p19Arf sensitizes B16 melanoma cells to interferon-ß delivered via mesenchymal stem cells in vitro.

The immune stimulatory and anti-neoplastic functions of type I interferon have long been applied for the treatment of melanoma. However, the systemic application of high levels of this recombinant protein is often met with toxicity. An approach that provides localized, yet transient, production of type I interferon may overcome this limitation. We propose that the use of mesenchymal stem cells (MSCs) as delivery vehicles for the production of interferon-ß (IFNß) may be beneficial when applied together with our cancer gene therapy approach. In our previous studies, we have shown that adenovirus-mediated gene therapy with IFNß was especially effective in combination with p19Arf gene transfer, resulting in immunogenic cell death. Here we showed that MSCs derived from mouse adipose tissue were susceptible to transduction with adenovirus, expressed the transgene reliably, and yet were not especially sensitive to IFNß production. MSCs used to produce IFNß inhibited B16 mouse melanoma cells in a co-culture assay. Moreover, the presence of p19Arf in the B16 cells sensitizes them to the IFNß produced by the MSCs. These data represent a critical demonstration of the use of MSCs as carriers of adenovirus encoding IFNß and applied as an anti-cancer strategy in combination with p19Arf gene therapy.

p19Arf sensitizes B16 melanoma cells to interferon-β delivered via mesenchymal stem cells in vitro

The immune stimulatory and anti-neoplastic functions of type I interferon have long been applied for the treatment of melanoma. However, the systemic application of high levels of this recombinant protein is often met with toxicity. An approach that provides localized, yet transient, production of type I interferon may overcome this limitation. We propose that the use of mesenchymal stem cells (MSCs) as delivery vehicles for the production of interferon-β (IFNβ) may be beneficial when applied together with our cancer gene therapy approach. In our previous studies, we have shown that adenovirus-mediated gene therapy with IFNβ was especially effective in combination with p19Arf gene transfer, resulting in immunogenic cell death. Here we showed that MSCs derived from mouse adipose tissue were susceptible to transduction with adenovirus, expressed the transgene reliably, and yet were not especially sensitive to IFNβ production. MSCs used to produce IFNβ inhibited B16 mouse melanoma cells in a co-culture assay. Moreover, the presence of p19Arf in the B16 cells sensitizes them to the IFNβ produced by the MSCs. These data represent a critical demonstration of the use of MSCs as carriers of adenovirus encoding IFNβ and applied as an anti-cancer strategy in combination with p19Arf gene therapy.

Efeito da inclusão da torta de macaúba na população de protozoários do rúmen de vacas leiteiras / The effect of macauba meal inclusion in the ruminal protozoa population of dairy cows

Avaliaram-se os efeitos da inclusão da torta de macaúba (TM) na dieta sobre a concentração e o perfil da população de protozoários ciliados do rúmen. Foram utilizadas oito vacas Holandesas, dispostas em dois quadrados latinos 4x4 e alimentadas com diferentes níveis de inclusão da TM (0, 100, 200 e 300g kg-1 de matéria seca na dieta total). Para quantificação e identificação dos protozoários, amostras de 1mL do líquido ruminal de cada animal foram diluídas em 9mL de solução de formaldeído a 10%. Após diluições decimais sucessivas em solução salina, os protozoários foram quantificados em câmara de Sedgewick. A identificação foi realizada sob a luz da microscopia óptica, utilizando-se a objetiva de 40x para caracterizar o mínimo de 100 indivíduos por amostra. As concentrações de protozoários pequenos (P<0,01), médios (P=0,05), grandes (P=0,01) e o número total (P<0,01) apresentaram comportamento linear decrescente com o aumento dos níveis de inclusão da TM nas dietas. Foram encontradas características morfológicas típicas de 16 gêneros de protozoários ciliados, sendo os gêneros Entodinium e Eodinium os mais frequentes no líquido ruminal dos animais avaliados. O perfil da população de protozoários ciliados sofreu pequenas alterações com a inclusão da TM, sendo observada alteração na participação percentual dos gêneros Charonina (P = 0,03) e Polyplastron (P<0,01). A utilização da TM na dieta de bovinos promove redução na concentração e modificação no perfil da população de protozoários ciliados do rúmen.

