A novel proline-rich M-superfamily conotoxin that can simultaneously affect sodium, potassium and calcium currents

Background Conotoxins have become a research hotspot in the neuropharmacology field for their high activity and specificity in targeting ion channels and neurotransmitter receptors. There have been reports of a conotoxin acting on two ion channels, but rare reports of a conotoxin acting on three ion channels. Methods Vr3a, a proline-rich M-superfamily conotoxin from a worm-hunting Conus varius, was obtained by solid-phase synthesis and identified by mass spectrometry. The effects of synthesized Vr3a on sodium, potassium and calcium currents were tested on rat DRG cells by patch clamp experiments. The further effects of Vr3a on human Cav1.2 and Cav2.2 currents were tested on HEK293 cells. Results About 10 μM Vr3a has no effects on the peak sodium currents, but can induce a ~10 mV shift in a polarizing direction in the current-voltage relationship. In addition, 10 μM Vr3a can increase 19.61 ± 5.12% of the peak potassium currents and do not induce a shift in the current-voltage relationship. An amount of 10 μM Vr3a can inhibit 31.26% ± 4.53% of the peak calcium currents and do not induce a shift in the current-voltage relationship. The IC50 value of Vr3a on calcium channel currents in rat DRG neurons is 19.28 ± 4.32 μM. Moreover, 10 μM Vr3a can inhibit 15.32% ± 5.41% of the human Cav1.2 currents and 12.86% ± 4.93% of the human Cav2.2 currents. Conclusions Vr3a can simultaneously affect sodium, potassium and calcium currents. This novel triple-target conotoxin Vr3a expands understanding of conotoxin functions.(AU)

Mannitol-induced drought stress on calli of Trigonella foenum-graecum L. Var. RMt-303.

Different explants of fenugreek, T. foenum-graecum L. (Var. RMt-303), were compared for their callus induction and subsequent shoot regeneration capabilities on Murashige and Skoog media supplemented with different phytohormones in varying concentration. The highest percentage of callus induction frequency was observed in 1ppm benzylaminopurine (BAP). Maximum shoots were induced on media supplemented with 0.5ppm BAP using leaf and stem tissues as explants. However, root tissues showed only callusing with no subsequent shooting. Cotyledonary node responded better than hypocotyls in terms of shoot induction on media supplemented with thidiazuron (0.1ppm). The callus was subjected to drought stress as simulated by reduced water potential of growth media due to addition of mannitol. Calli could withstand -2 MPa water potential till 30 days indicating that the drought stress tolerance mechanisms are functional in this variety. Chlorophyll a and b and total chlorophyll, proline and total phenolic contents, total peroxidase and catalase activities increased under stress conditions suggesting the tolerance of callus to drought stress. However, ascorbate peroxidase, guaiacol peroxidase activities were found to decrease slightly. Malondialdehyde and H2O2 contents were found to decrease while only a slight disturbance was found in membrane stability index. These results underline the mechanisms that are crucial for drought stress tolerance in fenugreek.

Prolina livre em alecrim-pimenta sob estresse hídrico antes da colheita / Free proline in Lippia sidoides Cham. under water stress before harvest

O estresse hídrico é considerado um dos fatores limitantes para o desenvolvimento das plantas, com isso, os vegetais desenvolvem mecanismos para tolerar a deficiência hídrica do solo. Entre esses mecanismos, está o acúmulo de prolina livre nos tecidos, tendo como função a regulação osmótica dos tecidos. A influência do estresse hídrico em plantas medicinais ainda é pouco estudado, principalmente em plantas nativas, como é o caso do alecrim-pimenta Lippia sidoides Cham. Assim, o trabalho teve por objetivo avaliar a prolina livre e o teor de flavonoides totais na referida espécie sob condições de estresse hídrico antes da colheita. O trabalho foi realizado no Instituto de Ciências Agrárias da UFMG, Campus Montes Claros, delineamento em blocos casualizados, com 5 tratamentos e 4 repetições, sendo eles 8, 6, 4, 2 e 0 dias de supressão da irrigação antes da colheita. Os teores encontrados de prolina variaram entre 0,2489 a 3,2421 ìM.g-1 de matéria seca e de flavonoides entre 0,0139 a 0,008 g.g-1. Os resultados observados indicam que a planta diminui seu teor de flavonoides e aumenta o teor de prolina com o aumento do período de estresse.

