The curcumin analog DM-1 induces apoptotic cell death in melanoma

The main difficulty in the successful treatment of metastatic melanoma is that this type of cancer is known to be resistant to chemotherapy. Chemotherapy remains the treatment of choice, and dacarbazine (DTIC) is the best standard treatment. The DM-1 compound is a curcumin analog that possesses several curcumin characteristics, such as antiproliferative, antitumor, and antimetastatic properties. The objective of this study was to evaluate the signaling pathways involved in melanoma cell death after treatment with DM-1 compared to the standard agent for melanoma treatment, DTIC. Cell death was evaluated by flow cytometry for annexin V and iodide propide, cleaved caspase 8, and TNF-R1 expression. Hoechst 33342 staining was evaluated by fluorescent microscopy; lipid peroxidation and cell viability (MTT) were evaluated by colorimetric assays. The antiproliferative effects of the drugs were evaluated by flow cytometry for cyclin D1 and Ki67 expression. Mice bearing B16F10 melanoma were treated with DTIC, DM-1, or both therapies. DM-1 induced significant apoptosis as indicated by the presence of cleaved caspase 8 and an increase in TNF-R1 expression in melanoma cells. Furthermore, DM-1 had antiproliferative effects in this the same cell line. DTIC caused cell death primarily by necrosis, and a smaller melanoma cell population underwent apoptosis. DTIC induced oxidative stress and several physiological changes in normal melanocytes, whereas DM-1 did not significantly affect the normal cells. DM-1 antitumor therapy in vivo showed tumor burden decrease with DM-1 monotherapy or in combination with DTIC, besides survival rate increase. Altogether, these data confirm DM-1 as a chemotherapeutic agent with effective tumor control properties and a lower incidence of side effects in normal cells compared to DTIC

Basic Red 51, a permitted semi-permanent hair dye, is cytotoxic to human skin cells: Studies in monolayer and 3D skin model using human keratinocytes (HaCaT)

The use of hair dyes is closely associated with the increase of cancer, inflammation and other skin disorders. The recognition that human skin is not an impermeable barrier indicates that there is the possibility of human systemic exposure. The potential carcinogenicity of hair dye ingredients has attracted the attention of toxicologists for many decades, mainly due to the fact that some ingredients belong to the large chemical family of aromatic amines. Herein, we investigated the cytotoxicity of Basic Red 51 (BR51) in immortalized human keratinocytes (HaCaT). BR51 is a temporary hair dye that belongs to the azo group (NN); the cleavage of this bond may result in the release of toxic aromatic amines. The half maximal effective concentration (EC50) in HaCaT cells is 13mug/mL. BR51 induced a significant decrease on expression of p21 in a dose dependent manner. p53 was not affected, whereas BR51 decreased procaspase 8 and cleaved procaspase 9. These results proved that caspase 3 is fully involved in BR51-induced apoptosis. The dye was also able to stop this cell cycle on G2 in sub-toxic doses. Moreover, we reconstructed a 3D artificial epidermis using HaCaT cells; using this model, we observed that BR51 induced cell injury and cells were undergoing apoptosis, considering the fragmented nuclei. Subsequently, BR51 induced reactive oxygen species (ROS) leading to an increase on the levels of 8-oxo-dG. In conclusion, we provide strong evidence that consumer and/or professional exposure to BR51 poses risk to human health

Allergens of permanent hair dyes induces epidermal damage, skin barrier loss and IL-1 α increase in epidermal in vitro model

Allergic and irritant skin reactions caused by topical exposure to permanent hair dyes are a common problem. For regulatory and ethnical purposes, it is required to perform chemical safety assessment following the replacement, reduction, and refinement of animal testing (3Rs). Permanent hair dyes are formed by a mixture of ingredients that vary from low to extreme skin sensitizing potency and that inter-react to form unknown by-products. Because of the complex reaction, this cytotoxic mechanism has not yet been elucidated and is the subject of this study. Here, we topically exposed p-phenylenediamine (PPD), Resorcinol (RES), Hydrogen Peroxide (H2O2) alone or as a mixture to RhE and evaluated parameters related to skin irritation such as epidermal viability, keratinocytes damage, barrier loss and IL-1 α. Our data indicates that ingredients tested alone did not lead to an increase of cytotoxic parameters related to skin irritation. However, when the mixture of PPD/H2O2/RES and PPD/H2O2 was applied to the RhE, some of the parameters such as morphological changes including the presence of apoptotic cells, barrier loss and increased IL- 1 α release were observed. The results indicate that the mixture of ingredients used in permanent hair dyes have an irritant effect in RhE while the ingredients alone not

The oxidation of p-phenylenediamine, an ingredient used for permanent hair dyeing purposes, leads to the formation of hydroxyl radicals: Oxidative stress and DNA damage in human immortalized keratinocytes

The hair-dyeing ingredient, p-phenylenediamine (PPD), was previously reported to be mutagenic, possibly by inducing oxidative stress. However, the exact mechanism of PPD in inducing oxidative stress upon skin exposure during hair-dyeing in human keratinocytes remains unknown. The aim of our studies was therefore to investigate the toxicity of PPD and its by-products in human immortalized keratinocytes (HaCaT) after auto-oxidation and after reaction with hydrogen peroxide (H2O2). We found that the PPD half maximal effective cytotoxic concentration (EC50) to HaCaT is 39.37 and 35.63mug/mL after 24 and 48h, respectively, without addition of H2O2 to induce oxidation. When PPD (10 or 100mug/mL) is combined with 10.5mug/mL of H2O2, intracellular ROS production by HaCaT after 1h was significantly increased and enhanced levels of DNA damage were observed after 4h of exposure. After 24h incubations, 20mug/mL of PPD increased the level of DNA oxidation in HaCaT. Also, we found that the in vitro reaction between PPD and H2O2, even below the maximum allowance by cosmetic industries, released hydroxyl radicals which can damage DNA. Taken together, we conclude that PPD alone and when combined with H2O2 increases the formation of reactive oxygen species in human keratinocytes, leading to oxidative stress and subsequent DNA damage. These alterations suggest that the mechanism by which PPD exposure, alone or combined with H2O2, damages keratinocytes by the formation of the high reactive HO radicals

Microencapsulation and tissue engineering as an alternative treatment of diabetes

In the 70's, pancreatic islet transplantation arose as an attractive alternative to restore normoglycemia; however, the scarcity of donors and difficulties with allotransplants, even under immunosuppressive treatment, greatly hampered the use of this alternative. Several materials and devices have been developed to circumvent the problem of islet rejection by the recipient, but, so far, none has proved to be totally effective. A major barrier to transpose is the highly organized islet architecture and its physical and chemical setting in the pancreatic parenchyma. In order to tackle this problem, we assembled a multidisciplinary team that has been working towards setting up the Human Pancreatic Islets Unit at the Chemistry Institute of the University of São Paulo, to collect and process pancreas from human donors, upon consent, in order to produce purified, viable and functional islets to be used in transplants. Collaboration with the private enterprise has allowed access to the latest developed biomaterials for islet encapsulation and immunoisolation. Reasoning that the natural islet microenvironment should be mimicked for optimum viability and function, we set out to isolate extracellular matrix components from human pancreas, not only for analytical purposes, but also to be used as supplementary components of encapsulating materials. A protocol was designed to routinely culture different pancreatic tissues (islets, parenchyma and ducts) in the presence of several pancreatic extracellular matrix components and peptide growth factors to enrich the beta cell population in vitro before transplantation into patients. In addition to representing a therapeutic promise, this initiative is an example of productive partnership between the medical and scientific sectors of the university and private enterprises