Can obstructive intralymphatic granulomas be the cause of cheilitis granulomatosa?

Cheilitis granulomatosa (ChG), also known as Miescher's cheilitis, is an uncommon, immunologically mediated nonnecrotizing granulomatous inflammatory disease characterized by recurrent, painless swelling of the lips. The aim of the study was a pathomorphological and immunohistochemical assessment of cases clinically classified as ChG to investigate potential pathological mechanisms of ChG symptoms and to verify the hypothesis of intravascular granulomas as a cause of lymphatic vessel obstruction and localized edema in ChG. We report 6 patients with ChG who clinically presented localized edema of the lips. Lip biopsy with pathomorphological and immunohistochemical examination was performed in all cases. We found discrete, non-necrotizing granulomas which were adjacent to numerous blood and lymphatic vessels. The lumen of lymphatic channels was dilated and was either empty or contained lymph and few macrophages or was completely occluded by nearby granulomas. All patients demonstrated a characteristic pattern of lymphangiectasia and perivascular lymphatic aggregates with evidence of non-necrotizing granulomas. None manifested intralymphatic granulomas. These results do not support the view that lymphatic vessel obstruction is caused by intravascular histiocytic granulomas described as the main part in the etiology of lymphatic edema in ChG. However, perivascular granulomas and dilation of lymphatic vessels confirm presence of inflammatory lymphostasis in all studied cases of ChG.

A novel canine B-cell leukaemia cell line. Establishment, characterisation and sensitivity to chemotherapeutics.

We established a new B-cell leukaemia cell line CLB70 from a dog with chronic lymphocytic leukaemia. This cell line is positive for CD20, CD45, CD79a, MHC class II, IgG, IgM; weakly positive for CD21; and negative for CD3, CD4, CD5, CD8, CD14, CD34, CD117. PCR for antigen receptor gene rearrangement (PARR) analysis revealed a biclonal immunoglobulin heavy chain (IgH) gene rearrangement and negative result for TCRγ. Western blot analysis of anti- and pro-apoptotic proteins showed increased expression of Bcl-2, Mcl-1, NF-kB, and Ras, and decreased expression of p53. CLB70 cells grow rapidly in vitro and are tumourigenic in nude mice. The CLB70 line is highly sensitive to doxorubicin, less sensitive to etoposide and imatinib, and resistant to piroxicam, celecoxib and dexamethasone. Our results indicate that CLB70 cells are derived from mature B-cells and they may be a useful tool for the development of new therapeutic strategies for both dogs and humans.

Hemoglobin - a novel ligand of hepatocyte ectopic F1-ATPase.

The liver is largely responsible for free hemoglobin uptake, but the molecular mechanism of this phenomenon has never been revealed. This paper presents the results of the study on hemoglobin binding components of the hepatocyte membrane that were purified using affinity chromatography on a hemoglobin matrix and identified by peptide mass fingerprinting. Both F1-ATPase alpha and beta subunits were retrieved. The binding was confirmed via an intrinsic fluorescence quenching study using a purified recombinant F1-ATPase beta subunit, and the dissociation constant for the complex was estimated from the saturation binding curve (Kd = 7.5 x 10(-7) M). The results indicate that haemoglobin binds to hepatocyte ectopic F1-ATPase. We suggested the plausible role of the receptor in endocytosis of haemoglobin by the hepatocyte.

The role of inflammation in the progression of ocular surface damage in children following allogeneic haematopoietic cell transplantation.

The aim of the study was to evaluate HLA-DR expression and cellular morphology of the conjunctival epithelium cells in children who underwent haematopoietic cell transplantation, and to assess the relation between HLA-DR expression and cellular morphology. Impression cytology with staining was used to visualize epithelium cells, whereas immunohistochemistry was applied to assess HLA-DR expression. Elevated HLA-DR expression and increased cytological abnormalities were observed in the study group when compared to the controls. An increase in HLA-DR expression was accompanied by a decrease in the number of eyes with normal epithelium morphology together with the increase in squamous metaplasia features. We can conclude that inflammation of conjunctiva can follow stem cell allotransplantation. Ocular surface inflammation may lead to squamous metaplasia of the conjunctiva.

