The significance of immunohistochemical expression of Ki-67, p53, p21, and p16 in meningiomas tissue arrays

Although histologic grading of meningiomas has prognostic and clinical implications, it is difficult in some cases to predict the outcome of patients. There have been several efforts to evaluate the use of different immunohistochemical markers for predicting meningioma prognosis. We analyzed the immunohistochemical expression of Ki-67, p53, p21, p16, and PTEN proteins in 130 meningiomas (64 benign, 39 atypical, and 27 malignant meningiomas) using tissue microarray. The tumors were graded according to the World Health Organization classification. There was a statistically significant correlation between the expression of Ki-67, p53, p21, p16, and the grade of meningiomas (p0.001). By ordinal logistic regression, p53 and Ki-67 were significantly associated with grade, and an increase of 1% in the labeling index of these markers resulted in an increase in the risk of raising the grade by 2.17 and 1.49, respectively. Histological grade, p53, Ki-67 labeling indices, and overexpression of p16 were strongly associated with decreased event-free survival in univariate analysis. In contrast, multivariate analysis revealed that only tumor grade is an independent factor for predicting meningioma recurrence. We conclude that the Ki-67 and p53 labeling indices are useful additional tools in discriminating atypical from benign or anaplastic meningiomas, especially in histological borderline cases.     

Correlation between SPARC (Osteonectin) expression with immunophenotypical and invasion characteristics of pituitary adenomas

Pituitary adenomas are the third most common intracranial tumors. Invasive adenomas account for only 0.1–0.2% of pituitary tumors. SPARC is a matrix glycoprotein that plays a role in progression and invasiveness of neoplasms. In this study, we examined the potential role of SPARC in invasive pituitary adenomas. Forty pituitary adenomas have been examined with histopathological and immunohistochemical techniques. The cohort has been classified into two groups as invasive (n = 25) and non‐invasive (n = 15) utilizing the Hardy classification. Formalin fixed tissues have been stained with hematoxylin eosin. Ki‐67, p53, and SPARC monoclonal antibodies have been used. We did not detect any significant difference on Ki‐67, SPARC, and p53 expression patterns correlating with the pathological subtype or invasiveness. Only 24% of invasive adenomas had Ki‐67 levels over 1%. A total of 67.7% non‐invasive adenomas had Ki‐67 levels below 1%. We did not detect any relation between SPARC levels and invasiveness of pituitary adenomas. Absence of significant SPARC expression in tumor progression, sellar dilatation, erosion and destruction suggest that SPARC scores are not related with invasiveness or progressiveness of pituitary adenomas.     

Over-expression of p53, p21 and Cdc2 in histologically negative surgical margins is correlated with local recurrence of laryngeal squamous cell carcinoma

The aim of this study was to explore the relationship between the expression of p53, p21 and Cdc2 in the early laryngeal cancer with negative pathological margins and its local recurrence. During 2004-2010, a total of 85 patients with early laryngeal cancer were selected in Tangshan Union Hospital, Hebei, China, and immunohistochemical method was used to detect the expression of p53, p21 and Cdc2 in the negative pathological margin tissues. All patients were followed up for two years to collect pathological data for evaluating the survival and tumor recurrence. Two years after surgery 14 of 85 patients with laryngeal cancer presented with recurrence (recurrent group), while 71 patients without recurrence (none recurrent group). The positive rate of p53, p21 and Cdc2 protein in laryngeal cancer tissues was 60.0% (51/85), 38.8% (33/85) and 70.6% (60/85), respectively, while that of the three proteins in the cancer adjacent tissues was 36.5% (31/85), 21.2% (18/85) and 29.4% (25/85), respectively. The differentiation and TNM stage of tumor had no correlation with the three proteins. The positive rate of p53 in the surgical margin of the recurrent group and non recurrent group was 71.4% (10/14) and 29.6% (21/71) (P = 0.003), that of p21 was 50.0% (7/14) and 15.5% (11/71), (P = 0.011) and Cdc2 was 57.1% (8/14) and 23.9% (17/71) (P = 0.030), respectively. In conclusion, p53, p21 and Cdc2 may be involved in the occurrence, development and recurrence of laryngeal squamous cell carcinoma. Overexpression of p53, p21 and Cdc2 in the surgical margin of early laryngeal cancer is closely related to local recurrence of tumor.     

