Subcutaneous Application of a Gelatin/Hyaluronic Acid Hydrogel Induces the Production of Skin Extracellular Matrix.

The development of injectable hydrogels with natural biopolymers such as gelatin (Ge) and hyaluronic acid (Ha) is widely performed due to their biocompatibility and biodegradability. The combination of both polymers crosslinked with N-Ethyl-N'-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC) can be used as an innovative dermal filler that stimulates fibroblast activity and increases skin elasticity and tightness. Thus, crosslinked Ge/Ha hydrogels with different concentrations of EDC were administered subcutaneously to test their efficacy in young and old rats. At higher EDC concentrations, the viscosity decreases while the particle size of the hydrogels increases. At all concentrations of EDC, amino and carboxyl groups are present. The histological analysis shows an acute inflammatory response, which disappears seven days after application. At one and three months post-treatment, no remains of the hydrogels are found, and the number of fibroblasts increases in all groups in comparison with the control. In addition, the elastic modulus of the skin increases after three months of treatment. Because EDC-crosslinked Ge/Ha hydrogels are biocompatible and induce increased skin tension, fibroblast proliferation, and de novo extracellular matrix production, we propose their use as a treatment to attenuate wrinkles and expression lines.

Gelatin/Hyaluronic Acid Scaffold Coupled to CpG and MAGE-A5 as a Treatment against Murine Melanoma.

The half-time of cells and molecules used in immunotherapy is limited. Scaffolds-based immunotherapy against cancer may increase the half-life of the molecules and also support the migration and activation of leukocytes in situ. For this purpose, the use of gelatin (Ge)/hyaluronic acid (HA) scaffolds coupled to CpG and the tumor antigen MAGE-A5 is proposed. Ge and HA are components of the extracellular matrix that stimulate cell adhesion and activation of leucocytes; CpG can promote dendritic cell maturation, and MAGE-A5 a specific antitumor response. C57BL/6 mice were treated with Ge/HA/scaffolds coupled to MAGE-A5 and/or CpG and then challenged with the B16-F10 melanoma cell line. Survival, tumor growth rate and the immune response induced by the scaffolds were analyzed. Ge/HA/CpG and Ge/HA/MAGE-A5 mediated dendritic cell maturation and macrophage activation, increased survival, and decreased the tumor growth rate and a tumor parenchyma with abundant cell death areas and abundant tumor cells with melanin granules. Only the scaffolds coupled to MAGE-A5 induced the activation of CD8 T cells. In conclusion, Ge/HA scaffolds coupled to CpG or MAGE-A5, but not the mixture, can induce a successful immune response capable of promoting tumor cell clearance and increased survival.

Immunomodulatory Properties of Masticadienonic Acid and 3α-Hydroxy Masticadienoic Acid in Dendritic Cells.

Dendritic cells are antigen-presenting cells, which identify and process pathogens to subsequently activate specific T lymphocytes. To regulate the immune responses, DCs have to mature by the recognition of TLR ligands, TNFα or IFNγ. These ligands have been used as adjuvants to activate DCs in situ or in vitro, with toxic effects. It has been shown that some molecules affect the immune system, e.g., Masticadienonic acid (MDA) and 3α-hydroxy masticadienoic acid (3α-OH MDA) triterpenes naturally occurring in several medicinal plants, since they activate the nitric oxide synthase in macrophages and induce T lymphocyte proliferation. The DCs maturation induced by MDA or 3a-OH MDA was determined by incubating these cells with MDA or 3α-OH MDA, and their phenotype was afterwards analyzed. The results showed that only 3α-OH MDA was able to induce DCs maturation. When mice with melanoma were inoculated with DCs/3α-OH MDA, a decreased tumor growth rate was observed along with an extended cell death area within tumors compared to mice treated with DCs incubated with MDA. In conclusion, it is proposed that 3α-OH MDA may be an immunostimulant molecule. Conversely, it is proposed that MDA may be a molecule with anti-inflammatory properties.

Epicutaneous Administration of 17ß-Estradiol Induces Langerhans Cells Depletion.

