Producción de compuestos hipoglucemiantes en cultivos de células en suspensión de Cecropia obtusifolia (Bertol) "guarumbo"

Cecropia obtusifolia, popularmente conocida como “guarumbo”, es un árbol medicinal que se utiliza en México para el control de la diabetes mellitus tipo 2 (DM2). La investigación de esta especie, tanto a nivel experimental en animales de laboratorio como en el humano, ha demostrado las propiedades antidiabéticas que posee el vegetal, las cuales son atribuidas a los compuestos fenólicos ácido clorogénico (AC) e isoorientina (ISO). El presente estudio tuvo como objetivo establecer las condiciones in vitro para la propagación masiva de las plantas de C. obtusifolia y para el desarrollo de los cultivos de células en suspensión que conserven la capacidad de producción de los compuestos bioactivos de la planta AC e ISO. Para lo anterior, a partir de explantes de hoja de plantas aclimatadas se establecieron cultivos de callos en medio Murashige y Skoog (MS) empleando diferentes niveles de auxinas; la mejor respuesta se obtuvo utilizando 8.92 μM de ácido naftalenacético (ANA) y ácido 2,4-diclorofenoxiacético (2,4-D) en combinación con 2.22 μM de bencilaminopurina (BAP). En una etapa posterior, los explantes de hoja, tallo, hipocótilo y cotiledón derivados de plántulas in vitro, desarrollaron callos rizogénicos con 2,4-D; en tanto que los explantes de hoja, hipocótilo y cotiledón formaron callos rizogénicos en el medio de cultivo con ANA y los de tallo formaron callos con brotes. Los brotes se enraizaron en medio MS al 50% libre de hormonas y las plantas formadas se aclimatizaron exitosamente en las condiciones de invernadero; no se observaron variaciones morfológicas entre la población durante su desarrollo y las tasas de crecimiento fueron superiores a las reportadas para las plantas silvestres. En los callos se comprobó por HPLC la presencia de AC en concentraciones similares a las detectadas en las hojas de los árboles en campo, así como su precursor ácido caféico. El otro compuesto fenólico (ISO) no fue producido. Por otra parte, los cultivos de células en suspensión se establecieron a partir de callos friables derivados de explantes de hoja con el tratamiento hormonal de 2,4-D manejando inóculos de 4 y 5 % en peso fresco. Las suspensiones celulares se desarrollaron mejor con el inóculo del 5% y en estos cultivos se determinaron niveles de AC similares a los detectados en las hojas de los árboles silvestres. La ISO se detectó únicamente al final del crecimiento logarítmico. Al disminuir el contenido de los nitratos en el medio MS a 8.0 mM, las biomasas celulares máximas disminuyeron y la concentración de AC detectada se cuadruplicó con respecto a la del cultivo en suspensión con 27.4 mM de nitratos. Con respecto a la producción de ISO, la síntesis de este compuesto se indujo más temprano y por más tiempo, incrementándose sus niveles al final del cultivo. Las hojas del árbol silvestre, plantas aclimatada y obtenida por micropropagación con 6 meses en aclimatización, acumularon ambos compuestos en concentraciones similares entre ellas; sin embargo, las plantas derivadas por micropropagación presentaron diferencias significativas en la acumulación del AC a lo largo de su desarrollo. La aglícona de ISO, luteolina, también fue detectada. Tomando en consideración que las plantas micropropagadas conservaron los componentes y las propiedades hipoglucemiantes de la planta silvestre, se concluye que la propagación in vitro representa una alternativa y perspectiva viable para la propagación masiva y uniforme de los árboles de C. obtusifolia. Otra contribución importante, es el manejo del cultivo de células en suspensión de C. obtusifolia con restricción de nitratos como fuente de material vegetal para la obtención de extractos activos enriquecidos en AC e ISO.The Cecropia obtusifolia tree, popularly known as guarumbo in Mexico, is used in folk medicine for treatment of type 2 diabetes mellitus (DM2). C. obtusifolia’s antidiabtic effect was assessed in animal models, as well as in humans and attributed to chlorogenic acid (CA) and isoorientine (ISO). The aim of this study was to establish in vitro conditions for the micropropagation and development of cell suspension cultures that conserve the capacity to produce the CA and ISO bioactive compounds that exist in the wild plant. For that, callus cultures of leaf explants from aclimated plant were set up using Murashige and Skoog medium (MS) with different auxins levels; treatments with naphtalene acetic acid (NAA) and 2,4-dichorophenoxi acetic acid (2,4-D) to 8.92 μM with benzylaminopurine (BAP) to 2.22 µM stimulate highest callus production. Cotyledon, hypocotyl, leaf, and stem explants developed calluses bearing roots in a medium enriched with 2,4-D; when hypocotyl, cotyledon and leaf explants were grown on medium with NAA, they developed morphogenic calluses with the presence of abundant roots, while 75% of stem explants formed simple calluses and the remaining explants developed calluses with shoots. Shoots were rooted on free growth regulator MS medium, and whole regenerated plants were aclimatizated under greenhouse conditions; morphological changes were not observed among the population during their development, and growth rates were superior to those reported for wild plants. Recorded concentrations of CA in the different cell lines were similar to those detected in the leaves of wild plants; its precursor caffeic acid was also detected. The other phenolic compound (ISO) was not produced. On the other hand, cell cultures in suspension were established from friable calluses derived from leaf explants with 2,4-D hormonal treatment and employing 4 and 5 % inocula in fresh weight. Better development of cell suspensions was achieved with the 5% inoculum, and the determined CA levels in these cultures were similar to those detected in leaves of wild plants. ISO was detected only at the end of logarithmic growth. On diminishing nitrate content in the MS medium to 8.0 mM, maximum cell biomasses diminished and the CA concentration detected quadrupled with respect to that of the culture in suspension with 27.4 mM of nitrates. Regarding ISO production, synthesis of this compound was induced earlier and for a longer time period, increasing its levels at the end of culture. Leaves from wild tree, aclimated and micropropagated plants with 6 month of acclimatization produced both compounds at similar concentrations; however, micropropagation-derived plants presented significant differences in CA accumulation throughout their development. The aglycone of ISO, luteoline, was also detected. Considering that micropropagated plants retained their ability to produce bioactive compounds and hypoglycemic activity of the wild plant, we concluded that in vitro cultures represent an alternative and interesting perspective for C. obtusifolia tree multiplication. Another important contribution is the handling of cells suspension culture from C. obtusifolia with nitrate restriction like source of vegetal material for the obtaining of enriched active extracts in CA and ISO