Histopathologic indicators of recurrence in meningiomas: correlation with clinical and genetic parameters

Meningiomas in general are circumscribed slow-growing tumors. However, despite gross total resection, tumor relapse and patients’ outcome are still an issue. Risk stratification based on histomorphology alone remains problematic. This study explored the independent prognostic value of potential risk factors among 206 patients who underwent meningioma resection and followed-up until death or a median of 44 months. The statistical analysis considered clinical data, histomorphologic parameters, cytogenetic findings, Ki-67 immunoreactivity, and activity of tissue non-specific alkaline phosphatase (ALPL). Recurrence-free survival estimates were computed and prognostic factors were identified using Cox proportional hazards model. Independent predictors of recurrence included (1) anaplasia; (2) mitotic index ≥20/10 high-power fields; (3) subtotal tumor resection; (4) loss of short arm of chromosome 1 (1p−); and (5) Ki-67 labeling index (LI) >12%. Among totally resected WHO grade I meningiomas, neither histopathologic nor clinical parameters were predictive, whereas 1p− was the only independent prognostic factor. ALPL did not reach significance in the multivariate modeling, however, the fast and low-cost histochemical detection of ALPL expression could be proved as a highly sensitive screening method for 1p−. In particular, biologically aggressive meningiomas of histologically benign or “borderline” phenotype could be therefore identified by ALPL detection followed by 1p in situ hybridization.     

Loss of p53 expression is accompanied by upregulation of beta‐catenin in meningiomas: a concomitant reciprocal expression

Crosstalk between Wnt and p53 signalling pathways in cancer has long been suggested. Therefore in this study we have investigated the involvement of these pathways in meningiomas by analysing their main effector molecules, beta‐catenin and p53. Cellular expression of p53 and beta‐catenin proteins and genetic changes in TP53 were analysed by immunohistochemistry, PCR/RFLP and direct sequencing of TP53 exon 4. All the findings were analysed statistically. Our analysis showed that 47.5% of the 59 meningiomas demonstrated loss of expression of p53 protein. Moderate and strong p53 expression in the nuclei was observed in 8.5% and 6.8% of meningiomas respectively. Gross deletion of TP53 gene was observed in one meningioma, but nucleotide alterations were observed in 35.7% of meningiomas. In contrast, beta‐catenin, the main Wnt signalling molecule, was upregulated in 71.2%, while strong expression was observed in 28.8% of meningiomas. The concomitant expressions of p53 and beta‐catenin were investigated in the same patients. In the analysed meningiomas, the levels of the two proteins were significantly negatively correlated (P = 0.002). This indicates that meningiomas with lost p53 upregulate beta‐catenin and activate Wnt signalling. Besides showing the reciprocal relationship between proteins, we also showed that the expression of p53 was significantly (P = 0.021) associated with higher meningioma grades (II and III), while beta‐catenin upregulation was not associated with malignancy grades. Additionally, women exhibited significantly higher values of p53 loss when compared to males (P = 0.005). Our findings provide novel information about p53 involvement in meningeal brain tumours and reveal the complex relationship between Wnt and p53 signalling, they suggest an important role for beta‐catenin in these tumours.     

IGFII and MIB1 immunohistochemistry is helpful for the differentiation of benign from malignant adrenocortical tumours

AIMS: The differentiation of adrenocortical carcinomas from adenomas may be difficult based on morphology alone. Differential expression of insulin-like growth factor (IGF) II and cyclin-dependent kinase (CDK) 4 has recently been described in these tumours. The aim of this study was to investigate the diagnostic usefulness of these markers immunohistochemically. METHODS AND RESULTS: We examined 22 benign and 17 malignant adrenocortical tumours and compared IGFII and CDK4 expression with known immunohistochemical as well as morphological criteria of malignancy. Thirteen of 17 carcinomas showed immunohistochemical reactivity for IGFII, whereas all adenomas but one were negative. Intense CDK4 expression was detected in 11 of 17 carcinomas but was present in only three of 22 adenomas. The MIB1 index was >5% in 14 of 16 carcinomas and was <5% in all adenomas but one. The combination of IGFII immunohistochemistry with MIB1 index led to high sensitivity and specificity in detecting adrenocortical carcinomas. CONCLUSIONS: IGFII and MIB1 are helpful immunohistochemical markers to predict malignancy in adrenocortical neoplasms. These markers can be used in addition to clinical, gross and morphological features to establish a diagnosis in difficult cases.     

