Herb-drug interaction: Effect of aqueous extract of Bridelia ferruginea leaves on the pharmacokinetics of metformin

The concurrent use of herbal medicines and orthodox drugs, especially in the treatment and management of chronic ailments, may result in clinically significant herb-drug interactions. Bridelia ferruginea Benth (Euphorbiaceae) is a common medicinal plant with known anti-diabetic properties and has been reported to be taken alongside the orthodox medicine, metformin. The aim of this study was to investigate the effect of aqueous extract of B. ferruginea leaves on the pharmacokinetics of metformin in female Sprague-Dawley rats. Reconstituted freeze dried extract of B. ferruginea leaves (30 mg/kg), and metformin (7 mg/kg) were administered concurrently as a single dose to female Sprague-Dawley rats. Whole blood samples (1 ml) were aseptically withdrawn by tail bleeding at 1, 2, 4, 8, and 24 h after administration of the single dose for pharmacokinetics analyses. Concurrent administration of metformin and B. ferruginea significantly affected (P < 0.05) all the pharmacokinetics parameters of metformin except for the time to attain the maximum concentration (Tmax), which increased but insignificantly. Whereas the area under the curve, maximum whole blood concentration (Cmax) and half-life (T½) of metformin decreased significantly in the presence of B. ferruginea, the elimination rate constant (Kel), clearance (Cl), absorption rate constant (Ka), and volume of distribution (Vd) of metformin increased significantly in the presence of B. ferruginea. Therefore, in clinical practice, patients should be advised on the implication of concurrent administration of metformin and B. ferrruginea

ANTIHYPERGLYCAEMIC AND ANTIOXIDANT EFFECTS OF ADENIA LOBATA ENGL. (PASSIFLORACEAE) IN STREPTOZOTOCIN-INDUCED DIABETIC RATS

The antihyperglycaemic and antioxidant activities of a Ghanaian medicinal plant namely Adenia lobata Engl (Passifloraceae), used to treat diabetes mellitus in traditional medicine, was investigated. The dried stem powder of A. lobata was successively extracted by Soxhlet with petroleum ether and 70% ethanol to obtain the crude petroleum ether (PEAL: yield =1.1w/w %) and ethanol (EEAL: yield = 5.4 w/w %) extracts. The extracts were assessed for their antihyperglycaemic and antioxidant activities. The antihyperglycaemic activity of PEAL and EEAL were determined in streptozotocin-induced diabetic rats (70 mg/kg body weight). Five groups of diabetic rats were given 150, 300 and 600 mg/kg body weight of PEAL and EEAL orally once daily for 20 days. Glibenclamide (5 mg/kg body weight) was used as positive control while distilled water (5 ml) acted as the normal diabetic control. The blood glucose levels were monitored initially for 6 hours and subsequently over 24 days. Both extracts exhibited statistically significant (p< 0.001) antihyperglycaemic activity throughout the study period, with EEAL showing the greatest activity. The antioxidant properties of the petroleum ether and ethanol extracts of A. lobata (PEAL and EEAL) were evaluated using five assays; total phenolic content, total antioxidant capacity, reducing power, DPPH scavenging effect and lipid peroxidation activity. In all these assays, the antioxidant properties increased with increasing concentration of the extracts

EPIGENETIC AND POST-TRANSCRIPTIONAL ELIMINATION OF CELIAC-CAUSING WHEAT STORAGE PROTEINS

Celiac disease is an autoimmune condition caused by consumption of certain wheat, barley and rye grain storage proteins known as prolamins. In susceptible individuals, exposure to these proteins leads to a painful chronic erasure of the microvilli of the intestinal epithelium. At present, the only effective solution is a strict dietary abstinence from these prolamins. The aim of the current project was to develop `celiac-safe' wheat genotypes. With this goal in mind, first, a microspore transformation procedure for production of doubled haploid wheat transformants was optimized. The next step was to develop artificial microRNA (amiRNA) constructs for post-transcriptional silencing of the wheat DEMETER homoeologues identified as transcriptional activators of the immunogenic prolamins in the developing wheat endosperm. An alternative approach employing a `chimeric-hairpin' construct capable of producing multiple interfering RNAs corresponding with the transcripts of the prolamin super-family genes was also undertaken. The latter strategy required aligning the sequences of the wheat gliadins (/-, - and -types) and low molecular weight glutenins (LMWgs) available in the public domain. The alignment results allowed identification of five consensus sequences each representing a prolamin gene family, which were fused together to obtain the arms of the chimeric-hairpin. The amiRNA and chimeric-hairpin constructs were used in wheat transformation via electroporation of embryogenic microspores, and particle bombardment of scutellar calli in the varieties Louise, Hollis, WPB926, Farnum, Brundage 96 and Simon. Microspore transformation using the chimeric hairpin construct yielded 47 T2 transformants in Louise, Hollis, WPB926 and Farnum with sizable amounts of reductions in the content of immunogenic prolamins. Similarly, the 12 T2 lines characterized in the background of Brundage 96 obtained via particle bombardment of amiRNA constructs have shown reductions and specific-eliminations to variable degrees in the grain prolamin content. A compensatory increase in the amount of high molecular weight glutenins (HMWgs) was also observed in some cases, which is desirable for retaining end-use quality of the genotypes. These wheat transformants are intended for crossing with the aim of pyramiding their effects on prolamin content or composition, into a single wheat genotype. This will advance the goal of producing celiac-safe wheat cultivars

