Associations of testosterone and sex hormone binding globulin with adipose tissue hormones in midlife women

Objective:

Regulators of adipose tissue hormones remain incompletely understood, but may include sex hormones. As adipose tissue hormones have been shown to contribute to numerous metabolic and cardiovascular disorders, understanding their regulation in midlife women is of clinical importance. Therefore, we assessed the associations between testosterone (T) and sex hormone binding globulin (SHBG) with leptin, high molecular weight (HMW) adiponectin, and the soluble form of the leptin receptor (sOB‐R) in healthy midlife women.

Design and Methods:

Cross‐sectional analyses were performed using data from 1,881 midlife women (average age 52.6 (±2.7) years) attending the sixth Annual follow‐up visit of the multiethnic Study of Women's Health Across the Nation.

Results:

T was weakly negatively associated with both HMW adiponectin and sOB‐R (r = ?0.12 and r = ?0.10, respectively; P < 0.001 for both), and positively associated with leptin (r = 0.17; P < 0.001). SHBG was more strongly and positively associated with both HMW adiponectin and sOB‐R (r = 0.29 and r = 0.24, respectively; P < 0.001 for both), and more strongly and negatively associated with leptin (r = ?0.27; P < 0.001). Adjustment for fat mass, insulin resistance, or waist circumference only partially diminished associations with HMW adiponectin and sOB‐R, but attenuated associations with leptin. In conclusion, in these midlife women, lower SHBG values, and to a lesser extent, higher T levels, were associated with lower, or less favorable, levels of adiponectin and sOB‐R, independent of fat mass.

Conclusions:

These data suggest that variation in these adipose hormones resulting from lower SHBG levels, and possibly, though less likely, greater androgenicity, may contribute to susceptibility for metabolic and cardiovascular outcomes during midlife in women.

     

Trypsin inhibitors in xoconostle seeds (Opuntia joconostle Weber.)

Seed proteins recovered after heating a seed extract from Opuntia joconostle Weber (xoconostle, an acid cactus pear) were screened for different biochemical activities, detecting only trypsin-like inhibitory activity. Two trypsin-like inhibitor forms from seeds were separated by RP-HPLC and partially sequenced and characterized as an enriched mixture. They were evaluated for inhibition on several serine proteinases, but only trypsin-like inhibition was detected by the inhibitor extract. The two isolated forms, OjTI 1 and OjTI 2 showed low molecular weights of 4.26 and 4.17 kDa as determined by mass spectrometry. An enriched inhibitory fraction showed a high thermal stability by retaining the activity after heating the sample for 1 h at 90 °C, as well as after heating for at 120 °C under 1 kg/cm² for 15 min at different pH values. Partial sequence of the two forms was determined by mass spectrometry indicating that they were similar and after alignments analysis they showed the highest similarity with the trypsin inhibitor from O. streptacantha and to a lesser extent to other trypsin inhibitors of the MEROPS database families. The inhibitory spectrum was evaluated against several digestive enzymes from pests and beneficial insects from several taxonomic orders.     

Physiological cyclic stretch directs L-arginine transport and metabolism to collagen synthesis in vascular smooth muscle.

Application of cyclic stretch (10% at 1 hertz) to vascular smooth muscle cells (SMC) increased L-arginine uptake and this was associated with a specific increase in cationic amino acid transporter-2 (CAT-2) mRNA. In addition, cyclic stretch stimulated L-arginine metabolism by inducing arginase I mRNA and arginase activity. In contrast, cyclic stretch inhibited the catabolism of L-arginine to nitric oxide (NO) by blocking inducible NO synthase expression. Exposure of SMC to cyclic stretch markedly increased the capacity of SMC to generate L-proline from L-arginine while inhibiting the formation of polyamines. The stretch-mediated increase in L-proline production was reversed by methyl-L-arginine, a competitive inhibitor of L-arginine transport, by hydroxy-L-arginine, an arginase inhibitor, or by the ornithine aminotransferase inhibitor L-canaline. Finally, cyclic stretch stimulated collagen synthesis and the accumulation of type I collagen, which was inhibited by L-canaline. These results demonstrate that cyclic stretch coordinately stimulates L-proline synthesis by regulating the genes that modulate the transport and metabolism of L-arginine. In addition, they show that stretch-stimulated collagen production is dependent on L-proline formation. The ability of hemodynamic forces to up-regulate L-arginine transport and direct its metabolism to L-proline may play an important role in stabilizing vascular lesions by promoting SMC collagen synthesis.     

