Characterization of cross-bridge elasticity and kinetics of cross-bridge cycling during force development in single smooth muscle cells

Force development in smooth muscle, as in skeletal muscle, is believed to reflect recruitment of force-generating myosin cross-bridges. However, little is known about the events underlying cross-bridge recruitment as the muscle cell approaches peak isometric force and then enters a period of tension maintenance. In the present studies on single smooth muscle cells isolated from the toad (Bufo marinus) stomach muscularis, active muscle stiffness, calculated from the force response to small sinusoidal length changes (0.5% cell length, 250 Hz), was utilized to estimate the relative number of attached cross-bridges. By comparing stiffness during initial force development to stiffness during force redevelopment immediately after a quick release imposed at peak force, we propose that the instantaneous active stiffness of the cell reflects both a linearly elastic cross-bridge element having 1.5 times the compliance of the cross-bridge in frog skeletal muscle and a series elastic component having an exponential length-force relationship. At the onset of force development, the ratio of stiffness to force was 2.5 times greater than at peak isometric force. These data suggest that, upon activation, cross-bridges attach in at least two states (i.e., low-force-producing and high-force-producing) and redistribute to a steady state distribution at peak isometric force. The possibility that the cross-bridge cycling rate was modulated with time was also investigated by analyzing the time course of tension recovery to small, rapid step length changes (0.5% cell length in 2.5 ms) imposed during initial force development, at peak force, and after 15 s of tension maintenance. The rate of tension recovery slowed continuously throughout force development following activation and slowed further as force was maintained. Our results suggest that the kinetics of force production in smooth muscle may involve a redistribution of cross-bridge populations between two attached states and that the average cycling rate of these cross-bridges becomes slower with time during contraction.     

Panmixia across elevation in thermally sensitive Andean dung beetles

Janzen's seasonality hypothesis predicts that organisms inhabiting environments with limited climatic variability will evolve a reduced thermal tolerance breadth compared with organisms experiencing greater climatic variability. In turn, narrow tolerance breadth may select against dispersal across strong temperature gradients, such as those found across elevation. This can result in narrow elevational ranges and generate a pattern of isolation by environment or neutral genetic differentiation correlated with environmental variables that are independent of geographic distance. We tested for signatures of isolation by environment across elevation using genome‐wide SNP data from five species of Andean dung beetles (subfamily Scarabaeinae) with well‐characterized, narrow thermal physiologies, and narrow elevational distributions. Contrary to our expectations, we found no evidence of population genetic structure associated with elevation and little signal of isolation by environment. Further, elevational ranges for four of five species appear to be at equilibrium and show no decay of genetic diversity at range limits. Taken together, these results suggest physiological constraints on dispersal may primarily operate outside of a stable realized niche and point to a lower bound on the spatial scale of local adaptation.     

1-Methyl-3-nitropyridine: An Efficient Oxidant of NADH in Non-enzymatic and Enzyme-mediated Processes

It is shown that NADH can be effectively oxidized by 1-methyl-3-nitropyridine in non-enzymatic and enzyme-mediated processes. Mechanistic issues of these reactions are discussed. These processes seem to contribute to the observed cytotoxicity of 1-methyl-3-nitropyridine. A key role of 1-methyl-3-nitropyridinyl radical formed in the enzyme-mediated processes is emphasized.     

E0771 and 4T1 murine breast cancer cells and interleukin 6 alter gene expression patterns but do not induce browning in cultured white adipocytes

Breast cancer remains a substantial clinical problem worldwide, and cancer-associated cachexia is a condition associated with poor prognosis in this and other malignancies. Adipose tissue is involved in the development and progression of cancer-associated cachexia, but its various roles and mechanisms of action are not completely defined, especially as it relates to breast cancer. Interleukin 6 has been implicated in several mechanisms contributing to increased breast cancer tumorigenesis, as well as a net-negative energy balance and cancer-associated cachexia via adipose tissue remodeling in other models of cancer; however, its potential role in breast cancer-associated white adipose browning has not been explored. In this study, we demonstrate localized white adipose tissue browning in a spontaneous model of murine mammary cancer. We then used an in vitro murine adipocyte culture system with the E0771 and 4T1 cell lines as models of breast cancer. We demonstrate that while the E0771 and 4T1 secretomes and cross-talk with white adipocytes alter white adipocyte mRNA expression, they do not directly induce white adipocyte browning. Additionally, we show that neither exogenous administration of interleukin 6 alone or with its soluble receptor directly induce white adipocyte browning. Together, these results demonstrate that neither the E0771 or 4T1 murine breast cancer cell lines, nor interleukin 6, directly cause browning of cultured white adipocytes. This suggests that their roles in adipose tissue remodeling are more complex and indirect in nature.     

Decomposition processes interacting with microtopography maintain ecosystem heterogeneity in a subalpine grassland

Plant and Soil - Grasslands are among the largest ecosystems in the world and store up to 30% of the global reserves of carbon. Decomposition processes have a crucial role in maintaining carbon...     

