Molecular dynamics simulations of evolved collective motions of atoms in the myosin motor domain upon perturbation of the ATPase pocket

A crucial point for mechanical force generation in actomyosin systems is how the energy released by ATP hydrolysis in the myosin motor domain gives rise to the movement of the myosin head along the actin filament. We assumed the signal of the ATP hydrolysis to be transmitted as modulated atomic vibrations from the nucleotide-binding site throughout the myosin head, and carried out 1-ns all-atom molecular dynamics simulations for that signal transmission. We distributed the released energy to atoms located around the ATPase pocket as kinetic energies and examined how the effect of disturbance extended throughout the motor domain. The result showed that the disturbance signal extended over the motor domain in 150 ps and induced slowly varying collective motions of atoms at the actin-binding site and the junction with the neck, both of which are relevant to the movement of the myosin head along the actin filament. We also performed a principal component analysis of thermal atomic motions for the motor domain, and the first principal component was consistent with the response to the disturbance given to the ATPase pocket.     

Morphological examination during in vitro cartilage formation by human mesenchymal stem cells

The formation of the skeleton through endochondral ossification is one of the most complex processes in development. One approach to resolving this complexity is to examine simplified systems. In vitro cartilage formation by mesenchymal stem cells (MSCs) is observed when the cells are cultured as a micromass. Several studies have confirmed the molecular events, showing the usefulness of these cells as a differentiation model. We have elucidated the process of cartilage formation in MSCs from the morphological point of view by light and transmission electron microscopy and immunohistochemical examination. The morphology of the MSCs changed from spherical to spindle-shaped, and the cells aggregated and formed junctional complexes during Day 1. At Day 7, three layers were observed. The superficial zone consisted of several layers of elongated cells with junctional complexes. The middle zone was composed of apoptotic bodies, and the deep zone was occupied by chondrocyte-like cells excreting extracellular matrices. At Day 14, the middle zone had disappeared, and the chondrocyte-like cells in the deep zone were detected within cartilage lacuna. They were covered by cartilage matrices containing collagen types I, II, and X and chondroitin sulfate. By Day 21, the outer layer consisting of spindle-shaped cells had disappeared in places. As the pellet grew, the outer layer seemed to be unable to stretch to maintain a constant covering around the pellet. Our findings have thus revealed that MSCs change their morphology depending upon their microenvironment during differentiation. In vitro cartilage formation by MSCs makes it possible to clarify the detailed morphological events that occur during chondrogenesis. S. Ichinose and I. Sekiya contributed equally to this study     

Prediction of circulatory equilibrium in response to changes in stressed blood volume

Accurate prediction of cardiac output (CO), left atrial pressure (PLA), and right atrial pressure (PRA) is a prerequisite for management of patients with compromised hemodynamics. In our previous study (Uemura et al. Am J Physiol Heart Circ Physiol 286: H2376-H2385, 2004), we demonstrated a circulatory equilibrium framework, which permits the prediction of CO, PLA, and PRA once the venous return surface and integrated CO curve are known. Inasmuch as we also showed that the surface can be estimated from single-point CO, PLA, and PRA measurements, we hypothesized that a similar single-point estimation of the CO curve would enable us to predict hemodynamics. In seven dogs, we measured the PLA-CO and PRA-CO relations and derived a standardized CO curve using the logarithmic function CO = SL[ln(PLA - 2.03) + 0.80] for the left heart and CO = SR[ln(PRA - 2.13) + 1.90] for the right heart, where SL and SR represent the preload sensitivity of CO, i.e., pumping ability, of the left and right heart, respectively. To estimate the integrated CO curve in each animal, we calculated SL and SR from single-point CO, PLA, and PRA measurements. Estimated and measured CO agreed reasonably well. In another eight dogs, we altered stressed blood volume (-8 to +8 ml/kg of reference volume) under normal and heart failure conditions and predicted the hemodynamics by intersecting the surface and the CO curve thus estimated. We could predict CO [y = 0.93x + 6.5, r2 = 0.96, standard error of estimate (SEE) = 7.5 ml.min(-1).kg(-1)], PLA (y = 0.90x + 0.5, r2= 0.93, SEE = 1.4 mmHg), and PRA (y = 0.87x + 0.4, r2= 0.91, SEE = 0.4 mmHg) reasonably well. In conclusion, single-point estimation of the integrated CO curve enables accurate prediction of hemodynamics in response to extensive changes in stressed blood volume.     

