Barbara W. Chwirot

Ultraviolet laser-induced fluorescence of human stomach tissues : Detection of cancer tissues by imaging techniques

The background for this work was several literature reports on applications of the fluorescence methods to detection and localization of human cancers. The objective of our study has been to investigate if such an approach could be applied for the detection of gastric cancers.

Detection of colonic malignant lesions by digital imaging of UV laser-induced autofluorescence.

Abstract— The objective of our study was to investigate whether digital imaging of autofluorescence could be applied in the detection of colonic malignancies. Autofluorescence was excited with a 325 nm line from a He‐Cd laser. Images were recorded in vitro in six spectral bands. The material for study was 50 resected specimens for which images of 64 areas were recorded. The main result is the observation that for a majority of malignant and premalignant lesions the intensity of autofluorescence was lower than for the corresponding normal mucosa in all of the spectral bands selected for imaging. The spectral bands centered around 440 nm and 475 nm seem to be most promising in terms of possible future clinical applications.

JARID1B expression in human melanoma and benign melanocytic skin lesions.

It has been suggested that dynamically regulated expression of the JARID1B protein is required for the continuous growth of tumors and at the same time downregulated in melanoma. The majority of the data on a role of JARID1B in maintaining tumor growth has come from in-vitro and xenografting experiments, with only one immunohistochemical study involving human tissues. We compared JARID1B expression levels in human melanomas and benign nevi and analyzed patterns of spatial distributions of positive cells among different skin layers of the lesions. The expression of JARID1B was evaluated by immunohistochemistry in formalin-fixed paraffin-embedded samples of 30 nevi, 27 primary melanomas, four lymph node metastases, and one local recurrence of melanoma. Staining for JARID1B protein was stronger in melanomas compared with nevi. We also found a significant difference in the spatial distribution of positive cells in individual skin layers of nevi and melanomas. Staining of melanocytes located in granular and spinous layers of nevi was observed very rarely, whereas for melanomas, the mean percentage fractions of positive cells present in these layers exceeded the maximum values found for nevi. The spatial patterns and expression levels of JARID1B did not change significantly with melanoma progression and were similar for primary, metastatic, and recurrent melanomas. Contrary to earlier reports, this study shows enhanced expression of JARID1B by melanoma cells and indicates that such an enhancement may be an early event in the disease progression, is not correlated with melanoma invasiveness, and therefore may not be a suitable candidate as a prognostic marker.

The value of cyclooxygenase-2 expression in differentiating between early melanomas and histopathologically difficult types of benign human skin lesions.

Early cutaneous melanomas may present a substantial diagnostic challenge. We have already reported that expression of cyclooxygenase-2 (COX-2) may be useful for differentiating between cutaneous melanomas and naevi. The purpose of this study was to examine the value of COX-2 in a challenging task of differential diagnosis of early melanomas and melanocytic naevi considered by histopathologists as morphologically difficult to unequivocally diagnose as benign lesions. The material for the study comprised formalin-fixed paraffin-embedded samples of 46 naevi (including 27 cases of dysplastic, Spitz and Reed naevi) and 30 early human cutaneous melanomas. The expression of COX-2 was detected immunohistochemically. Melanomas expressed COX-2 significantly more strongly compared with naevi. The test, on the basis of determination of the percentage fractions of COX-2-positive cells, allows for differentiation of early skin melanomas and naevi with high sensitivity and specificity. Receiver operating characteristic analysis of the test results yielded areas under receiver operating characteristics curves (AUC)=0.946±0.030 for central regions and AUC=0.941±0.031 for the peripheries of the lesions. The performance of the test in differentiating between melanomas and the naevi group comprising dysplastic, Spitz and Reed naevi was also good, with AUC=0.933±0.034 and 0.923±0.037 for the central and the border regions of the lesions, respectively. Using a more complex diagnostic algorithm also accounting for the staining intensity did not result in an improvement in the resolving power of the assay. A diagnostic algorithm using differences in the percentage fractions of cells expressing COX-2 may serve as a useful tool in aiding the differential diagnosis of ‘histopathologically difficult’ benign melanocytic skin lesions and early melanomas.

Porcine Skin as a Model System for Studies of Ultraviolet a Effects in Human Skin

The range of diagnostic and therapeutic applications of ultraviolet A (UVA) radiation has been continuously expanding. UVA radiation is a well-known mutagenic factor capable of damaging both cells and tissues. At the same time there is a very limited information on long-term consequences of irradiating the skin with different doses of UVA and long-wavelength ultraviolet B (UVB) radiation used in therapies of skin disorders. It was demonstrated that for UVA doses of 0.1 to 1000 mJ/cm2 the sensitivity of the porcine skin to the UVA-induced breaking of nuclear DNA is similar to that of the human skin. Results indicate that porcine skin may serve as a model system for population studies of the deleterious effects of UVA irradiation of the skin cells.

Cyclooxygenase-2 immunohistochemistry in human melanoma: differences between results obtained with different antibodies.

