin complex immunohistochemistry. RESULTS: Sixty-three of the 86 astrocytic tumors (73%) showed positive PR immunoreactivity. PR expression was observed in 4 of 9 pilocytic astrocytomas, 13 of 24 Grade 2 astrocytomas, 15 of 20 anaplastic astrocytomas, and 31 of 33 glioblastomas. In addition to the tumor cells, cells of microvascular endothelial proliferation and the smooth muscle of tumor vessel walls were frequently PR positive. Glioblastomas had a significantly higher percentage of PR positive cells compared with anaplastic (P < 0.0008) and low grade (P < 0.0001) astrocytomas. Patients with PR positive astrocytomas were of an older age than patients with PR negative astrocytomas (48.71 +/- 21.95 years vs. 37.09 +/- 24.69 years; P < 0.04). The mean Ki-67 labeling index (LI) was significantly higher in the high grade (3-4) astrocytomas compared with low grade (1-2) astrocytomas (P < 0.0001). PR positive astrocytic tumors had higher Ki-67 LI than PR negative tumors. PR expression was not correlated with tumor recurrence and patient survival. CONCLUSIONS: The current study suggests that PR in the astrocytic tumors correlates with histologic grade and PR may participate in the growth of these tumors and tumor angiogenesis. The measurement of PR in these tumors may indirectly represent tumor growth potential.     

Enhanced expression of Ki-67, topoisomerase IIalpha, PCNA, p53 and p21WAF1/Cip1 reflecting proliferation and repair activity in UV-irradiated melanocytic nevi

To investigate the effect of ultraviolet (UV) irradiation on the expression of cell cycle-associated proteins, melanocytic nevi from healthy volunteers were partially covered, irradiated with a defined UV dose, and excised 1 week thereafter. The irradiated and the protected parts were examined separately by conventional microscopy and immunohistochemistry using the antibodies Ki-S11 (Ki-67), Ki-S7 (topoisomerase IIalpha), PC10 (proliferating cell nuclear antigen [PCNA]), DO-7 (p53), 6B6 (p21WAF1/Cip1), and the melanocytic marker HMB-45. DNA nick-end labeling was used as a marker of apoptosis. Irradiation resulted in morphological changes and increased HMB-45 reactivity. Proliferation, as assessed by Ki-67 and topoisomerase IIalpha expression, was also clearly enhanced in the UV-exposed areas. This was confirmed by the appearance of occasional mitotic figures. PCNA expression levels markedly exceeded those of the proliferation markers and did not correlate with the latter in most cases. p21 immunolabeling indices were also consistently augmented after UV exposure; hence it is likely that growth-inhibitory mechanisms partly compensate for the proliferative impulse, and the disproportional rise in PCNA expression probably reflects DNA repair activity. Enhanced p53 immunostaining in four cases suggests that the induction of p21 after irradiation may be p53 mediated, whereas no concomitant apoptotic events were observed. We conclude that UV light can stimulate the proliferative activity of melanocytes in melanocytic nevi, but that simultaneously cell cycle inhibitors are activated to permit DNA repair.     

Immunohistochemical expression of p53, MDM2, Ki-67 and PCNA in central giant cell granuloma and giant cell tumor

Central giant cell granuloma (CGCG) is a reactive bone lesion that occurs mainly in the jaws. The giant cell tumour (GCT) is a benign locally aggressive neoplasm located near the articular end of tubular bones. Both lesions are characterised histologically by multinucleated giant cells in a background of ovoid to spindle-shaped mesenchymal cells. There is a basic question whether both lesions are separate entities or variants of the same disease. The study of cell cycle-associated proteins may give insights into clarifying such question. The expression of these proteins is also important to determine the cell cycle regulation in both tumours. The purpose of this study was to evaluate the immunohistochemical expression of p53, MDM2, Ki-67 and PCNA in CGCG and GCT. The results demonstrated that, despite the lack of p53 immunoreactivity, all the samples showed wide expression of MDM2. The percentage of Ki-67- and PCNA-positive cells in CGCG was statistically higher than that of GCT Our findings show that CGCG has a higher proliferative activity compared with that of the GCT. Our results also suggest that p53 inactivation by MDM2 expression may be involved in the pathogenesis of giant cell lesions of the jaws and long bones.     

