Efecto de la escarificación y estratificación sobre la germinación in vitro de Aristotelia chilensis (Molina) Stuntz

Two in vitro assays were performing in order to improve Aristotelia chilensis germination rate. Scarification was tested in the first assay and compare to a control. In the second assay, cold stratification was applied during 2, 4, 6 and 8 weeks at 4 °C in darkness, plus an unstratified control. Seeds were germinated on Petri dish containing water and agar (WA), and incubated for 60 days in a controlled atmosphere at 25 ± 2 °C using cold white fluorescent light tubes (50 ?mol m-2 s-1 light intensity), under a photoperiod of 16 h light and 8 h of darkness. All treatments achieved higher average germination rates than the control. The highest germination rate was obtained with scarification (Gmáx, 92%) though the fastest growth was achieved with 8 weeks of stratification, reaching the lowest average germination time of 18 days. The results demonstrate that A. chilensis seeds present a moderate level of primary dormancy, both exogenous and endogenous.Con el fin de mejorar la germinación de semillas de Aristotelia chilensis, se establecieron dos ensayos in vitro. En el primero, se realizó escarificación y se comparó con un testigo. En el segundo, se aplicó estratificación fría por diferentes períodos de tiempo (2, 4, 6 y 8 semanas a 4 °C en oscuridad), más un testigo sin estratificar. Las semillas fueron germinadas en placas con agua y agar (AA) e incubadas por 60 días en cámara de cultivo a 25 ± 2 °C, con tubos fluorescentes de luz fría blanca bajo un fotoperíodo de 16 h luz, 8 h de oscuridad y una intensidad lumínica de 50 ?mol m-2 s-1. En todos los tratamientos se logró valores promedios de germinación superiores a los testigos. La mayor germinación se obtuvo con escarificación (Gmáx, 92%), no obstante la mayor velocidad inicial de germinación se registró con la estratificación por ocho semanas, alcanzando el menor tiempo medio de germinación (TM, 18 días). Los resultados demuestran que la semilla de A. chilensis presenta dormancia primaria de nivel moderado tanto exógena como endógena

Enraizamiento in vitro y ex vitro de microtallos de Ugni molinae Turcz., una especie nativa de Chile

Two trials were established to evaluate the in vitro and ex vitro rooting of selected microshoots of two selected clones of Ugnimolinae. In the in vitro experiment, nine treatments were compared, combination of three concentrations of Murashige and Skoogmedium (MS) (1, ½ and ¼ strength macronutrients) and three concentration of indole 3-butyric acid (IBA, at 0, 1 and 2 ?M). In theex vitro assay we established for IBA four concentrations (0; 4.9; 9.8 and 19.7 mM). After 30 days total survival, rooting percentage,root length and number of roots per explants were evaluated. Both clones responded differently to the treatments used. Under in vitroconditions, Clone 1 had an average of 97% rooting at ½ MS + 1 ?M IBA, whereas Clone 2 obtained a 93% of rooting with ¼ MS+ 2 ?M IBA. Our results indicate that the decrease in the strength of MS medium and the addition of IBA promotes rooting. Withregard to ex vitro conditions, the Clone 1 did not respond to IBA applications, while in Clone 2, IBA promoted the rooting at theconcentration of 19.7 mM, with percentages of 90 and 85%, respectively. Considering these results, it is generally recommended touse the ex vitro rooting system because both clones were able to obtain the same high percentage rooting (above 85%) in this system.Ex vitro propagation will also safe costs.Con el objetivo de evaluar el enraizamiento in vitro y ex vitro de microtallos de dos clones de Ugni molinae, se establecierondos ensayos. En el experimento de enraizamiento in vitro se contrastaron nueve tratamientos producto de la combinación de tresconcentraciones de medio Murashige y Skoog (MS) (1, ½ y ¼ de los macronutrientes diluidos) y tres de ácido 3-indolbutírico(AIB) (0, 1 y 2 ?M). En tanto, para el ensayo de enraizamiento ex vitro se contrastaron cuatro tratamientos correspondientes a lasconcentraciones de 0; 4,9; 9,8 y 19,7 mM de AIB. Después de 30 días se evaluó la supervivencia, enraizamiento, longitud de raíz ynúmero de raíces por explante. El tratamiento que logró el mayor enraizamiento difirió entre ambos clones. En condiciones in vitroel Clon 1 obtuvo un promedio de enraizamiento de 97% con ½ MS + 1 ?M de AIB, mientras que el Clon 2 obtuvo un 93% con ¼ deMS + 2 ?M de AIB. Nuestros resultados indican que la disminución de sales en el medio de cultivo MS y la adición de AIB favoreceel enraizamiento. Con respecto a las condiciones ex vitro, el Clon 1 no respondió a las aplicaciones de AIB, en cambio en el Clon 2, elAIB promovió el enraizamiento a la concentración de 19,7 mM, alcanzando porcentajes de 90 y 85%, respectivamente. En virtud delos resultados obtenidos, se sugiere un enraizamiento ex vitro para los clones 1 y 2, para evitar los costos asociados al enraizamientoin vitro, ya que independiente del sistema utilizado, ambos clones obtuvieron enraizamientos iguales o superiores a un 85% en elmejor de sus tratamientos

