Immunohistochemical identification of intracytoplasmic lumens by cytokeratin typing may differentiate renal oncocytomas from chromophobe renal cell carcinomas

Renal oncocytomas and chromophobe renal cell carcinomas (RCCs) share a common phenotype and both originate from the intercalated cells of the collecting duct. This makes it very difficult to differentiate between the two tumors immunohistochemically. Therefore, we studied the results of immunohistochemistry focusing on certain characteristic structures that are occasionally present in renal oncocytomas. We carried out Hale's colloidal iron staining and immunohistochemistry for various cytokeratins (cytokeratins 7, 8, 10, 10/13, 14, 18, 19 and 20, and AE1/AE3) in four oncocytomas and six chromophobe RCCs. In addition, one renal oncocytoma and one chromophobe RCC were studied using electron microscopy. Two renal oncocytomas and one chromophobe RCC were completely unstained by colloidal iron. There was no evident difference between the immunohistochemical characteristics of oncocytomas and those of chromophobe RCCs. However, in all four renal oncocytomas we identified intracytoplasmic ring-like positive reactions for some cytokeratins (at least 3 antigens of cytokeratins 7, 8 and 19, and AE1/AE3), which corresponded ultrastructurally to the intracytoplasmic lumens (ICLs). In contrast, no such structures were found in any of the chromophobe RCCs using the antibodies employed. Therefore, immunohistochemical identification of ICLs by cytokeratin typing may be useful for differentiating between renal oncocytomas and chromophobe RCCs and be more sensitive in this respect than colloidal iron staining.     

Gene expression profiling of chromophobe renal cell carcinomas and renal oncocytomas by Affymetrix GeneChip using pooled and individual tumours

Due to overlapping morphology, malignant chromophobe renal cell carcinomas (RCC) and benign renal oncocytomas (RO) may pose a diagnostic problem. In the present study, we have applied different algorithms to evaluate the data sets obtained by hybridisation of pooled and also individual samples of renal cell tumours (RCT) onto two different gene expression platforms. The two approaches revealed high similarities in the gene expression profiles of chromophobe RCCs and ROs but also some differences. After identifying the differentially expressed genes by statistic analyses, the candidate genes were further selected by a real time and normal RT-PCR and their products were analysed by immunohistochemistry. We have identified CD82 and S100A1 as valuable markers for chromophobe RCC as well as AQP6 for ROs. However, these genes are expressed at the protein level in other types of RCTs as well albeit at a low frequency and low intensity. As none of the selected genes marks exclusively one type of RCTs, for the differential diagnosis of chromophobe RCCs and ROs, a set of markers such as CD82, S100A1 and AQP6 as well as some others would be an option in routine histological laboratories.     

The participation of myofibroblasts in the capsular formation of human conventional and chromophobe renal cell carcinomas

The presence of myofibroblasts has been elucidated in neoplastic capsules of various organs. In the present article, we examine the presence of myofibroblasts in the capsule of renal cell carcinoma (RCC) and discuss the origin of the myofibroblasts. Nineteen renal tumors (conventional RCC, n=17; chromophobe RCC, n=2) with evident and totally surrounded fibrous capsule were selected. Abundant myofibroblasts were immunohistochemically observed in the capsule of the RCCs. These findings were confirmed by electron and immunoelectron microscopic studies of three conventional RCCs. Type III and I collagens were predominant in the outer and inner layers of the RCC capsule, respectively. The cytoplasm of the tubular epithelial cells in the tissue surrounding the neoplastic capsule stained positively for transforming growth factor (TGF)-beta 1. In situ hybridization detected type I collagen mRNA in myofibroblasts of the capsule. Myofibroblasts may participate in the capsular formation of conventional and chromophobe RCCs through the collagen production.     