Immunohistochemical expression of Retinoblastoma gene product in normal,hyperplastic and malignant endometrium. Correlation with p53 protein expression,c-erbB-2, hormone receptors' status and proliferative activity

Alterations of the retinoblastoma (Rb) gene have been described in several human neoplasms and recently, it has been suggested that these alterations may play a role in the development of endometrial carcinomas. Paraffin sections from 31 cases of normal endometrium (16 proliferative, 15 secretory), 35 hyperplastic lesions and 89 endometrial carcinomas were investigated immunohistochemically for Rb protein (pRb) expression. The results were compared with p53 and c-erbB-2 protein expression, estrogen (ER) and progesterone (PR) receptors' status and with clinicopathological prognostic factors. pRb was expressed in normal, hyperplastic and neoplastic epithelium. Proliferative endometrium showed more intense and extensive pRb staining than secretory endometrium. pRb reactivity was heterogeneous in the hyperplastic endometrial cells. Lack or focal (< 10% of endometrial cells) pRb immunostaining was noted in 56.2% and 27% of carcinomas, respectively. In the remaining cases (16.8%) pRb staining was heterogeneous or diffuse. The absence or presence of pRb expression was independent of grade and stage. In normal proliferative and secretory endometrium, pRb expression was correlated with PR (p = 0.006 and p = 0.001, respectively), PCNA (p = 0.04 and p = 0.01, respectively) and MIB1 (p = 0.02 and p<0.0001, respectively) expression. In hyperplasias, pRb was related to PR (p = 0.016) and MIB1 (p < 0.0001) expression. In carcinomas, a relationship of pRb expression with p53 (p = 0.0015), ER (p = 0.0002), PR (p = 0.0004) and PCNA (p = 0.013) status was detected. We suggest that the absence or presence of pRb expression does not seem to be associated with the progression of endometrioid carcinoma. In addition, pRb seems to be normally regulated in relation to the proliferative growth fraction of the tumours.     

High expression of meningioma 1 is correlated with reduced survival rates in colorectal cancer patients

The identification of prognostic markers for colorectal cancer (CRC) has important clinical implications. However, the association between meningioma 1 (MN1) expression and clinical outcomes of CRC has not been fully investigated. The aim of this study was to investigate the expression of MN1 in the clinical context of CRC. We first used immunohistochemistry (IHC) staining to examine and compare MN1 expression between multiple human cancer tissues and normal tissues. Initial screening revealed that the expression of MN1 proteins was significantly higher in tumor tissues of the breast, colon, and liver than in normal tissues. In further testing conducted on 59 paired CRC samples, we observed that the expression of MN1 in CRC tissue samples was significantly higher than in adjacent normal tissues. Moreover, high MN1 expression was not significantly associated with clinicopathological characteristics. Kaplan-Meier survival analysis revealed that high expression of MN1 mRNA or MN1 protein was significantly associated with poor CRC prognosis. Furthermore, univariate Cox analysis revealed that a high MN1 score was significantly associated with prognostic factors. Multivariate Cox analysis further indicated that gender, histologic grade, tumor-node-metastasis (TNM) stage, and a high MN1 score were independent factors of overall CRC survival rates. Finally, MN1 and PCNA protein levels were positively correlated, which suggests that MN1 may be involved in the cell proliferation process during CRC formation. Our results, which confirm those of other studies, indicate that (1) high levels of MN1 expression contribute to poor CRC prognosis and (2) MN1 can serve as a novel potential biomarker in predicting the prognosis of CRC patients.     

Apoptosis is associated with atypical or malignant change in meningiomas. An in situ labelling and immunohistochemical study

Aims : Although necrosis is an important phenomenon with implications for grading and prognostication in meningiomas, the alternate form of cell death, apoptosis, has not been extensively studied. In this series, we aimed to determine whether apoptosis in meningiomas correlated with histological types and grading. We also looked for a relationship between expression of apoptosis-related genes bcl-2 , p53 , c-myc and apoptosis in meningiomas.  