Podoplanin expression in cancer-associated fibroblasts correlates with VEGF-C expression in cancer cells of invasive ductal breast carcinoma.

Podoplanin (D2-40) was shown to be expressed in cancer-associated fibroblasts (CAFs) of various malignancies. The study aimed at examining its impact on angiogenesis and lymphangiogenesis markers in invasive ductal carcinoma of the breast (IDC). The studies were performed on 104 archival cases of IDC using immunohistochemical technique. Podoplanin expression in CAFs correlated positively with cancer cell VEGF-C expression (r=0.19, p=0.0495) and intratumoral microvessel count (MVC) of CD31 positive vessels (r=0.30, p=0.0018), whereas negative correlations were observed with peritumoral MVC of D2-40 and Lyve-1 positive lymphatic vessels (r=-0.26, p=0.008 and r=-0.27, p=0.0058, respectively). Podoplanin expression in CAFs did not correlate with VEGF-A and VEGF-D expression in cancer cells, nor exerted any prognostic significance. Podoplanin expression in CAFs may have impact on angio- and lymphangiogenesis processes in IDC.

Chyloperitoneum, chylothorax and lower extremity lymphedema in woman with sporadic lymphangioleiomyomatosis successfully treated with sirolimus: a case report.

Lymphangioleiomyomatosis (LAM) is a rare disease characterized by diffuse thin-walled cysts throughout the lungs on computed tomography and diffuse proliferation of abnormal smooth muscle-like cells (LAM cells) on lung biopsy. LAM affects women almost exclusively, predominantly in their reproductive age. The most typical presenting symptoms include dyspnea, spontaneous pneumothorax, cough and chylothorax. Abdominal findings represent less common initial manifestations of the disease and may pose diagnostic difficulties. The treatment of LAM has not been fully established. Recent studies report effectiveness of sirolimus in LAM patients. We report the case of a 45-year-old woman with sporadic LAM, successfully treated with sirolimus, in whom the first manifestation of the disease was chyloperitoneum and after three and nine years, respectively, lymphedema of the left lower extremity and right sided chylothorax occurred.

Adventitial lymphatics and atherosclerosis.

Lymphatic vessels are important in reverse cholesterol transport and play a crucial role in regression of atherosclerotic plaque in experimental animal models. Therefore, we attempted to analyze adventitial microcirculation including lymphatic vessels and adventitial macrophages in large human arteries in various stages of atherosclerosis. Eighty-one arterial segments of large arteries (iliac arteries and abdominal aortas) were obtained from deceased organ donors. Lymphatic vessels were identified using anti-LYVE-1 and anti-D2-40/podoplanin immunohistochemical staining. Adventitial blood vessels and macrophages were visualized using anti-CD-31 and anti-CD-68. Intimal thickness was measured under 100x magnification with an Olympus BX 41 light microscope using the visual mode analySIS 3.2 software. Lymphatic vessels were counted in each cross section of the examined arteries, and adventitial blood vessels (CD31+) were counted using the "hot spot" method. Statistical analysis was performed with Statistica 9.1 PL software (StatSoft, Cracow, Poland). Mann-Whitney, F-Cox, Chi-square, and Spearman's correlation tests were performed and the differences were considered significant at p < 0.05. Lymphatic and blood vessels in the adventitia of examined arteries were identified and quantified. Significant positive correlations were found between the number of adventitial lymphatics (LYVE-L +) and intimal thickness (r = 0.37; p < 0.05) as well as with age of the subjects (r = 0.3; p < 0.05). Thus, lymphatic vessels are present in the adventitia of large arteries in humans and the number of adventitial lymphatic vessels increases with progression of atherosclerosis as assessed by intimal thickness.

Ceramide galactosyltransferase (UGT8) is a molecular marker of breast cancer malignancy and lung metastases.

