Leiomyosarcoma of the vulva.

Malignant tumors of the vulva soft tissue are uncommon. About 1-3% are sarcomas. They can be mistaken as benign lesions, leading to misdiagnosis and mistreatment. A case of a 71-year-old woman with a leiomyosarcoma of the vulva is presented. The surgical excision of the lesion is described and there were no additional malignancies or lesions found. There was no need for adjuvant therapy.

Peritoneal enterobiasis causing endometriosis-like symptoms.

PURPOSE: Enterobiasis is the most common parasitic disease of the temperate zones and infects the human intestinal tract. In rare cases extraintestinal infections with Enterobius vermicularis may occur and can affect the female genital tract and peritoneal cavity. In most cases the infection is asymptomatic, but there are also cases described in which peritoneal enterobiasis can cause abdominal pain. METHODS: A case report and review of the pertinent literature. RESULTS: A 32-year-old patient was admitted with cyclical lower abdominal pain. With suspected endometriosis a diagnostic autofluorescence laparoscopy (DAFE) was performed. At surgery extensive peritoneal deposits were seen. Macroscopically these deposits were not typical for endometriosis. The histological examination showed granuloma caused by E. vermicularis eggs. The patient was treated with mebendazole. After completion of treatment the patient was asymptomatic. At the second-look laparoscopy no more peritoneal changes were detected. CONCLUSION: Enterobius vermicularis may cause symptoms similar to endometriosis. In cases with reasonable suspicion it should therefore be considered in the differential diagnosis.

Regression of human metastatic renal cell carcinoma after vaccination with tumor cell-dendritic cell hybrids.

Reports of spontaneous regressions of metastases and the demonstration of tumor-reactive cytotoxic T lymphocytes indicate the importance of the host's immune system in controlling the devastating course of metastatic renal cell carcinoma. Recent research indicates that immunization with hybrids of tumor and antigen presenting cells results in protective immunity and rejection of established tumors in various rodent models. Here, we present a hybrid cell vaccination study of 17 patients. Using electrofusion techniques, we generated hybrids of autologous tumor and allogeneic dendritic cells that presented antigens expressed by the tumor in concert with the co-stimulating capabilities of dendritic cells. After vaccination, and with a mean follow-up time of 13 months, four patients completely rejected all metastatic tumor lesions, one presented a 'mixed response', and two had a tumor mass reduction of greater 50%. We also demonstrate induction of HLA-A2-restricted cytotoxic T cells reactive with the Muc1 tumor-associated antigen and recruitment of CD8+ lymphocytes into tumor challenge sites. Our data indicate that hybrid cell vaccination is a safe and effective therapy for renal cell carcinoma and may provide a broadly applicable strategy for other malignancies with unknown antigens.

Betulinic acid induces apoptosis and inhibits hedgehog signalling in rhabdomyosarcoma.

BACKGROUND: Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma in childhood with the ability to resist apoptosis by the activation of survival promoting and anti-apoptotic proteins. METHODS: Efficacy of the apoptosis-inducing agent betulinic acid (BA) was determined in RMS cell cultures and in vivo by measuring cell viability, survival, apoptosis, hedgehog signalling activity, and neovascularisation. RESULTS: Betulinic acid had a strong cytotoxic effect on RMS cells in a dose-dependent manner. The BA treatment caused a massive induction of apoptosis mediated by the intrinsic mitochondrial pathway, which could be inhibited by the broad-range caspase inhibitor zVAD.fmk. Exposure of hedgehog-activated RMS-13 cells to BA resulted in a strong decrease in GLI1, GLI2, PTCH1, and IGF2 expression as well as hedgehog-responsive luciferase activity. Intraperitoneal injection of 20 mg BA per kg per day significantly retarded growth of RMS-13 xenografts in association with markedly higher counts of apoptotic cells and down-regulation of GLI1 expression compared with control tumours, while leaving microvascular density, cell proliferation, and myogenic differentiation unaffected. CONCLUSION: Our data show that induction of apoptosis and inhibition of hedgehog signalling are important features of the anti-tumourigenic effect of BA in RMS and advices this compound for the use in a multimodal therapy of this highly aggressive paediatric tumour.

Protein profiling of non-malignant and malignant ascites by SELDI-TOF MS: proof of principle.