Number of nucleoli and nucleolar organizer regions per nucleus and nucleolus--prognostic value in canine mammary tumors

The carbohydrate residues of the surface coat of 20 axenic cultures of Blastocystis hominis were studied using FITC-labelled lectins (ConA, WGA, DBA, HPA, SBA, PNA, UEAI and LPA). The specific affinity of reactive lectins was determinated by competitive inhibition assay with specific carbohydrates or by enzymatic pre-treatment of cells. All stocks strongly bound ConA and HPA; WGA, UEAI and LPA were partially reactive, and the remaining lectins were nonreactive. Inhibition assays showed abolition (WGA, LPA, UEAI and HPA) or partial reduction (ConA) of lectin affinity, which demonstrated the specificity of binding assay. These results indicate that B. hominis has surface components containing alpha-D-mannose, alpha-D-glucose, N-acetyl-alpha-D-glucosamine, alpha-L-fucose, chitin and sialic acid.     

Accumulation of p53 and Ki-67 expression do not predict survival in patients with fibrillary astrocytomas or the response of these tumors to radiotherapy

OBJECTIVE: Although radiotherapy is often used in the treatment of patients with low-grade astrocytomas, its value is still uncertain. Radiotherapy carries a risk of morbidity for patients and has time and cost implications for health services. We have assessed the value of two histological variables, p53 accumulation and Ki-67 expression, in predicting the response of astrocytomas to radiotherapy. The former antigen was assessed because many astrocytic tumors show mutations in the p53 gene, the function of which is crucial for mediating cell death after radiotherapy, and the latter was assessed because it is expressed only in proliferating tumor cells, which may show greater radiosensitivity than nonproliferating cells. METHODS: Immunohistochemistry was used to detect the accumulation of p53 and expression of Ki-67 in a retrospective series of 96 patients with supratentorial fibrillary astrocytomas, 58 of whom had received postoperative radiotherapy. The immunohistochemical data were correlated with survival after radiotherapy. RESULTS: There was no significant difference in survival between the patients who did and those who did not receive radiotherapy. The p53 and Ki-67 labeling indices did not correlate with survival in either the irradiated or the nonirradiated cohort, nor with overall survival in the series as a whole. CONCLUSION: Immunohistochemical assessment of p53 accumulation and Ki-67 expression does not help in predicting the survival of patients with supratentorial fibrillary astrocytomas or in predicting whether particular patients are likely to benefit from radiotherapy.     

Ki-67 staining in benign, borderline, malignant primary and metastatic ovarian tumors: correlation with steroid receptors, epidermal-growth-factor receptor and cathepsin D

Ki-67 immunoreactivity was studied in relation to immunohistochemically assessed expression of epidermal-growth-factor receptor (EGFR), estrogen receptor (ER) progesterone receptor (PgR) and cytosolic levels of cathepsin D in advanced human ovarian adenocarcinomas, borderline and benign cystadenomas and normal ovaries. A significantly higher number of Ki-67-positive cells were found in metastatic tumors vs. primary adenocarcinomas and in the total group of adenocarcinomas vs. benign/borderline cystadenomas. Cathepsin-D levels were also significantly higher in metastatic tumors than in primary adenocarcinomas, which in turn presented higher levels than were found in normal ovaries. However, no significant difference was observed between cathepsin-D levels in malignant adenocarcinomas and borderline/benign cystadenomas. Immunohistochemically assessed expression of ER and PgR was detected in variable percentages of epithelial tumor cells, and stromal cells were occasionally positive as well. In the group of primary adenocarcinomas, 46% were ER-positive and 34% were PgR-positive, although there was no significant difference between primary and metastatic lesions with respect to ER or PgR expression. Concordance between immunohistochemically assessed ER or PgR data and cytosolic ER and PgR levels measured with enzyme immunoassay was relatively low. EGFR, immunohistochemically assessed with MAb-EGFRI, was positive in 76% of the primary and in 78% of the metastatic adenocarcinomas. A strong positive association was detected between ER and PgR, and EGFR was observed to present a weak positive correlation with Ki-67 and ER. Cathepsin-D levels were not found to be significantly correlated with the expression of ER, PgR, EGFR or Ki-67.     

