Discovery, Structure-Activity Relationships, and In Vivo Evaluation of Novel Aryl Amides as Brain Penetrant Adaptor Protein 2-Associated Kinase 1 (AAK1) Inhibitors for the Treatment of Neuropathic Pain.

Effective treatment of chronic pain, in particular neuropathic pain, without the side effects that often accompany currently available treatment options is an area of significant unmet medical need. A phenotypic screen of mouse gene knockouts led to the discovery that adaptor protein 2-associated kinase 1 (AAK1) is a potential therapeutic target for neuropathic pain. The synthesis and optimization of structure-activity relationships of a series of aryl amide-based AAK1 inhibitors led to the identification of 59, a brain penetrant, AAK1-selective inhibitor that proved to be a valuable tool compound. Compound 59 was evaluated in mice for the inhibition of µ2 phosphorylation. Studies conducted with 59 in pain models demonstrated that this compound was efficacious in the phase II formalin model for persistent pain and the chronic-constriction-injury-induced model for neuropathic pain in rats. These results suggest that AAK1 inhibition is a promising approach for the treatment of neuropathic pain.

Beneficial effects of hydro-alcoholic extract of Caralluma fimbriata against high-fat diet-induced insulin resistance and oxidative stress in Wistar male rats.

The main aim of this study was to investigate the beneficial effects of hydro-alcoholic extract of Caralluma fimbriata (CFE) on the effects of high-fat diet feeding on insulin resistance and oxidative stress in Wistar rats. High-fat diet (60% of fat) and CFE (200 mg/kg body weight/day) were given concurrently to the rats for a period of 90 days. Feeding with high-fat diet resulted in the development of hyperglycemia, hyperinsulinemia, hyperleptinemia, and hypertriglyceridemia and impaired insulin sensitivity (P < 0.05). Administration of CFE to high-fat diet-fed rats for 90 days resulted in a significant improvement in plasma glucose, insulin, leptin, and triglycerides. Regarding liver antioxidant status, high-fat fed rats showed higher levels of lipid peroxidation, protein oxidation and lower GSH levels and lower activities of enzymatic antioxidants, while CFE treatment prevented all these observed abnormalities. In conclusion, intake of CFE may be beneficial for the suppression of high-fat diet-induced insulin resistance and oxidative stress.

Protective effect of ethanolic extract of Commiphora mukul gum resin against oxidative stress in the brain of streptozotocin induced diabetic Wistar male rats.

The objective of the study was to investigate the possible neuroprotective effect of ethanolic extract of Commiphora mukul gum resin (EtCMGR) against oxidative stress in the brain of streptozotocin (STZ) induced diabetic Wistar rats. The experimental animals were divided into four groups: control (C), control treated with EtCMGR (C+CM), diabetic (D) and diabetic treated with EtCMGR (D+CM). Diabetes was induced by a single intraperitoneal injection of STZ (55 mg/kg body weight). Plant extract treated groups (C+CM and D+CM) were administered EtCMGR at a dose of 200 mg/kg body weight/day by gavage for 60 days. Diabetic rats showed hyperglycemia, hypoinsulinemia with impaired insulin sensitivity. EtCMGR treatment to diabetic (D+CM group) rats prevented the rise in glucose level by 96.7 %, while enhancing insulin level (77.7 %) and improving insulin sensitivity (27.3 %) compared to D group. The brain antioxidant status of D group rats showed higher levels of lipid peroxidation (77.9 %), protein glycation (100 %), and increased activities of xanthine oxidase (47.1 %) and sorbitol dehydrogenase (101.9 %) and lowered concentration of reduced glutathione (38.2 %) and decreased activities of antioxidant enzymes i.e., glutathione reductase (24 %), glutathione peroxidase (24.4 %) and superoxide dismutase (42.1 %) and increased activities of catalase (87.4 %) and glutathione-S-transferase (45.3 %) compared to control group. While EtCMGR treatment for 60 days in D+CM group prevented the observed abnormalities of antioxidant status of D group. This study demonstrates that EtCMGR is a potent neuroprotective agent against oxidative damage induced under diabetes.

Deciphering the regulatory mechanisms of abiotic stress tolerance in plants by genomic approaches.