Ascites is a common clinical symptom in liver cirrhosis, inflammatory disorders of the abdomen and a major late manifestation of metastatic malignancies. Standard cytopathological techniques and immunocytochemistry have specificities and sensitivities of approximately 95 and 60%, respectively for the presence of tumor cells. Development of faster and more accurate screening methods would be of great clinical utility. In this work we examined differential analysis of the unbound proteins in the supernatant of ascites fluid by Protein-Chip SELDI mass spectrometry. There were 21 tumor cell-positive and 34 tumor cell-negative samples. We used principal component analysis coupled with linear regression applied to the mass spectra of the samples to distinguish between the sample groups. Two sample sets for statistical analysis were created after randomization, a training set with 37 samples and a validation set with 18 samples resulting in a specificity of 93% and a sensitivity of 83% on the training set. The validation set yielded a specificity and sensitivity of 75%. This study suggests that SELDI-TOF mass spectrometry appears to have great potential as a surrogate diagnostic tool to evaluate effusion specimens.

[Determination of renal carcinoma progression in small animals by means of flat-panel volumetric computer tomography]. / Die Erfassung der Progression des Nierenzellkarzinoms in Kleintieren mittels Flachdetektor-Volumen-CT.

PURPOSE: We investigated the feasibility of using flat panel volumetric computer tomography (fpVCT) for the detection of orthotopically implanted renal carcinomas in nude mice. MATERIALS AND METHODS: One million renal cell carcinoma cells [A-498 line (Braunschweig, Germany), in 0.2 ml phosphate-buffered solution (PBS), pH 7.4] were injected into the left kidney of each of the eight nude mice. Each mouse was imaged twice (12 and 16 weeks after implantation) with fpVCT (GE prototype with circular gantry with two 1024 x 1024, 200 microm pixel size, aSi/CsI flat panel detector) after injection of 200 microl contrast medium to check for tumour spread. After 16 weeks the mice were killed and dissected, and the imaging findings in liver, kidneys and lung were compared with the macroscopic findings. RESULTS: No local evidence of tumour or of metastatic spread was seen on fpVCT after 12 weeks in any of the mice. After 16 weeks fpVCT revealed tumour growth in 6 of the 16 kidneys. Two mice had each developed a multifocal renal cell carcinoma and one mouse, a bilateral renal tumour manifestation. In one mouse liver metastases were seen. The fpVCT findings correlated well with the observations recorded in the pathological examination. CONCLUSION: fpVCT is an innovative and noninvasive imaging procedure that can be used for longitudinal investigation of tumour progression following orthotopic implantation of renal cell carcinoma to small animals. The use of a system of this kind will make a decisive contribution to reducing the number of animals used in experimental test projects.

Heterogeneity of angiogenesis and blood vessel maturation in human tumors: implications for antiangiogenic tumor therapies.

Microvessel density (MVD) counting techniques have been widely used to assess the vasculature in tumors. MVD counts assess the presence of blood vessels but do not give an indication of the degree of angiogenesis and the functional status of the tumor neovasculature. To analyze angiogenesis and the functional status of the tumor vascular bed, we have quantitated endothelial cell proliferation and the recruitment of pericytes in human tumors [glioblastomas (n = 30), renal cell carcinomas (n = 22), colon carcinomas (n = 18), mammary carcinomas (n = 24), lung carcinomas (n = 15), and prostate carcinomas (n = 19)]. These findings were compared to the physiological angiogenesis in the cyclic bovine ovarian corpus luteum. Tissue sections were examined applying double-labeling immunohistochemical techniques to detect proliferating endothelial cells and to colocalize endothelial cells and pericytes. The following parameters were quantitated: (a) MVD count; (b) proliferating capillary index (PCI); (c) proliferating tumor versus endothelial cell index; and (d) microvessel pericyte coverage index (MPI). Based on endothelial cell proliferation, angiogenesis was found to be present in all tumors with characteristic and significant differences between the tumor types (glioblastomas, PCI = 9.6 +/- 6.1%; renal cell carcinomas, PCI = 9.4 +/- 5.2%; colon carcinomas, PCI = 7.8 +/- 5.2%; mammary carcinomas, PCI = 5.0 +/- 4.8%; lung carcinomas, PCI = 2.6 +/- 2.5%; prostate carcinomas, PCI = 2.0 +/- 1.4%). There was a considerable degree of heterogeneity in the intensity of angiogenesis within each tumor group, as indicated by large standard deviations. Even in the most angiogenic tumors, angiogenesis was found to be 4 to 20 times less intense as compared with the physiological angiogenesis in the growing ovarian corpus rubrum (PCI = 40.6 +/- 6.2%). Varying degrees of pericyte recruitment to the tumor microvasculature were determined in the different tumor types (glioblastomas, MPI = 12.7 +/- 7.9%; renal cell carcinomas, MPI = 17.9 +/- 7.8%; colon carcinomas, MPI = 65.4 +/- 10.5%; mammary carcinomas, MPI = 67.3 +/- 14.2%; lung carcinomas, MPI = 40.8 +/- 14.5%; prostate carcinomas, MPI = 29.6 +/- 9.5%). The data demonstrate distinct quantitative variations in the intensity of angiogenesis in malignant human tumors. Furthermore, the varying degrees of pericyte recruitment indicate differences in the functional status of the tumor vasculature in different tumors that may reflect varying degrees of maturation of the tumor vascular bed.