Measurement of mandibular movements in patients with temporomandibular disorders and in asymptomatic subjects

The aim of the study was to investigate the range of mandibular movements and to analyze the difference in range of mouth opening, right and left lateral movements, and protrusive movement between patients with clinical diagnoses of temporomandibular disorders and asymptomatic subjects (control group) in a young male population. A total of 240 subjects, aged 19-28, were included in the study. The TMD sample comprised 180 patients (60 patients with muscle disorders; 60 patients with disc displacement with reduction; and 60 patients with muscle disorders and disc displacement with reduction) and was compared with 60 healthy control subjects. All participants were evaluated by the attending dentists at baseline by means of a physical examination of the masticatory system and a history questionnaire which included the Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD) Axis I measures. Analysis of variance (ANOVA) with the post hoc Bonferroni criteria showed significant difference in ranges of mandibular movements between and within the groups of asymptomatic subjects and TMD patients for active mouth opening (p = 0.001), right lateral movement (p = 0.002), left lateral movement (p = 0.006), and protrusive movement (p = 0.05). It has been found that there are statistically significant differences in the range of mandibular movements that separate asymptomatic subjects and patients with muscle disorders and disc displacements with reduction in this young male population. However, we cannot conclude that measurements of active mandibular movements can discriminate one group (TMD patients) from the other (asymptomatic subjects), because the mean ranges of these active movements between the groups were measured in clinically "normal" values.     

Cigarette smoke extract induces oxidative stress and apoptosis in human lung fibroblasts

Cigarette smoke is a mixture of chemicals having direct and/or indirect toxic effects on different lung cells. We investigated the effect of cigarette smoke on human lung fibroblasts (HFL-1) oxidation and apoptosis. Cells were exposed to various concentrations (1, 5, and 10%) of cigarette smoke extract (CSE) for 3 h, and oxidative stress and apoptosis were assessed by fluorescence-activated cell sorting and confocal laser fluorescence microscopy. Both oxidative stress and apoptosis exhibited a dose-response relationship with CSE concentrations. Lung fibroblasts also showed marked DNA fragmentation at the Comet assay after exposure to 10% CSE. Coincubation of HLF-1 cells with N-acetylcysteine (1 mM) during CSE exposure significantly reduced oxidative stress, apoptosis, and DNA fragmentation, whereas preincubation (3 h) with the glutathione-depleting agent buthionine sulfoximine (125 microM) produced a significant increase of oxidative stress. Cigarette smoke is a potent source of oxidative stress, DNA damage, and apoptosis for HFL-1 cells, and we speculate that this could contribute to the development of pulmonary emphysema in the lungs of smokers.     

The satisfaction with the removable partial denture therapy in the Croatian adult population

Little is known about the outcomes of treatment and patient's satisfaction with removable partial dentures in adult Croatian population. Therefore patient's satisfaction with their partial dentures in relation to some socio-economic variables was studied. Patient's satisfaction with denture retention, speech, aesthetics, comfort of wearing dentures, chewing ability was also studied in relation to different denture classification, construction, material, denture base shape (major connectors), denture support and the number of missing teeth. A total of 165 patients, 59 males and 105 females between 38 and 87 years took part in this study. A questionnaire, devised for a purpose of the study, was divided into three parts. In the first part, patients answered questions about age, gender, marital status, education, general health, socio-economic status, self-supporting life, period of tooth loss and number of previous denture experiences and in the second part, patients graded their partial dentures, depending on the level of satisfaction, by using a scale from 1 to 5. In the third part a dentist determined Kennedy classification and their modifications, denture material and denture support, denture base shape and the number of missing teeth and graded a denture construction. Influence of these factors on patient's satisfaction was analyzed. A majority of the examined patients were satisfied with the partial prosthesis, but a small amount of dissatisfaction existed. More then half of them scored all the examined parameters to the best score category. Considering chewing with lower partial dentures, women were more satisfied than men (p < 0.05). Patients with more missing teeth gave lower grades for the comfort of wearing dentures (p < 0.05). Patients of higher education gave lower grades (p < 0.05) for the aesthetics. Patients were not satisfied with speech if the dentist graded a construction of a lower partial denture low (p < 0.05). Dissatisfaction was related to mastication, aesthetics, number of missing teeth and ability of speech. These findings can aid a clinician in discussing a treatment plan and help a patient understand the risk of dissatisfaction in the presence of certain factors.     

A fluorescent alpha-factor analogue exhibits multiple steps on binding to its G protein coupled receptor in yeast

The yeast alpha-factor receptor encoded by the STE2 gene is a member of the extended family of G protein coupled receptors (GPCRs) involved in a wide variety of signal transduction pathways. We report here the use of a fluorescent alpha-factor analogue [K(7)(NBD), Nle(12)] alpha-factor (Lys(7) (7-nitrobenz-2-oxa-1,3-diazol-4-yl), norleucine(12) alpha-factor) in conjunction with flow cytometry and fluorescence microscopy to study binding of ligand to the receptor. Internalization of the fluorescent ligand following receptor binding can be monitored by fluorescence microscopy. The use of flow cytometry to detect binding of the fluorescent ligand to intact yeast cells provides a sensitive and reproducible assay that can be conducted at low cell densities and is relatively insensitive to fluorescence of unbound and nonspecifically bound ligand. Using this assay, we determined that some receptor alleles expressed in cells lacking the G protein alpha subunit exhibit a higher equilibrium binding affinity for ligand than the same alleles expressed in isogenic cells containing the normal complement of G protein subunits. On the basis of time-dependent changes in the intensity and shape of the emission spectrum of [K(7)(NBD),Nle(12)] alpha-factor during binding, we infer that the ligand associates with receptors via a two-step process involving an initial interaction that places the fluorophore in a hydrophobic environment, followed by a conversion to a state in which the fluorophore moves to a more polar environment.