Cinnamaldehyde regulates H2O 2-induced skeletal muscle atrophy by ameliorating the proteolytic and antioxidant defense systems

Skeletal muscle atrophy/wasting is associated with impaired protein metabolism in diverse physiological and pathophysiological conditions. Elevated levels of reactive oxygen species (ROS), disturbed redox status, and weakened antioxidant defense system are the major contributing factors toward atrophy. Regulation of protein metabolism by controlling ROS levels and its associated catabolic pathways may help in treating atrophy and related clinical conditions. Although cinnamaldehyde (CNA) enjoys the established status of antioxidant and its role in ROS management is reported, impact of CNA on skeletal muscle atrophy and related pathways is still unexplored. In the current study, the impact of CNA on C2C12 myotubes and the possible protection of cultured cells from H ₂O ₂-induced atrophy is examined. Myotubes were treated with H ₂O ₂ in the presence and absence of CNA and the changes in the antioxidative, proteolytic systems, and mitochondrial functions were scored. Morphological analysis showed significant protective effects of CNA on length, diameter, and nuclei fusion index of myotubes. The evaluation of biochemical markers of atrophy; creatine kinase, lactate dehydrogenase, succinate dehydrogenase along with the study of muscle-specific structural protein (i.e., myosin heavy chain-fast [MHCf] type) showed significant protection of proteins by CNA. CNA pretreatment not only checked the activation of proteolytic systems (ubiquitin-proteasome E3-ligases [MuRF1/Atrogin1]), autophagy [Beclin1/LC3B], cathepsin L, calpain, caspase), but also prevented any alteration in the activities of antioxidative defense enzymes (catalase, glutathione- S-transferase, glutathione-peroxidase, superoxide dismutase, glutathione reductase). The results suggest that CNA protects myotubes from H ₂O ₂-induced atrophy by inhibiting/resisting the amendments in proteolytic systems and maintains cellular redox-balance.     

Accumulation of heavy metals in epidermal glands of the waterlily (Nymphaeaceae)

This study investigates the anatomical aspects of heavy-metal accumulation in the waterlily (Nymphaea `Aurora', Nymphaeaceae). Epidermal glands were identified by light microscopy on the abaxial side of the leaf laminae and on the epidermis of the rhizome; glandular trichomes were observed in the petiole epidermis. Glands were not observed in the roots. Accumulation of heavy metals in these glands was monitored using a scanning electron microscope equipped for energy-dispersive spectroscopy. Further experiments showed maximal cadmium and calcium accumulation in the mature leaf lamina in daylight, and this accumulation was inhibited by the herbicide 3-(3′,4′-dichlorophenyl)-1,1-dimethylurea. These results suggest that, in Nymphaea, heavy metals are accumulated primarily in association with glands found in plant organs that have direct contact with water or mud. Deposition and storage of heavy metals by these glands may represent a stage in the sequestration and detoxification of the metals. Our results raise the possibility of utilizing waterlilies for the removal of heavy metals from polluted environments. Received: 29 April 2000 / Accepted: 8 June 2000     

Formation of the hydrophobic core of ribonuclease A through sequential coordinated conformational transitions

With steady-state and time-resolved fluorescence energy-transfer measurements, we determined the distributions of intramolecular distances in nine mutants to study the conformations of wild-type ribonuclease A in the reduced state under folding conditions. Although far-UV-CD measurements show no evidence for a secondary-structure transition, temperature- and GdnHCl-induced changes in intramolecular distance distributions in the reduced state revealed evidence for long-range subdomain structures in the denatured protein. These poorly defined structures, reflected here by wide distributions corresponding to a wide range of energies, form during refolding in a complex sequence of multiple subdomain transitions. A more well-defined structure emerges only when this structural framework, which directs the successive steps in the folding process, matures and is reinforced by stronger interactions such as disulfide bonds.     

Utility of genetic screening in hypertrophic cardiomyopathy: prevalence and significance of novel and double (homozygous and heterozygous) beta-myosin mutations

Genetic screening of the beta-myosin heavy chain gene (MYH7) was evaluated in 100 consecutive unrelated patients with hypertrophic cardiomyopathy (HCM) and 200 normal unrelated subjects. Seventeen beta-myosin mutations were identified in 19 patients. Notably, 13, or 76%, were novel. Mutations were detected in both alleles in two patients: homozygous for Lys207Gln in one, and heterozygous for Pro211 Leu and Arg663His in another. No mutation was detected in the controls. MYH7-associated HCM was associated with more marked left atrial enlargement and syncope than non-MYH7-related HCM. Our findings indicate that: (1) screening methods should allow identification of novel mutations; and (2) more than one sarcomeric mutation may be present in a patient more commonly than is appreciated. Further studies are necessary to ascertain the clinical consequences of the novel and compound gene abnormalities, and to determine whether correlating functional domain to phenotype provides more useful information about the clinical significance of the molecular defects.     