Continuous recruitment, co-expression of tumour necrosis factor-alpha and matrix metalloproteinases, and apoptosis of macrophages in gout tophi.

Gout tophi are characterised by foreign body granulomas consisting of mono- and multinucleated macrophages surrounding deposits of monosodium urate microcrystals. After primary formation, granulomas grow associated with degradation of the extracellular matrix. Based on this background, we have sought (1) to investigate whether during granuloma's growth new macrophages are recruited into the tophi, (2) to find in situ evidence for macrophages' active role in matrix degradation and (3) to examine whether shrunk cells seen within gout tophi are apoptotic. Immunohistochemistry showed that perivascular localised mononuclear cells are CD68+, S100A8+, S100A9+, 25F9-, representing freshly migrated monocytes/macrophages. In contrast, almost all CD68+ mono- and multinucleated cells arranged within granulomas were S100A8-, S100A9-, 25F9+, representing mature (non-migrating) macrophages. Serial sections revealed that macrophages co-express tumour necrosis factor (TNF)-alpha and matrix metalloproteinases (MMPs) 2 and 9. In situ end-labelling of fragmented DNA demonstrated that CD68+ macrophages undergo apoptosis within gout tophi. Our data show that macrophages are continuously recruited into the gout tophi. These macrophages co-produce the proinflammatory cytokine TNF-alpha and two TNF-alpha inducible lytic enzymes, MMP-2 and MMP-9, suggesting that TNF-alpha may induce MMP production followed by matrix degradation within foreign body granulomas. In parallel, macrophages undergo apoptosis, a phenomenon that may restrict the destructive potential of inflammatory macrophages.

Vascular endothelial growth factor expression, angiogenesis, and necrosis in renal cell carcinomas.

Rapidly growing tumors often develop necrosis. In the present study the expression of vascular endothelial growth factor (VEGF) was investigated and compared to microvessel density and necrosis of renal cell carcinomas. In the tumor-host interface the microvessel density was significantly increased compared to central tumor areas. Tumor necrosis was associated with a decrease of microvessel density and an increase of the VEGF protein expression within the perinecrotic rim. VEGF protein was focally upregulated in vital tumor tissue. An increase of the apoptotic rate of endothelia and vital tumor tissue in tumors with necrosis could not be detected. VEGF(121,165) mRNA was decreased in proliferatively active carcinomas compared to less proliferative tumors. Multicellular renal cell cancer spheroids as a model of chronic hypoxia developed central apoptosis but no necrosis. VEGF was upregulated in the spheroid. Tumor microvessels expressed matrix metalloproteinase -2 and -9 and an incomplete pericyte covering in comparison to tumor-free tissue indicating immature active angiogenesis. We conclude that highly proliferative renal cell carcinomas outgrow their vascular supply and develop chronic hypoxia inducing a decrease of proliferation and an increase of VEGF expression. However, chronic hypoxia does not cause significant necrosis or apoptosis. Tumor necrosis is more likely induced by acute hypoxia due to immature microvessels. Furthermore, VEGF expression associated with concomitant tumor necrosis may help identify renal cell carcinomas susceptible to antiangiogenic therapy.

Polyploidization and losses of chromosomes 1, 2, 6, 10, 13, and 17 in three cases of chromophobe renal cell carcinomas.

