Mitochondrial Membrane Studies Using Impedance Spectroscopy with Parallel pH Monitoring

A biological microelectromechanical system (BioMEMS) device was designed to study complementary mitochondrial parameters important in mitochondrial dysfunction studies. Mitochondrial dysfunction has been linked to many diseases, including diabetes, obesity, heart failure and aging, as these organelles play a critical role in energy generation, cell signaling and apoptosis. The synthesis of ATP is driven by the electrical potential across the inner mitochondrial membrane and by the pH difference due to proton flux across it. We have developed a tool to study the ionic activity of the mitochondria in parallel with dielectric measurements (impedance spectroscopy) to gain a better understanding of the properties of the mitochondrial membrane. This BioMEMS chip includes: 1) electrodes for impedance studies of mitochondria designed as two- and four-probe structures for optimized operation over a wide frequency range and 2) ion-sensitive field effect transistors for proton studies of the electron transport chain and for possible monitoring other ions such as sodium, potassium and calcium. We have used uncouplers to depolarize the mitochondrial membrane and disrupt the ionic balance. Dielectric spectroscopy responded with a corresponding increase in impedance values pointing at changes in mitochondrial membrane potential. An electrical model was used to describe mitochondrial sample’s complex impedance frequency dependencies and the contribution of the membrane to overall impedance changes. The results prove that dielectric spectroscopy can be used as a tool for membrane potential studies. It can be concluded that studies of the electrochemical parameters associated with mitochondrial bioenergetics may render significant information on various abnormalities attributable to these organelles.     

Warm-up optimization in amateur male soccer players: A comparison of small-sided games and traditional warm-up routines on physical fitness qualities

The aim of this study was to compare the effects of small-sided soccer games (SSSGs) and traditional warm-up (TWU) routines on physical fitness qualities in soccer players. Following a between-subject, randomized design, amateur-level soccer players were assigned to a SSSG warm-up (n = 10; age: 19.3 ± 2.8 years) or TWU group (n = 10; age: 19.3 ± 2.4 years). Players completed multiple trials of 10-m and 30-m linear sprints, change-of-direction speed (CODS) tests, and countermovement jumps (CMJ) prior to and following the warm-up routine. Separate mixed ANOVAs were performed to assess group effects (SSSG vs. TWU), time effects within each group (pre- vs. post-warm-up), and their interaction for each physical fitness quality. No significant interaction effects were observed for any dependent variable. Significant improvements were evident between baseline and follow-up measurements for 10-m sprint time (p = 0.002, Hedges’ g effect size [g] = 0.59) and CMJ variables (height: p = 0.016, g = 0.20; power: p = 0.003, g = 0.19; force: p = 0.002, g = 0.14) in the TWU group and for CODS performance time (p = 0.012, g = 0.51) and CMJ variables (height: p < 0.001, g = 0.46; power: p = 0.002, g = 0.35; force: p = 0.001, g = 0.27) in the SSSG warm-up group. Both SSSG and TWU protocols improved selected physical fitness qualities with SSSG more effective at improving CODS and CMJ performance, and TWU more effective at improving linear speed. Soccer coaches may choose between SSSG or traditional warm-up activities according to player needs and preferences; however, the superior effects of SSSG suggest it might offer greater benefits than TWU in preparing players for optimal physical output.     

Isorhamnetin Inhibits Proliferation and Invasion and Induces Apoptosis through the Modulation of Peroxisome Proliferator-activated Receptor γ Activation Pathway in Gastric Cancer

Gastric cancer (GC) is a lethal malignancy and the second most common cause of cancer-related deaths. Although treatment options such as chemotherapy, radiotherapy, and surgery have led to a decline in the mortality rate due to GC, chemoresistance remains as one of the major causes for poor prognosis and high recurrence rate. In this study, we investigated the potential effects of isorhamnetin (IH), a 3′-O-methylated metabolite of quercetin on the peroxisome proliferator-activated receptor γ (PPAR-γ) signaling cascade using proteomics technology platform, GC cell lines, and xenograft mice model. We observed that IH exerted a strong antiproliferative effect and increased cytotoxicity in combination with chemotherapeutic drugs. IH also inhibited the migratory/invasive properties of GC cells, which could be reversed in the presence of PPAR-γ inhibitor. We found that IH increased PPAR-γ activity and modulated the expression of PPAR-γ regulated genes in GC cells. Also, the increase in PPAR-γ activity was reversed in the presence of PPAR-γ-specific inhibitor and a mutated PPAR-γ dominant negative plasmid, supporting our hypothesis that IH can act as a ligand of PPAR-γ. Using molecular docking analysis, we demonstrate that IH formed interactions with seven polar residues and six nonpolar residues within the ligand-binding pocket of PPAR-γ that are reported to be critical for its activity and could competitively bind to PPAR-γ. IH significantly increased the expression of PPAR-γ in tumor tissues obtained from xenograft model of GC. Overall, our findings clearly indicate that antitumor effects of IH may be mediated through modulation of the PPAR-γ activation pathway in GC.     

