Purification of fat body glutathione S‐transferase from the desert locust Schistocerca gregaria: investigation of flavonoid inhibitory effects on enzyme activity

Focus on the development of botanical insecticides such as polyphenols may represent an alternative method to chemical control. In the present study, total glutathione concentration and its related antioxidant enzymes in foregut, midgut, hindgut and fat body homogenates of the desert locust Schistocerca gregaria are examined. Glutathione S‐transferase (GST) activity exhibits a significantly higher value in fat bodies compared with other tissues. A simple and reproducible procedure for the purification of S. gregaria fat body GST is established and the purified enzyme is shown to be homogenous. The purified GST displays a typical Michaelis behaviour with respect to its substrates. Characterization of the GST, including optimum pH, substrate specificity and inhibitor effects, is carried out. The ability of some flavonoids to inhibit S. gregaria fat body GST activity is examined. High‐performance liquid chromatography analysis indicates that the major components in Glycyrrhiza glabra roots are 18α‐glycyrrhetinic acid, quercetin and rutin, and the major components in Hibiscus sabdariffa calyx are cyanidin 3‐O‐glucoside chloride and delphinidin. Quercetin and delphinidin chloride exhibit strong GST inhibition and the inhibition type is determined for both. Rutin shows a smaller inhibitory effect, whereas 18α‐glycyrrhetinic acid and cyanidin have no effect. Inhibition of S. gregaria fat body GST activity would be expected to prevent, or at least delay, the development of resistance to chemical pesticides. Among the examined levels of the antioxidant enzymes, total glutathione concentration and its related enzymes in foregut, midgut, hindgut and fat body crude homogenates of S. gregaria GST activity exhibit a significantly higher value in fat bodies compared with other tissues. Some flavonoids that are detected in H. sabdariffa calyx and G. glabra root extracts are the most effective inhibitors of the purified S. gregaria fat body GST activity. Inhibition of S. gregaria fat body GST activity by quercetin and delphinidin (major compounds detected by HPLC) would be expected to prevent, or at least delay, the development of resistance to chemical pesticides.     

Development of Molecular Tools for Characterization and Genetic Diversity Analysis in Tunisian Fig (Ficus carica) Cultivars

Fig, Ficus carica L., is a useful genetic resource for commercial cultivation. In this study, RAPD (60), ISSR (48), RAMPO (63), and SSR (34) markers were compared to detect polymorphism and to establish genetic relationships among Tunisian fig tree cultivars. The statistical procedures conducted on the combined data show considerable genetic diversity, and the tested markers discriminated all fig genotypes studied. The identification key established on the basis of SSR permitted the unambiguous discrimination of cultivars and confirmed the reliability of SSR for fingerprinting fig genotypes. The study findings are discussed in relation to the establishment of a national reference collection that will aid in the conservation of Tunisian fig resources.     

Health Impact of Fasting in Saudi Arabia during Ramadan: Association with Disturbed Circadian Rhythm and Metabolic and Sleeping Patterns

Background

Muslims go through strict Ramadan fasting from dawn till sunset for one month yearly. These practices are associated with disturbed feeding and sleep patterns. We recently demonstrated that, during Ramadan, circadian cortisol rhythm of Saudis is abolished, exposing these subjects to continuously increased cortisol levels.

Hypothesis

Secretory patterns of other hormones and metabolic parameters associated with cortisol, and insulin resistance, might be affected during Ramadan.

Protocol

Ramadan practitioners (18 males, 5 females; mean age ±SEM = 23.16±1.2 years) were evaluated before and two weeks into Ramadan. Blood was collected for measurements of endocrine and metabolic parameters at 9 am (±1 hour) and again twelve hours later.

Results

In Ramadan, glucose concentration was kept within normal range, with a significant increase in the morning. Mean morning concentration of leptin was significantly higher than pre-Ramadan values (p = 0.001), in contrast to that of adiponectin, which was significantly lower (p<0.001). These changes were associated with increased insulin resistance in morning and evening. Concentrations of hsCRP were lower during Ramadan than those during regular living conditions, however, normal circadian fluctuation was abolished (p = 0.49). Even though means of liver enzymes, total bilirubin, total protein and albumin were all decreased during Ramadan, statistically lower means were only noted for GGT, total protein, and albumin (p = 0.018, 0.002 and 0.001 respectively).

