Search

There is increasing evidence that Annexin AI (ANX AI) expression is dysregulated in several carcinomas and tumour cell lines. In order to gain insight into the putative role of ANX AI in tumorigenesis, clinical outcome and metastatic potential of conventional renal cell carcinomas (CRCCs) we investigated the expression of ANX AI in CRCCs and metastases. Furthermore, it was elucidated whether ANX AI overexpression affects migratory potential in Caki-1 cells. ANX AI immunohistochemistry was performed on 33 samples of CRCCs and 10 metastases. ANX AI expression was assessed in 12 samples by 2-dimensional gelelectrophoresis (2-DE), subsequent mass spectrometry and RT-PCR. Immunohistochemical data were statistically correlated with pathological parameters, amount of eosinophilic cells and clinical outcome. Furthermore, a haptotactic migration assay was done on Caki-1 cells transfected with ANX AI. Immunostaining for ANX AI was found in 18 tumours and all metastases investigated. Intensity of immunohistochemical staining correlated to Fuhrman grade, amount of eosinophilic cells and clinical outcome. 2-DE and RT-PCR confirmed the presence of ANX AI in neoplastic tissue. Overexpression of ANX AI did not significantly influence cell migration. From these findings ANX AI expression seems to be related to Fuhrman grade, clinical outcome and metastatic potential of CRCCs. Thus ANX AI could serve as a prognostic marker for tumour progression.

Clinical outcome of penile squamous cell carcinoma (PSCC) largely depends on the presence of lymph node metastasis. In search of a valuable marker predicting the risk for metastasis, the expression of Ki67 was investigated immunohistochemically in primary tumors and compared to presence of inguinal lymph node metastasis. As human papilloma virus (HPV) is thought to affect Ki67 expression, we evaluated whether occurrence of HPV DNA correlates to Ki67 score or metastatic potential. Samples originated from patients subjected to resection of invasive SCC of penis. Immunohistochemistry was done on paraffin-embedded sections using a monoclonal antibody against Ki67. After DNA isolation from paraffin embedded tissue the presence of HPV 6/11, HPV 16 and HPV 18 DNA was analyzed by PCR. Statistical analysis was done using two tail unpaired t test and Chi-square test. Four of 28 patients showed a weak Ki67 expression, without displaying lymph node metastasis. Among 17 patients showing an intermediate Ki67 index, eight exhibited metastases while in all seven patients with a strong expression of Ki67 lymph node metastases were found. The median Ki67 expression in metastastic lesions was significantly different (50.3%) from tumors without lymph node metastasis (31.8%) (p=0.024). Furthermore, a correlation between presence of HPV DNA and strong Ki67 expression was determined (p=0.009). Since our study demonstrated a strong Ki67 labeling index significantly associated to positive lymph nodes, we suggest Ki67 expression as a prognostic marker for lymph node metastasis in penile squamous carcinoma.

Annexins (ANXs) represent a family of calcium and phospholipid binding proteins that are involved in several physiological processes e.g. signal transduction, cellular differentiation and proliferation. Since they are known to be dysregulated in a variety of cancers we investigated the immunolocalization of ANXs in whole prostate sections containing benign prostatic epithelium (BPE), benign prostatic hyperplasia (BPH), prostatic intraepithelial neoplasia (PIN) and prostate cancer (PCa) in order to evaluate their possible role during tumorigenesis. Samples were obtained from 28 patients undergoing radical prostatectomy. Gross sections of whole prostates were examined immunohistochemically for the distribution of ANX I, II, IV and VII. In BPE all ANXs were localized to the cell membranes and the cytoplasm of all gland cells. In BPH the immunoreactivity of ANX I and II was restricted to the basal cells of glands and expression pattern of ANX IV and VII was similar to BPE. In PIN only basal cells expressed ANX II. In PCa ANX II immunoreactivity was absent and weak ANX I and ANX IV immunoreactivity was restricted to the cytoplasm of tumor cells. ANX VII immunoreactivity was seen in some but not all tumor cells. Since ANX IV and VII expression did not show significant changes in PCa compared to non-neoplastic tissue and PIN an essential role during prostate tumourigenesis seems unlikely. In contrast, as progression from PIN to PCa is characterized by a reduction of ANX I and II this suggests that downregulation of these proteins could represent an important event in prostate carcinogenesis.