Methods and results

Fifty-one meningiomas of diverse histological subtypes and grades were investigated with in-situ end-labelling of DNA fragments as well as immunohistochemical analysis of three apoptosis-related genes: p53 , bcl-2 and c-myc . Our results showed that the apoptosis index was significantly higher in high-grade meningiomas (0.12%, n  = 12) in than the benign meningiomas (0.023%, n  = 39) ( P  = 0.001) but there was no difference among the different histological subtypes of the benign meningiomas ( P  = 0.125). There is no obvious relationship between p53, bcl-2 and c-myc staining and apoptosis index in this group of meningiomas.  

Conclusion

We conclude that apoptosis is an important phenomenon in meningiomas and that it is associated with atypical or malignant changes in meningiomas. Apoptosis in meningiomas has no clearcut relationship with expression of p53 , bcl-2 and c-myc .     

Increased mucin 1 expression in recurrence and malignant transformation of salivary gland pleomorphic adenoma

Soares A B, Demasi A P D, Altemani A & de Araújo V C
(2011) Histopathology 58 , 377–382
Increased mucin 1 expression in recurrence and malignant transformation of salivary gland pleomorphic adenoma Aims: Pleomorphic adenoma (PA) is the most common salivary gland tumour with a tendency to recur (RPA) and a risk of malignant transformation. Mucin 1 (MUC‐1) plays a role in the progression of many tumours and may be a marker to predict RPA. The aim of this study was to evaluate MUC‐1 expression in different phases of the adenoma to carcinoma sequence. Methods and results: Twenty‐one cases of PA, 18 cases of RPA, three cases of RPA with focal transformation (TRPA) and 11 cases of carcinoma ex‐pleomorphic adenoma (CXPA) were analysed immunohistochemically for MUC1 expression using an antibody to MUC1/DF3. MUC1 reactivity in RPA was stronger than that observed in PA and, in all the different carcinoma groups, MUC‐1 expression was significantly higher in carcinoma than in RPA and PA. Conclusion: This study has confirmed that MUC‐1 is related to the recurrence of PA and that this molecule is associated with malignant transformation of PA with carcinoma cells overexpressing MUC‐1.     

Abundant immunohistochemical expression of dopamine D2 receptor and p53 protein in meningiomas: follow-up,relation to gender,age, tumor grade,and recurrence

Meningiomas are common, usually benign tumors, with a high postoperative recurrence rate. However, the genesis and development of these tumors remain controversial. We aimed to investigate the presence and implications of a mutated p53 protein and dopamine D₂ receptor in a representative series of meningiomas and to correlate these findings with age, gender, tumor grade, and recurrence. Tumor tissue samples of 157 patients diagnosed with meningioma (37 males and 120 females, mean age 53.6±14.3 years) who underwent surgical resection between 2003 and 2012 at our institution were immunohistochemically evaluated for the presence of p53 protein and dopamine D₂ receptor and were followed-up to analyze tumor recurrence or regrowth. Tumors were classified as grades I (n=141, 89.8%), II (n=13, 8.3%), or grade III (n=3, 1.9%). Dopamine D₂ receptor and p53 protein expression were positive in 93.6% and 49.7% of the cases, respectively. Neither of the markers showed significant expression differences among different tumor grades or recurrence or regrowth statuses. Our findings highlight the potential role of p53 protein in meningioma development and/or progression. The high positivity of dopamine D₂ receptor observed in this study warrants further investigation of the therapeutic potential of dopamine agonists in the evolution of meningiomas.     

Telomerase activity, MIB-1, PCNA, HPV 16 and p53 as diagnostic markers for cervical intraepithelial neoplasia