BACKGROUND: It was shown recently on the level of gene expression that UGT8, coding UDP-galactose:ceramide galactosyltransferase, is one of six genes whose elevated expression correlated with a significantly increased the risk of lung metastases in breast cancer patients. In this study primary tumours and their lung metastases as well as breast cancer cell lines were analysed for UGT8 expression at the protein level. METHODS: Expression of UGT8 in breast cancer tissue specimens and breast cancer cell lines was analysed using IHC, real-time PCR and Western blotting. RESULTS: Comparison of the average values of the reaction intensities (IRS scale) showed a significant difference in UGT8 expression between (1) primary and metastatic tumours (Mann-Whitney U, P<0.05), (2) tumours of malignancy grades G3 and G2 (Mann-Whitney U, P<0.01) as well as G3 and G1 (Mann-Whitney U, P<0.001) and (3) node-positive and node-negative tumours (Mann-Whitney U, P<0.001). The predictive ability of increased expression of UGT8 was validated at the mRNA level in three independent cohorts of breast cancer patients (721). Similarly, breast cancer cell lines with the 'luminal epithelial-like' phenotype did not express or weakly expressed UGT8, in contrast to malignant, 'mesenchymal-like,' cells forming metastases in nude mice. CONCLUSION: Our data suggest that UGT8 is a significant index of tumour aggressiveness and a potential marker for the prognostic evaluation of lung metastases in breast cancer.

Tumor-associated carbohydrate antigens: Sialyl Lea and T/Tn antigens in canine mammary tumors.

Twenty-eight canine mammary tubulopapillary carcinomas and 14 simple adenomas were studied by immunohistochemistry for the expressions of the tumor-associated carbohydrate antigens. Sialyl Le(a) was detected in 71.42% of the malignant and 92.84% of the benign tumors. Staining with anti-T and anti-Tn monoclonal antibodies revealed that 85.70% of the tubulopapillary carcinomas expressed T and Tn antigens. In contrast, 50% of the adenomas did not express T antigen, and 42.85% of them were only weakly stained for this carbohydrate structure. In the case of Tn antigen, the majority (57.14%) of samples was weakly stained, and no binding was observed in 35.71% of the analyzed specimens. Comparison of average values of reaction intensity (IRS) scale for malignant versus benign tumors by the Mann-Whitney U-test revealed a significant relationship between T and Tn antigens expression and type (malignant vs. benign) mammary tumors. Based on the results obtained, it is suggested that each of the studied antigens can be treated as a tumor-associated antigen of canine mammary tumors. However, only the T and Tn antigens seem to be associated with malignant transformation of mammary gland cells and to be of potential value as diagnostic markers.

Multivariate analysis of oestrogen receptor alpha, pS2, metallothionein and CD24 expression in invasive breast cancers.

Determination of oestrogen receptor alpha (ER) represents at present the most important predictive factor in breast cancers. Data of ours and of other authors suggest that promising predictive/prognostic factors may also include pS2, metallothionein (MT) and CD24. Present study aimed at determining prognostic and predictive value of immunohistochemical determination of ER, pS2, MT, and CD24 expression in sections originating from 104 patients with breast cancer. An univariate and multivariate analysis was performed. Both univariate and multivariate analyses demonstrated that cytoplasmic-membranous expression of CD24 (CD24c-m) represents a strong unfavourable prognostic factor in the entire group and in most of the subgroups of patients. In several subgroups of the patients also a prognostic value was demonstrated of elevated expression of pS2 and of membranous expression of CD24. Our studies demonstrated that all patients with good prognostic factors (higher ER and pS2 expressions, lower MT expression, CD24c-m negativity) survived total period of observation (103 months). The study documented that cytoplasmic-membranous expression of CD24 represented an extremely strong unfavourable prognostic factor in breast cancer. Examination of the entire panel of the studied proteins permitted to select a group of patients of an exceptionally good prognosis.

Immunohistochemical identification method of tumour cells in the S phase of mitotic cycle and its usefulness in diagnostics of mammary gland adenocarcinomas in bitches.

The studies aimed at identification of neoplastic cells at the S phase of mitotic cycle in mammary gland adenocarcinomas of bitches. The material was sampled from bitches of various races, aging 6 to 12 years, in which the mammary gland tumours developed spontaneously. The tumours were verified histopathologically and, then, immunohistochemical reactions were performed in order to detect cells which had incorporated BrdU (bromodeoxyuridine), contained Ki-67 or PCNA antigen. The histological preparations were photographed and obtained pictures were subjected to computer-assisted image analysis using Axiophot microscope (Carl Zeiss) coupled to a computer and the Multi-ScaneBase V 8.08 software, working under Windows. Fifty percent of sections from mammary gland adenocarcinomas demonstrated BrdU labelling index of 4-5%, 40% of 1-3%, while in the remaining 10% of examined tumours no BrdU incorporation could be demonstrated. No evident relationship could be detected between the presence of BrdU incorporation and Ki-67 or PCNA antigen presence but a significant correlation was demonstrated between the expression of Ki-67 and PCNA.