Sublethal doses of dinophysistoxin-1 and okadaic acid stimulate secretion of inflammatory factors on innate immune cells: Negative health consequences

© 2016 Elsevier LtdOne of the proposed mechanisms to explain why Diarrhetic Shellfish Poison (DSP) toxins are tumor promoters is founded on the capacity of these toxins to increase TNF-α secretion. Although macrophages are the principal cells in the activation of the inflammatory response, the immune profile that Okadaic acid (OA) and Dinophysistoxin-1 (DTX-1) trigger in these cells has not been fully explored. We have therefore investigated the effect of various concentrations of both toxins on the activity of several inflammatory factors. Our results demonstrate that OA and DTX-1, at sublethal doses, stimulate secretion of inflammatory factors. Nevertheless DTX-1 was more potent than OA in increasing TNF-α and IL-6 as well as their dependent chemokines KC, MCP-1, LIX, MIP-1 α, MIP-1 β and MIP-2. On the other hand, secretion of IFN-γ and the anti-inflammatory cytokines, IL-4 and IL-10, was unaffected. In addition, DTX-1 also raises matrix metalloproteinase-9 (MMP-9) activity. In thi

Effect of scarification and stratification on the in vitro germination of Aristotelia chilensis (Molina) Stuntz

Two in vitro assays were performing in order to improve Aristotelia chilensis germination rate. Scarification was tested in the first assay and compare to a control. In the second assay, cold stratification was applied during 2, 4, 6 and 8 weeks at 4 degrees C in darkness, plus an unstratified control. Seeds were germinated on Petri dish containing water and agar (WA), and incubated for 60 days in a controlled atmosphere at 25 +/- 2 degrees C using cold white fluorescent light tubes (50 mu mol m(-2) s(-1) light intensity), under a photoperiod of 16 h light and 8 h of darkness. All treatments achieved higher average germination rates than the control. The highest germination rate was obtained with scarification (G(max), 92%) though the fastest growth was achieved with 8 weeks of stratification, reaching the lowest average germination time of 18 days. The results demonstrate that A. chilensis seeds present a moderate level of primary dormancy, both exogenous and endogenous

In vitro and ex vitro rooting of Ugni molinae Turcz. microshoots, a native species to Chile

Two trials were established to evaluate the in vitro and ex vitro rooting of selected microshoots of two selected clones of Ugni molinae. In the in vitro experiment, nine treatments were compared, combination of three concentrations of Murashige and Skoog medium (MS) (1, 1/2 and 1/4 strength macronutrients) and three concentration of indole 3-butyric acid (IBA, at 0, 1 and 2 mu M). In the ex vitro assay we established for IBA four concentrations (0; 4.9; 9.8 and 19.7 mM). After 30 days total survival, rooting percentage, root length and number of roots per explants were evaluated. Both clones responded differently to the treatments used. Under in vitro conditions, Clone 1 had an average of 97% rooting at 1/2 MS + 1 mu M IBA, whereas Clone 2 obtained a 93% of rooting with 1/2 MS + 2 mu M IBA. Our results indicate that the decrease in the strength of MS medium and the addition of IBA promotes rooting. With regard to ex vitro conditions, the Clone 1 did not respond to IBA applications, while in Clone 2, IBA promoted the rooting at the concentration of 19.7 mM, with percentages of 90 and 85%, respectively. Considering these results, it is generally recommended to use the ex vitro rooting system because both clones were able to obtain the same high percentage rooting (above 85%) in this system. Ex vitro propagation will also safe costs