Discussion

Saudi Ramadan practitioners have altered adipokine patterns, typical of insulin resistance. The noted decreases of hsCRP, liver enzymes, total protein, and albumin, are most likely a result of fasting, while loss of circadian rhythmicity of hsCRP is probably due to loss of circadian cortisol rhythm.

Conclusions

Modern Ramadan practices in Saudi Arabia, which are associated with evening hypercortisolism, are also characterized by altered adipokines patterns, and an abolished hsCRP circadian rhythm, all likely to increase cardiometabolic risk.     

Latrunculin and cytochalasin decrease chondrocyte matrix retention.

The proteoglycan-rich extracellular matrix (ECM) directly associated with the cells of articular cartilage is anchored to the chondrocyte plasma membrane via interaction with the hyaluronan receptor CD44. The cytoplasmic tail of CD44 interacts with the cortical cytoskeleton. The objective of this study was to determine the role of the actin cytoskeleton in CD44-mediated matrix assembly by chondrocytes and cartilage matrix retention and homeostasis. Adult bovine articular cartilage tissue slices and isolated chondrocytes were treated with latrunculin or cytochalasin. Tissues were processed for histology and chondrocytes were examined for CD44 expression and pericellular matrix assembly. Treatments that disrupt the actin cytoskeleton reduced chondrocyte pericellular matrix assembly and the retention of proteoglycan within cartilage explants. There was enhanced detection of a neoepitope resulting from proteolysis of aggrecan. Cytoskeletal disruption did not reduce CD44 expression, as monitored by flow cytometry, but detergent extraction of CD44 was enhanced and hyaluronan binding was decreased. Thus, disruption of the cytoskeleton reduces the anchorage of CD44 in the chondrocyte membrane and the capacity of CD44 to bind its ligand. The results suggest that cytoskeletal disruption within cartilage uncouples chondrocytes from the matrix, resulting in altered metabolism and deleterious changes in matrix structure.     

Hyperglycemia induces elevated expression of thyroid hormone binding protein in vivo in kidney and heart and in vitro in mesangial cells

During a search for glucose-regulated abundant mRNAs in the diabetic rat kidney, we cloned thyroid hormone binding protein (THBP), also known as μ-crystallin or CRYM. The aim of this study was to investigate the effect of hyperglycemia/high glucose on the expression of THBP. THBP mRNA copy numbers were determined in kidneys and hearts of diabetic GK rats vs normoglycemic Wistar rats, and in human mesangial cells (HMCs) exposed to high glucose using real-time qPCR, and THBP protein levels were measured by Western blotting and immunofluorescence. Intracellular ROS was measured in THBP transfected cells using DCF fluorescence. Hyperglycemia significantly increased THBP mRNA in GK rat kidneys (326 ± 50 vs 147 ± 54, p < 0.05), and hearts (1583 ± 277 vs 191 ± 63, p < 0.05). Moreover, the levels of THBP mRNA increased with age and hyperglycemia in GK rat kidneys, whereas in normoglycemic Wistar rat kidneys there was a decline with age. High glucose significantly increased THBP mRNA (92 ± 37 vs 18 ± 4, p < 0.005), and protein in HMCs. The expression of THBP as a fusion protein in transfected HMCs resulted in reduction of glucose-induced intracellular ROS. We have shown that THBP mRNA is increased in diabetic kidney and heart, is regulated by high glucose in renal cells, and appears to attenuate glucose-induced intracellular ROS. These data suggest that THBP may be involved in the cellular pathways activated in response to glucose. This is the first report linking hyperglycemia with THBP and suggests that the role of THBP in diabetic complications should be further investigated.     

Design,synthesis, anti-inflammatory activity and molecular docking of potential novel antipyrine and pyrazolone analogs as cyclooxygenase enzyme (COX) inhibitors

As a part of a directed program for development of new active agents, novel heterocyclic derivatives with antipyrine and pyrazolone moieties -incorporated in- have been designed and synthesized. Starting with 4-arylidene-3-methyl-1-phenyl-5-pyrazolone derivative 2a,b novel Mannich bases derivatives have been synthesized and biologically evaluated for their anti-inflammatory activity. Furthermore, the activity of such compounds has been tested interestingly as COX-1 and COX-2 inhibitors. Structure elucidation of the synthesized compounds was attained by the use of elemental analysis, IR, ¹H NMR, ¹³C NMR, and Mass spectrometry techniques. Compounds 3b, 3d and 4b represent the high % inhibition values for both COX-1 and COX-2. On the other hand, compound 8 showed little selectivity against COX-2 while compound 10 showed good selectivity against COX-1 only. Structure activity relationship has been discussed and the results were confirmed by molecular docking calculations.     