Temporary blockade of contractility during reperfusion elicits a cardioprotective effect of the p38 MAP kinase inhibitor SB-203580

p38 MAP kinase activation is known to be deleterious not only to mitochondria but also to contractile function. Therefore, p38 MAP kinase inhibition therapy represents a promising approach in preventing reperfusion injury in the heart. However, reversal of p38 MAP kinase-mediated contractile dysfunction may disrupt the fragile sarcolemma of ischemic-reperfused myocytes. We, therefore, hypothesized that the beneficial effect of p38 MAP kinase inhibition during reperfusion can be enhanced when contractility is simultaneously blocked. Isolated and perfused rat hearts were paced at 330 rpm and subjected to 20 min of ischemia followed by reperfusion. p38 MAP kinase was activated after ischemia and early during reperfusion (<30 min). Treatment with the p38 MAP kinase inhibitor SB-203580 (10 microM) for 30 min during reperfusion, but not the c-Jun NH(2)-terminal kinase inhibitor SP-600125 (10 microM), improved contractility but increased creatine kinase release and infarct size. Cotreatment with SB-203580 and the contractile blocker 2,3-butanedione monoxime (BDM, 20 mM) or the ultra-short-acting beta-blocker esmorol (0.15 mM) for the first 30 min during reperfusion significantly reduced creatine kinase release and infarct size. In vitro mitochondrial ATP generation and myocardial ATP content were significantly increased in the heart cotreated with SB-203580 and BDM during reperfusion. Dystrophin was translocated from the sarcolemma during ischemia and reperfusion. SB-203580 increased accumulation of Evans blue dye in myocytes depleted of sarcolemmal dystrophin during reperfusion, whereas cotreatment with BDM facilitated restoration of sarcolemmal dystrophin and mitigated sarcolemmal damage after withdrawal of BDM. These results suggest that treatment with SB-203580 during reperfusion aggravates myocyte necrosis but concomitant blockade of contractile force unmasks cardioprotective effects of SB-203580.     

Transgenic prolactin-/- mice: effect of trauma-hemorrhage on splenocyte functions

Prolactin (PRL) is involved in the regulation of immune functions under normal and pathological conditions. Trauma-hemorrhage (T-H) produces profound immunosuppression in male mice but not in proestrus female mice. Administration of PRL in males after T-H, however, restores immune functions. In this study, PRL^+/+ and transgenic (PRL^–/–) male and female mice were used to assess immune suppression after T-H and to determine the reasons for the hormone's beneficial effect. In vitro lymphoproliferation assay with Nb2 cells showed complete absence of PRL in the circulation of the transgenic PRL^–/– mice of both sexes, whereas very high levels of the hormone were detected in the wild-type PRL^+/+ mice of both sexes. Moreover, T-H resulted in the appearance of significant levels of the hormone in circulation, but only in PRL^+/+ mice. Splenocyte proliferation in male PRL^–/– mice was significantly lower than in PRL^+/+ mice after T-H. Marginal differences between PRL^+/+ and PRL^–/– mice were observed in the release of IL-2 and IFN- by splenocytes, while the release of IL-10 was significantly higher in PRL^–/– than in PRL^+/+ mice. A significant observation of our study is the release of a 25-kDa protein in the concanavalin A-stimulated splenocytes of male PRL^+/+ and PRL^–/– mice that was active in the in vitro lymphoproliferation assay with Nb2 cells. It is unlikely that this protein is PRL because it is also present in the splenocyte extracts of PRL^–/– transgenic mice. Nonetheless, because control of lymphoid cell proliferation is considered one of the characteristics of the immune system, the local release of this protein may be significant in the differences observed in splenocyte cytokine release after T-H in wild-type as well as transgenic mice. Nb2 cells; cytokines; immune functions     

Suppression of thymic lymphoma induction by life-long low-dose-rate irradiation accompanied by immune activation in C57BL/6 mice

The induction of thymic lymphomas by whole-body X irradiation with four doses of 1.8 Gy (total dose: 7.2 Gy) in C57BL/6 mice was suppressed from a high frequency (90%) to 63% by preirradiation with 0.075 Gy X rays given 6 h before each 1.8-Gy irradiation. This level was further suppressed to 43% by continuous whole-body irradiation with 137Cs gamma rays at a low dose rate of 1.2 mGy/h for 450 days, starting 35 days before the challenging irradiation. Continuous irradiation at 1.2 mGy/h resulting in a total dose of 7.2 Gy over 258 days yielded no thymic lymphomas, indicating that this low-dose-rate radiation does not induce these tumors. Further continuous irradiation up to 450 days (total dose: 12.6 Gy) produced no tumors. Continuously irradiated mice showed no loss of hair and a greater body weight than unirradiated controls. Immune activities of the mice, as measured by the numbers of CD4+ T cells, CD40+ B cells, and antibody-producing cells in the spleen after immunization with sheep red blood cells, were significantly increased by continuous 1.2-mGy/h irradiation alone. These results indicate the presence of an adaptive response in tumor induction, the involvement of radiation-induced immune activation in tumor suppression, and a large dose and dose-rate effectiveness factor (DDREF) for tumor induction with extremely low-dose-rate radiation.     

A modified error backpropagation algorithm for complex-value neural networks

The complex-valued backpropagation algorithm has been widely used in fields of dealing with telecommunications, speech recognition and image processing with Fourier transformation. However, the local minima problem usually occurs in the process of learning. To solve this problem and to speed up the learning process, we propose a modified error function by adding a term to the conventional error function, which is corresponding to the hidden layer error. The simulation results show that the proposed algorithm is capable of preventing the learning from sticking into the local minima and of speeding up the learning.