Water stress is considered a limiting factor for plant growth, thus, the plants develop mechanisms to tolerate the soil water deficit. Among these mechanisms is the accumulation of free proline in tissues, with the poke the osmotic regulation of tissues. The influence of water stress in medicinal plants is still poorly studied, mainly on native plants such as rosemary-pepper Lippia sidoides Cham. Thus, the study aimed to evaluate the free proline and total flavonoid content in that species under conditions of water stress before harvest. The study was conducted at the Institute of Agricultural Sciences, UFMG, Campus Montes Claros, randomized block design with 5 treatments and 4 replicates, and they 8, 6, 4, 2 and 0 days of withholding irrigation before harvest. The content of proline ranged from 0.2489 to 3.2421 ìM.g-1 of dry matter and flavonoids from 0.0139 to 0.008 gg-1. The results indicate that the plant reduces its content of flavonoids and proline content increases with increasing stress time.

Salinity effects on protein content, lipid peroxidation, pigments, and proline in Paulownia imperialis (Siebold & Zuccarini) and Paulownia fortunei (Seemann & Hemsley) grown in vitro

We evaluated the effects of saline stress on soluble proteins, lipid peroxidation (TBAR), chlorophyll a, chlorophyll b, beta-carotene, violaxanthin, and proline in Paulownia imperialis and Paulownia fortunei plants grown in vitro. When the propagated plants reached a determined size, they were transferred aseptically to WPM culture medium containing different sodium chloride concentrations (0, 20, 40, 60, 80, and 160 mM) and were sampled at 15 and 30 days. Proline content was determined at 30 days after transfer only. Protein concentration significantly decreased with the highest salt levels in P. imperialis compared to controls in which no sodium chloride was added. In both P. imperialis and P. fortunei, lipid peroxidation significantly increased at 15 days but decreased at 30 days. Chlorophyll a, chlorophyll b, beta-carotene, and violaxanthin significantly decreased with exposure to higher sodium chloride concentrations at 15 and 30 days in both species. Proline content in P. imperialis significantly increased in plants grown in 20 and 40 mM of sodium chloride and decreased in higher sodium chloride concentrations. In P. fortunei, this measure significantly decreased proline content at all salt concentrations in plants exposed to all levels of sodium chloride compared to controls. Our results show that P. imperialis is more tolerant to salt stress at the salinity conditions tested.

Current concepts of the relationship between cholesterol fed rats and endogenous amino acids

The effect of long term feeding of cholesterol 100 mg/kg/day for 20 weeks on free amino acid patterns in plasma and liver contents in rats was investigated using LKB-amino acid analyzer [Biochrom Ltd., Cambridge, England] and correlated these results with their effects on plasma levels of cholesterol, triglyceride, lipoprotein and Gamma Glutamyl Transferase [yGT]. indicate that all amino acid contents in the liver were significantly decreased in the cholesterol feed animal group. The amplitude of reductions varied between 40-100%. Glutamine and tryptophan were not detected in the liver of cholesterol feed group. The plasma concentrations of taurine, glutamate, alanine, valine and phenylalanine were elevated whose mean percentage increases were 48 +/- 4, 40 +/- 3, 25 +/- 3 and 35 +/- 4 respectively. These increases were associated with significant decrease in the concentration of ornithine [37%]. Meanwhile proline was not detected in the plasma of treated animals. Also, plasma cholesterol, triglyceride, lipoproteins and yGT were determined by colorimetric methods using Kits from Boehringer Mannheim [GmbH]. Results indicated that feeding cholesterol significantly increased the plasma yGT activity. In these experimental conditions the chronic intake of cholesterol had no significant effects on plasma cholesterol or other plasma lipids parameters tested except plasma triglyceride which was significantly increased, these results indicated that there are interactions between cholesterol intake and hepatic glutamine and tryptophan as well as plasma proline and these interaction mechanisms may be considered the factor generating metabolic events, which play physiological functions in the regulation of plasma cholesterol. Thus, under pathological conditions there is an imbalance between these interaction mechanisms which cause an increase in the circulating levels of cholesterol, leading to pathological processes such as hyperlipidemias, atherosclerosis and bile stones

