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Identification of therapeutic targets for treating fibrotic diseases and cancer remains challenging. Our study aimed to investigate the effects of TGF-β1 and TGF-β3 on myofibroblast differentiation and extracellular matrix deposition in different types of fibroblasts, including normal/dermal, cancer-associated, and scar-derived fibroblasts. When comparing the phenotype and signaling pathways activation we observed extreme heterogeneity of studied markers across different fibroblast populations, even within those isolated from the same tissue. Specifically, the presence of myofibroblast and deposition of extracellular matrix were dependent on the origin of the fibroblasts and the type of treatment they received (TGF-β1 vs. TGF-β3). In parallel, we detected activation of canonical signaling (pSMAD2/3) across all studied fibroblasts, albeit to various extents. Treatment with TGF-β1 and TGF-β3 resulted in the activation of canonical and several non-canonical pathways, including AKT, ERK, and ROCK. Among studied cells, cancer-associated fibroblasts displayed the most heterogenic response to TGF-β1/3 treatments. In general, TGF-β1 demonstrated a more potent activation of signaling pathways compared to TGF-β3, whereas TGF-β3 exhibited rather an inhibitory effect in keloid- and hypertrophic scar-derived fibroblasts suggesting its clinical potential for scar treatment. In summary, our study has implications for comprehending the role of TGF-β signaling in fibroblast biology, fibrotic diseases, and cancer. Future research should focus on unraveling the mechanisms beyond differential fibroblast responses to TGF-β isomers considering inherent fibroblast heterogeneity., (© 2023. The Author(s).)

Recent evidence indicates that targeting IL-6 provides broad therapeutic approaches to several diseases. In patients with cancer, autoimmune diseases, severe respiratory infections [e.g. coronavirus disease 2019 (COVID-19)] and wound healing, IL-6 plays a critical role in modulating the systemic and local microenvironment. Elevated serum levels of IL-6 interfere with the systemic immune response and are associated with disease progression and prognosis. As already noted, monoclonal antibodies blocking either IL-6 or binding of IL-6 to receptors have been used/tested successfully in the treatment of rheumatoid arthritis, many cancer types, and COVID-19. Therefore, in the present review, we compare the impact of IL-6 and anti-IL-6 therapy to demonstrate common (pathological) features of the studied diseases such as formation of granulation tissue with the presence of myofibroblasts and deposition of new extracellular matrix. We also discuss abnormal activation of other wound-healing-related pathways that have been implicated in autoimmune disorders, cancer or COVID-19., (© 2022. The Author(s).)

The incidence of cutaneous malignant melanoma is increasing worldwide. While the treatment of initial stages of the disease is simple, the advanced disease frequently remains fatal despite novel therapeutic options . This requires identification of novel therapeutic targets in melanoma. Similarly to other types of tumours, the cancer microenvironment plays a prominent role and determines the biological properties of melanoma. Importantly, melanoma cell-produced exosomes represent an important tool of intercellular communication within this cancer ecosystem. We have focused on potential differences in the activity of exosomes produced by melanoma cells towards melanoma-associated fibroblasts and normal dermal fibroblasts. Cancer-associated fibroblasts were activated by the melanoma cell-produced exosomes significantly more than their normal counterparts, as assessed by increased transcription of genes for inflammation-supporting cytokines and chemokines, namely IL-6 or IL-8. We have observed that the response is dependent on the duration of the stimulus via exosomes and also on the quantity of exosomes. Our study demonstrates that melanoma-produced exosomes significantly stimulate the tumour-promoting proinflammatory activity of cancer-associated fibroblasts. This may represent a potential new target of oncologic therapy ., (© 2021. The Author(s).)