Exercise-induced apoptosis in rat kidney is mediated by both angiotensin II AT1 and AT2 receptors.

Excessive physical exercise may lead to disturbance of the entire homeostasis in the body, including damage not only in skeletal muscles but also in many distant organs. The mechanisms responsible for the exercise-induced changes could include oxidative stress or angiotensin II. We previously showed that acute exercise led to apoptosis in kidney but not as a result of oxidative stress. In this study, we examined the role of angiotensin II and its AT1 and AT2 receptors in mediation of exercise-induced apoptosis in kidney. We clearly demonstrated that acute physical exercise induced apoptosis in renal cells of distal convoluted tubuli and cortical and medullary collecting ducts. Moreover, the cells displayed an increased expression of both AT1 and AT2 angiotensin II receptors and of p53 protein. The results suggest that angiotensin II could upregulate p53 expression in renal distal convoluted tubular cells and in the cells collecting ducts via both AT1 and AT2 receptors, which might be the crucial apoptosis-mediating mechanism in kidneys after excessive exercise.

LYVE-1 expression on high endothelial venules (HEVs) of lymph nodes.

LYVE-1 (lymphatic endothelium hyaluronan receptor) has been identified as a powerful marker for lymphatic endothelium. Apart from lymphatic endothelium, LYVE-1 is expressed in normal liver blood sinusoids, spleen endothelium and activated tissue macrophages. LYVE-1 has not been detected in blood vascular endothelium with the exception of blood vessels in the lung. High endothelial venules (HEVs) belong to the vascular compartment of lymph nodes. They are the major site of entry for circulating lymphocytes into the node. HEVs are characterized by cuboidal endothelial cells, the existence of discontinuous junctions between these endothelial cells, and the presence of large numbers of lymphocytes within their walls. 40 paraffin-embedded lymph node biopsy specimens from newly diagnosed patients with non-Hodgkin lymphoma were evaluated as well as 10 lymph node biopsy specimens from adult patients with reactive lymphadenitis, and 10 normal, non-metastatic lymph nodes obtained from adult patients during cancer surgery served as controls. Samples were fixed in 10% buffered formalin, paraffin embedded, and stained with hematoxylin and eosin for histopathological evaluation. Sections were also evaluated with mouse monoclonal antibodies against LYVE-1 and CD34, and expression of both LYVE-1 and CD34 was demonstrated in HEVs. LYVE-1 expression was also found on the endothelial cells of the lymphatic sinus and in reticular cells in the lymph nodes.

Elevated metallothionein (MT) expression in invasive ductal breast cancers predicts tamoxifen resistance.

Elevated expression of the low molecular weight metallothionein (MT) proteins can be found typically in breast cancer cases with less favourable prognosis. The MT gene has been described to be potentially down-regulated by estrogen receptor alpha. The present study is aimed at examining the predictive value of MT expression for results of tamoxifen treatment in breast cancer in relation to steroid receptor status. Sixty patients with primary invasive ductal breast cancers with post-operative tamoxifen treatment were enrolled in the study. In paraffin sections of the studied tumours immmunohistochemical reactions were performed using antibodies directed against MT, estrogen receptors (ER) and progesterone receptors (PgR). Results of the immunohistochemical reactions and of clinical observations were analysed using multivariate progression analysis based on the Cox proportional hazard model. Elevated MT expression was demonstrated to be typical for cases with documented relapse of the disease (P<0.001) or terminated by death (P=0.03). Decreased ER expression was found to be typical for cases of a higher grade (P=0.02) and cases terminated by death (P=0.006). The multivariate analysis showed that elevated MT expression was characteristic for cases with shorter overall survival time (P=0.04). The data showed that MT carried an independent, and also independent from ER status, unfavourable predictive value as far as results of tamoxifen treatment were concerned.

