Ghrelin induces proliferation in human aortic endothelial cells via ERK1/2 and PI3K/Akt activation

The direct ghrelin (Ghr) involvement in cardiovascular (CV) system homeostasis has been suggested by the expression of its receptor in CV tissues and by evidence that ghrelin mediates CV activities in animals and in humans. Moreover, low Ghr plasma levels have been reported in pathological conditions characterized by high cardiovascular risk. In the present study, we investigated Ghr effect on proliferation of human aortic endothelial cell (HAEC) and involved transduction pathways. Our results indicate that ghrelin elicited proliferation in a dose-dependent manner (EC(50) about of 5nmol/L) in cultured HAEC, and that this effect was inhibited by the receptor antagonist (D-Lys3)-GHRP-6. Western blot experiments documented an activation of external receptor activated kinases (ERK1/2) and Akt in a dose-dependent fashion, as well as involvement of the cAMP pathway in ERK1/2 phosphorylation. Experiments conducted with appropriate pharmacological inhibitors to investigate Ghr-induced HAEC proliferation confirmed the involvement of ERK1/2 and I3P/Akt pathways, as well as the role of AMP cyclase/PKA pathway in ERK1/2 phosphorylation. Our results indicate that Ghr promotes HAEC proliferation, and thus may be a protective factor against vascular damage. The low ghrelin serum levels reported in insulin-resistant states may not be able to effectively counteract endothelial cell injury.     

Monohydrocalcite in calcareous corpuscles of Mesocestoides corti

Mesocestoides corti (syn. vogae), as many other cestode platyhelminthes, contains abundant mineralized structures called calcareous corpuscles. These concretions may constitute as much as 40% of the dry weight of the organisms, but their function remains poorly understood. In this work, we reviewed the mineral composition of the calcareous corpuscles of M. corti. X-ray diffraction pattern showed that the major mineral component of the corpuscles is a hydrated form of calcium carbonate, monohydrocalcite, also confirmed by infrared spectrometry. The baseline shift of the X-ray diffraction spectra suggested the presence of amorphous calcium carbonate, accordingly to previous reports, and an organic matrix was confirmed by FTIR. Monohydrocalcite is a rare mineral unusually found in biominerals. Although the significance of monohydrocalcite in biominerals has not been determined, the knowledge of corpuscles composition is of relevance to establish their function and for the elucidation of the mechanisms involved in mineralization processes.     

Development of Overweight Associated with Childbearing Depends on Smoking Habit: The Coronary Artery Risk Development in Young Adults (CARDIA) Study

Objective: To prospectively evaluate whether childbearing leads to development of overweight in women and to evaluate the role of other known risk factors. Research Methods and Procedures: A prospective, multicenter observational study, the Coronary Artery Risk Development in Young Adults (CARDIA) Study from 1986 to 1996, examined subjects at baseline and in follow‐up years 2, 5, 7, and 10. Included were 998 (328 black and 670 white) nulliparous women, age 18‐30 years, who were not overweight at baseline. Relative odds for incident overweight (BMI ≥ 25 kg/m²) associated with parity change (0, 1, or 2+) and risk factors were estimated using discrete‐time survival models adjusted for baseline and time‐dependent covariates. Results: Parity change‐association with development of overweight depended on smoking habit (interaction, p < 0.001). In multivariate adjusted models, 1 and 2+ births vs. 0, respectively, were associated with increased risk for development of overweight among never smokers [odds ratio (OR) = 2.66; 95% confidence interval (CI): 1.80, 3.93, and 2.10, 95% CI: 1.24, 3.56] and decreased risk among current smokers (OR = 0.41; 95% CI: 0.17, 0.96, and 0.36, 95% CI: 0.08, 1.65). Risk was increased for black vs. white race (OR = 3.49; 95% CI: 2.59, 4.69), frequent weight cycling (OR = 1.45; 95% CI: 1.03, 2.04), and high school education or less (OR = 2.21; 95% CI: 1.50, 3.26) and was decreased for highest physical activity quartile (OR = 0.62; 95% CI: 0.43, 0.90). Discussion: Childbearing contributes to development of overweight in nonsmokers but not in smokers, where development of overweight is less likely in women who bear children. Race, education, and behaviors are important factors in development of overweight in young women.     

Inducible defences and the paradox of enrichment

In order to evaluate the effects of inducible defences on community stability and persistence, we analyzed models of bitrophic and tritrophic food chains that incorporate consumer-induced polymorphisms. These models predict that intra-specific heterogeneity in defence levels resolves the paradox of enrichment for a range of top-down effects that affect consumer death rates and for all possible levels of primary productivity. We show analytically that this stability can be understood in terms of differences in handling times on the different prey types. Our predictions still hold when defences also affect consumer attack rates. The predicted stability occurs in both bitrophic and tritrophic food chains.
Inducible defences may promote population persistence in tritrophic food chains. Here the minimum densities of cycling populations remain bound away from zero, thus decreasing the risk of population extinctions. However, the reverse can be true for the equivalent bitrophic predator–prey model. This shows that theoretical extrapolations from simple to complex communities should be made with caution. Our results show that inducible defences are among the ecological factors that promote stability in multitrophic communities.     