Fine needle aspiration of chromophobe renal cell carcinoma

OBJECTIVE: To describe the cytomorphologic findings of chromophobe renal cell carcinoma (CRCC) in order to preoperatively distinguish this rare neoplasm from other primary or secondary tumors arising from the kidney or presenting as retroperitoneal masses. STUDY DESIGN: Clinical data, fine needle aspiration (FNA) and follow-up surgical specimens from 4 patients with CRCC (3 primaries and 1 metastatic to the liver) were reviewed. Electron microscopy was available for 2 histologic specimens. RESULTS: Two tumors (1 primary and 1 metastatic case) were readily identified as CRCC on FNA. The 2 remaining cases were diagnosed as renal cell carcinoma (RCC) consistent with CRCC. All tumors showed aspirates with moderate to high cellularity, with the cells arranged in small clusters and single cells. Neoplastic cells had abundant heterogeneous cytoplasm, a thickened cell membrane, nuclear hyperchromasia, nuclear outline irregularity, significant nuclear size variation, intranuclear inclusions and frequent binucleation. Histology of the 4 renal tumors was characteristic of CRCC, with positivity for Hale's colloidal iron in all cases. Ultrastructurally, characteristic cytoplasmic microvesicles were observed in the 2 cases that we studied. CONCLUSION: In the adequate clinicoradiologic setting, CRCC has distinctive cytologic features that may allow an accurate preoperative FNA diagnosis.     

Renal oncocytoma. I. Cytochrome c oxidase in normal and neoplastic renal tissue as detected by immunohistochemistry--a valuable aid to distinguish oncocytomas from renal cell carcinomas

Using a polyclonal antibody raised against bovine heart cytochrome c oxidase, the occurrence of this mitochondrial marker enzyme has been investigated in 63 kidney tumors (ten renal oncocytomas, 43 renal cell carcinomas and ten tubulopapillary adenomas) as well as in normal renal tissue by an immunoperoxidase method (PAP-technique). The differentiation between renal oncocytomas and mitochondria-rich carcinomas represents a problem of histopathology since these tumors have a different prognosis and require different patient managements. The strong immunoreactivity in renal oncocytomas contrasted with the much weaker reactivity in renal carcinomas and adenomas. Even mitochondria-rich (granular cell type) carcinomas exhibited only moderate staining intensity. Furthermore, single strongly stained oncocytes or small complexes were sometimes detected in normal renal tissue. The demonstration of marked differences in enzyme content between renal oncocytomas and granular cell carcinomas renders this method suitable for unequivocal distinction between these renal neoplasms. The antibody proved to be a valuable marker for detecting "true" oncocytic transformation in renal tumors and was useful in defining even single oncocytes or small oncocytic lesions.     

嫌色性肾细胞癌17例临床病理学分析

目的探讨嫌色性肾细胞癌的临床病理特征、诊断与鉴别诊断要点。方法对17例嫌色性肾细胞癌进行组织形态学、免疫组化染色及Hale’s胶样铁染色观察,结合文献对其临床表现、病理形态特点及鉴别诊断进行探讨。结果嫌色性肾细胞癌17例,大体肿瘤直径3-10.5cm。镜下肿瘤由嫌色细胞和嗜酸细胞构成,呈片状、梁状和腺泡状分布。嫌色细胞体积较大,多角形,胞膜清晰,胞质半透明细网状,胞核皱缩,可见核沟及核异型,核仁不明显;而嗜酸细胞胞质嗜酸,可见明显的核周空晕。免疫组化:EMA 100%阳性,CD10 52.9%阳性,Vimentin阴性,CK7 88.2%阳性,P504S29.4%阳性,CD11794.1%阳性。Hale’s胶样铁染色100%阳性。17例中12例随访6个月到3年,仅1例在术后15个月发现肝脏转移,其余均未发现复发及转移。结论嫌色性肾细胞癌是一种少见的肾肿瘤,恶性程度相对较低,预后良好。掌握该肿瘤独特的病理学特征,对鉴别其他肾上皮性肿瘤有重要帮助。     

嫌色性肾细胞癌与嗜酸细胞瘤的临床病理分析与比较

目的探讨嫌色性肾细胞癌(chromophobe renal cell carcinoma,CRCC)和嗜酸细胞瘤(oncocytoma)的临床病理学特征,提高对二者的认识和诊疗水平。方法对手术切除的5例CRCC和2例Oncocytoma进行肉眼和光镜观察、免疫组织化学染色,分析其临床病理学差异。结果 CRCC以实体结构为主、胞质半透明或嗜酸性颗粒状、核皱缩且核周有空晕;Oncocytoma以巢状结构为主,胞质嗜酸颗粒状、核圆形、核仁小。免疫组化染色显示两者均呈E-cadherin、EMA、CK20、vimentin阳性;CRCC瘤细胞CD10和CK7强阳性而S-100蛋白阴性,5名患者随访8~32个月,其中1例手术12个月后由于转移死亡;Oncocytoma瘤细胞S-100蛋白强阳性、CD10和CK7阴性,2名患者随访41个月,均未出现复发和转移。结论 CRCC和oncocytoma二者形态学各有特征,免疫表型相似,二者鉴别诊断主要在于生长方式和细胞学特征。综上所述:CRCC为惰性肿瘤,Oncocytoma为良性肿瘤,预后较好,手术切除后极少发生复发和转移。     

Cytokeratin 20 immunoreactivity in renal oncocytomas.