Use of in situ detection of histone mRNA in the assessment of epidermal proliferation: comparison with the Ki67 antigen and BrdU incorporation

The labelling index is commonly used as a measure of proliferation. However, the use of tritiated thymidine or BrdU labelling of S-phase cells is limited to prospective samples. We have employed an oligonucleotide cocktail complementary to the mRNA species encoding the replication-dependent histones H2B, H3 and H4 for non-isotopic in situ hybridization (NISH), and have compared the resultant proliferation indices in normal skin with those obtained by bromodeoxyuridine (BrdU) incorporation and by Ki67 immunohistochemistry (IHC) using the monoclonal antibody MIB1. In addition, we compared the staining characteristics of histone NISH and Ki67 IHC in a further 25 samples from a variety of inflammatory dermatoses and neoplastic conditions, as well as from normal skin. In normal skin, S-phase (histone NISH and BrdU) and cycling (Ki67) cells were confined to the basal and low suprabasal layers. The labelling indices determined by histone NISH and BrdU incorporation were similar, whereas that of Ki67 IHC was four times greater. In biopsies from hyperproliferative dermatoses and dysplastic or malignant lesions, the number of histone NISH- and Ki67 IHC-positive cells was generally elevated; in accordance with the differential expression of these two markers during the cell cycle, MIB1 consistently gave higher results. The advantage of histone NISH over Ki67 IHC is that it is a marker of the same part of the cell cycle as BrdU incorporation. However, the combined use of both histone NISH and Ki67 IHC to measure two cell cycle parameters, namely S-phase and the number of cycling cells, allows more detailed retrospective study of epidermal proliferation than has been possible previously.     

An observation of the effect of tanshinone on cancer cell proliferation by Brdu and PCNA labeling

The aim of this study was to find out the anti-cancer activity of Tanshinone and its mechanism of action. Human hepatic carcinoma cell line (SMMC-7721) and leukemia cell line (HL60) were treated with tanshinone the cancer cell proliferation indices were measured by Brdu Labeling and immuno-histochemical stain of PCNA. The Brdu labeling rates of human hepatic carcinoma and leukemia cells treated with tanshinone were 8.95% and 19.01%, which were lower than those of controls (28.0%, 25.57%) respectively (P < 0.01), PCNA positive rates were 57.0% and 30.32%, which were significantly lower than those of controls (74.3%, 47.05%) (P < 0.01). The results indicate that Brdu labeling and PCNA detection may have important utility in the studies of tumor cell proliferation and the relative factors affecting the cell proliferation. The inhibitory effect of tanshinone on cancer cell proliferation might be associated with inhibiting DNA synthesis, PCNA expression and activity of DNA polymerase delta of the tumor cells.     

The proliferation of mouse mammary epithelial cells in response to specific mitogens is modulated by the mammary fat pad in vitro

A volume-preserving three-dimensional smoothing approach is described that can be directly applied to 3D medical image data consisting of sets of 2D image slices, e.g., segmented intravascular ultrasound image sequences. Two local smoothing filters ℱ and 𝒢 were designed according to different smoothing goals and their performance was compared. Filtering with the ℱ filter of a relatively large frequency window keeps the important local characteristics of the object and results in little shrinkage while removing noise. Filtering with the Gaussian filter G that has an added volume compensation step results in no global shrinkage and may be used for multiscale filtering. The two filters can be easily extended to n-dimensional filtering.     

Chemoprevention of radiation-induced mammary tumors in rats by bezafibrate administered together with diethylstilbestrol as a promoter

Pregnant Wistar-MS strain rats were irradiated with 2.6 Gy of gamma-rays at day 20 of pregnancy. Rats in the control group (n = 48) were then implanted with a diethylstilbestrol (DES) pellet at 35 days after weaning, while being fed a control (MB-1) diet. The incidence of mammary tumors was 89.6% within 1 year. In the experimental group (n = 22), a bezafibrate (0.15%) diet was initiated immediately after weaning, and 35 days after weaning a DES pellet was implanted. Administration of dietary bezafibrate together with DES-implantation continued for a period of 1 year, at which time the experiment was terminated. The incidence (27.3%) of the mammary tumors in the bezafibrate-fed rats was less than one-third of that in the control rats. Compared with the control group, the number of mammary tumors per tumor-bearing rat in the bezafibrate-treated group was reduced. For clarification of the mechanism of the chemopreventive effects of bezafibrate, lipid and hormone concentrations in serum were measured. Bezafibrate-fed rats showed a significant decrease in serum prolactin (56%) and triglyceride (63%) concentrations, and a significant increase in serum estradiol-17beta (3.8-fold), cholesterol ester (2.0-fold) and TSH (2.0-fold) concentrations in comparison with the control rats. The bezafibrate diet inhibited the formation of DES-induced pituitary tumors. However, the development of mammary glands in the bezafibrate-fed rats was stimulated more than that in the control rats treated with DES alone. The present results demonstrate that bezafibrate is effective in preventing mammary tumors induced by radiation together with DES, possibly by reducing prolactin and triglyceride concentrations.     