Environmental constraints that include abiotic stress factors such as salt, drought, cold and extreme temperatures severely limit crop productivity. Improvement of crop plants with traits that confer tolerance to these stresses was practiced using traditional and modern breeding methods. Molecular breeding and genetic engineering contributed substantially to our understanding of the complexity of stress response. Mechanisms that operate signal perception, transduction and downstream regulatory factors are now being examined and an understanding of cellular pathways involved in abiotic stress responses provide valuable information on such responses. This review presents genomic-assisted methods which have helped to reveal complex regulatory networks controlling abiotic stress tolerance mechanisms by high-throughput expression profiling and gene inactivation techniques. Further, an account of stress-inducible regulatory genes which have been transferred into crop plants to enhance stress tolerance is discussed as possible modes of integrating information gained from functional genomics into knowledge-based breeding programs. In addition, we envision an integrative genomic and breeding approach to reveal developmental programs that enhance yield stability and improve grain quality under unfavorable environmental conditions of abiotic stresses.

Differential gene expression during seed germination in barley (Hordeum vulgare L.).

A barley cDNA macroarray comprising 1,440 unique genes was used to analyze the spatial and temporal patterns of gene expression in embryo, scutellum and endosperm tissue during different stages of germination. Among the set of expressed genes, 69 displayed the highest mRNA level in endosperm tissue, 58 were up-regulated in both embryo and scutellum, 11 were specifically expressed in the embryo and 16 in scutellum tissue. Based on Blast X analyses, 70% of the differentially expressed genes could be assigned a putative function. One set of genes, expressed in both embryo and scutellum tissue, included functions in cell division, protein translation, nucleotide metabolism, carbohydrate metabolism and some transporters. The other set of genes expressed in endosperm encodes several metabolic pathways including carbohydrate and amino acid metabolism as well as protease inhibitors and storage proteins. As shown for a storage protein and a trypsin inhibitor, the endosperm of the germinating barley grain contains a considerable amount of residual mRNA which was produced during seed development and which is degraded during early stages of germination. Based on similar expression patterns in the endosperm tissue, we identified 29 genes which may undergo the same degradation process.

Identification of genes specifically expressed in maternal and filial tissues of barley caryopses: a cDNA array analysis.

Developing seeds consist of genetically distinct maternal and filial tissues, whose interactions during development are largely unknown. To better understand the molecular physiology of developing seed tissues in barley, we created a high-density cDNA macroarray bearing 711 cDNA fragments from 691 clones representing at least 620 unique genes mainly derived from a cDNA library constructed with mRNA from the early stages of caryopsis development. This array has been used to compare gene expression patterns in maternal pericarp and filial embryo sac tissues of caryopses sampled 1-7 days after flowering (DAF). The profiles obtained for both tissues revealed that at least 26 genes in pericarp and 12 genes in embryo sac tissues were up-regulated by more than a factor of two during this period. RNAs expressed at high levels in the pericarp mainly encode enzymes involved in carbohydrate and lipid metabolism, but also include mRNA for a transcription factor related to FILAMENTOUS FLOWER (FIL). Genes preferentially expressed in the embryo sac are mainly related to degradation and/or processing of proteins or are involved in the process of starch accumulation, which begins in the seed at this time. Some of the most conspicuously regulated genes were studied in more detail by Northern analysis and in situ hybridization. The mRNA with the highest apparent signal intensity encodes a methionine synthase (MSY). MSY is highly expressed throughout the pericarp and to a lower extent in the transfer cell layer of the endosperm. Of special interest is a gene of unknown function because its high-level expression is restricted to the nucellar projection, the maternal transfer tissue of the caryopsis. This gene, represented by clone HY09L21, may play a central role in transport processes and thus in embryo growth.

Stress responses in two genotypes of mulberry (Morus alba L.) under NaCl salinity.

Changes in biomass yield rates, cell membrane stability (CMS), malondialdehyde (MDA) content and in the levels of physiological stress markers such as proline and glycine betaine in two high yielding genotypes (S1 and ATP, salt tolerant and salt sensitive, respectively) of mulberry under NaCl salinity were studied. Biomass yield rates and CMS were significantly decreased in both the genotypes under stress conditions. Per cent of decrease in biomass yield rate and CMS was relatively less in S1 than in ATP. Salt stress results a significant increase in the accumulation of proline, by 6-fold in S1 and 4-fold in ATP. Glycine betaine content was also increased significantly in stressed plants. However, the per cent increase was more in S1 than in ATP. The level of lipid peroxidation as indicated by MDA formation was greater in ATP than in S1. These results clearly support the better salt tolerant nature of S1 compared to ATP genotype.