Antitumor activity and carrier properties of novel hemocyanins coupled to a mimotope of GD2 ganglioside

Conjugation to carrier proteins is a way to improve the immunogenicity of peptides. Such is the case for peptides mimicking carbohydrate tumor-associated antigens in cancer vaccine development. The most used protein for this purpose is the keyhole limpet hemocyanin (KLH) from Megathura crenulata. Its limited bioavailability has prompted interest in finding new candidates; nevertheless, it is not known whether other hemocyanins might be equally efficient as carrier of carbohydrate peptide mimotopes to promotes anti-tumor responses. Here, we evaluated the carrier and antitumor activity of novel hemocyanins with documented immunogenicity obtained from Concholepas concholepas (CCH) and Fissurella latimarginata (FLH), coupled through sulfo-SMCC to P10, a mimetic peptide of GD2, the major ganglioside constituent of neuroectodermal tumors, and incorporating AddaVax as an adjuvant. The humoral immune responses of mice showed that CCH-P10 and FLH-P10 conjugates elicited specific IgM and IgG antibodies against P10 mimotope, similar to those obtained with KLH-P10, which was used as a positive control. The CCH-P10 and FLH-P10 antisera, exhibited cross-reactivity with murine and human melanoma cells, like anti-CCH and anti-FLH sera suggesting a cross-reaction of CCH and FLH glycosylations with carbohydrate epitopes on the tumor cell surfaces, similar to the KLH antisera. When mice were primed with each hemocyanin-P10 and challenged with melanoma cells, better antitumor effects were observed for FLH-P10 than for CCH-P10 and, as for KLH-P10, irrespective of conjugation. These data demonstrate that CCH and FLH are useful carriers of carbohydrate mimotopes; however, the best antitumor activity of FLH preparations, indicate that is a suitable candidate for further cancer vaccines research. (C) 2018 Elsevier Masson SAS. All rights reserved.FONDECYT 1110651 115133

A Novel Immunomodulatory Hemocyanin from the Limpet Fissurella latimarginata Promotes Potent Anti- Tumor Activity in Melanoma

Artículo de publicación ISIHemocyanins, the huge oxygen-transporting glycoproteins of some mollusks, are used as immunomodulatory proteins with proven anti-cancer properties. The biodiversity of hemocyanins has promoted interest in identifying new anti-cancer candidates with improved immunological properties. Hemocyanins promote Th1 responses without known side effects, which make them ideal for long-term sustained treatment of cancer. In this study, we evaluated a novel hemocyanin from the limpet/gastropod Fissurella latimarginata (FLH). This protein has the typical hollow, cylindrical structure of other known hemocyanins, such as the keyhole limpet hemocyanin (KLH) and the Concholepas hemocyanin (CCH). FLH, like the KLH isoforms, is composed of a single type of polypeptide with exposed N- and O-linked oligosaccharides. However, its immunogenicity was significantly greater than that of KLH and CCH, as FLH induced a stronger humoral immune response and had more potent anti-tumor activity, delaying tumor growth and increasing the survival of mice challenged with B16F10 melanoma cells, in prophylactic and therapeutic settings. Additionally, FLH-treated mice demonstrated increased IFN-c production and higher numbers of tumor-infiltrating CD4+ lymphocytes. Furthermore, in vitro assays demonstrated that FLH, but not CCH or KLH, stimulated the rapid production of pro-inflammatory cytokines (IL-6, IL-12, IL-23 and TNF-a) by dendritic cells, triggering a pro-inflammatory milieu that may explain its enhanced immunological activity. Moreover, this effect was abolished when deglycosylated FLH was used, suggesting that carbohydrates play a crucial role in the innate immune recognition of this protein. Altogether, our data demonstrate that FLH possesses increased anti-tumor activity in part because it activates a more potent innate immune response in comparison to other known hemocyanins. In conclusion, FLH is a potential new marine adjuvant for immunization and possible cancer immunotherapy