BACKGROUND: Langerhans cells (LC) number and function in mouse vaginal mucosa are affected by 17ß-estradiol (E2) application; nonetheless, its effect on epidermal LC has not been studied. The purpose of this study was to evaluate the effect of topical administration of E2 on the number, phenotype, and migratory ability of LC in mouse skin. METHODS: Ears of adult CD1 male mice were topically treated once with several doses. Immunohistochemical staining for CD207 and TUNEL staining were performed. LC migration to lymph nodes and the effect on the expression of costimulatory molecules on cultured dendritic cells (DC) were also evaluated. RESULTS: E2 decreased the number of CD207+ LC in a dose-dependent manner. One hour after treatment, 1 and 10 µg/mL E2 significantly reduced the LC number by 21% and 26%, respectively, after two hours, the reduction was 23% and 41%, respectively. After 48 hours, LC recovered, and after 96 hours of treatment, the CD207+/MHCII+ DC numbers were increased in regional lymph nodes. However, CD86 and CD40 molecules were expressed at lower levels than in positive control. The TUNEL assay did not show apoptotic cells. Furthermore, in cultured DC, E2 promoted a decrease in CD40 and CD86 expression and an increase in CD273, CD274, MHCII, and CCR7. CONCLUSIONS: The topical administration of E2 induced a transitory local diminution of LC population and a tolerogenic phenotype. This decrease in epidermal LC suggests that E2 may affect skin immune responses, inducing an inhibitory response, which should be considered when prescribing topical E2 medications.

GK-1 improves the immune response induced by bone marrow dendritic cells loaded with MAGE-AX in mice with melanoma.

The aim of dendritic cell (DC) vaccination in cancer is to induce tumor-specific effector T cells that may reduce and control tumor mass. Immunostimulants that could drive a desired immune response are necessary to be found in order to generate a long lasting tumor immune response. GK-1 peptide, derived from Taenia crassiceps, induces not only increase in TNFα, IFNγ, and MCP-1 production in cocultures of DCs and T lymphocytes but also immunological protection against influenza virus. Moreover, the aim of this investigation is the use of GK-1 as a bone marrow DCs (BMDCs) immunostimulant targeted with MAGE antigen; thus, BMDC may be used as immunotherapy against murine melanoma. GK-1 induced in BMDCs a meaningful increment of CD86 and IL-12. In addition, the use of BMDCs TNFα/GK-1/MAGE-AX induced the highest survival and the smallest tumors in mice. Besides, the treatment helped to increase CD8 lymphocytes levels and to produce IFNγ in lymph nodes. Moreover, the histopathological analysis showed that BMDCs treated with GK-1/TNFα and loaded with MAGE-AX induced the apparition of more apoptotic and necrotic areas in tumors than in mice without treatment. These results highlight the properties of GK-1 as an immunostimulant of DCs and suggest as a potential candidate the use of this immunotherapy against cancer disease.

Inhaled vanadium pentoxide decrease gamma-tubulin of mouse testes at different exposure times.

Vanadium is an important environmental and industrial pollutant whose concentrations have increased in the last decades. Due to its status as reproductive toxicant and a microtubule damaging agent, the present study investigated by immunohistochemistry the effect of the inhalation of vanadium pentoxide on gamma-tubulin within somatic and testicular germ cells. Male mice inhaled vanadium pentoxide (V2O5) (0.02 M) 1 h/twice a week for 12 weeks. Our results demonstrated that vanadium accumulates in the testes starting with the initial inhalation (24 h), and this pattern remained until the last week of treatment. In general, vanadium was capable of significantly decreasing the percentage of gamma-tubulin in all analyzed testicular cells (Sertoli, Leydig and germ cells) starting with the first week of treatment. For all cell types studied, regression analysis revealed a negative and significant relationship between the percentage of immunopositive cells to gamma-tubulin and exposure time, showing a time dependent response in all cases. Our findings suggest that alterations on this protein might imply changes in microtubule-involved function such as cell division, which in the testes might lead to damage in the spermatogenesis, leading probably to infertility.