Considerations of AOX Functionality Revealed by Critical Motifs and Unique Domains

An understanding of the genes and mechanisms regulating environmental stress in crops is critical for boosting agricultural yield and safeguarding food security. Under adverse conditions, response pathways are activated for tolerance or resistance. In multiple species, the alternative oxidase (AOX) genes encode proteins which help in this process. Recently, this gene family has been extensively investigated in the vital crop plants, wheat, barley and rice. Cumulatively, these three species and/or their wild ancestors contain the genes for AOX1a, AOX1c, AOX1e, and AOX1d, and common patterns in the protein isoforms have been documented. Here, we add more information on these trends by emphasizing motifs that could affect expression, and by utilizing the most recent discoveries from the AOX isoform in Trypanosoma brucei to highlight clade-dependent biases. The new perspectives may have implications on how the AOX gene family has evolved and functions in monocots. The common or divergent amino acid substitutions between these grasses and the parasite are noted, and the potential effects of these changes are discussed. There is the hope that the insights gained will inform the way future AOX research is performed in monocots, in order to optimize crop production for food, feed, and fuel

Genome-wide identification and analysis of the ALTERNATIVE OXIDASE gene family in diploid and hexaploid wheat

A comprehensive understanding of wheat responses to environmental stress will contribute to the long-term goal of feeding the planet. ALERNATIVE OXIDASE (AOX) genes encode proteins involved in a bypass of the electron transport chain and are also known to be involved in stress tolerance in multiple species. Here, we report the identification and characterization of the AOX gene family in diploid and hexaploid wheat. Four genes each were found in the diploid ancestors Triticum urartu, and Aegilops tauschii, and three in Aegilops speltoides. In hexaploid wheat (Triticum aestivum), 20 genes were identified, some with multiple splice variants, corresponding to a total of 24 proteins for those with observed transcription and translation. These proteins were classified as AOX1a, AOX1c, AOX1e or AOX1d via phylogenetic analysis. Proteins lacking most or all signature AOX motifs were assigned to putative regulatory roles. Analysis of protein-targeting sequences suggests mixed localization to the mitochondria and other organelles. In comparison to the most studied AOX from Trypanosoma brucei, there were amino acid substitutions at critical functional domains indicating possible role divergence in wheat or grasses in general. In hexaploid wheat, AOX genes were expressed at specific developmental stages as well as in response to both biotic and abiotic stresses such as fungal pathogens, heat and drought. These AOX expression patterns suggest a highly regulated and diverse transcription and expression system. The insights gained provide a framework for the continued and expanded study of AOX genes in wheat for stress tolerance through breeding new varieties, as well as resistance to AOX-targeted herbicides, all of which can ultimately be used synergistically to improve crop yield

Generation of doubled haploid transgenic wheat lines by microspore transformation.

Microspores can be induced to develop homozygous doubled haploid plants in a single generation. In the present experiments androgenic microspores of wheat have been genetically transformed and developed into mature homozygous transgenic plants. Two different transformation techniques were investigated, one employing electroporation and the other co-cultivation with Agrobacterium tumefaciens. Different tissue culture and transfection conditions were tested on nine different wheat cultivars using four different constructs. A total of 19 fertile transformants in five genotypes from four market classes of common wheat were recovered by the two procedures. PCR followed by DNA sequencing of the products, Southern blot analyses and bio/histo-chemical and histological assays of the recombinant enzymes confirmed the presence of the transgenes in the T0 transformants and their stable inheritance in homozygous T1∶2 doubled haploid progenies. Several decisive factors determining the transformation and regeneration efficiency with the two procedures were determined: (i) pretreatment of immature spikes with CuSO4 solution (500 mg/L) at 4°C for 10 days; (ii) electroporation of plasmid DNA in enlarged microspores by a single pulse of ∼375 V; (iii) induction of microspores after transfection at 28°C in NPB-99 medium and regeneration at 26°C in MMS5 medium; (iv) co-cultivation with Agrobacterium AGL-1 cells for transfer of plasmid T-DNA into microspores at day 0 for <24 hours; and (v) elimination of AGL-1 cells after co-cultivation with timentin (200-400 mg/L)

Alignment of select TaAOX (hexaploid wheat) proteins with TbAOX (<i>T</i>. <i>brucei</i>).

<p>Yellow highlights indicate conserved motifs. Red font indicates residues proposed to coordinate the diiron center of the active site. Blue font indicates residues experimentally tested for loss of activity by previous researchers. Underlined residues are involved in the TbAOX hydrophobic cavity. Splice variants were identical for the protein region analyzed. The “reg” proteins were not analyzed due to the absence of the conserved motifs.</p

Putative cis-elements for hormonal and developmental responses in promoter regions.

<p>(A) <i>TaAOX</i> gene family and (B) diploid <i>AOX</i> gene families. The promoter for <i>put</i>.<i>regTaAOX-6BL</i> was not analyzed due to the majority of the promoter sequence being unresolved.</p

Transmembrane topologies of wheat AOX proteins predicted by Phyre2.

<p>Transmembrane topologies of wheat AOX proteins predicted by Phyre2.</p

Heat map of expression profiles for high-confidence <i>TaAOX</i> genes under biotic and abiotic stresses.

<p>Heat map of expression profiles for high-confidence <i>TaAOX</i> genes under biotic and abiotic stresses.</p