Characterization of the rat pituitary capsule: Evidence that the cerebrospinal fluid filled the pituitary cleft and the inner side of the capsule.

In humans, the pituitary gland is covered by a fibrous capsule and is considered a continuation of the meningeal sheath. However, in rodents some studies concluded that only the pars tuberalis (PT) and pars nervosa (PN) are enwrapped by the pia mater, while others showed that the whole gland is covered by this sheath. At PT the median eminence subarachnoid drains cerebrospinal fluid (CSF) to its cisternal system representing a pathway to the hypothalamus. In the present study we examined the rat pituitary capsule to elucidate its configuration, its physical interaction with the pituitary border and its relationship with the CSF. Furthermore, we also revisited the histology of the pituitary cleft and looked whether CSF drained in it. To answer such questions, we used scanning and transmission electron microscopy, intracerebroventricular infusion of Evan´s blue, fluorescent beads, and sodium fluorescein. The latter was measured in the pars distalis (PD) and various intracranial tissues. We found a pituitary capsule resembling leptomeninges, thick at the dorsal side of the pars intermedia (PI) and PD, thicker at the level of PI in contiguity with the PN and thinner at the rostro-ventral side as a thin membrane of fibroblast-like cells embedded in a fibrous layer. The capsule has abundant capillaries on all sides. Our results showed that the CSFs bathe between the capsule and the surface of the whole gland, and ciliate cells are present in the pituitary border. Our data suggest that the pituitary gland intercommunicates with the central nervous system (CNS) through the CSF.

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.

Antimetastatic, antineoplastic, and toxic effects of 4-hydroxycoumarin in a preclinical mouse melanoma model.

PURPOSE: We have previously reported that in vitro treatment of B16-F10 melanoma cells with 4-hydroxycoumarin (4-HC) decreases their metastatic potential. However, the antimetastatic efficacy of 4-HC in vivo is unknown; therefore, we investigated the antimetastatic and antineoplastic effects of 4-HC in a mouse melanoma model. Based on the findings, the immunomodulatory and toxic effects of 4-HC were also studied. METHODS: Experimental metastasis assay was performed in C57BL/6 mice that received 4-HC before intravenous injection of B16-F10 cells. Antitumor and antimetastatic efficacy of 4-HC was assessed in mice implanted subcutaneously with melanoma cells. Possible immunostimulant and toxic effects of 4-HC were studied in healthy mice. RESULTS: 4-HC reduced the number of experimental lung metastases. Moreover, 4-HC diminished primary tumor growth and increased survival time in mice bearing melanoma tumors. Treatments also decrease spontaneous lung metastases in the same animals. Different to other coumarins, the antitumor effect of 4-HC seems to be unrelated to immunostimulation, since plasma concentrations of cytokines remained unchanged. In contrast, toxic histological changes in nephrons and bronchiolar epithelium and a pronounced anticoagulant effect were found in 4-HC treated animals. CONCLUSIONS: These results show that 4-HC not only exhibit antimetastatic effect in vivo, but also effectively reduces tumor growth and improves survival, even when it produce toxic effects. Although the molecular mechanism of 4-HC actions needs to be further defined, our data suggest that 4-HC may lead to the development of agents that could be used as adjuvants in the therapy of melanoma.

Combined therapy with amphotericin B and caspofungin in an experimental model of disseminated histoplasmosis.

OBJECTIVE: To assess the effect of amphotericin B and caspofungin, as well as their combinations in the therapy of experimental disseminated histoplasmosis. MATERIAL AND METHODS: BALB/c mice were intraperitoneally infected with four different strains of Histoplasma capsulatum and given to antifungal treatments. The response to intraperitoneal therapy with amphotericin B (0.5, 1.0, and 2.0 mg/kg of body weight) or caspofungin (10 mg/kg of body weight) and their combinations, was evaluated by the quantification of yeast colony-forming units (CFU) per gram of spleen or lung, from each animal. Additionally, the pathogen was monitored histopathologically in the excised organs. Data were analyzed with the Kruskall-Wallis and Tukey tests. RESULTS: Caspofungin was more effective than amphotericin B in reducing the CFU/ g. A synergistic effect was observed when caspofungin (10 mg/ kg) was combined with amphotericin B (0.5 or 1.0 mg/kg). Significant differences in CFU values, H = 119.78 (P = 0.00001), were found among the treatment groups. However, statistical analyses did not reveal significant differences, H = 2.837 (P = 0.428), in the therapeutic responses with the four H. capsulatum strains tested. CONCLUSION: Combined therapy with amphotericin B and caspofungin could represent an alternative treatment to be explored in severe human histoplasmosis.

Efficient post-exposure prophylaxis against rabies by applying a four-dose DNA vaccine intranasally.

We tested two post-exposure prophylaxes (PEPs) for rabies in laboratory animals; one was a traditional antirabies vaccine for humans via intramuscular route (IM), and the other was a DNA vaccine administered by intranasal route (IN). In contrast to The World Health Organization's recommended five-dose PEP, we gave only four doses without hyper-immune antirabies sera, making the PEP more rigorous. All animals were challenged with challenge virus strain (CVS); 16h later, PEP was applied. All animals that received the PEP with DNA/IN survived, and 87% of the rabbits and 80% of the mice that received the PEP with traditional antirabies vaccine/IM survived. Negative controls succumbed to infection. The expression of G protein was detected in the NALT, cerebellum, cerebral cortex (neocortex), cerebellum and hippocampus, mainly in the glial cells (microglia) and microvessels. On the other hand, plasmid construct was detected in brain and its mRNA expression in medium and posterior encephalon. The efficiency of this DNA/IN PEP is probably due to the early expression of the antigen in the brain stimulating the immune system locally.