The globally increasing incidence of cancer, including melanoma, requires novel therapeutic strategies. Development of successful novel drugs is based on clear identification of the target mechanisms responsible for the disease progression. The specific cancer microenvironment represents a critically important aspect of cancer biology, which cannot be properly studied in simplistic cell culture conditions. Among other traditional options, the study of melanoma cell growth on the chicken chorioallantoic membrane offers several significant advantages. This model offers increased complexity compared to usual in silico culture models and still remains financially affordable. Using this model, we studied the growth of three established human melanoma cell lines: A2058, BLM, G361. The combination of histology, immunohistochemistry with the application of human-specific antibodies, intravascular injection of contrast material such as filtered Indian ink, Mercox solution and phosphotungstic acid, and X-ray micro-CT and live-cell monitoring was employed. Melanoma cells spread well on the chicken chorioallantoic membrane. However, invasion into the stroma of the chorioallantoic membrane and the limb primordium graft was rare. The melanoma cells also significantly influenced the architecture of the blood vessel network, resulting in the orientation of the vessels to the site of the tumour cell inoculation. The system of melanoma cell culture on the chorioallantoic membrane is suitable for the study of melanoma cell growth, particularly of rearrangement of the host vascular pattern after cancer cell implantation. The system also has promising potential for further development.

Melanoma represents a malignant disease with steadily increasing incidence. UV-irradiation is a recognized key factor in melanoma initiation. Therefore, the efficient prevention of UV tissue damage bears a critical potential for melanoma prevention. In this study, we tested the effect of UV irradiation of normal keratinocytes and their consequent interaction with normal and cancer-associated fibroblasts isolated from melanoma, respectively. Using this model of UV influenced microenvironment, we measured melanoma cell migration in 3-D collagen gels. These interactions were studied using DNA microarray technology, immunofluorescence staining, single cell electrophoresis assay, viability (dead/life) cell detection methods, and migration analysis. We observed that three 10 mJ/cm 2 fractions at equal intervals over 72 h applied on keratinocytes lead to a 50% increase (p < 0.05) in in vitro invasion of melanoma cells. The introduction cancer-associated fibroblasts to such model further significantly stimulated melanoma cells in vitro invasiveness to a higher extent than normal fibroblasts. A panel of candidate gene products responsible for facilitation of melanoma cells invasion was defined with emphasis on IL-6, IL-8, and CXCL-1. In conclusion, this study demonstrates a synergistic effect between cancer microenvironment and UV irradiation in melanoma invasiveness under in vitro condition.

Tumour microenvironment plays a critical role in cell invasion and metastasis. To investigate the role of cancer-associated fibroblasts (CAFs) in melanoma cell invasiveness, we used 3D spheroid invasion assay. The effect of conditioned media from normal fibroblasts and CAFs cultivated alone or co-cultivated with melanoma cells on BLM or A2058 melanoma spheroid invasion was analysed. We found that conditioned media from CAFs and CAFs co-cultured with melanoma cells, especially, promote invasion and migration, without significant effect on melanoma cell proliferation. We further analysed the expression of pro-invasive cytokines IL-8 and IL-6 in media and found that melanoma cells are dominant producers of IL-8 and fibroblasts are dominant producers of IL-6 in 2D monocultures, while co-cultivation of CAFs with melanoma cells induces production/secretion of IL-6 and IL-8 into the media. The analyses of IL-6 levels in 3D cultures and human melanoma samples, however, revealed that at least in some cases IL-6 is also produced directly by melanoma cells. Analysis of the role of IL-6 and IL-8 in CAF-induced melanoma invasion, using neutralising antibodies, revealed that simultaneous blocking of IL-6 and IL-8 is sufficient to fully inhibit CAF-induced human melanoma cell invasiveness. In summary, these experiments indicate the important role of CAFs and IL-8 and IL-6 cytokines in melanoma cell invasiveness.

Epithelial-mesenchymal interaction between stromal fibroblasts and cancer cells influences the functional properties of tumor epithelium, including the tumor progression and spread. We compared fibroblasts prepared from stroma of squamous cell carcinoma and normal dermal fibroblasts concerning their biological activity toward normal keratinocytes assessed by immunocytochemistry and profiling of gene activation for growth factors/cytokines by microarray chip technology. IGF-2 and BMP-4 were determined as candidate factors responsible for tumor-associated fibroblast activity that influences normal epithelia. This effect was confirmed by addition of recombinant IGF-2 and BMP4, respectively, to the culture medium. This hypothesis was also verified by inhibition experiments where blocking antibodies were employed in the medium conditioned by cancer-associated fibroblast. Presence of these growth factors was also detected in tumor samples.