Effects of Isolated and Combined Exposure of the Brain and Lungs to a Laser-Induced Shock Wave(s) on Physiological and Neurological Responses in Rats.

Blast-induced traumatic brain injury (bTBI) has been suggested to be caused by direct head exposure and by torso exposure to a shock wave (thoracic hypotheses). It is unclear, however, how torso exposure affects the brain in real time. This study applied a mild-impulse laser-induced shock wave(s) (LISW[s]) only to the brain (Group 1), lungs (Group 2), or to the brain and lungs (Group 3) in rats. Because LISWs are unaccompanied by a dynamic pressure in principle, the effects of acceleration can be excluded, allowing analysis of the pure primary mechanism (the effects of a shock wave). For all rat groups, real-time monitoring of the brain and systemic responses were conducted for up to 1 h post-exposure and motor function assessments for up to seven days post-exposure. As reported previously, brain exposure alone caused cortical spreading depolarization (CSD), followed by long-lasting hypoxemia and oligemia in the cortices (Group 1). It was found that even LISW application only to the lungs caused prolonged hypoxemia and mitochondrial dysfunction in the cortices (Group 2). Importantly, features of CSD and mitochondrial dysfunction were significantly exacerbated by combined exposure (Group 3) compared with those caused by brain exposure alone (Group 1). Motor dysfunction was observed in all exposure groups, but their time courses differed depending on the groups. Rats with brain exposure alone exhibited the most evident motor dysfunction at one day post-exposure, and after that, it did not change much for up to seven days post-exposure. Alternatively, two groups of rats with lung exposure (Group 2 and Group 3) exhibited continuously aggravated motor functions for up to seven days post-exposure, suggesting different mechanisms for motor dysfunction caused by brain exposure and that caused by lung exposure. As for the reported thoracic hypotheses, our observations seem to support the volumetric blood surge and vagovagal reflex. Overall, the results of this study indicate the importance of the torso guard to protect the brain and its function.

Evaluation of Cerebral Hemodynamics and Tissue Morphology of In Vivo Rat Brain Using Spectral Diffuse Reflectance Imaging.

We investigated a quantitative imaging of reduced scattering coefficients µs'( λ) and the absorption coefficients µa( λ) of in vivo cortical tissues in the range from visible to near-infrared (NIR) wavelengths based on diffuse reflectance spectral imaging technique. In this method, diffuse reflectance images of in vivo cortical tissue are acquired at nine wavelengths (500, 520, 540, 560, 570, 580, 600, 730, and 760 nm). A multiple regression analysis aided by the Monte Carlo simulation for the absorbance spectra is then utilized to estimate the optical coefficients of cortical tissue. This analysis calculates the concentration of oxygenated hemoglobin and that of deoxygenated hemoglobin, the scattering amplitude a and the scattering power b. The spectrum of absorption coefficient is deduced from the estimated concentrations of oxygenated hemoglobin and deoxygenated hemoglobin. The spectrum of reduced scattering coefficient is determined by the estimated scattering amplitude and scattering power. The particle size distribution of microstructure is calculated from the estimated scattering power b for evaluating the morphological change in brain tissue quantitatively. Animal experiments with in vivo exposed brain of rats demonstrated that the responses of the absorption properties to hyperoxic and anoxic conditions are in agreement with the expected well-known cortical hemodynamics. The average particle size was significantly reduced immediately after the onset of anoxia and then it was changed into an increase, which implied the swelling and shrinkage of the cellular and subcellular structures induced by loss of tissue viability in brain tissue.

Inhibition of Rho-associated kinases disturbs the collective cell migration of stratified TE-10 cells.