Focus: A Multifaceted Battle Against Cancer: Extracting Knowledge from Chemical Imaging Data Using Computational Algorithms for Digital Cancer Diagnosis

Fourier transform infrared (FTIR) spectroscopic imaging is an emerging microscopy modality for clinical histopathologic diagnoses as well as for biomedical research. Spectral data recorded in this modality are indicative of the underlying, spatially resolved biochemical composition but need computerized algorithms to digitally recognize and transform this information to a diagnostic tool to identify cancer or other physiologic conditions. Statistical pattern recognition forms the backbone of these recognition protocols and can be used for highly accurate results. Aided by biochemical correlations with normal and diseased states and the power of modern computer-aided pattern recognition, this approach is capable of combating many standing questions of traditional histology-based diagnosis models. For example, a simple diagnostic test can be developed to determine cell types in tissue. As a more advanced application, IR spectral data can be integrated with patient information to predict risk of cancer, providing a potential road to precision medicine and personalized care in cancer treatment. The IR imaging approach can be implemented to complement conventional diagnoses, as the samples remain unperturbed and are not destroyed. Despite high potential and utility of this approach, clinical implementation has not yet been achieved due to practical hurdles like speed of data acquisition and lack of optimized computational procedures for extracting clinically actionable information rapidly. The latter problem has been addressed by developing highly efficient ways to process IR imaging data but remains one that has considerable scope for progress. Here, we summarize the major issues and provide practical considerations in implementing a modified Bayesian classification protocol for digital molecular pathology. We hope to familiarize readers with analysis methods in IR imaging data and enable researchers to develop methods that can lead to the use of this promising technique for digital diagnosis of cancer.     

Photoperiodic regulation of seasonal responses in Indian weaver bird (Ploceus philippinus)

Photoperiod (=day length) is the vital factor for the regulation of behavioral and physiological activities in many avian species. This study investigated the seasonal cycles of testicular growth and secondary sexual characteristics of Indian weaver bird under natural day length (NDL) and the effects of duration and intensity of light on photoperiodic induction. In the first experiment, groups of birds (n = 7 each) were exposed to under NDL in April 2008 and May 2009 for 8 and 12 months, respectively. In second and third experiment, birds (n = 6 each group) were exposed to different photoperiods (11.5L:12.5D, 12L:12D, 13L:11D, and 15L:9D) at the same (500 lux) light intensity, and to 13L:11D at different light intensities (10-, 50-, 500-, and 800-lux). Observations on testis size, molt, and plumage score were recorded 2-week (molt and plumage) or at 4-week intervals (testes). Both the NDL groups showed similar seasonal cycles of testicular growth-regression and secondary sexual characteristics. Second and third experiments suggest that the photoperiodic induction was depending upon duration and intensity of the light. Birds showed testicular growth-regression cycle followed by molt and plumage color change only under 13L:11D and 15L:9D and only 500- and 800-lux under 13L:11D photoperiod but not under 11.5L:12.5D and 12L:12D and 10- and 50-lux light intensities. Pre- and post-nuptial molting on body feathers were progressed with gonadal stimulation–maturation and regression cycle under 13L:11D and 15L:9D. Results under different light–dark cycles suggest that day length of about 12 h or more and above the threshold level of light intensity are essential for the induction of photoperiodic responses.     

Alterations in radiation induced cell cycle perturbations by 2-deoxy-D-glucose in human tumor cell lines

In the present studies, effects of glucose analogue, 2-deoxy-D-glucose (2-DG) on radiation-induced cell cycle perturbations were investigated in human tumor cell lines. In unirradiated cells, the levels of cyclin B1 in G2 phase were significantly higher in both the glioma cell lines as compared to squamous carcinoma cells. Upon irradiation with Co60 gamma-rays (2 Gy), the cyclin B1 levels were reduced in U87 cells, while no significant changes could be observed in other cell lines, which correlated well with the transient G2 delay observed under these conditions by the BrdU pulse chase measurements. 2-DG (5 mM, 2 hr) induced accumulation of cells in the G2 phase and a time-dependent increase in the levels of cyclin B1 in both the glioma cell lines, while significant changes could not be observed in any of the squamous carcinoma cell lines. 2-DG enhanced the cyclin B1 level further in all the cell lines following irradiation, albeit to different extents. Interestingly, an increase in the unscheduled expression of B1 levels in G1 phase 48 hr after irradiation was observed in all the cell lines investigated. 2-DG also increased the levels of cyclin D1 at 24 hr in BMG-1 cell line. These observations imply that 2-DG-induced alterations in the cell cycle progression are partly responsible for its radiomodifying effects.