Cytokeratins (CKs) are a group of 20 antigenically distinct intermediate filaments, generally confined to epithelia and their neoplasms. Immunostaining for CKs, in particular coordinate staining for CK7 and CK20, has become a useful tool in diagnostic pathology. Although studies defining CK distribution in neoplasms identify 0--7.7% of renal cell carcinomas (RCCs) positive for CK20, none has described the incidence of CK20 immunopositivity in renal oncocytomas (ROs). Distinction between RCC and RO may be difficult but this distinction is clinically significant, prompting us to establish the incidence of CK20 positivity in RO. We selected fifteen surgical cases of RO from our archives and studied their immunoreactivity for CKs including CK7 and CK20; 12/15 (80%) were positive for CK20, with variation in the number of cells staining. There was also variation in the distribution of CKs within the cells, including diffuse cytoplasmic, perinuclear, and a punctate or dot-like pattern. Such punctate staining corresponds to cytoplasmic balls of intermediate filaments and has been described with CAM 5.2 in RO and CK20 in Merkel cell carcinomas. Our findings suggest that CK20 immunohistochemistry is a useful tool for distinguishing RCCs from ROs. (J Histochem Cytochem 49:919-920, 2001)     

Diagnostic value of cytokeratin 7 and parvalbumin in differentiating chromophobe renal cell carcinoma from renal oncocytoma

OBJECTIVE: To investigate the diagnostic value of cytokeratin 7 (CK7) and parvalbumin at mRNA and protein levels. STUDY DESIGN: CK7 and parvalbumin mRNA expression levels in 23 oncocytomas and 32 chromophobe renal cell carcinomas (RCCs) were examined using gene expression microarrays. Immunohistochemistry was performed using monoclonal antibodies specific for CK7 or parvalbumin in 41 chromophobe RCCs and 55 oncocytomas. RESULTS: CK7 mRNA was overexpressed in 18 of 32 chromophobe RCCs but only 3 of 23 oncocytomas. Parvalbumin mRNA was overexpressed in 15 of 32 chromophobe RCCs and only 4 of 23 oncocytomas. In contrast, CK7 mRNA underexpression was noted in 13 of 23 oncocytomas and only 6 of 32 chromophobe RCCs, while parvalbumin underexpression was seen in 14 of 23 oncocytomas but only 6 of 32 chromophobe RCCs. By immunohistochemistry, 27 of 41 (66%) chromophobe RCCs expressed CK7 diffusely compared to only 3 of 55 (5%) oncocytomas. Diffuse parvalbumin expression was seen in all 41 of 41 (100%) chromophobe RCCs and only in 26 of 55 (47%) oncocytomas. CONCLUSION: Both mRNA and protein expression levels of CK7 appear significantly higher in chromophobe RCC compared to oncocytoma (p < 0.001). Parvalbumin expression is less specific but often displays a patchy pattern in oncocytomas. Our study provides further evidence that CK7 and parvalbumin immunostains may be useful in differentiating oncocytoma from chromophobe RCC in problematic cases. Negative or patchy staining (< 50% cells) for CK7 and/or parvalbumin strongly favors the diagnosis of oncocytoma.     

Cytoplasmic microvesicles in chromophobe cell renal carcinoma demonstrated by freeze fracture

In the chromophobe cell type of renal carcinoma, cytoplasmic microvesicles (frequently with "inner vesicles") demonstrable by transmission electron microscopy are one of the most important diagnostic features. The present paper reports on these microvesicles in freeze fracture replicas. Their diameter is mainly between 140 and 300 micron, but smaller and very much larger vesicles may also occur. The vesicle membrane is devoid of, or contains only scanty intramembranous particles. Cytoplasmic invaginations, probably the precursors of "inner vesicles" can also be detected. Connections with the agranular endoplasmic reticulum, mitochondria or other cell components could not be documented. Larger vacuoles limited by a membrane and containing large numbers of microvesicles have been interpreted as autophagic vacuoles. In freeze fracture replicas, there are marked differences between the chromophobe cell and the clear cell type of renal carcinoma, providing further evidence that the chromophobe cell type is a distinct entity.     