Clonal chromosome aberrations identified after short-term culture are presented for three cases of chromophobe renal cell carcinomas (RCC). All tumors revealed abnormal karyotypes with a varying proportion of polyploid tumor cells. Common numerical abnormalities were combined losses of chromosomes 1, 2, 6, 10, 13, and 17. Clonal karyotypic evolution was demonstrated in one case in which several related clones could be identified. An additional balanced translocation t(3;14)(p24;q22) observed in this case proved to be of constitutional nature by cytogenetic analysis of normal kidney cells and peripheral blood lymphocytes. These cytogenetic findings provide further evidence that chromophobe renal cell carcinomas are characterized by a highly specific combination of chromosomal losses most commonly including chromosomes 1, 2, 6, 10, 13, and 17.

Quantification by competitive quantitative RT-PCR of VEGF121 and VEGF165 in renal cell carcinoma.

The field of angiogenesis and its mediators in tumour tissue is gaining more and more interest because of their therapeutic implications to arrest progressive growth and metastasis. We examined the expression status of the proangiogenic vascular endothelial growth factor (VEGF) in tumour tissue and adjacent tumour-free tissue from renal cell carcinoma (RCC) as well as in cell cultures deriving from tumour tissue. Quantification of both secreted isoforms ot VEGF by competitive RT-PCR revealed a marked increase of VEGF message in tumours and especially in cell cultures from RCC. We found a significant correlation of VEGF expression and microvessel density which was investigated immunohistochemically. As further compared to other urological neoplasms we investigated for VEGF mediated vascularization, RCC is the most promising candidate for anti-angiogenic therapies.

Expression of IFNgamma, coexpression of TNFalpha and matrix metalloproteinases and apoptosis of T lymphocytes and macrophages in granuloma annulare.

Granuloma annulare, a prototype noninfectious granulomatous dermatitis, is morphologically characterized by a necrobiotic core surrounded by a cellular infiltrate. Because of many morphological similarities to tuberculosis, granuloma annulare has been suggested to represent a delayed-type hypersensitivity (Th1) reaction in the course of which inflammatory cells elicit matrix degradation. In the present study we (1) investigated the expression of interferon-gamma as the most important Th1-associated cytokine, (2) sought in situ evidence for the coexpression of the proinflammatory cytokine tumor necrosis factor-alpha and cytokine-regulated matrix metalloproteinases 2 (gelatinase A) and 9 (gelatinase B), and (3) sought to determine whether shrunken cells seen within necrobiotic areas of granuloma annulare are apoptotic cells. In situ hybridization combined with immunofluorescence showed that large numbers of infiltrating CD3+ lymphocytes express interferon-gamma. Application of catalyzed signal amplification in immunodetection revealed that the vast majority of CD3+ lymphocytes and CD68+ macrophages contained tumor necrosis factor-alpha. Immunohistochemistry demonstrated that macrophages producing tumor necrosis factor-alpha coexpress matrix metalloproteinases 2 and 9. In situ end-labeling combined with immunofluorescence detected few apoptotic T cells in perivascular regions and numerous apoptotic macrophages within necrobiotic areas. These results suggest that in granuloma annulare interferon-gamma+ Th-1 lymphocytes may cause a delayed-type hypersensitivity reaction whereby macrophages are differentiated to aggressive effector cells expressing tumor necrosis factor-alpha and matrix metalloproteinases. In parallel, activation-induced apoptosis in lymphocytes and macrophages may serve to restrict the destructive potential of the inflammatory cells.

The value of anti-calretinin antibody in the differential diagnosis of normal and reactive mesothelia versus metastatic tumors in effusion cytology.

In effusion cytology the distinction of reactive mesothelia from metastatic carcinoma cells may be a diagnostic challenge. Immunocytochemistry using antibodies suitable to detect epithelial cells must be considered carefully due to limited sensitivity and specificity of these antibodies. Efficient results in histological differential diagnosis of malignant mesothelioma versus lung-adenocarcinoma applying a novel antiserum against the calcium binding protein calretinin inspirated us to investigate the value of anti-calretinin antibody in effusion cytology combined with an epithelial marker. Cytoslides prepared by cytocentrifugation from 42 malignant and 65 reactive effusion specimens were immunostained using antibodies against calretinin and the epithelial marker Ber-EP4. Positive immunoreaction for calretinin in normal and reactive mesothelial cells was noted in 93% of the cases, whereas immunoreaction for calretinin was completely negative in the metastatic cells in 95% of the malignant effusions. Metastatic carcinoma cells were detected with anti-Ber-EP4 in 83% of malignant effusions. Non-specific positive reactions for Ber-EP4 in single mesothelial cells were observed in 16% of all cases and, moreover, frequently with macrophages or neutrophilic granulocytes. Our results demonstrate high sensitivity and specificity of anti-calretinin antibody for mesothelial cells in effusion specimens. They support its application to improve the diagnostic reliability of epithelial markers, especially because anti-calretinin antibody could be helpful in the assessment of false positive and false negative reactions of epithelial markers.