Prognostic significance of augmented metallothionein (MT) expression correlated with Ki-67 antigen expression in selected soft tissue sarcomas.

In soft tissue sarcomas, the most important prognostic criteria include extent of malignancy (G), size of the tumour and intensity of Ki-67 antigen expression. In recent times expression of metallothionein (MT) in cells of some malignant processes of epithelial origin was found to correlate with intensity of Ki-67 antigen expression and to carry a possible prognostic significance. The present study aimed at a demonstration of prognostic value of MT expression and at comparing it with Ki-67 antigen expression and G grade in selected soft tissue sarcomas. Immunohistochemical studies were performed on paraffin sections in 54 cases of malignant fibrous histiocytoma (MFH), 18 cases of liposarcoma and 20 cases of synovial sarcoma. The extent of MT and Ki-67 antigen expression was evaluated and an attempt was made to correlate the results with each other and with grade of the tumour. Expression of MT was evident both in the cytoplasm and in cell nuclei of all studied sarcomas. The most pronounced MT expression was noted in MFH-type tumours. The extent of Ki-67 antigen expression was similar in MFH and liposarcoma and was the lowest in synovial sarcoma. In MFH, liposarcoma and synovial sarcoma a pronounced positive correlation was documented between expression of MT and Ki-67 antigen (r=0.85; p<0.001; r=0.93, p<0.0001; r=0.79, p<0.0001). In all types of the tumours a positive relation was detected between MT expression, expression of Ki-67 and G grade of malignancy in the tumour. Moreover, patients with higher MT expression in the studied tumours demonstrated a shorter survival. MT expression in soft tissue tumours of MFH, liposarcoma and synovial sarcoma type strongly correlated with intensity of proliferation (Ki-67) and G grade and could be useful in defining the extent of malignancy and in prognostic appraisal in the tumours.

Intensity of apoptosis as related to the expression of metallothionein (MT), caspase-3 (cas-3) and Ki-67 antigen and the survival time of patients with primary colorectal adenocarcinomas.

The present study aimed at analysing the intensity of apoptosis, as related to the expression of pro- and anti-apoptotic cellular markers (cas-3, MT, Ki-67 antigen), and at evaluating their expression in relation to the survival time of patients with colorectal adenocarcinoma. Material for the studies was obtained from 40 patients with primary colorectal adenocarcinomas (G2, T3N0M0), treated at the Lower Silesia Centre of Oncology in Wroclaw. Tumour samples were fixed in 4% buffered formalin and embedded in paraffin blocks. In obtained paraffin sections, TUNEL reaction (detection of apoptosis) and immunocytochemical reactions were performed (detection of cas-3, MT and Ki-67 antigen expression). The results disclosed a weak correlation between the intensity of apoptosis and the expression of MT, cas-3 and Ki-67 antigen (r = 0.18; r = 0.33; r = 0.15, respectively) in cells of colorectal adenocarcinomas. The survival time of the patients was shorter, when the apoptosis and expression of Ki-67 antigen were highly pronounced. The time periods of the patients' survival showed no correlation with the expression of cas-3 or MT. The obtained results point to the key role of apoptosis and proliferation processes in the clinical course of colorectal adenocarcinomas.

Role of metallothionein expression in non-small cell lung carcinomas.

Metallothionein (MT) is a low molecular weight protein, which participates in differentiation and proliferation of normal and tumour cells. In some malignant tumours (mammary, renal, ovarian cancers), its increased expression is thought to represent an unfavourable prognostic factor. Non-small-cellular lung cancers (mainly squamocellular cancer and adenocarcinoma) are characterised by ill-defined prognosis, which poses problems in the selection of effective post-surgical therapy. The present study aimed at demonstration of the prognostic significance of MT expression in cells of non-small cell lung cancers, attempting to correlate the intensity of MT expression with G grade and with the intensity of proliferation-associated antigen, Ki-67 expression. The studies were performed on archival paraffin blocks with samples of 25 cases of non-small cell lung cancers (5 squamous cell cancers, 20 adenocarcinomas). In paraffin sections of the studied tumours, immunocytochemical reactions were performed, using mouse monoclonal anti-MT and anti-Ki-67 antibodies. The expressions of MT and Ki-67 were demonstrated in all the studied tumours. An analysis of correlation between the expression of MT, Ki-67 antigen and G grade demonstrated a strong positive relation between the latter two parameters (r=0.70; p<0.05). Less pronounced positive correlations were disclosed between MT expression and G grade (r=0.44; p<0.05) and between MT expression and the expression of Ki-67 antigen (r=0.41; p<0.05). In addition, in 15 cases of examined tumours, survival analysis was performed, which disclosed a shorter survival in patients with high MT expression. The obtained results confirmed the relationship between MT expression and Ki-67 antigen expression, indicating an involvement of the proteins in processes of tumour cell proliferation. In turn, the shorter survival of patients with high expression of MT pointed to prognostic significance of the protein in non-small cell lung cancers.

