Functional morphology of pituitary -thyroid and -adrenocortical axes in middle-aged male rats treated with Vitex agnus castus essential oil.

We previously reported that Vitex agnus-castus L. essential oil (VACEO), when administered to middle-aged males, exerts a bone-protective effect, induces silencing of locomotor activities and decreases pituitary prolactin immunopositivity. To further assess the putative endocrine effects of VACEO, we examined the pituitary-thyroid and -adrenocortical axes in our model. Sixteen-month-old Wistar rats were subcutaneously administered 60mg/kg of VACEO dissolved in sterile olive oil, while the control group received the same amount of vehicle alone for three weeks. Pituitaries, thyroids and adrenals were analyzed by qualitative and quantitative histological approaches. Concentration of thyroid stimulating hormone (TSH), total thyroxine and triiodothyronine (TH), adrenocorticotrophic hormone (ACTH), corticosterone in serum and in adrenal tissue were measured. In VACEO-treated rats, the relative volume density of pituitary thyrotrophs increased (p<0.001), while intensity of cytoplasmic TSHß immunostaining decreased (p<0.001), consistent with elevated TSH in serum (p<0.01). The thyroid tissue was characterized by a micro-follicular structure, increased relative volume of follicular epithelium (p<0.05), decreased volume of luminal colloid (p<0.001) and increased basolateral expression of sodium-iodide symporter-immunopositivity (p<0.05). Serum TH also increased (p<0.01). The relative volume density of pituitary corticotrophs decreased (p<0.05), compatible with decline in circulating ACTH (p<0.05). Neither tissue nor serum corticosterone levels were affected by VACEO treatment. In conclusion, the observed changes in TSH and ACTH strongly indicate central endocrine effects of prolonged VACEO treatment. In this respect, production of ACTH decreased without impact on corticosterone production. Increase in serum concentration of both TH and TSH are not compatible with a negative feedback loop and suggest a major change in set-point regulation of the hypothalamic-pituitary-thyroid axis.

Plant regeneration from callus cultures of Vitex trifolia (Lamiales: Lamiaceae): a potential medicinal plant.

Vitex trifolia is a shrub species with popular use as a medicinal plant, for which leaves, roots and flowers have been reported to heal different distresses. The increasing exploitation of these plants has endangered its conservation, and has importantly justified the use of biotechnological tools for their propagation. Our aim was to present an efficient protocol for plant regeneration through organogenesis; and simultaneously, to analyze the genetic homogeneity of the established clonal lines by Randomly Amplified Polymorphic DNA (RAPD) and Inter Simple Sequence Repeat (ISSR) markers. Plantlet regeneration was achieved in callus cultures derived from stem, leaf and petiole explants of V. trifolia on a differently supplemented Murashige & Skoog medium, and incubated at 25 +/-2 degrees C under a light intensity of 61 micromol/m2s from cool white fluorescent lamps and a 16 h photoperiod. The rate of shoot bud regeneration was positively correlated with the concentration of hormones in the nutrient media. Shoot buds regenerated more rapidly from stem and petiole explants as compared to leaf explants on medium containing 11.10 microM BAP in combination with 0.54 microMNAA. Addition of 135.74-271.50 microM adenine sulphate (Ads) and 0.72-1.44 microM gibberellic acid (GA3) to the culture medium increased the growth of shoot buds. The highest rate of shoot bud regeneration responses was obtained in stem explants using 11.10 microM BAP in combination with 0.54 microM NAA, 271.50 microM Ads and 1.44 microM GA3. In vitro rooting of the differentiated shoots was achieved in media containing 1.23 microM indole butyric acid (IBA) with 2% (w/v) sucrose. Regenerated plantlets were successfully established in soil with 86% survival under field condition. Randomly Amplified Polymorphic DNA and Inter Simple Sequence Repeat markers analyses have confirmed the genetic uniformity of the regenerated plantlets derived from the second up to fifth subcultures. This protocol may help in mass propagation and conservation of this important medicinal plant of great therapeutic potential.