[Influence of DNA ploidy and grading on survival in primary metastatic prostate carcinoma]. / Einfluss von DNA-Ploidie und Grading auf die Uberlebenszeit beim primär metastasierten Prostatakarzinom.

An aneuploid pattern of prostatic cancer defined by flow cytometry was shown to be of value in predicting progression rates and patient survival times. We evaluated the long-term value of DNA analysis in prostatic cancer in 61 patients with advanced disease. We performed flow cytometry on 61 fresh prostate specimens obtained-from a transrectal needle biopsy or a transurethral resection. All patients received antihormonal therapy. Time until death was evaluated in all patients. Of the DNA histograms analyzed, 37% showed a diploid pattern, 63% an non-diploid pattern, and 37% a tetraploid or hypertetraploid pattern. Kaplan-Meier plots were generated for analysis of the probability of survival. Mean survival time was 44 months for patients with diploid (range 1-126 months) and 40 months for patients with non-diploid tumors (range 1-96 months). This difference is not statistically significant. However, the combination of non-diploid and low-differentiated (G3) tumors reduced survival time significantly (mean 20 months, range 1-62 months). There was no patient with combination of a diploid and highly differentiated tumor.

[Treosulfan displays cytotoxic effect on spheroids of primary cell cultures of renal cell carcinoma independent of p-glycoprotein expression]. / Treosulfan zeigt zytoxische Wirkung an Sphäroiden von primären Zellkulturen des Nierenzellkarzinoms unabhängig von der P-Glykoproteinexpression.

Therapy of advanced renal cell carcinoma remains difficult. New therapeutic schemes besides cytokine treatment should be evaluated. The following study analyzes the in vitro toxicity of treosulfan on spheroids of 8 primary cultures of renal cell carcinoma cells. these data were compared to the toxicity of vinblastine. All investigations were performed in regard to the P-glycoprotein (Pgp) expression of the cells, which is one of the main causes of multidrug resistance. Four Pgp positive and four Pgp negative spheroids were incubated with the drugs in increasing doses. Toxicity was measured using the MTT toxicity assay as well as trypan blue exclusion. Significantly higher toxicity of treosulfan compared to vinblastine could be demonstrated. In addition, the effects of treosulfan were not related to Pgp expression. These results are encouraging and a phase II study analyzing the efficacy of treosulfan in patients with advanced renal cell carcinoma has been initiated in our institution.

High expression levels of erythropoietin and its receptor are not correlated with shorter survival in human glioblastoma.

Erythropoietin (EPO) is used to treat anemia in neoplastic disease. EPO also exerts neuroprotective effects on neuronal cells, making a prophylactic use against the neurocognitive effects of radiochemotherapy probable. However, EPO/EPO-receptor (EPOR) signalling has been also detected in glioblastoma cells. Data collected in vitro and in vivo show conflicting results on the effect of EPO on malignant gliomas. The association between EPO and EPOR expression and the prognosis of human glioblastomas was analyzed. Probes of human glioblastomas with complete documentation of clinical course and treatment were assessed by immunohistochemistry for the expression of EPO and EPOR (n=80). Using univariate and multivariate survival analysis, the association with age, gender, radiation, chemotherapy and extent of resection was determined. High levels of EPOR were correlated with a median survival advantage of 7 months (p<0.01). By univariate, but not multivariate, analysis, high levels of EPO and EPOR were associated with a significant prolongation of 7 months median survival when compared to low levels of both molecules. In patients treated with radiochemotherapy adjuvant to surgery, the median survival was 6.5 months longer in patients with high levels of EPOR (p<0.04). According to previous studies, longer patient survival is associated with EPOR expression. Therefore, EPO appears to be safe for the treatment of anemia in glioblastoma patients. However, a prophylactic use, i.e., for neuroprotection, is not recommended in light of the functional studies described in the literature.

Potential role of fractalkine receptor expression in human renal fibrogenesis.