Estudios etnofarmacológicos de Cecropia obtusifolia (Urticaceae) y su importancia en el tratamiento de la diabetes mellitus tipo 2 (DM-2): una mini-revisión

Background and Aims: Diabetes mellitus type 2 (DM-2) is one of the most recurrent chronic diseases worldwide, it is usually treated with synthetic medications, many of which have important repercussions on the patient's body. For this reason, ethnopharmacology has become more important in recent years, because a large number of plant resources origin are used in medicine and are shown as positive effect in the treatment of different diseases, including DM-2. Therefore, the aim of this review is to highlight how previous studies are adding to the understanding and knowledge of the biological effects reported in Cecropia obtusifolia, a tropical plant included in the Herbal Pharmacopoeia of the United Mexican States (FHEUM), which is commonly used to treat DM-2.Methods: A profound literature review was carried out on C. obtusifolia, focusing on diverse phytochemical, pharmacological, clinical and toxicological studies, as well as on some other relevant research findings.Key results: It is recognized that this species is able to decrease serum glucose, cholesterol and triglyceride levels due to its hypoglycemic and hypolipidemic properties, which has been demonstrated both in animal and human experimental models, attributing these effects to chlorogenic acid and isoorientin. These previous reports suggest that C. obtusifolia is a promising candidate for the development of a phytopharmaceutical which could be used in the treatment of DM-2.Conclusions: We provide an updated and complete overview of the phytochemistry, traditional uses, and pharmacological activities of C. obtusifolia. Regarding its pharmacological activities, we focus mainly on the hypoglycemic effect of this plant which supports its traditional use in DM-2 control. Moreover, the present knowledge was critically assessed to provide some evidence and justifications and propose future research prospects.Antecedentes y Objetivos: La diabetes mellitus tipo 2 (DM-2) es una de las enfermedades crónicas recurrentes más comunes, generalmente se trata con medicamentos sintéticos, muchos de los cuales tienen importantes repercusiones en el cuerpo del paciente. Por esta razón, la etnofarmacología se ha vuelto más importante en los últimos años, porque una gran cantidad de recursos de origen vegetal son utilizados en medicina y muestran un efecto positivo en el tratamiento de diferentes enfermedades, incluyendo DM-2. Por lo tanto, el objetivo de esta revisión es destacar cómo los estudios previos se suman a la comprensión y el conocimiento de los efectos biológicos informados en Cecropia obtusifolia, una planta tropical incluida en la Farmacopea Herbolaria de los Estados Unidos Mexicanos (FHEUM), comúnmente usada para tratar DM-2.Métodos: Se realizó una revisión profunda de la literatura sobre C. obtusifolia, centrándose en diversos estudios fitoquímicos, farmacológicos, clínicos y toxicológicos, así como en otros hallazgos relevantes de investigación.Resultados clave: Se reconoce que esta especie es capaz de disminuir los niveles séricos de glucosa, colesterol y triglicéridos, debido a sus propiedades hipoglucémicas e hipolipidémicas reductoras de lípidos, que se han demostrado tanto en modelos experimentales animales como en humanos, atribuyendo estos efectos al ácido clorogénico y la isoorienina. Estos informes previos sugieren que C. obtusifolia es un candidato prometedor para el desarrollo de un fitofarmacéutico que podría usarse en el tratamiento de DM-2.Conclusiones: Brindamos una visión general actualizada y completa de la fitoquímica, los usos tradicionales y las actividades farmacológicas de C. obtusifolia. En cuanto a sus actividades farmacológicas, nos centramos principalmente en el efecto hipoglucémico de esta planta que apoya su uso tradicional en el control de DM-2. Además, el conocimiento actual se evaluó críticamente para proporcionar algunas pruebas y justificaciones y proponer futuras perspectivas de investigación