Individual and combined effect of mercury and manganese on phenol and proline content in leaf and stem of mungbean seedlings.

Mungbean (Vigna radiata L. Wilczelk cv. Pusa Baisakhi seedlings were raised in individual (0, 1, 10, 100 and 1000 ppm) and combined solutions (1 : 1, 10: 1, 1: 0 ppm Hg : Mn) of mercury and manganese for 6 days. Phenol and proline were found to accumulate in leaves in response to treatment with heavy metals. The magnitude of accumulation correlated with concentration of metals. However, a reverse trend was noticed in stem for phenol. Accumulation of phenol in response to heavy metal treatment was organ specific and occurred at higher rate in plant parts, which faced the stress mostly. However, accumulation of proline helped the plant to survive stress situation. In combined solutions, amelioration of mercurial toxicity by manganese was recorded.

Padronização e variações do teste da atividade da prolina aminopeptidase: novo método de diagnóstico das vaginoses bacterianas / Padronization and variations of the proline aminopeptidase activity tests: a new method of a bacterial vaginosis diagnosis

A vaginose bacteriana (VB) pode ser diagnosticada clinicamente e através de métodos laboratoriais que utilizem a microscopia óptica convencional (exame a fresco, método de Gram e Papanicolaou), cultura e análises cromatográficas. O teste da atividade da prolina aminopeptidase (TAPA) é uma técnica que envolve uma reação enzimática e foi testada com o objetivo de esclarecer os casos de diagnósticos duvidosos ou indeterminados para substituir a cultura e as análises cromatográficas que são técnicas morosas e dispendiosas. O TAPA foi comparado com a cultura semi-quantitativa para Gardnerella vaginalis demonstrando valor “p” significativo, sensibilidade igual a 95%, especificidade igual a 78% e valor preditivo positivo igual a 96%.

La deficiencia de vitamina C durante la gestación como un factor de riesgo para la ruptura prematura de las membranas fetales / Vitamin C deficiency during pregnancy as a risk factor for premature fetal membrane rupture

Se analizan las características de las membranas amnióticas de partos con ruptura prematura de membranas (RPM) y con ruptura tardía de las membranas fetales (RTM), en cuanto a su composición aminoacídica y relación prolina/hidroxiprolina y la importancia de la situación dietética y nutricional de las gestantes, con respecto a la vitamina C en relación con este accidente obstétrico, así como demostrar, mediante el uso de un modelo experimental, que la membrna fetal es también un sitio sensible de la deficiencia de vitamina C. A las mujeres estudiadas se les determinó acido ascórbico en plasma en el segundo y tercer trimestre de gestación, cortisol en plasma en las mismas etapas, acido ascórbico en leucocitos al final de la gestación, en las membranas amnióticas colectadas durante el trabajo de parto. Se determinó prolina, hidroxiprolina y aminoacidos totales individuales y se aisló el colágeno en el cual se realizaron las mismas determinaciones que en la membrana. Se realizó una encuesta dietétic. El modelo experimental utilizado fue la curiela gestante, los animales se dividieron en 3 grupos, uno de los cuales ingirió una dieta escorbutigénica durante la última etapa de la gestación, al fianl de la misma los animales fueron sacrificados y en las membranas fetales se realizaron las mismas determinaciones descritas para humanos. Se encontró una mayor incidencia de RPM en mujeres con bajos niveles de vitamina C