PDQ Wizard: automated prioritization and characterization of gene and protein lists using biomedical literature

SUMMARY: PDQ Wizard automates the process of interrogating biomedical references using large lists of genes, proteins or free text. Using the principle of linkage through co-citation biologists can mine PubMed with these proteins or genes to identify relationships within a biological field of interest. In addition, PDQ Wizard provides novel features to define more specific relationships, highlight key publications describing those activities and relationships, and enhance protein queries. PDQ Wizard also outputs a metric that can be used for prioritization of genes and proteins for further research. AVAILABILITY: PDQ Wizard is freely available from http://www.gti.ed.ac.uk/pdqwizard/.     

Propolis protects against high glucose-induced vascular endothelial dysfunction in isolated rat aorta

While propolis is known to have abundant bioactive constituents and a variety of biological activities, it is not clear whether propolis has beneficial effects on high glucose-mediated vascular endothelial impairment. The aim of the present study was to investigate the potential protective effect of propolis extract against the acute vascular endothelial dysfunction resulting from exposure to high glucose load and to elucidate its underlying mechanism. Rat aortic rings were incubated with normal glucose (11 mM), high glucose (44 mM), or mannitol (44 mM) for 3 h with or without propolis extract (400 μg/ml). Contraction to phenylephrine (Phe, 10^?9–10^?5 M) and relaxation to acetylcholine (ACh, 10^?9–10^?5 M) and sodium nitroprusside (SNP, 10^?9–10^?5 M) were measured before and after incubation. Changes in malondialdehyde (MDA), reduced glutathione (GSH), and superoxide dismutase (SOD) were also measured. Phe-induced contraction was impaired by high glucose as the E _max decreased from 138.87?±?11.43 to 103.65?±?11.5 %. In addition, ACh-induced relaxation was impaired as the E _max decreased from 99.80?±?7.25 to 39.20?±?6.5 %. SNP-induced relaxation was not affected. Furthermore, high glucose decreased the levels of both SOD (by 6 U/ml) and GSH (by 68 %) and increased levels of MDA (by 85 %). Propolis extract prevented high glucose-induced impairment of Phe and ACh responses and increased both SOD and GSH, leading to decreased MDA levels. In conclusion, propolis can protect against high glucose-induced vascular dysfunction by reducing oxidative stress.     

Optimal Use of MRSASelect and PCR to Maximize Sensitivity and Specificity of MRSA Detection

Suspected colonies of methicillin-resistant Staphylococcus aureus (MRSA) on chromogenic, MRSASelect (BioRad) medium were confirmed using routine microbiological methods, and a multiplex real-time PCR (n = 108). Although the specificity of MRSASelect assessed at 24 h of incubation was much higher than that of 48 h (91.4 vs. 60 %), extending the incubation time to 48 h, along with PCR confirmation, increased the total number of true positive samples by 27.8 %. These results provide a cost effective method for sensitive and specific detection of MRSA.     

Cardiac stem cells: Current knowledge and future prospects

Regenerative medicine is the field concerned with the repair and restoration of the integrity of damaged human tissues as well as whole organs.Since the inception of the field several decades ago,regenerative medicine therapies,namely stem cells,have received significant attention in preclinical studies and clinical trials.Apart from their known potential for differentiation into the various body cells,stem cells enhance the organ's intrinsic regenerative capacity by altering its environment,whether by exogenous injection or introducing their products that modulate endogenous stem cell function and fate for the sake of regeneration.Recently,research in cardiology has highlighted the evidence for the existence of cardiac stem and progenitor cells(CSCs/CPCs).The global burden of cardiovascular diseases’morbidity and mortality has demanded an in-depth understanding of the biology of CSCs/CPCs aiming at improving the outcome for an innovative therapeutic strategy.This review will discuss the nature of each of the CSCs/CPCs,their environment,their interplay with other cells,and their metabolism.In addition,important issues are tackled concerning the potency of CSCs/CPCs in relation to their secretome for mediating the ability to influence other cells.Moreover,the review will throw the light on the clinical trials and the preclinical studies using CSCs/CPCs and combined therapy for cardiac regeneration.Finally,the novel role of nanotechnology in cardiac regeneration will be explored.