Microbiological and Pharmacological Evaluation of the Micropropagated Rubus liebmannii Medicinal Plant

Rubus liebmannii is an endemic species from Mexico used in traditional medicine primarily to treat dysentery and cough. The in vitro activity against Giardia lamblia and Entamoeba histolytica that produces the ethanolic extract of the aerial parts of the plant led us to expand the pharmacological and phytochemical research of this species. Gastrointestinal disorders including amebiasis remain one of the health problems that need to be addressed and it is of interest to find alternatives that improve their treatment. Also, it is important to emphasize that R. liebmannii grows wild in the country and is not found in abundance; therefore, alternatives that avoid overexploitation of the natural resource are mandatory. Ongoing with the evaluation of the potentialities that R. liebmannii possesses for treating infectious gastrointestinal diseases, the aim of the present study was to evaluate the biological effects and the chemical composition of the micropropagated plant

Geographical and temporal distribution of SARS-CoV-2 clades in the WHO European Region, January to June 2020

We show the distribution of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) genetic clades over time and between countries and outline potential genomic surveillance objectives. We applied three genomic nomenclature systems to all sequence data from the World Health Organization European Region available until 10 July 2020. We highlight the importance of real-time sequencing and data dissemination in a pandemic situation, compare the nomenclatures and lay a foundation for future European genomic surveillance of SARS-CoV-2

The Journey of a Medicinal Plant throughout Science: <i>Sphaeralcea angustifolia</i> (Cav.) G. Don (Malvaceae)

Using herbal medicine is an ancestral cultural practice among Mexicans. A broad sector of society turns to plants to treat priority health problems, a reality that leads scientists to explore the healing value attributed to them. Advances in the experimental research of Sphaeralcea angustifolia confirmed the anti-inflammatory activity of the species; therefore, an analysis of the scope of these studies is now warranted. As such, this paper is a compendium of the advances published in the scientific literature (from 2004 to 2021) on the anti-inflammatory properties of this plant. The promise offered by the species as a potential therapeutic agent is also considered, without dismissing aspects necessary for the preservation of this resource and its cultural and physical environment. The chemical–pharmacological aspects of the wild plant and its in vitro culture are highlighted. The plant’s anti-inflammatory properties support its clinical application as an anti-inflammatory phytopharmaceutical to treat arthritic conditions. The sustained therapeutic potential of S. angustifolia is reinforced by the biotechnological processes designed to conserve the resource, thus contributing to the protection of biodiversity and cultural diversity, aspects distinctive of a megadiverse country such as Mexico

Pharmacokinetic Study of Anti-osteoarthritic Compounds of a Standardized Fraction from Sphaeralcea Angustifolia