Prognostic significance of metallothionein expression in correlation with Ki-67 expression in adenocarcinomas of large intestine.

The study aimed at determining levels of metallothionein (MT) and Ki-67 antigen expression in adenocarcinomas of large intestine and examining relation of the expression levels with various clinical and pathological variables. The studies were performed on 81 cases of large intestine adenocarcinoma. Using immunocytochemistry, expressions of MT (positive reaction in 73 cases) and of Ki-67 (positive reaction in 79 cases) antigen were examined and the obtained results were compared with, i.a., grade (G) of the tumour and depth to which intestinal wall was infiltrated by individual tumours. Patient survival analysis was also performed, as correlated to expression levels of the two antigens. The obtained results permitted to disclose that the lower was grade of histological differentiation (G2, G3), the more pronounced was expression of MT and Ki-67. Also, the deeper was neoplastic infiltration of intestinal wall, the more pronounced was MT and Ki-67 expression. Despite the relatively strong correlation between MT expression and Ki-67 expression (r=0.536; p<0.05), only Ki-67 antigen expression in large intestine adenocarcinomas was inversely correlated to survival of the patients. Ki-67 proved to be a better prognostic marker, as compared to MT, in large intestine adenocarcinomas.

Correlation of histopathological and biochemical appraisal of anthracyclin-induced myocardium damage.

Treatment of various tumor types with anthracycl in cytostatic drugs (DNR--daunorubicin and DOX--doxorubicin) is limited by their high cardiotoxicity. This study was aimed at examining effectiveness of histopathological appraisal of myocardial cell injury induced by the cytostatic drugs and evaluated according to Billingham and by MTS methods as compared to results of parallel biochemical assessment of lipid peroxidation indices (MDA- malonyldialdehyde, 4-HDA-4-hydroxyalkenes). The experiments were performed on rats intoxicated with DNR or DOX in an acute manner (1 x 10 mg/kg body weight, i.v.) or a subchronic manner (3 x 3 mg/kg body weight. i.v.). Significant positive correlations were demonstrated between results of histological and biochemical appraisal in rats intoxicated in the acute manner with DNR (r=(0.91), intoxicated in the subchronic manner with DNR (r=0.90) and in rats intoxicated in the acute or subchronic manner with DOX (r=0.91, r=0.77, respectively). The obtained results have confirmed the free radical mechanism of cardiomyocyte injury induced by anthracyclines and the applied techniques of evaluating the destruction may be used independently of each other.

Immunocytochemical evaluation of metallothionein (MT) expression in myoepithelial cells of ductal mammary carcinoma and its relation to survival time: analysis of 7-year course of the disease.

Myoepithelial cells participate in the development of mammary glands and have been suggested to play a role in the biology of mammary cancer. Recent studies demonstrated in the cells expression and overexpression of metallothionein (MT): a low molecular weight, cystein-rich protein which participates, i.a., in the multidrug resistance to cvtostatic drugs. The present study was aimed at examining the relation between results of immunocytochemical (ICC) detection of MT expression in myoepithelial cells, present in structures of a ductal mammary carcinoma, and survival of the patients. In sections originating from 43 patients with ductal mammary carcinoma ICC reactions were performed to detect MT and to confirm the presence of myoepithelial cells (using antibodies to smooth muscle actin). Survival of the patients was also determined in the course of 7-year observations. Statistical analysis using the Coxe's model did not detect relations between MT expression intensity, in the myoepithelium on one hand and patient survival on the other (chi2=0.003 p=0.96).