Distinction of basaloid carcinoma of the prostate from benign basal cell lesions by using immunohistochemistry for bcl-2 and Ki-67

The distinction of rare basaloid carcinomas (BC) of the prostate from more common basal cell hyperplasia may be difficult, because basal cell hyperplasia (BCH) may have prominent nucleoli and may appear infiltrative. Using immunohistochemistry, we studied bcl-2 and p53 expression and Ki-67 proliferation index in eight cases of typical BCH, eight cases of BCH with nucleoli, and six cases of BC. Bcl-2 expression (P < .0001) and Ki-67 index (P=.005) were elevated in BC compared with typical BCH or BCH with nucleoli, whereas there was no significant difference between typical BCH and BCH with nucleoli. P53 was not discriminative in separating benign from malignant basal cell lesions of the prostate. Bcl-2 may play a role in the pathogenesis of basal cell lesions of the prostate. Elevated expression of bcl-2 and higher Ki-67 index may aid in the diagnosis of basal cell proliferative lesions of the prostate.     

Immunohistochemical study on the prognostic value of the expression of laminin and Ki-67 receptors in advanced gastric cancer

The expression of 67-KDa laminin receptor (LR) was investigated in a group of 75 patients who underwent curative gastrectomy for advanced gastric cancer, with special reference to the possible role in the tumor progression and in the overall survival. In 56 out of these 75 patients also the prognostic significance of proliferative activity was investigated using the monoclonal antibody Ki-67. The tumor LR expression and the Ki-67 labeling index (Ki-67 LI) were immunohistochemically determined in paraffin-embedded sections using the avidin-biotin immunoperoxidase method. The cumulative 5-years survival rate was 75.1% for patients without expression of LR, 52.6% for those with positive LR expression. Significant association between LR expression and depth of tumor invasion (p = 0.022) was found. By univariate analysis the presence of laminin receptor seemed to be associated with an higher risk of death (RR1.73-95% C.I. 0.71-4.20), but this effect disappeared after controlling for depth of tumor invasion. There was no significant relationship between the Ki-67 LI and wall invasion (p = 0.80) or nodal status (p = 0.73). The cumulative 5-year survival rates (95% CI) were 61.0% (35.3-79.2) in patients with Ki-67 index < 10%, 52.4% (29.7-70.9) with Ki-67 index = 10%-40%, 52.9% (27.6-73.0) with Ki-67 index > 40% and the differences were not statistically significant (p = 0.93). Also in multivariate analysis the proliferative activity did not independently affect survival (p = 0.98). An interaction between Ki-67 index and age was found and Ki-67 index > 40% was significantly associated with a poor prognosis in patients over 70 years old old (p = 0.002). In conclusion, tumor expression of laminin receptor could be correlated with gastric cancer aggressiveness, however its prognostic significance is already provided by depth of tumor invasion. The proliferative activity, determined with the monoclonal antibody Ki-67, does not seems to influence the survival except in elderly patients (> or = 70 years old).     

Hydroxyurea synchronization of cellular proliferation in the esophageal epithelium of mice with tumors

The synchronizing effect of hydroxyurea on the passage of the esophagus epithelia cells through the stage of the DNA synthesis and mitosis was investigated in albino mice bearing sarcoma. Dinrnal variations in the number of epithelial cells at these stages of the mitotic cycle were taken into consideration. Two mutually complementary indices for the measurement of the shifts in the cell population and the rate of the synchronization are used. Two cell groups which passed the S-phase and mitosis synchronously were found in artificial synchronization in the esophageal epithelium. The synchronization indices characterizing the first group were lower than control. In the second group the cell count during the DNA synthesis was double as compared with the number of cells synthesizing DNA in natural synchronization; however, the rate of the changes in the synchronism was the same in the experimental and control groups.     

Ras gene analysis in mammary tumors of dogs by means of PCR-SSCP and direct genomic analysis

The oncogenic capacities of RAS family genes (Ha-ras, Ki-ras, and N-ras) are usually activated by point mutations in the conserved regions (codons 12, 13, and 61), resulting in single amino acid substitution in the specific proteins (p21). In order to verify the involvement of RAS genes in dog mammary tumors we analyzed the genomic DNA from 20 mammary tumors of dog by means of the Polymerase Chain Reaction-Single Strand Conformation Polymorphism (PCR-SSCP) method and the direct genomic sequencing. The absence of point mutations in the "hot spots" of RAS genes suggests a lack or a low frequency of such a pattern of RAS genes activation in dog mammary tumors. The results are also in agreement to what reported in human mammary tumors. However, the presence of genetic alterations in other functional areas of the RAS genes or other mechanisms of activations cannot be ruled out.