The present investigation evaluated the relationship between dysplasia of the uterine cervix and telomerase activity, expression of p53, MIB-1 and PCNA. Telomerase activity was measured on cervical cytobrush material from 126 women suspected of having dysplasia and 61 controls using the telomeric repeat amplification protocol. Immunohistochemistry was used to detect the tumor suppressor protein p53 and cell proliferation, the latter by MIB-1 and PCNA expression. Infection with human papillomavirus 16 was detected by PCR amplification and Southern blot hybridization of DNA extracted from the same brush material. Positive telomerase activity was found in 5 of 43 (11.6%) normal samples, 12 of 57 (21.1%) samples with inflammation or koilocytosis, 7 of 17 (41.2%) CIN 1 (cervical intraepithelial neoplasia, grade 1), 8 of 20 (40.0%) CIN 2, and 25 of 42 (59.5%) CIN 3/ CIS. Telomerase activity was significantly related to the level of dysplasia (p<0.001) and proliferation measured by MIB-1 (p=0.019), but not to the level of PCNA (p=0.445), HPV 16 status (p=0.098) or staining for p53 (p=0.271). Dysplasia was also related to PCNA, MIB1, p53, and presence of HPV 16. A sequential increase in the examined parameters, paralleling the progression of abnormality, was observed. PCNA and telomerase showed an increase in CIN 1, MIB-1 and HPV16 in CIN 2, and finally p53 in CIN 3/CIS.     

p53 expression is common in malignant mesothelioma

The p53 tumour suppressor gene has been shown to be frequently mutated in a wide range of human neoplasms. This is accompanied by increased levels of p53 protein which become immunologically detectable in pathological material. We have investigated the possibility that the differential diagnosis between reactive and neoplastic mesothelium might be resolved using a polyclonal serum raised to human p53 protein, CM-1. None of 20 cases of reactive mesothelial proliferation showed p53 immunoreactivity while 70% (14 of 20) of cases of malignant mesothelioma showed p53 staining. We can thus infer that abnormalities of p53 appear to be a common event in malignant mesothelioma and that p53 immunostaining may be of value in the distinction of malignant mesothelioma from reactive hyperplasia.     

SPARC基因RNAi慢病毒载体的构建及其在SKM-1细胞中的表达

目的:构建SPARC基因RNAi慢病毒载体获得稳定产毒的细胞,并观察其对人MDS细胞株SKM-1细胞的转染效率及其对SPARC基因的抑制效率。方法:针对已经筛选确定的SPARC基因RNAi有效靶序列,合成靶序列的OligoDNA,退火形成双链DNA,与经AgeⅠ和EcoRⅠ双酶切后的pGCSIL-GFP载体连接产生GC-shSPARC慢病毒载体,PCR筛选阳性克隆并进行测序鉴定。用GC-shSPARC、pHelper1.0和pHelper 2.0载体共转染包装细胞293T细胞,包装产生慢病毒,以293T细胞GFP蛋白的表达水平测定病毒滴度,并将获得的重组慢病毒GC-shSPARC转染SKM-1细胞,通过荧光显微镜检测转染后GFP表达情况,测定转染效率;RT-PCR和Western blot分别验证转染后SKM-1细胞SPARC mRNA及蛋白的表达。结果:经测序证实,构建出了SPARC shRNA的慢病毒载体GC-sh SPARC。包装、浓缩病毒悬液的滴度为1×109TU/mL。荧光显微镜下能直接观察到转染组细胞的GFP表达,转染效率为70%,RT-PCR、Western blot技术分别检测到GC-shSPARC慢病毒转染SKM-1细胞SPARC mRNA、SPARC蛋白表达水平较空白组明显降低(P<0.05)。结论:成功构建SPARC基因RNAi慢病毒载体,其能高效干扰SKM-1细胞SPARC基因的表达。     

Immunohistochemical analysis of p53 protein overexpression in normal,premalignant, and malignant tissues of the cervix uteri

Two hundred and thirty-eight cervical lesions ranging from normal to malignant were examined for overexpression of p53 protein. Whereas p53 protein was identified in 62 per cent of invasive squamous cell carcinomas, 11 per cent of invasive adenocarcinomas, and 7 per cent of squamous cell carcinomas in situ, no staining was found in adenocarcinoma in situ, dysplastic tissue, condyloma, and normal tissue. In 9 per cent of the positive cases of invasive squamous cell carcinomas. 5-50 per cent of the tumour ceiis were immunoreactive for p53 protein, whereas the other positive specimens were characterized by only rare p53-positive cells. We conclude that in invasive cervical carcinomas widespread overexpression of p53 protein is unusual, but occasional positive nuclei can be found frequently. Furthermore, our results indicate that altered expression of p53 protein may be involved in the progression of cervical carcinomas.     

