Essential Oil of Bursera morelensis Promotes Cell Migration on Fibroblasts: In Vitro Assays.

Essential oils (EOs) are complex mixtures of volatile natural compounds. We have extensively studied the EO of Bursera morelensis, which demonstrates antibacterial, antifungal, anti-inflammatory, and wound-healing activities. The objective of this work was to determine the effect of this EO on fibroblast migration in a three-dimensional in vitro model. For the three-dimensional in vitro model, a series of fibrin hydrogel scaffolds (FSs) were built in which fibroblasts were cultured and subsequently stimulated with fibroblast growth factor (FGF) or EO. The results demonstrated that these FSs are appropriate for fibroblast culture, since no decrease in cell viability or changes in cell proliferation were found. The results also showed that this EO promotes cell migration four hours after stimulation, and the formation of cell projections (filopodia) outside the SF was observed. From these results, we confirmed that part of the mechanism of action of the essential oil of B. morelensis during the healing process is the stimulation of fibroblast migration to the wound site.

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.

Wound Healing Activity of α-Pinene and α-Phellandrene.

Bursera morelensis is used in Mexican folk medicine to treat wounds on the skin. Recently, it was shown that the essential oil (EO) of B. morelensis has wound healing activity, accelerating cutaneous wound closure and generating scars with good tensile strength. α-pinene (PIN) and α-phellandrene (FEL) are terpenes that have been found in this EO, and it has been shown in different studies that both have anti-inflammatory activity. The aim of this study was to determine the wound healing activity of these two terpenes. The results of in vitro tests demonstrate that PIN and FEL are not cytotoxic at low concentrations and that they do not stimulate fibroblast cell proliferation. In vivo tests showed that the terpenes produce stress-resistant scars and accelerate wound contraction, due to collagen deposition from the early stages, in wounds treated with both terpenes. Therefore, we conclude that both α-pinene and α-phellandrene promote the healing process; this confirms the healing activity of the EO of B. morelensis, since having these terpenes as part of its chemical composition explains part of its demonstrated activity.

Wound Healing Activity of the Essential Oil of Bursera morelensis, in Mice.

Bursera morelensis is used in Mexican folk medicine to treat wounds on the skin. It is an endemic tree known as "aceitillo", and the antibacterial and antifungal activity of its essential oil has been verified; it also acts as an anti-inflammatory. All of these reported biological activities make the essential oil of B. morelensis a candidate to accelerate the wound-healing process. The objective was to determine the wound-healing properties of B. morelensis' essential oil on a murine model. The essential oil was obtained by hydro-distillation, and the chemical analysis was performed by gas chromatography-mass spectrometry (GC-MS). In the murine model, wound-healing efficacy (WHE) and wound contraction (WC) were evaluated. Cytotoxic activity was evaluated in vitro using peritoneal macrophages from BALB/c mice. The results showed that 18 terpenoid-type compounds were identified in the essential oil. The essential oil had remarkable WHE regardless of the dose and accelerated WC and was not cytotoxic. In vitro tests with fibroblasts showed that cell viability was dose-dependent; by adding 1 mg/mL of essential oil (EO) to the culture medium, cell viability decreased below 80%, while, at doses of 0.1 and 0.01 mg/mL, it remained around 90%; thus, EO did not intervene in fibroblast proliferation, but it did influence fibroblast migration when wound-like was done in monolayer cultures. The results of this study demonstrated that the essential oil was a pro-wound-healing agent because it had good healing effectiveness with scars with good tensile strength and accelerated repair. The probable mechanism of action of the EO of B. morelensis, during the healing process, is the promotion of the migration of fibroblasts to the site of the wound, making them active in the production of collagen and promoting the remodeling of this collagen.

Histological Analysis of Intra-Abdominal Adhesions Treated with Sodium Hyaluronate and Carboxymethylcellulose Gel.

PURPOSE: To evaluate macro and microscopically the adhesions developed after using the anti-adherence compound sodium hyaluronate and carboxymethylcellulose (SH-CBMC) gel and to determine the volume of the adhesions using a stereological estimation. METHODS: The study was experimental, random, comparative, and prospective. The subjects of the study were male Wistar rats divided in three groups (n = 10). Group I (control) included rats with no peritoneal injury. Group II rats had a 2 cm diameter injury created bilaterally in the parietal peritoneum at 3 cm from the abdominal midline with electrocautery coated with physiological solution. Group III rats were given the same injuries and coated with SH-CBMC gel. All groups were followed up postoperatively for 30 days, after which a laparotomy was performed to macroscopically determine the presence and type of adhesions. Experimental models were euthanized with anesthetic overdose and biopsies were taken for histopathological examination and stereological estimate of the volume of adhesions. RESULTS: Macroscopic adhesions were 20% less prevalent in Group III compared to Group II, which presented 40% more multiple and firm adhesions, unlike in Group III, in which they were unique and lax. There was a statistically significant decrease in the presence and number of adhesions in rats treated with SH-CBMC gel. Inflammatory infiltrate was significantly lower in rats treated with SH-CBMC gel, but there were no differences in connective tissue, fibrosis, and angiogenesis among groups. There was no statistical difference in the overall volume of adhesions among the treatment groups. CONCLUSIONS: SH-CBMC gel reduces macroscopic presence and number of adhesions and the severity of the inflammatory infiltrate.

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.

Role of epidermal dendritic cells in drug-induced cutaneous adverse reactions.

Drug-induced adverse reactions (ADR) include any undesirable pharmacological effect that occurs following drug administration at therapeutic doses. The appearance of ADR significantly limits the use of drugs in as much as their clinical symptoms may range from very mild discomfort such as cutaneous rash, up to very severe, or even fatal tissue necrolysis such as the Stevens Johnson syndrome.One of the most frequently involved organ during ADR is the skin. Drug-induced cutaneous reactions (CDR) incidence is variable but they may appear in 2-3% of ambulatory patients, and it may increase to 10-15% when patients are hospitalized, or even be as high as 60% when co morbidity includes the presence of virus, bacteria, or parasites.Due to the fact that skin is one of the organs most frequently involved in ADR, in this work we analyze and propose a mechanism by which epidermal dendritic cells operating as the sentinels of the skin neuro-immune-endocrine system may contribute to CDR via either immunogenic or tolerogenic immune responses towards drugs, whenever they are administered topic or systemically.