Synthesis,characterization, and biological activity of poly(arginine)‐derived cancer‐targeting peptides in HepG2 liver cancer cells

The solid‐phase synthesis, structural characterization, and biological evaluation of a small library of cancer‐targeting peptides have been determined in HepG2 hepatoblastoma cells. These peptides are based on the highly specific Pep42 motif, which has been shown to target the glucose‐regulated protein 78 receptors overexpressed and exclusively localized on the cell surface of tumors. In this study, Pep42 was designed to contain varying lengths (3–12) of poly(arginine) sequences to assess their influence on peptide structure and biology. Peptides were effectively synthesized by 9‐fluorenylmethoxycarbonyl‐based solid‐phase peptide synthesis, in which the use of a poly(ethylene glycol) resin provided good yields (14–46%) and crude purities >95% as analyzed by liquid chromatography–mass spectrometry. Peptide structure and biophysical properties were investigated using circular dichroism spectroscopy. Interestingly, peptides displayed secondary structures that were contingent on solvent and length of the poly(arginine) sequences. Peptides exhibited helical and turn conformations, while retaining significant thermal stability. Structure–activity relationship studies conducted by flow cytometry and confocal microscopy revealed that the poly(arginine) derived Pep42 sequences maintained glucose‐regulated protein 78 binding on HepG2 cells while exhibiting cell translocation activity that was contingent on the length of the poly(arginine) strand. In single dose (0.15 mM) and dose‐response (0–1.5 mM) cell viability assays, peptides were found to be nontoxic in human HepG2 liver cancer cells, illustrating their potential as safe cancer‐targeting delivery agents. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.     

Expression profile and function of Wnt signaling mechanisms in malignant mesothelioma cells

Malignant mesothelioma (MM) is an uncommon and particularly aggressive cancer associated with asbestos exposure, which currently presents an intractable clinical challenge. Wnt signaling has been reported to play a role in the neoplastic properties of mesothelioma cells but has not been investigated in detail in this cancer. We surveyed expression of Wnts, their receptors, and other key molecules in this pathway in well established in vitro mesothelioma models in comparison with primary mesothelial cultures. We also tested the biological response of MM cell lines to exogenous Wnt and secreted regulators, as well as targeting β-catenin. We detected frequent expression of Wnt3 and Wnt5a, as well as Fzd 2, 4 and 6. The mRNA of Wnt4, Fzd3, sFRP4, APC and axin2 were downregulated in MM relative to mesothelial cells while LEF1 was overexpressed in MM. Functionally, we observed that Wnt3a stimulated MM proliferation while sFRP4 was inhibitory. Furthermore, directly targeting β-catenin expression could sensitise MM cells to cytotoxic drugs. These results provide evidence for altered expression of a number of Wnt/Fzd signaling molecules in MM. Modulation of Wnt signaling in MM may prove a means of targeting proliferation and drug resistance in this cancer.     

Microorganisms under high pressure — Adaptation,growth and biotechnological potential

Hydrostatic pressure is a well-known physical parameter which is now considered an important variable of life, since organisms have the ability to adapt to pressure changes, by the development of resistance against this variable. In the past decades a huge interest in high hydrostatic pressure (HHP) technology is increasingly emerging among food and biosciences researchers. Microbial specific stress responses to HHP are currently being investigated, through the evaluation of pressure effects on biomolecules, cell structure, metabolic behavior, growth and viability. The knowledge development in this field allows a better comprehension of pressure resistance mechanisms acquired at sub-lethal pressures. In addition, new applications of HHP could arise from these studies, particularly in what concerns to biotechnology. For instance, the modulation of microbial metabolic pathways, as a response to different pressure conditions, may lead to the production of novel compounds with potential biotechnological and industrial applications. Considering pressure as an extreme life condition, this review intends to present the main findings so far reported in the scientific literature, focusing on microorganisms with the ability to withstand and to grow in high pressure conditions, whether they have innated or acquired resistance, and show the potential of the application of HHP technology for microbial biotechnology.     

19F Nuclear Magnetic Resonance as a Tool To Investigate Microbial Degradation of Fluorophenols to Fluorocatechols and Fluoromuconates

A method was developed to study the biodegradation and oxidative biodehalogenation of fluorinated phenols by ¹⁹F nuclear magnetic resonance (NMR). Characterization of the ¹⁹F NMR spectra of metabolite profiles of a series of fluorophenols, converted by purified phenol hydroxylase, catechol 1,2-dioxygenase, and/or by the yeast-like fungus Exophiala jeanselmei, provided possibilities for identification of the ¹⁹F NMR chemical shift values of fluorinated catechol and muconate metabolites. As an example, the ¹⁹F NMR method thus defined was used to characterize the time-dependent metabolite profiles of various halophenols in either cell extracts or in incubations with whole cells of E. jeanselmei. The results obtained for these two systems are similar, except for the level of muconates observed. Altogether, the results of the present study describe a ¹⁹F NMR method which provides an efficient tool for elucidating the metabolic pathways for conversion of fluorine-containing phenols by microorganisms, with special emphasis on possibilities for biodehalogenation and detection of the type of fluorocatechols and fluoromuconates involved. In addition, the method provides possibilities for studying metabolic pathways in vivo in whole cells.     