Plant regeneration from callus cultures of Vitex trifolia (Lamiales: Lamiaceae): a potential medicinal plant / Regeneración de plantas a partir de cultivos de callos de Vitex trifolia (Lamiales: Lamiaceae): una planta medicinal potencial

Vitex trifolia is a shrub species with popular use as a medicinal plant, for which leaves, roots and flowers have been reported to heal different distresses. The increasing exploitation of these plants has endangered its conservation, and has importantly justified the use of biotechnological tools for their propagation. Our aim was to present an efficient protocol for plant regeneration through organogenesis; and simultaneously, to analyze the genetic homogeneity of the established clonal lines by Randomly Amplified Polymorphic DNA (RAPD) and Inter Simple Sequence Repeat (ISSR) markers. Plantlet regeneration was achieved in callus cultures derived from stem, leaf and petiole explants of V. trifolia on a differently supple mented Murashige & Skoog medium, and incubated at 25±2ºC under a light intensity of 61µmol/m2s from cool white fluorescent lamps and a 16h photoperiod. The rate of shoot bud regeneration was positively correlated with the concentration of hormones in the nutrient media. Shoot buds regenerated more rapidly from stem and petiole explants as compared to leaf explants on medium containing 11.10µM BAP in combination with 0.54µMNAA. Addition of 135.74-271.50µM adenine sulphate (Ads) and 0.72-1.44µM gibberellic acid (GA3) to the culture medium increased the growth of shoot buds. The highest rate of shoot bud regeneration responses was obtained in stem explants using 11.10µM BAP in combination with 0.54µM NAA, 271.50µM Ads and 1.44µM GA3. In vitro rooting of the differentiated shoots was achieved in media containing 1.23µM indole butyric acid (IBA) with 2% (w/v) sucrose. Regenerated plantlets were successfully established in soil with 86% survival under field condition. Randomly Amplified Polymorphic DNA and Inter Simple Sequence Repeat markers analyses have confirmed the genetic uniformity of the regenerated plantlets derived from the second up to fifth subcultures. This protocol may help in mass propagation and conservation of this important medicinal plant of great therapeutic potential.

Vitex trifolia es una especie arbustiva de uso popular como planta medicinal, sus hojas, raíces y flores se han reportado para la cura de diferentes aflicciones. El aumento de la explotación de estas plantas ha puesto en peligro su conservación y ha justificado el uso de herramientas biotecnológicas para su propagación. El objetivo de esta investigación fue presentar un protocolo eficiente para la regeneración de estas plantas a través de la organogénesis, y analizar la homogeneidad genética de las líneas clonales establecidas por ADN polimórfico amplificado aleatoriamente (RAPD) mediante la repetición de marcadores de inter secuencia simple (ISSR). La regeneración de plántulas se logró en cultivos de callos derivados de explantes de tallo, hoja y pecíolo de V. trifolia en un medio diferenciado Murashige & Skoog, que se incubaron a 25±2ºC bajo una intensidad de luz de 61μmol/m2s con lámparas fluorescentes blancas y un fotoperíodo de 16h. La tasa de regeneración de brotes se correlacionó positivamente con la concentración de las hormonas en el medio nutritivo. Los brotes se regeneraron más rápidamente a partir de explantes de tallo y pecíolos en comparación con explantes de hoja. La mayor tasa de regeneración de brotes se obtuvo en los explantes de tallo utilizando 11.10μM BAP en combinación con 0.54μM NAA, 271.50μM Ads y 1.44μM GA3. Este protocolo puede ayudar a la propagación masiva y conservación de esta importante planta medicinal de gran potencial terapéutico.

Role of TDZ in the quick regeneration of multiple shoots from nodal explant of Vitex trifolia L.--an important medicinal plant.

The effect of thidiazuron (TDZ) has been investigated in shoot multiplication for a simple, efficient, rapid, and commercially applicable regeneration protocol of an important medicinal plant, Vitex trifolia. Multiple shoots were induced in nodal explants obtained from a mature tree on Murashige and Skoog (MS) medium supplemented with TDZ in various concentrations (0.5, 1.0, 2.5, 5.0, 7.5, or 10.0 µM). Prolonged exposure of the culture to TDZ had an adverse affect. To avoid this, the cultures were transferred to TDZ-free MS medium or MS medium fortified with various concentrations of 6-benzyladenine (BA) alone or in combination with α-naphthalene acetic acid (NAA) to enhance multiplication, proliferation, and elongation of induced shoots. Optimum shoot multiplication and elongation was achieved when TDZ-exposed explants were repeatedly subcultured on MS media containing a combination of 1.0 µM BA and 0.5 µM NAA. The highest shoot regeneration frequency (90 %) and maximum number (22.3 ± 0.2) of shoots per explant with shoot length of (5.2 ± 0.2 cm) was recorded on MS medium fortified with 5.0 µM TDZ. In vitro rooting of isolated shoots was achieved best in half-strength MS medium containing 0.5 µM NAA. Properly rooted plantlets were successfully hardened off and acclimatized in thermocol cups containing sterile Soilrite. These plantlets were then transferred to pots containing different potting substrate; percentage survival of the plantlets was highest in vermiculite/garden soil mixture (1:1) and successfully transfer to greenhouse under sunlight.