Resolution of Two Sub-Populations of Conformers and Their Individual Dynamics by Time Resolved Ensemble Level FRET Measurements

Most active biopolymers are dynamic structures; thus, ensembles of such molecules should be characterized by distributions of intra- or intermolecular distances and their fast fluctuations. A method of choice to determine intramolecular distances is based on Förster resonance energy transfer (FRET) measurements. Major advances in such measurements were achieved by single molecule FRET measurements. Here, we show that by global analysis of the decay of the emission of both the donor and the acceptor it is also possible to resolve two sub-populations in a mixture of two ensembles of biopolymers by time resolved FRET (trFRET) measurements at the ensemble level. We show that two individual intramolecular distance distributions can be determined and characterized in terms of their individual means, full width at half maximum (FWHM), and two corresponding diffusion coefficients which reflect the rates of fast ns fluctuations within each sub-population. An important advantage of the ensemble level trFRET measurements is the ability to use low molecular weight small-sized probes and to determine nanosecond fluctuations of the distance between the probes. The limits of the possible resolution were first tested by simulation and then by preparation of mixtures of two model peptides. The first labeled polypeptide was a relatively rigid Pro₇ and the second polypeptide was a flexible molecule consisting of (Gly-Ser)₇ repeats. The end to end distance distributions and the diffusion coefficients of each peptide were determined. Global analysis of trFRET measurements of a series of mixtures of polypeptides recovered two end-to-end distance distributions and associated intramolecular diffusion coefficients, which were very close to those determined from each of the pure samples. This study is a proof of concept study demonstrating the power of ensemble level trFRET based methods in resolution of subpopulations in ensembles of flexible macromolecules.     

Dynamics of a Kalahari long-lived mega-lake system: hydromorphological and limnological changes in the Makgadikgadi Basin (Botswana) during the terminal 50 ka

The Kalahari features a long-lived lacustrine system which may exist since the Early Pleistocene. The emergence of an extant cichlid fish radiation from this (palaeo-) lake during the Middle Pleistocene indicates an ancient lake character. The early history of the system remains speculative, but it is established that lake extensions matching modern Lake Victoria in size have occurred during the Late Pleistocene. It has been assumed that the hydrographical dynamics chiefly depended on the inflow from the Okavango River and thus on ITCZ-controlled precipitation. Our studies, which focused the hydromorphological and palaeolimnological development of the Makgadikgadi Basin during the last 50 ka, suggest that from c. 46–16 ka it did not receive water from the Okavango River but from palaeo-rivers located in the northern and south-western catchment. A northward shift of the winter rainfall zone during the Last Glacial Maximum sustained a high lake level for a period of c. 6 ka. During Heinrich Event 1 (17–16 ka) the lake probably desiccated abruptly and completely. Higher lake levels, controlled by water from the Okavango river system, were reached again during the Holocene before the lake dried up in the middle of the last millennium.     

Inhibition of mycobacterial arylamine N-acetyltransferase contributes to anti-mycobacterial activity of Warburgia salutaris

In this study, we show that extracts and a purified compound of Warburgia salutaris exhibit anti-mycobacterial activity against Mycobacterium tuberculosis H37Rv and Mycobacterium bovis BCG Pasteur. The extracts did not inhibit growth of Escherichia coli and were not toxic to cultured mammalian macrophage cells at the concentrations at which anti-mycobacterial activity was observed. The extract and pure compound inhibited pure recombinant arylamine N-acetyltransferase (NAT), an enzyme involved in mycobacterial cell wall lipid synthesis. Moreover, neither extract nor pure compound inhibited growth of a strain of M. bovis BCG in which nat has been deleted suggesting that NAT may indeed be a target within the mycobacterial cell. The purified compound is a novel drimane sesquiterpenoid lactone, 11alpha-hydroxycinnamosmolide. These studies show that W. salutaris is a useful source of anti-tubercular compounds for further analysis and supports the hypothesis of a link between NAT inhibition and anti-mycobacterial activity.     

A confocal microscopic study of solitary pulmonary neuroendocrine cells in human airway epithelium

Background

Pulmonary neuroendocrine cells (PNEC) are specialized epithelial cells that are thought to play important roles in lung development and airway function. PNEC occur either singly or in clusters called neuroepithelial bodies. Our aim was to characterize the three dimensional morphology of PNEC, their distribution, and their relationship to the epithelial nerves in whole mounts of adult human bronchi using confocal microscopy.

Methods

Bronchi were resected from non-diseased portions of a lobe of human lung obtained from 8 thoracotomy patients (Table ​(Table1)1) undergoing surgery for the removal of lung tumors. Whole mounts were stained with antibodies to reveal all nerves (PGP 9.5), sensory nerves (calcitonin gene related peptide, CGRP), and PNEC (PGP 9.5, CGRP and gastrin releasing peptide, GRP). The analysis and rendition of the resulting three-dimensional data sets, including side-projections, was performed using NIH-Image software. Images were colorized and super-imposed using Adobe Photoshop.

Table 1

Patient Demographic Data