The effects of meal macaúba (MM) inclusion on the concentration and profile of the ciliate protozoa of the bovine rumen were evaluated. Eight Holstein cows were used during second lactation, with average 480 kg of body weight, arranged in two 4x4 Latin squares simultaneously and fed four diets with different concentrations of the MM (0, 100, 200 and 300 g kg-1). For identification and quantification of protozoa, samples of ruminal fluid from each animal with 1 ml were diluted in 9mL of 10% formaldehyde. After consecutive decimal dilutions in saline solution, protozoa were assessed in a Sedgewick chamber. The identification was carried out under the optical light microscope using a 40x objective to characterize at least 100 individuals per sample. The concentrations of small (P<0.01), medium (P=0.05), large (P= 0.01), and the total number (P<0.01) of protozoa decreased linearly with the increasing MM inclusion. Morphological characteristics typical of 16 genera of ciliates were found, being the Entodinium spp. and Eodinium spp. the most frequent in the rumen fluid of animals evaluated. The profile of the population of ciliated protozoa suffer minor changes with the addition of MM, with a change in the percentage share of Charonina spp. (P = 0.03) and Polyplastron spp. (P <0.01). The use of the MM in the cattle diet promotes reduction in the concentration and change in the profile of the population of rumen ciliates protozoa.

Mutagenic potential and modulatory effects of the medicinal plant Luehea divaricata (Malvaceae) in somatic cells of Drosophila melanogaster: SMART/wing.

Luehea divaricata is a native plant of the Brazilian Cerrado, known as "açoita-cavalo". It is used as a popular herbal medicine in the treatment of dysentery, bleeding, arthritis, tumors, ulcers, and gangrenous wounds. Considering that herbal medicines sometimes provoke tumors and/or may prevent mutational events, it is important to study the action of these natural drugs on DNA. Aqueous extract of the bark of L. divaricata was evaluated at three different concentrations (0.10, 0.30, 0.50 mg/mL), individually and in combination with the neoplastic drug doxorubicin (DXR), by the somatic mutation and recombination test (SMART/wing) in Drosophila melanogaster. Distilled water was included as a negative control. The mutation frequency in the treatments with L. divaricata extract alone was not significantly higher than in the negative control for standard (ST) and high bioactivation (HB) crosses. When L. divaricata extract was combined with DXR, there was a significant reduction in the frequency of spots when compared to DXR alone, in both crosses. Further studies with other experimental models would be useful to confirm that L. divaricata extract is not harmful and that it could be used in the prevention of cancer.

Modulatory effect of Palicourea coriacea (Rubiaceae) against damage induced by doxorubicin in somatic cells of Drosophila melanogaster.

Palicourea coriacea, popularly known as "douradinha", is a medicinal plant from the Brazilian Cerrado region used in folk medicine to treat kidney and urethral stones and kidney inflammation. We evaluated the cytotoxic, genotoxic, and possible antigenotoxic activities of an aqueous extract of P. coriacea on somatic cells of Drosophila melanogaster, using the somatic mutation and recombination test. We used third-stage larvae of D. melanogaster from a standard cross and a high bioactivation cross and tested 10 different doses of P. coriacea aqueous extract (5, 15, 25, 35, 50, 65, 80, 95, 110, and 125 mg/mL). Doxorubicin (0.125 mg/mL) was used as a positive control and distilled water as a negative control. None of the doses was lethal to the larvae.There was no genotoxic effect at 5, 10, or 15 mg extract/mL. However, a significant decrease in the frequency of spots induced by doxorubicin was observed when administered with P. coriacea aqueous extract at these same doses. We conclude that P. coriacea aqueous extract is not cytotoxic or genotoxic at these doses, but it does protect against the genotoxic action of doxorubicin.