Sphaeralcea angustifolia has been widely used in inflammatory conditions such as blows, bruises, fractures, and wounds. The compounds identified as active in plants and suspension cell culture of S. angustifolia were tomentin, scopoletin, and sphaeralcic acid. To consolidate the integral use of knowledge about the S. angunstifolia and strengthen its pharmacological use in patients with knee osteoarthritis, the pharmacokinetic behavior of the active compounds was characterized. The SaTSS (S. angustifoloia standardized in Tomentin, Scopoletin, and Sphaeralcic acid) anti-ostearthritic fraction was obtained from cell suspension. The analytical method of High-Performance Liquid Chromatography (HPLC) for tomentin, scopoletin, and sphaeralcic acid were validated determining the accuracy, precision linearity, sensibility, specificity, detection limits, and quantification time-range parameters, as well as extraction efficiency and stability of compounds. The pharmacokinetic assay was performed with ICR mice strain, in which the mice were administrated with a single oral or intravenous dose (400 mg/kg with 7.1 mg/kg of scopoletin and tomentin in mixture and 34.6 mg/kg of sphaeralcic acid) of the SaTSS standardized active fraction. The results of the validated analytical methods allowed establishing, in a validated manner, that a coumarin mixture and sphaeralcic acid present in the SaTES fraction were detected in plasma. According to the values of Akaike Information Criteria (AIC), Sum of Squares (SS), Schwarz Criteria (SC), and by the determination coefficient (R2), the compounds follow a two-compartment model

Pharmacokinetics and Tissue Distribution of Coumarins from Tagetes lucida in an LPS-Induced Neuroinflammation Model

Tagetes lucida has been widely used as a folk remedy in illnesses associated with the central nervous system and inflammatory ailments. Among the chemical compounds that stand out in the plant against these conditions are coumarins, such as 7-O-prenylscopoletin (PE), scoparone (SC), dimethylfraxetin (DF), herniarin (HR), and 7-O-prenylumbelliferone (PU), considered potential anti-neuroinflammatory compounds. Therefore, the relationship between the therapeutic effect and the dose can be evaluated through pharmacokinetic&ndash;pharmacodynamic (PK&ndash;PD) studies under a model of neuroinflammation induced by lipopolysaccharide (LPS). Nonetheless, accomplishing those studies requires an accurate and robust analytical method for the detection of these compounds in different biological matrices of interest. Due to the above, in the present study, a bioanalytical method was established by HPLC&ndash;DAD-UV for the simultaneous quantification of the coumarins present in the hexane extract of T. lucida, which was able to determine the temporal concentration profiles of each of the coumarins in the plasma, brain, kidney, and spleen samples of healthy and damaged mice. Coumarins showed an increase in plasma concentrations of up to three times in the neuroinflammation model, compared to healthy mice, so it was possible to quantify the therapeutic agents in the main target organ, the brain. The ability of compounds to cross the blood&ndash;brain barrier is an advantage in the treatment of diseases associated with neuroinflammation processes that can be studied in future PK&ndash;PD evaluations

Integrated Analysis of the Transcriptome and Metabolome of Cecropia obtusifolia: A Plant with High Chlorogenic Acid Content Traditionally Used to Treat Diabetes Mellitus

This investigation cultured Cecropia obtusifolia cells in suspension to evaluate the effect of nitrate deficiency on the growth and production of chlorogenic acid (CGA), a secondary metabolite with hypoglycemic and hypolipidemic activity that acts directly on type 2 diabetes mellitus. Using cell cultures in suspension, a kinetics time course was established with six time points and four total nitrate concentrations. The metabolites of interest were quantified by high-performance liquid chromatography (HPLC), and the metabolome was analyzed using directed and nondirected approaches. Finally, using RNA-seq methodology, the first transcript collection for C. obtusifolia was generated. HPLC analysis detected CGA at all sampling points, while metabolomic analysis confirmed the identity of CGA and of precursors involved in its biosynthesis. Transcriptome analysis identified differentially expressed genes and enzymes involved in the biosynthetic pathway of CGA. C. obtusifolia probably expresses a key enzyme with bifunctional activity, the hydroxycinnamoyl-CoA quinate hydroxycinnamoyl transferase and hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyl transferase (HQT/HCT), which recognizes shikimic acid or quinic acid as a substrate and incorporates either into one of the two routes responsible for CGA biosynthesis