Review of chromophobe renal cell carcinoma with focus on clinical and pathobiological aspects

In recent years, the concept of chromophobe renal cell carcinoma (RCC) has been established. Chromophobe RCCs account for about 4-6% of all renal tumors. Macroscopically, the cut surface of the tumor is generally grey-beige in color. Histologically, there are two variants (typical and eosinophilic). In the typical variant, large tumor cells with architecture of a compact tubulo-cystic pattern proliferate. The cytoplasm is abundant and shows a fine reticular translucent pattern. The cell border is thick, prominent and eosinophilic. In the eosinophilic variant, tumor cells are smaller and markedly eosinophilic, and a perinuclear halo is often seen. Histochemically, the tumor cells generally show a diffuse and strong reaction for Hale's colloidal iron staining. Ultrastructurally, tumor cells contain many cytoplasmic microvesicles (150-300 nm). In chromosomal analysis, a low chromosome number is characteristic of chromophobe RCCs, due to the frequent occurrence of a combined loss of chromosomes 1, 2, 6, 10, 13, 17, and 21. In differential diagnosis, histological distinction from oncocytomas, which share a common phenotype (intercalated cells of the collecting duct system), is most important. In this diagnostic setting, recent studies have given rise to several problems. Firstly, some cases of coexistent chromophobe RCC and oncocytoma (so-called renal oncocytosis) or cases of oncocytoma with metastasis have recently been reported. Secondly, the existence of chromophobe adenoma, which is the benign counterpart of chromophobe RCC, and an oncocytic variant of chromophobe RCC has recently been suggested. Therefore, further studies are needed to elucidate the relationship between chromophobe RCCs and oncocytomas, to confirm whether chromophobe adenoma actually exists or not, and to identify the key gene that causes chromophobe RCCs.     

Nomograms-based prediction of overall and cancer-specific survivals for patients with chromophobe renal cell carcinoma

This study built and tested two effective nomograms for the purpose of predicting cancer-specific survival and overall survival of chromophobe renal cell carcinoma (chRCC) patients. Multivariate Cox regression analysis was employed to filter independent prognostic factors predictive of cancer-specific survival and overall survival, and the nomograms were built based on a training set incorporating 2901 chRCC patients in a retrospective study (from 2004 to 2015) downloaded from the surveillance, epidemiology, and end results (SEER) database. The nomograms were verified on a validation cohort of 1934 patients, subsequently the performances of the nomograms were examined according to the receiver operating characteristic curve, calibration curves, the concordance (C-index), and decision curve analysis. The results showed that tumor grade, AJCC and N stages, race, marital status, age, histories of chemotherapy, radiotherapy and surgery were the individual prognostic factors for overall survival, and that AJCC, N and SEER stages, histories of surgery, radiotherapy and chemotherapy, age, tumor grade were individual prognostic factors for cancer-specific survival. According to C-indexes, receiver operating characteristic curves, and decision curve analysis outcomes, the nomograms showed a higher accuracy in predicting overall survival and OSS when compared with TNM stage and SEER stage. All the calibration curves were significantly consistent between predictive and validation sets. In this study, the nomograms, which were validated to be highly accurate and applicable, were built to facilitate individualized predictions of the cancer-specific survival and overall survival to patients diagnosed with chRCC between 2004 and 2015.     

Identification of biomarkers of chromophobe renal cell carcinoma by weighted gene co-expression network analysis

Background

Chromophobe renal cell carcinoma (ChRCC) is the second common subtype of non-clear cell renal cell carcinoma (nccRCC), which accounting for 4–5% of renal cell carcinoma (RCC). However, there is no effective bio-marker to predict clinical outcomes of this malignant disease. Bioinformatic methods may provide a feasible potential to solve this problem.

Methods

In this study, differentially expressed genes (DEGs) of ChRCC samples on The Cancer Genome Atlas database were filtered out to construct co-expression modules by weighted gene co-expression network analysis and the key module were identified by calculating module-trait correlations. Functional analysis was performed on the key module and candidate hub genes were screened out by co-expression and MCODE analysis. Afterwards, real hub genes were filter out in an independent dataset GSE15641 and validated by survival analysis.

Results

Overall 2215 DEGs were screened out to construct eight co-expression modules. Brown module was identified as the key module for the highest correlations with pathologic stage, neoplasm status and survival status. 29 candidate hub genes were identified. GO and KEGG analysis demonstrated most candidate genes were enriched in mitotic cell cycle. Three real hub genes (SKA1, ERCC6L, GTSE-1) were selected out after mapping candidate genes to GSE15641 and two of them (SKA1, ERCC6L) were significantly related to overall survivals of ChRCC patients.

