Matrix metalloproteinases in human melanoma cell lines and xenografts: increased expression of activated matrix metalloproteinase-2 (MMP-2) correlates with melanoma progression

Matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) are involved in tumour progression and metastasis. In this study, we investigated the in vitro and in vivo expression patterns of MMP-1, MMP-2, MMP-3, MMP-9, TIMP-1 and TIMP-2 mRNA and protein in a previously described human melanoma xenograft model. This model consists of eight human melanoma cell lines with different metastatic behaviour after subcutaneous (s.c.) injection into nude mice. MMP-1 mRNA was detectable in all cell lines by reverse transcription polymerase chain reaction (RT-PCR), but the expression was too low to be detected by Northern blot analysis. No MMP-1 protein could be found using Western blotting. MMP-2 mRNA and protein were present in all cell lines, with the highest expression of both latent and active MMP-2 in the highest metastatic cell lines MV3 and BLM. MMP-3 mRNA was expressed in MV3 and BLM, and in the non-metastatic cell line 530, whereas MMP-3 protein was detectable only in MV3 and BLM. None of the melanoma cell lines expressed MMP-9. TIMP-1 and TIMP-2 mRNA and protein, finally, were present in all cell lines. A correlation between TIMP expression level and metastatic capacity of cell lines, however, was lacking. MMP and TIMP mRNA and protein expression levels were also studied in s.c. xenograft lesions derived from a selection of these cell lines. RT-PCR analysis revealed that MMP-1 mRNA was present in MV3 and BLM xenografts, and to a lesser extent in 530. Positive staining for MMP-1 protein was found in xenograft lesions derived from both low and high metastatic cell lines, indicating an in vivo up-regulation of MMP-1. MMP-2 mRNA was detectable only in xenografts derived from the highly metastatic cell lines 1F6m, MV3 and BLM. In agreement with the in vitro results, the highest levels of both latent and activated MMP-2 protein were observed in MV3 and BLM xenografts. With the exception of MMP-9 mRNA expression in 530 xenografts, MMP-3, MMP-9, and TIMP-1 mRNA and protein were not detectable in any xenograft, indicating a down-regulated expression of MMP-3 and TIMP-1 in vivo. TIMP-2 mRNA and protein were present in all xenografts; interestingly, the strongest immunoreactivity of tumour cells was found at the border of necrotic areas. Our study demonstrates that of all tested components of the matrix metalloproteinase system, only expression of activated MMP-2 correlates with increased malignancy in our melanoma xenograft model, corroborating an important role of MMP-2 in human melanoma invasion and metastasis. © 1999 Cancer Research Campaig

Understanding psychiatric institutionalization: a conceptual review

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited

CD13/Aminopeptidase N overexpression by basic fibroblast growth factor mediates enhanced invasiveness of 1F6 human melanoma cells

CD13/Aminopeptidase N (CD13) is known to play an important role in tumour cell invasion. We examined whether basic fibroblast growth factor (bFGF) is involved in the regulation of CD13 expression in human melanoma cells. 1F6 human melanoma cells were stably transfected with constructs encoding either the 18 kDa (18kD) or all (ALL) bFGF isoform proteins. We observed highly increased CD13 mRNA and protein expression in the 1F6 clones regardless of the overexpression of either the 18kD or all isoform proteins. Neutral aminopeptidase activity was increased five-fold and could be inhibited by bestatin and the CD13-neutralising antibody WM15. The enhanced invasion through Matrigel, but not migration in a wound assay, was efficiently abrogated by both bestatin and WM15. Upregulation of CD13 expression was the result of increased epithelial and myeloid promoter activity up to 4.5-fold in 1F6-18kD and 1F6-ALL clones. Interestingly, in a panel of human melanoma cell lines, a significant correlation (r2=0.883, P<0.05) between bFGF and CD13 mRNA and protein expression was detected. High bFGF and CD13 expression were clearly related with an aggressive phenotype. Taken together, our data indicate that high bFGF expression upregulates CD13 expression in human melanoma cells by activating both the myeloid and the epithelial CD13 promoter. In addition, we show that high bFGF and CD13 expression results in enhanced invasive capacity and metastatic behaviour of human melanoma cells

Reproducibility of detection of tyrosinase and MART-1 transcripts in the peripheral blood of melanoma patients: a quality control study using real-time quantitative RT-PCR

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Induction of reactive oxygen intermediates in human monocytes by tumour cells and their role in spontaneous monocyte cytotoxicity

The present study examined the ability of human monocytes to produce reactive oxygen intermediates after a contact with tumour cells. Monocytes generated oxygen radicals, as measured by luminol-enhanced chemiluminescence and superoxide anion production, after stimulation with the tumour, but not with untransformed, cells. The use of specific oxygen radical scavengers and inhibitors, superoxide dismutase, catalase, dimethyl sulphoxide and deferoxamine as well as the myeloperoxidase inhibitor 4-aminobenzoic acid hydrazide, indicated that chemiluminescence was dependent on the production of superoxide anion and hydroxyl radical and the presence of myeloperoxidase. The tumour cell-induced chemiluminescent response of monocytes showed different kinetics from that seen after activation of monocytes with phorbol ester. These results indicate that human monocytes can be directly stimulated by tumour cells for reactive oxygen intermediate production. Spontaneous monocyte-mediated cytotoxicity towards cancer cells was inhibited by superoxide dismutase, catalase, deferoxamine and hydrazide, implicating the role of superoxide anion, hydrogen peroxide, hydroxyl radical and hypohalite. We wish to suggest that so-called ‘spontaneous’ tumoricidal capacity of freshly isolated human monocytes may in fact be an inducible event associated with generation of reactive oxygen intermediates and perhaps other toxic mediators, resulting from a contact of monocytes with tumour cells. © 1999 Cancer Research Campaig