BACKGROUND: The collective cell migration of stratified epithelial cells is considered to be an important phenomenon in wound healing, development, and cancer invasion; however, little is known about the mechanisms involved. Furthermore, whereas Rho family proteins, including RhoA, play important roles in cell migration, the exact role of Rho-associated coiled coil-containing protein kinases (ROCKs) in cell migration is controversial and might be cell-type dependent. Here, we report the development of a novel modified scratch assay that was used to observe the collective cell migration of stratified TE-10 cells derived from a human esophageal cancer specimen. RESULTS: Desmosomes were found between the TE-10 cells and microvilli of the surface of the cell sheet. The leading edge of cells in the cell sheet formed a simple layer and moved forward regularly; these rows were followed by the stratified epithelium. ROCK inhibitors and ROCK small interfering RNAs (siRNAs) disturbed not only the collective migration of the leading edge of this cell sheet, but also the stratified layer in the rear. In contrast, RhoA siRNA treatment resulted in more rapid migration of the leading rows and disturbed movement of the stratified portion. CONCLUSIONS: The data presented in this study suggest that ROCKs play an important role in mediating the collective migration of TE-10 cell sheets. In addition, differences between the effects of siRNAs targeting either RhoA or ROCKs suggested that distinct mechanisms regulate the collective cell migration in the simple epithelium of the wound edge versus the stratified layer of the epithelium.

A magnetic anti-cancer compound for magnet-guided delivery and magnetic resonance imaging.

Research on controlled drug delivery for cancer chemotherapy has focused mainly on ways to deliver existing anti-cancer drug compounds to specified targets, e.g., by conjugating them with magnetic particles or encapsulating them in micelles. Here, we show that an iron-salen, i.e., µ-oxo N,N'- bis(salicylidene)ethylenediamine iron (Fe(Salen)), but not other metal salen derivatives, intrinsically exhibits both magnetic character and anti-cancer activity. X-Ray crystallographic analysis and first principles calculations based on the measured structure support this. It promoted apoptosis of various cancer cell lines, likely, via production of reactive oxygen species. In mouse leg tumor and tail melanoma models, Fe(Salen) delivery with magnet caused a robust decrease in tumor size, and the accumulation of Fe(Salen) was visualized by magnetic resonance imaging. Fe(Salen) is an anti-cancer compound with magnetic property, which is suitable for drug delivery and imaging. We believe such magnetic anti-cancer drugs have the potential to greatly advance cancer chemotherapy for new theranostics and drug-delivery strategies.

In vivo estimation of light scattering and absorption properties of rat brain using a single-reflectance fiber probe during cortical spreading depression.

Diffuse reflectance spectroscopy using a fiber optic probe is a promising technique for evaluating the optical properties of biological tissue. We herein present a method for determining the reduced scattering coefficient, µ's, the absorption coefficient, µa, and the tissue oxygen saturation, StO2, of in vivo brain tissue using a single-reflectance fiber probe with two source-collector geometries. We performed in vivo recordings of diffuse reflectance spectra and of the electrophysiological signals for exposed rat brain during the cortical spreading depression evoked by the topical application of KCl. The time courses of µa at 500, 570, and 584 nm indicated the hemodynamic change in the cerebral cortex as well as StO2. At 570 nm, the time course of µ's was well correlated with that of µa, which also reflects the scattering by RBCs. On the other hand, increases in µ's at 500 and 584 nm and a decrease in µ's at 800 nm were observed before the profound increase in µa, and these occurrences were synchronized with the negative dc shift of the local field potential. The resultant change in the slope of µ's ðλÞ is indicative of the morphological changes in the cellular and subcellular structures induced by the depolarization due to the temporal depression of the neuronal bioelectrical activity. The results of the present study indicate the potential application of the proposed method in evaluating the pathophysiological conditions of in vivo brain.

Multispectral imaging of absorption and scattering properties of in vivo exposed rat brain using a digital red-green-blue camera.