Conclusions

In summary, our findings identified molecular markers correlated with progression and prognosis of ChRCC, which might provide new implications for improving risk evaluation, therapeutic intervention, and prognosis prediction in ChRCC patients.

     

Syncytial giant cell component. Review of 55 renal cell carcinomas

Different types of multinucleated giant cells (MGC) have been documented in tumors with osteoclast-like appearance, with trophoblastic differentiation or as tumoral malignant giant cells. A new variety of MGC has been described in renal cell carcinoma. In order to study the frequency, nature and significance of this cellular type, we have reviewed our files. To assess the presence, nature and significance of these MGC in renal cell carcinomas and associated histologic subtype. To review all malignant renal tumors diagnosed in the last 5 years in our hospital and to carry out a morphologic and immunohistochemical study in renal cell carcinomas with syncytial type MGC. 55 renal cell carcinomas were reviewed. Clear cell (conventional) renal cell carcinoma was the most common type encountered (40 cases); two of these cases showed syncytial type MGC and low grade malignancy. Microscopically the MGC contained from 5 to 40 nuclei. Immunohistochemically, mononucleated and multinucleated cells were positive for cytokeratin CAM 5.2, cytokeratin AE1/AE3 and weakly positive for vimentin. Histiocytic, muscular, neural markers, beta-HCG and alpha-fetoprotein were negative. The presence of syncytial type MGC in renal cell carcinomas is an exceptional event. Among 55 renal cell carcinomas we found two cases, both of which were of clear cell subtype and low grade malignancy. The MGC proved positive for epithelial markers and probably are the result of mononucleated tumoral cell fusion. We are unaware of the impact of this MGC in the outcome of patients; such cells appear in low grade carcinomas and do not seem to be of dismal prognosis.     

Cystinyl aminopeptidase activity is decreased in renal cell carcinomas

The involvement of peptidases in carcinogenetic processes of several tumor types has been researched in recent years. Although kidney is one of the major tissues known to express cystinyl-aminopeptidase (CAP), little is known about its role in renal neoplasia. This study analyzes fluorimetrically membrane-bound and soluble CAP activity in the three main renal cancers: clear cell (CCRCC), papillary (PRCC), and chromophobe (ChRCC) renal cell carcinomas. Overall, a marked decrease of membrane-bound CAP activity in all the three renal cell carcinomas was detected when compared with their respective surrounding non-tumor tissues. So, the tumor vs. non-tumor CAP ratios (units of peptidase per mg of protein) was as follows: 926+/-111 vs. 3778+/-276 for CCRCCs, 737+/-181 vs. 4351+/-950 for PRCCs, and 592+/-118 vs. 4905+/-935 for ChRCCs. In contrast, the soluble fraction of this enzyme displayed minor and non-significant changes when comparing tumor and non-tumor CAP activities in the whole series. After stratification by stage and grade, CCRCCs displayed significant differences: pT3 category had significantly higher levels of membrane-bound activity than pT1, and high grade cases (G3-4) had higher soluble CAP activity than low grade ones (G1-2). These data may open additional possibilities in the study of renal cell carcinoma with regard to the prognosis of patients.     

Proteomic analysis of primary cell lines identifies protein changes present in renal cell carcinoma

New markers/targets for renal cell carcinoma (RCC) are needed to enable earlier detection and monitoring of disease and therapeutic targeting. To identify such molecules, normal and RCC-derived primary cell lines have been used as a simplified model system for studying changes that accompany tumorigenesis. Short-term cultures allow enrichment of relevant cell types from tissue samples, which is balanced against the potential for in vitro changes. Examination of 3 proteins with altered expression in RCC tissue showed the maintenance of normal-tumour differences in culture, although some changes were apparent, including alteration in the isoform of aldolase. Comparative analysis of primary cell lines by 2-DE found 43 proteins up-regulated and 29 down-regulated in at least three out of five tumour cell lines. Many of the observed changes have been previously reported in RCC, including up-regulation of several glycolytic enzymes, vimentin and heat shock protein 27, validating the approach. Additionally, several novel changes in protein expression were found, including up-regulation of several proteins involved in actin cytoskeleton organisation such as radixin and moesin, two members of the septin family, and the actin bundling protein, fascin. Validation studies using Western blotting and immunohistochemistry indicate that several of these molecules may be useful as markers for RCC.