Analysis of Genes Involved in Body Weight Regulation by Targeted Re-Sequencing

Introduction

Genes involved in body weight regulation that were previously investigated in genome-wide association studies (GWAS) and in animal models were target-enriched followed by massive parallel next generation sequencing.

Methods

We enriched and re-sequenced continuous genomic regions comprising FTO, MC4R, TMEM18, SDCCAG8, TKNS, MSRA and TBC1D1 in a screening sample of 196 extremely obese children and adolescents with age and sex specific body mass index (BMI) ≥ 99^th percentile and 176 lean adults (BMI ≤ 15^th percentile). 22 variants were confirmed by Sanger sequencing. Genotyping was performed in up to 705 independent obesity trios (extremely obese child and both parents), 243 extremely obese cases and 261 lean adults.

Results and Conclusion

We detected 20 different non-synonymous variants, one frame shift and one nonsense mutation in the 7 continuous genomic regions in study groups of different weight extremes. For SNP Arg695Cys (rs58983546) in TBC1D1 we detected nominal association with obesity (p_TDT = 0.03 in 705 trios). Eleven of the variants were rare, thus were only detected heterozygously in up to ten individual(s) of the complete screening sample of 372 individuals. Two of them (in FTO and MSRA) were found in lean individuals, nine in extremely obese. In silico analyses of the 11 variants did not reveal functional implications for the mutations. Concordant with our hypothesis we detected a rare variant that potentially leads to loss of FTO function in a lean individual. For TBC1D1, in contrary to our hypothesis, the loss of function variant (Arg443Stop) was found in an obese individual. Functional in vitro studies are warranted.     

Morphological and Physiological Changes on Rhizopus stolonifer by Effect of Chitosan,Oligochitosan or Essential Oils

Effects of chitosan, oligochitosan and the essential oils of clove and cinnamon were evaluated on hyphal morphology, cell wall thickness, minimum medium pH changes and respiration of Rhizopus stolonifer. Changes in hyphal morphology were observed due to chitosan or oligochitosan treatment in this fungus. Mycelial branching, abnormal shapes and swelling were showed on hyphae of R. stolonifer treated with chitosan, whereas the development of hyphae was markedly inhibited by the effect of oligochitosan. Clove and cinnamon oils caused few morphological changes in the hyphae of R. stolonifer. Cell wall thickness was increased approximately 2‐ to 3‐fold by effect of chitosan, oligochitosan and the essential oil of clove. R. stolonifer grown in minimum medium generated a decrease in the medium's pH. However, the addition of chitosan or oligochitosan caused increases in pH of medium culture. The highest pH value (5.4) was observed in the presence of chitosan. The respiration of R. stolonifer was stimulated at low concentrations of chitosan, oligochitosan or essential oils. Significant changes in morphology and physiology of this fungus were demonstrated by the effect of all evaluated compounds. The most important changes were induced on cells of R. stolonifer treated with chitosan and oligochitosan.     

Response to oxidative stress as a welfare parameter in swine

In pigs, the genetic selection for lean, large muscle blocks and fast growth has been linked to an increased prevalence of metabolic diseases such as porcine stress syndrome and mulberry heart disease. These diseases are associated with cardiovascular inadequacy, which may lead to oxidative stress. In the present study, reactive oxygen metabolites (ROMs) and the anti-oxidant power (OXY) in sera of different swine groups were investigated. The following groups were selected (each around 80 kg body weight): wild boars (WB), Cinta Senese (CS), and Landrace x Large White (LxLW), the latter as both specific pathogen-free (SPF) and intensively farmed animals. In addition, a group of LxLW agonic sows (AS) was also investigated; this group is known to be under oxidative stress. Two colorimetric micro-methods were used to measure ROMs and OXY; ROMs were expressed as mM H(2)O(2) and OXY as microM HOCl neutralised. Between groups, average ROM and OXY values were found to be significantly different by one-way ANOVA (P < 0.001). ROM levels were lower in WB (13.41 +/- 1.85) and CS (19.27 +/- 1.68), and highest in LxLW (42.00 +/- 1.36). OXY values ranged from 260.10 +/- 22.13 (WB) to 396.90 +/- 9.83 (LxLW). Only one swine group (the CS group) showed a significant, positive correlation between ROM and OXY values. The AS group even showed a negative correlation between ROM and OXY values. These results imply satisfactory environmental coping occurred only within the CS group. Results are discussed in the light of animal welfare legislation, food safety and consumers' protection.