Skp2 (S-phase kinase associated protein 2) controls progression from G- to S-phase by promoting the proteolysis of the cyclin dependent kinase inhibitor p27KIP1. Despite the fact that a p27KIP1 decrease has been documented in melanoma progression, the role of Skp2 in these tumours is unknown. We therefore examined by immunohistochemistry the expression of Skp2, p27KIP1 and Ki-67 in 10 naevi (Ns), 15 superficial spreading melanomas (SSMs), 10 nodular melanomas (NMs) and 14 melanoma metastases (Ms). Nuclear Skp2 expression augmented with increasing malignancy (Ns: 1.4%, SSMs: 5.6%, NMs: 17.3%, Ms: 19.1%). In all tumours nuclear Skp2 expression correlated with Ki-67 (p=0.024) and inversely with p27KIP1 (p=0.007). A cytoplasmic reaction for Skp2 was also observed in most tumours and its expression decreased from Ns (12.3%) to SSMs (7.9%) and NMs (4.5%). In contrast, Ms showed an increase of cytoplasmic Skp2 (11.9%) that correlated with its nuclear expression (p=0.016). While nuclear Skp2 expression correlated with the pT-level (p=0.023), Clarklevel (p=0.023) and Breslow index (p=0.019), the cytoplasmic Skp2 expression might be of biological significance only in NMs since it correlated with tumour depth (p=0.02) and pT-level (p=0.025). Our data suggests that Skp2 could contribute to melanoma progression. This is further highlighted by the fact that vertical growth phase (VGP) melanomas show significant higher nuclear Skp2 expressions when compared with the harmless radial growth phase (RGP) (p=0.047). Also nuclear Skp2 expression correlates with a reduced survival time (p=0.025) in melanoma.

The detection of structural and numerical chromosomal aberrations is an important part of the characterization of tumors and genetic diseases. The direct demonstration of DNA sequences in interphase nuclei and metaphases by fluorescence in situ hybridization (FISH) has been termed interphase cytogenetics. It has been proven as a powerful technique to detect specific aberrations in a wide variety of cell types, including paraffin-embedded tissue. Nowadays a standard method in leukemia and lymphoma, interphase cytogenetics contributes mainly to the diagnosis in these tumors and helps to classify soft tissue tumors. Therefore FISH is mandatory for the choice of therapy in these tumors. In contrast to the aforementioned, up to now, the value of FISH in solid tumors is mostly limited to pure research and contributes in this way to our understanding of tumor biology. But with the use of paraffin-embedded tissue and the first results obtained, it seems very likely that a direct correlation between histological classification and cytogenetic characteristics of solid tumors can be achieved in the near future. This information might not only provide insights into tumor biology, but could also contribute to a different tumor classification, a sort of risk estimation, where we might predict the possible biological behavior of solid tumors. This could greatly influence further therapeutic decisions thus establishing the FISH technique as an indisputable part in the diagnosis of solid tumors.

Oxygen radicals and oxidatively modified proteins seem to participate in degenerative vascular and inflammatory diseases. Factors that contribute to the development of atherosclerosis, eg, oxidation of low-density lipoproteins (LDLs), may also contribute to glomerulosclerosis. Although the nature of the in vivo oxidants remains unknown, recent findings indicated that the myeloperoxidase (MPO)-H2O2-halide system could play an important role in modification of (lipo)proteins in human tissues. MPO, the enzyme responsible for hypochlorite (HOCl/OCl-) formation, is present in human atherosclerotic lesions and in inflammatory conditions. In the present study, MPO was identified by Western blot analysis and immunohistochemical technique in diseased human kidney either with primarily sclerotic or inflammatory lesions. Furthermore, the presence of HOCl-modified proteins was demonstrated in diseased renal tissues using a specific monoclonal antibody (clone 2D10G9), raised against HOCl-modified LDL, that does not cross-react with native LDL or Cu(2+)-, 4-hydroxynonenal-, or malondialdehyde-modified LDL. The antibody recognized HOCl-modified proteins in glomerular and tubulointerstitial inflammatory and fibrotic lesions and pronounced immunostaining was demonstrated in mononuclear cells. LDL or human serum albumin oxidized by HOCl in vitro, but not native LDL or human serum albumin, effectively competed with epitopes in diseased kidney for antibody binding. Western blot analysis in diseased kidney protein samples revealed at least two major proteins recognized by the anti-HOCl-modified protein monoclonal antibody. Densitometric evaluation of immunoreactive bands obtained under these conditions demonstrated that expression of HOCl-modified proteins is tightly coupled to expression of immunoreactive MPO in the same tissue samples. From our studies it is proposed that oxidation of proteins by HOCl might be a leading event in glomerular and tubulointerstitial injury. By this mechanism, mononuclear cells, a permanent source for MPO, may play a key role in the development of nephrosclerosis, glomerulo-clerosis, and tubulointerstitial fibrosis, respectively.