Mutations in APC, CTNNB1 and K-ras genes and expression of hMLH1 in sporadic colorectal carcinomas from the Netherlands Cohort Study

BACKGROUND: The early to intermediate stages of the majority of colorectal tumours are thought to be driven by aberrations in the Wnt (APC, CTNNB1) and Ras (K-ras) pathways. A smaller proportion of cancers shows mismatch repair deficiency. The aim of this study was to analyse the co-occurrence of these genetic alterations in relation to tumour and patient characteristics. METHODS: In a group of 656 unselected sporadic colorectal cancer patients, aberrations in the APC, K-ras, CTNNB1 genes, and expression of hMLH1 were investigated. Additionally, tumours were divided in groups based on molecular features and compared with respect to patient's age at diagnosis, sex, family history of colorectal cancer, tumour sub-localisation, Dukes' stage and differentiation. RESULTS: Mutations at the phosphorylation sites (codons 31, 33, 37, and 45) in the CTNNB1 gene were observed in tumours from only 5/464 patients. Tumours with truncating APC mutations and activating K-ras mutations in codons 12 and 13 occurred at similar frequencies (37% (245/656) and 36% (235/656), respectively). Seventeen percent of tumours harboured both an APC and a K-ras mutation (109/656). Nine percent of all tumours (58/656) lacked hMLH1 expression. Patients harbouring a tumour with absent hMLH1 expression were older, more often women, more often had proximal colon tumours that showed poorer differentiation when compared to patients harbouring tumours with an APC and/or K-ras mutation. CONCLUSION: CTNNB1 mutations seem to be of minor importance in sporadic colorectal cancer. The main differences in tumour and patient characteristics are found between groups of patients based on mismatch repair deficiency

Identifying Key Factors Associated with Aggression on Acute Inpatient Psychiatric Wards

Aggressive behaviour is a critical issue for modern acute psychiatric services, not just because of the adverse impact it has on patients and staff, but also because it puts a financial strain on service providers. The aim of this study was to assess the relationship of patient violence to other variables: patient characteristics, features of the service and physical environment, patient routines, staff factors, the use of containment methods, and other patient behaviours. A multivariate cross sectional design was utilised. Data were collected for a six month period on 136 acute psychiatric wards in 26 NHS Trusts in England. Multilevel modelling was conducted to ascertain those factors most strongly associated with verbal aggression, aggression toward objects, and physical aggression against others. High levels of aggression were associated with a high proportion of patients formally detained under mental health legislation, high patient turnover, alcohol use by patients, ward doors being locked, and higher staffing numbers (especially qualified nurses). The findings suggest that the imposition of restrictions on patients exacerbates the problem of violence, and that alcohol management strategies may be a productive intervention. Insufficient evidence is available to draw conclusions about the nature of the link between staffing numbers and violence

Duloxetine compared with fluoxetine and venlafaxine: use of meta-regression analysis for indirect comparisons

BACKGROUND: Data comparing duloxetine with existing antidepressant treatments is limited. A comparison of duloxetine with fluoxetine has been performed but no comparison with venlafaxine, the other antidepressant in the same therapeutic class with a significant market share, has been undertaken. In the absence of relevant data to assess the place that duloxetine should occupy in the therapeutic arsenal, indirect comparisons are the most rigorous way to go. We conducted a systematic review of the efficacy of duloxetine, fluoxetine and venlafaxine versus placebo in the treatment of Major Depressive Disorder (MDD), and performed indirect comparisons through meta-regressions. METHODS: The bibliography of the Agency for Health Care Policy and Research and the CENTRAL, Medline, and Embase databases were interrogated using advanced search strategies based on a combination of text and index terms. The search focused on randomized placebo-controlled clinical trials involving adult patients treated for acute phase Major Depressive Disorder. All outcomes were derived to take account for varying placebo responses throughout studies. Primary outcome was treatment efficacy as measured by Hedge's g effect size. Secondary outcomes were response and dropout rates as measured by log odds ratios. Meta-regressions were run to indirectly compare the drugs. Sensitivity analysis, assessing the influence of individual studies over the results, and the influence of patients' characteristics were run. RESULTS: 22 studies involving fluoxetine, 9 involving duloxetine and 8 involving venlafaxine were selected. Using indirect comparison methodology, estimated effect sizes for efficacy compared with duloxetine were 0.11 [-0.14;0.36] for fluoxetine and 0.22 [0.06;0.38] for venlafaxine. Response log odds ratios were -0.21 [-0.44;0.03], 0.70 [0.26;1.14]. Dropout log odds ratios were -0.02 [-0.33;0.29], 0.21 [-0.13;0.55]. Sensitivity analyses showed that results were consistent. CONCLUSION: Fluoxetine was not statistically different in either tolerability or efficacy when compared with duloxetine. Venlafaxine was significantly superior to duloxetine in all analyses except dropout rate. In the absence of relevant data from head-to-head comparison trials, results suggest that venlafaxine is superior compared with duloxetine and that duloxetine does not differentiate from fluoxetine