Immunohistochemical study of MIB1 expression in phyllodes tumor and fibroadenoma

The histogenesis of phyllodes tumor (PT) and fibroadenoma (FA) is closely related, and discrimination between them by histopathological analysis is sometimes problematic. Moreover, objective criteria by which to categorize the grade of malignancy in PT are still controversial. The aim in this study is to clarify whether immunohistochemical evaluation using the MIB1 antibody, which reacts with the ki-67 antigen, correlates with the histological grade of malignancy in PT, and can discriminate between PT and FA. The 47 cases of phyllodes tumor (PT) were categorized into three groups (malignant, 4 cases; borderline, 6 cases; benign, 37 cases) according to the criteria proposed by Azzopardi (1979) and were investigated by immunohistochemistry. There were significant differences in stromal MIB1-index among the three groups (P < 0. 0001), and, unexpectedly, benign PT was easily divided into two groups according only to the MIB1-index. There were significant differences in the stromal MIB1-index (P < 0.001) and stromal cellularity (P < 0.01) between the two benign PT groups. A total of 478 cases of FA was reviewed and these were divided into 403 conventional fibroadenomas (CFA), 36 cellular fibroadenomas (CEFA) and 39 fibroadenomas with focal phyllodes structure (FAPS). All cases of CEFA and FAPS, and 140 cases of CFA were studied by immunohistochemistry. The 21/215 (9.8%) cases of FA, which were designated as FAMIB, showed a high stromal MlB1-index (more than 10/0.0625 mm2). Conversely, 77% cases of FA showed no MIB1-positive stromal cells. The incidence of MIB1-positive epithelium of FAMIB was much higher than that of FA. These results suggest that high proliferative activity may be present in both stromal and epithelial cells of FAMIB. Our study suggests that immunohistochemical evaluation using MIB1 antibody correlates with the histological grade of malignancy in PT, and can select FA with high proliferative activity. However, new objective diagnostic factors useful for discriminating FA from benign PT with a low MIB1-index should be developed.     

Higher expression of beclin 1 in human meningiomas is related to better clinical outcome and pathological grade

Meningiomas are common intracranial tumors, and most exhibit optimal prognosis; however, some meningiomas still recur and even develop malignant transformation in the following years, regardless of initial pathological grade. During these years, autophagy raises its significance in tumorigenesis and tumor suppression, both important for tumor development. The aim of this study was to elucidate the relationship between two autophagy markers, LC3B and beclin 1, with clinical and pathological parameters in patients with meningiomas. A total of 77 thin‐sectioned slides, retrospectively collected from meningioma patients, were analyzed and correlated with clinicopathological parameters. We found that expression of beclin 1 rather than LC3B correlated to better prognosis, lower pathological grade, and longer survival. Furthermore, intensity of beclin 1 was also found to be significantly related to the pathological grade. These findings indicated that beclin 1 as a protective factor predicts better prognosis and plays the role of tumor suppression in meningiomas.     

Co-expression patterns of Notch1, Snail,and p53 in grade III hepatocellular carcinoma with postoperative recurrence: a preliminary study

Background/Aims

We aimed to determine the association between the co-expression patterns of Notch1, Snail, and p53 proteins (NSP) and the postoperative prognosis of hepatocellular carcinoma (HCC).

Methods

The immunoblot data for molecular expression (147 HCC/corresponding non-HCC tissues and 15 dysplastic nodules) and the sequencing data for p53 mutations (110 HCCs) were obtained from our previous study. Data analyses were restricted to cases with HCC differentiation grade III (n=47), due to its high p53 mutation rate.

Results

Nineteen of the 47 patients (40.4%) -comprising 12 in the liver and 7 in distant organs-had relapsed at 1-2 years after surgery. There was no relationship between p53 mutation and postoperative recurrence in the grade III HCCs. Seven (87.5%) of the eight relapsed cases with Notch1, Snail, and p53 (wild) co-expression experienced recurrence only within the liver, and all tumors were smaller than 5 cm in diameter. Extrahepatic relapse occurred mostly in HCC patients with tumors larger than 5 cm in diameter, without any deviation in the NSP pattern.

Conclusions

The results of this preliminary study suggest that the co-expression of Notch1, Snail, and p53 (wild) is not inferior to the patterns with p53 mutation as an indicator of postoperative recurrence of grade III HCC.