Characterization of agonist-induced motilin receptor trafficking and its implications for tachyphylaxis

The motilin receptor (MR) is a member of the seven-transmembrane receptor family and is expressed throughout the gastrointestinal tract of humans and other species. Motilin, the natural MR peptide ligand, has profound stimulatory effects on gastrointestinal contractility, indicating a therapeutic potential for MR modulators. However, long-term clinical use of certain MR agonists is limited by tachyphylaxis, a reduced responsiveness to repeated compound exposure. This study was meant to characterize the ligand-induced endocytosis of MR and to test whether receptor trafficking contributes to tachyphylaxis. A cell-based assay was developed by fusing a green fluorescent protein (GFP) moiety to the motilin receptor, and high-content biology instrumentation was used to quantify time and dose dependence of MR-GFP endocytosis. Maximal internalization of MR-GFP was induced after 45 min of constant exposure to 80 nM motilin. This process was disrupted by nocodazole, suggesting an essential role for microtubules. Internalized MR-GFP vesicles disappeared within 15 to 45 min of motilin withdrawal but did not overlap with the lysosomal compartment, indicating that MR-GFP escaped degradation and was recycled back to the plasma membrane. It is noteworthy that the kinetics of MR-GFP redistribution varied substantially when stimulated with motilin, erythromycin, 6,9-hemiacetal 8,9-anhydro-4'-deoxy-3'-N-desmethyl-3'-N-ethylerythromycin B (ABT-229), or N-[(1S)-1-[[[(1S)-1-(aminocarbonyl)-3-phenylpropyl]amino]carbonyl]-3-phenylpropyl]-2'-(1,3-benzodioxol-5-ylmethyl)tetrahydro-1',3'-dioxo-spiro[piperidine-4,5'(6'H)-[1H][1,2,4]triazolo[1,2-a]pyridazine]-8'-carboxamide (BMS-591348) at equipotent doses for Ca(2+)-mobilization. Retardation of the intracellular MR-GFP sorting cycle seemed to correlate with the tachyphylaxis-inducing properties of each compound, but not its EC(50). These results indicate that MR internalization, desensitization, and resensitization are ligand-dependent and that appropriate screening strategies may enable the development of small molecule agonists with ideal combinations of these distinct properties.     

Footbath as a safe,simple, and non-pharmacological method to improve sleep quality of menopausal women

Due to the adverse effects of medication we decided to provide a safe, economic, and easy intervention to decrease sleep problems in menopausal women. This trial aimed to determine the effect of footbath on sleep disturbance in postmenopausal women. One hundred menopausal women were randomly stratified using a permuted block randomization procedure by Random Allocation Software, and assigned to one of two study groups: control (without intervention, n = 50) and intervention (footbath, n = 50). The footbath group was asked to lie their feet into the warm water for 20 min in a container with a depth of 10 cm one hour before the usual sleeping time for 6 weeks. To assess sleep quality, we used the Pittsburgh sleep quality index (PSQI). To evaluate severity of menopause symptoms, the Greene scale was applied. Before the intervention, 96% of the subjects in the footbath group and 94% in the control group were poor sleepers. The results of analysis of covariance showed significant improvement in sleep quality and decrease in severity of menopause signs in the footbath group compared to the control group. At the baseline, there was a statistically significant correlation between the severity of menopause signs and the global score of PSQI in all study individuals (p ≤ .001, r = .464). The present study findings showed that footbath intervention enhanced sleep quality in menopausal women. Thus, footbath as a safe, simple, and non-pharmacological application can improve quality of life in postmenopausal women and inhibit problems due to inadequate sleep quality.     

Effects of rutin on acrylamide-induced neurotoxicity

Background

Rutin is an important flavonoid that is consumed in the daily diet. The cytoprotective effects of rutin, including antioxidative, and neuroprotective have been shown in several studies. Neurotoxic effects of acrylamide (ACR) have been established in humans and animals. In this study, the protective effects of rutin in prevention and treatment of neural toxicity of ACR were studied.

Results

Rutin significantly reduced cell death induced by ACR (5.46 mM) in time- and dose-dependent manners. Rutin treatment decreased the ACR-induced cytotoxicity significantly in comparison to control (P <0.01, P < 0.001). Rutin (100 and 200 mg/kg) could prevent decrease of body weight in rats. In combination treatments with rutin (50, 100 and 200 mg/kg), vitamin E (200 mg/kg) and ACR, gait abnormalities significantly decreased in a dose-dependent manner (P < 0.01 and P < 0.001). The level of malondialdehyde significantly decreased in the brain tissue of rats in both preventive and therapeutic groups that received rutin (100 and 200 mg/kg).

Conclusion

It seems that rutin could be effective in reducing neurotoxicity and the neuroprotective effect of it might be mediated via antioxidant activity.     

Comparison of Three Commonly Used Genetic Markers for Detection of Leishmania Major: An Experimental Study

BackgroundLeishmaniasis is a vector-borne disease caused by an intracellular protozoan parasite called Leishmania spp. Different species produce different clinical outcomes; the majority of cases are cutaneous forms. Leishmania major is one of the main causative agents of cutaneous leishmaniasis (CL). Various methods are being using to diagnose CL, including microscopic examination, culture, and molecular detection of the parasite genome.MethodIn the current study, we tried to compare three common molecular markers, including Kinetoplast DNA (kDNA), Cytochrome b (Cyt b), and Internal transcribed space 1 (ITS1), for the detection of Leishmania major. After cultivation of standard strain of L. major MHOM/IR/75/ER in RPMI 1640, certain number of promastigotes was subjected to DNA extraction and different PCR reactions.ResultsThe lowest number of the parasite (5 promastigotes) can be detected by kDNA-PCR, followed by Cyt b-PCR (10 promastigotes), and ITS1-PCR (50 promastigotes).ConclusionIn conclusion, kDNA-PCR was the most sensitive marker and may provide more reliable data in the initial screening, especially in false-negative results provided by parasitological methods due to the low number of parasites.