The inhibition of several chemokine/chemokine receptors has been shown to reduce progressive renal interstitial fibrosis. In this study, we examined the expression of the CX(3)C receptor in human renal biopsies with interstitial fibrosis and from normal kidneys by real-time polymerase chain reaction (PCR) and immunohistochemistry. The CX(3)C receptor was not only detected in mononuclear, tubular epithelial, and dendritic cells but also in alpha-smooth muscle actin and vimentin-positive interstitial myofibroblasts in fibrotic kidneys. Real-time PCR indicated a significant upregulation of CX(3)C receptor mRNA in fibrotic kidneys compared with non-fibrotic nephropathies or donor biopsies. In renal fibroblasts in vitro, hydrogen peroxide increased the expression of the CX(3)C receptor, an increase that was inhibited by N-acetylcysteine and catalase. However, neither proinflammatory nor profibrotic cytokines resulted in this upregulation. Stimulation of fibroblasts by CX(3)C ligand led to a significant enhancement of migration, which was abrogated by pre-incubation with a blocking anti-CX(3)C receptor antibody. Our studies indicate that renal fibrosis is associated with the expression of CX(3)C receptors on human renal fibroblasts. The expression is induced by reactive oxygen species suggesting a role of oxidative stress.

Expression of gastrin releasing Peptide receptor in renal cell carcinomas: a potential function for the regulation of neoangiogenesis and microvascular perfusion.

PURPOSE: Gastrin releasing peptide (GRP) is a growth factor for renal cell carcinoma (RCC) and it has vasoactive properties. Blockade of GRP receptor inhibits the growth of GRP receptor positive and negative tumors in nude mice, suggesting GRP effects other than those related to tumor epithelium. Therefore, in this study we analyzed the effects of GRP receptor blockade on neoangiogenesis in RCC. MATERIALS AND METHODS: GRP receptor expression was determined in human RCC and corresponding normal tissue by real-time reverse transcriptase-polymerase chain reaction, immunohistochemistry and confocal laser scanning microscopy. Multicellular spheroids of the A498 RCC line were implanted into dorsal skin fold chambers of athymic nude mice. Neoangiogenesis was measured by intravital microscopy after blockade of GRP receptors by the GRP antagonist RC-3095. The influence of GRP on vascular endothelial growth factor secretion in A498 cells was studied in vitro. RESULTS: GRP receptor expression was immunolocalized in tumor cells and microvessels. Implanted tumor cell spheroids and spheroid microvessels of the chamber also expressed GRP receptors. Spheroid neoangiogenesis was significantly inhibited by RC-3095 when given immediately after spheroid implantation. Vascular endothelial growth factor secretion of A498 cells was not affected by GRP. CONCLUSIONS: RCC angiogenesis is sensitive to GRP receptor blockade. Therefore, GRP receptors may not only stimulate tumor cell proliferation, but also affect tumor microcirculation.

Expression of matrix metalloproteinases MMP-2, MMP-9 and their tissue inhibitors TIMP-1, -2, and -3 in benign and malignant tumours of the salivary gland.

AIMS: The balance between matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) is involved in the morphogenesis of the normal salivary gland as well as in mechanisms of tumour invasion and metastasis. Our aim was to analyse protein and mRNA expression of MMPs and TIMPs in normal salivary gland tissue and in various salivary gland neoplasms. METHODS AND RESULTS: Immunohistochemistry for MMP-2, MMP-9, TIMP-1, -2 and -3 was performed in 20 malignant and six benign salivary gland tumours. The immunoscores of MMP-2 were significantly higher in carcinomas compared with adenomas (P = 0.0028). The MMP/TIMP ratio was significantly higher in carcinomas than in adenomas (P = 0.0097). In mucoepidermoid carcinomas MMP-2 expression was preferentially observed in the intermediate cell type. Neoplastic acinar cells of acinic cell carcinoma demonstrated de novo expression of MMP-2, MMP-9, TIMP-1, and TIMP-2. Immunoscores of TIMP-3 were not decreased in malignant tumours compared with adenomas. In accordance with the immunostaining of MMP-2 and -9, gelatinolytic activity could be demonstrated by in-situ zymography. The mRNA expression of MMPs and TIMPs analysed by real-time reverse transcriptase-polymerase chain reaction in three malignant and two benign tumours and their corresponding normal tissue did not generally correlate with their protein expression. CONCLUSIONS: Our findings suggest a disturbed balance between MMP and TIMP in malignant salivary gland tumours. MMP-2 expression in particular seems to be related to the invasive properties and the malignant potential of these tumours.