Several groups have reported that cyclooxygenase-2 (COX-2) expression is significantly enhanced in human melanomas, and that the expression of this protein may be useful as diagnostic and prognostic marker for the disease. At the same time, collective analysis of immunohistochemical data on the COX-2 expression in melanomas, presented by different researchers, shows a clear lack of consistency of reported results commonly assigned to differences in protocols used for the staining. This paper describes a study involving the parallel use of three different primary anti-COX-2 antibodies targeting different COX-2 epitopes. A surprising outcome is that although the three antibodies gave very consistent results for the COX-2 expression in keratinocytes, they showed significant differences in immunoreactivity for both melanocytic naevi and melanomas. This phenomenon has not been described before, and has implications for the selection of antibodies for studies on the diagnostic potential of COX-2 for melanoma.

Different expression of cyclooxygenase-2 (COX-2) in selected nonmelanocytic human cutaneous lesions.

The aim of our study was to elucidate the possible involvement of COX-2 in the development and/or progression of nonmelanocytic skin lesions. To evaluate the usefulness of that enzyme as a potential molecular marker, we examined the intensity and spatial distribution of COX-2 expression in selected types of such tumors using the same immunohistochemical procedure as in our earlier studies of melanocytic cancers. We examined 20 benign epithelial lesions, 11 precancerous lesions, 21 basal cell carcinomas (BCC), 14 squamous cell carcinomas (SCC) and eight fibromas. The levels of COX-2 expression detected in benign lesions and in normal skin were comparable. Elevated expression of this protein may play a role in the development of SCC, as indicated by strong immunostaining both in SCCs and precancerous lesions. Significantly stronger staining in SCCs compared to BCCs may indicate a role of COX-2 in cancer malignancy and serve as an indicator useful for differential diagnostics of the two types of cancer. Strong staining in all skin layers of SCC may help in detecting cancer cells infiltrating surrounding skin layers.

Different Susceptibility of Cells of Porcine Skin and Internal Organs to Ultraviolet A–Induced Breaking of Nuclear DNA¶

Abstract There is a continuously growing interest in medical applications of ultraviolet radiation (UV-A and long-wavelength UV-B) especially for laser surgery, phototherapy and photodiagnostics of human internal organs. UV-B and UV-A radiation is potentially mutagenic, however, there has been very little information published to date concerning the significance of possible deleterious action of such photons on cells of internal tissues. The aim of this study is to compare the sensitivities of skin cells to those of internal organs upon exposure to UV-A. To assess this sensitivity we have determined the UV-A dose-dependent frequency of nuclear DNA breaks detected with the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end-labeling (TUNEL) technique. The materials for the study were macroscopic samples of porcine skin, colon and esophagus. The UV-A dose ranged from 0.1 to 1000 mJ/cm2, which is similar to doses received by cells in regions examined with laser-induced fluorescence or by cells surrounding areas subject to a laser ablation. To reduce the influence of DNA repair processes the tissue samples were kept at a low temperature during the irradiation and were deep frozen immediately after completing the irradiation procedure. The cells of the internal organs are much more susceptible to UV-A–induced breaking of DNA than the skin cells. The percentage fractions and the spatial distributions of the damaged cells and the characteristics of the UV-A dose dependence seem to vary by type of internal organ.

Porcine Skin as a Model System for Studies of Adverse Effects of Narrow-Band UVB Pulses on Human Skin

Ultraviolet (UV) radiation has been widely used in medicine, and in recent years there has been a growing interest in narrow-band UVB therapies, especially those employing pulses of the 308-nm line of XeCl excimer lasers. Comparative studies in several skin pathologies showed that narrow-band UVB was more effective than classical broad-band UVB radiation. Simultaneously, UVB is carcinogenic and there is a need for data to establish the risk associated with phototherapies involving irradiations of human skin with different doses of narrow- and broad-band UVA and/or UVB radiation. Relevant data are sparse predominantly due to a lack of suitable model systems for study of this phenomenon. Our comparative study of human and porcine skin responses to pulses of narrow-band UVB radiation demonstrated that for doses ranging from 5 to 10,000 mJ/cm2 both skin types have similar susceptibility to UVB-induced breaking of nuclear DNA, indicating that pig skin might serve as good model for studies of sensitivity of human skin to UVB radiation.

Stromal, rather than epithelial cyclooxygenase-2 (COX-2) expression is associated with overall survival of breast cancer patients

BackgroundPrognostic value of enhanced COX-2 expression in breast cancer has been controversial for a long time. The opinions vary widely between studies. Moreover, significant majority of studies considered only COX-2 expression in cancer epithelial cells.MethodsWe examined the prognostic value of COX-2 expression in both epithelial and stromal cells using three different antibodies and three algorithms of immunohistochemical scoring and categorizing the tumours into COX-2 overexpressing groups.ResultsOur results demonstrate that COX-2 expression in stromal cells is independent prognostic factor indicating worse overall survival of patients. Such a result was obtained using each of the three antibodies and two of the algorithms used for evaluations of COX-2 expression levels. We also show that immunohistochemical assessment of the prognostic value of COX-2 expression in cancer epithelial cells depends to a large extent on a combination of primary antibodies and algorithms used for determination of the COX-2 over-expressing tumours.ConclusionsOur results indicate that stromal expression of COX-2 is independent prognostic parameter relatively insensitive to variations in sensitivity of antibodies used for its determination. Wide scatter of the published results concerning prognostic value of COX-2 expression in breast cancer tissues seems to be due to a large extent to multitude of antibodies and scoring algorithms used by different groups.