Development and precise characterization of phospho-site-specific antibody of Ser(357) of IRS-1: elimination of cross reactivity with adjacent Ser(358)

Antibodies that recognize specifically phosphorylated sites on proteins are widely utilized for studying the regulation and biological function of phosphoproteins. The proposed strategy is a powerful, analytical tool allowing the generation of phospho-site specific antibodies albeit adjacent phosphorylation sites are present. Here, we demonstrate the assessment and elimination of cross reactivity of phospho-site-specific-Ser³⁵⁷ IRS-1 antibody. While determining the specificity of p-Ser³⁵⁷ antiserum we came across the cross reactivity of the antiserum with adjacent Ser³⁵⁸ which was successfully abolished by an improved immuno-purification method. The specificity of the purified antiserum was then verified by indirect ELISA, results of ELISA were also mirrored in the experiments carried out in BHK-IR cells using different mutants of IRS-1 carrying mutations at either Ser³⁵⁷/Ser³⁵⁸/Ser^357/358. Immuno-purified-p-Ser³⁵⁷ did not react with IRS-1 Ala³⁵⁷ and IRS-1 Ala^357/358. In conclusion, the present study describes generation and characterization of p-Ser³⁵⁷ IRS-1 antibody, which reacts with IRS-1 in site specific and phosphorylation state-dependent manner without showing cross reactivity to adjacent Ser³⁵⁸. This antibody can be effectively used to further clarify the inhibitory role of Ser³⁵⁷ in insulin signal transduction.     

Differences in Resting Metabolic Rate between White and African‐American Young Adults

Objective: A reported lower resting metabolic rate (RMR) in African‐American women than in white women could explain the higher prevalence of obesity in the former group. Little information is available on RMR in African‐American men. Research Methods and Procedures: We assessed RMR by indirect calorimetry and body composition by DXA in 395 adults ages 28 to 40 years (100 African‐American men, 95 white men, 94 African‐American women, and 106 white women), recruited from participants in the Coronary Artery Risk Development in Young Adults (CARDIA), Birmingham, Alabama, and Oakland, California, field centers. Results: Using linear models, fat‐free mass, fat mass, visceral fat, and age were significantly related to RMR, but the usual level of physical activity was not. After adjustment for these variables, mean RMR was significantly higher in whites (1665.07 ± 10.78 kcal/d) than in African Americans (1585.05 ± 11.02 kcal/d) by 80 ± 16 kcal/d (p < 0.0001). The ethnic × gender interaction was not significant (p = 0.9512), indicating that the difference in RMR between African‐American and white subjects was similar for men and women. Discussion: RMR is ～5% higher in white than in African‐American participants in CARDIA. The difference was the same for men and women and for lean and obese individuals. The prevalence of obesity is not higher in African‐American men than in white men. Because of these reasons, we believe that RMR differences are unlikely to be a primary explanation for why African‐American women are more prone to obesity than white women.     

Microplate-based screening for small molecule inhibitors of neuropilin-2/vascular endothelial growth factor-C interactions

Vascular endothelial growth factor-C (VEGF-C) is a secreted growth factor essential for lymphangiogenesis. VEGF-C functions in both physiological and pathological lymphangiogenesis, particularly in tumor metastasis, making it an attractive therapeutic target. Members of two families of cell surface receptors transduce VEGF-C signals: neuropilin-2 (Nrp2) and VEGF-receptor (VEGFR)-2/3. Nrp2 is a promising target for inhibition because it is highly expressed in lymphatic vessels. Here we describe a microplate-based assay for discovery of VEGF-C/Nrp2 inhibitors. We optimize this assay for use in screening an inhibitor library and identify three novel Nrp2/VEGF-C binding inhibitors from the National Institutes of Health (NIH) Clinical Collection small molecule library.     

Functional Integration of the Conserved Domains of Shoc2 Scaffold

Shoc2 is a positive regulator of signaling to extracellular signal-regulated protein kinases 1 and 2 (ERK1/2). Shoc2 is also proposed to interact with RAS and Raf-1 in order to accelerate ERK1/2 activity. To understand the mechanisms by which Shoc2 regulates ERK1/2 activation by the epidermal growth factor receptor (EGFR), we dissected the role of Shoc2 structural domains in binding to its signaling partners and its role in regulating ERK1/2 activity. Shoc2 is comprised of two main domains: the 21 leucine rich repeats (LRRs) core and the N-terminal non-LRR domain. We demonstrated that the N-terminal domain mediates Shoc2 binding to both M-Ras and Raf-1, while the C-terminal part of Shoc2 contains a late endosomal targeting motif. We found that M-Ras binding to Shoc2 is independent of its GTPase activity. While overexpression of Shoc2 did not change kinetics of ERK1/2 activity, both the N-terminal and the LRR-core domain were able to rescue ERK1/2 activity in cells depleted of Shoc2, suggesting that these Shoc2 domains are involved in modulating ERK1/2 activity.     

Gentamicin palmitate as a new antibiotic formulation for mixing with bone tissue and local release

During surgery with bone grafting, the impaction of bone tissue creates an avascular area where local circulation is disrupted. If infections arise, they may prevent systemically administered antibiotics from reaching the infected bone. In this study we evaluated gentamicin palmitate (GP) mixed with gentamicin sulfate (GS) as a coating for bone chips (BCh). The efficacy of the coated BCh was measured by gentamicin base release tests using B. subtilis, S. epidermidis and S. aureus. Gentamicin base release was evaluated in phosphate-buffered saline for up to 7 days using B. subtilis bioassay. Antimicrobial efficacy was tested with S. aureus and S. epidermidis. A significant difference on the release of gentamicin base between GS and GS + GP was observed. S. epidermidis are significantly more susceptible to GS + GP and GS than S. aureus. BCh can act as gentamicin carriers and showed efficacy against S. aureus and S. epidermidis.