In order to estimate multispectral images of the absorption and scattering properties in the cerebral cortex of in vivo rat brain, we investigated spectral reflectance images estimated by the Wiener estimation method using a digital RGB camera. A Monte Carlo simulation-based multiple regression analysis for the corresponding spectral absorbance images at nine wavelengths (500, 520, 540, 560, 570, 580, 600, 730, and 760 nm) was then used to specify the absorption and scattering parameters of brain tissue. In this analysis, the concentrations of oxygenated hemoglobin and that of deoxygenated hemoglobin were estimated as the absorption parameters, whereas the coefficient a and the exponent b of the reduced scattering coefficient spectrum approximated by a power law function were estimated as the scattering parameters. The spectra of absorption and reduced scattering coefficients were reconstructed from the absorption and scattering parameters, and the spectral images of absorption and reduced scattering coefficients were then estimated. In order to confirm the feasibility of this method, we performed in vivo experiments on exposed rat brain. The estimated images of the absorption coefficients were dominated by the spectral characteristics of hemoglobin. The estimated spectral images of the reduced scattering coefficients had a broad scattering spectrum, exhibiting a larger magnitude at shorter wavelengths, corresponding to the typical spectrum of brain tissue published in the literature. The changes in the estimated absorption and scattering parameters during normoxia, hyperoxia, and anoxia indicate the potential applicability of the method by which to evaluate the pathophysiological conditions of in vivo brain due to the loss of tissue viability.

Inhibition of Rho-associated kinases disturbs the collective cell migration of stratified TE-10 cells

BACKGROUND: The collective cell migration of stratified epithelial cells is considered to be an important phenomenon in wound healing, development, and cancer invasion; however, little is known about the mechanisms involved. Furthermore, whereas Rho family proteins, including RhoA, play important roles in cell migration, the exact role of Rho-associated coiled coil-containing protein kinases (ROCKs) in cell migration is controversial and might be cell-type dependent. Here, we report the development of a novel modified scratch assay that was used to observe the collective cell migration of stratified TE-10 cells derived from a human esophageal cancer specimen. RESULTS: Desmosomes were found between the TE-10 cells and microvilli of the surface of the cell sheet. The leading edge of cells in the cell sheet formed a simple layer and moved forward regularly; these rows were followed by the stratified epithelium. ROCK inhibitors and ROCK small interfering RNAs (siRNAs) disturbed not only the collective migration of the leading edge of this cell sheet, but also the stratified layer in the rear. In contrast, RhoA siRNA treatment resulted in more rapid migration of the leading rows and disturbed movement of the stratified portion. CONCLUSIONS: The data presented in this study suggest that ROCKs play an important role in mediating the collective migration of TE-10 cell sheets. In addition, differences between the effects of siRNAs targeting either RhoA or ROCKs suggested that distinct mechanisms regulate the collective cell migration in the simple epithelium of the wound edge versus the stratified layer of the epithelium.

Diesel NO(x) aftertreatment by combined process using temperature swing adsorption, NO(x) reduction by nonthermal plasma, and NO(x) recirculation: improvement of the recirculation process.

NO(x) emitted from a stationary diesel engine generator was treated with a hybrid system comprising NO(x) reduction by nonthermal plasma (NTP) and temperature swing adsorption (TSA) driven by engine waste heat. TSA produces a low-volume gas mixture of N(2) and highly concentrated NO(x), which is effectively reduced by NTP treatment. Improved treatment performance and efficiency are achieved by re-injecting the NTP-treated gas mixture into the engine intake. The system comprises two switchable adsorption chambers; the operation of this system was simulated by using a one-chamber system. The maximum energy efficiency for NO(x) treatment is 200 g(NO(2))/kWh. The respective contributions of NTP and injection of N(2) and NO(x) to the performance were theoretically analyzed. The analysis predicts that high energy efficiency and high NO(x)-removal efficiency can be simultaneously achieved with this system but miniaturization of the adsorption chambers will be a challenge.

Plasmodium induced by SU6656, an Src family kinase inhibitor, is accompanied by a contractile ring defect.

We have shown that SU6656, a potent Src family kinase inhibitor, has the ability to induce multinucleation at a high frequency in diverse cells: rat skin fibroblasts, bone marrow adherent cells, 5F9A mesenchymal stem cell-like clones, 2C5 tracheal epithelial cells and MDCK epithelial cells from dog kidney. To gain insight into the mechanism of multinucleation, we observed the process by time-lapse and confocal microscopy. These multinuclei generally seem to exist independently in one cell without any connections with each other. By time-lapse microscopy, multinucleated cells were found to be formed through the mechanism of plasmodium: karyokinesis without cytokinesis. The observation of EGFP-actin transfected cells by time-lapse confocal laser scanning microscopy suggested that plasmodium occurred with deficient contractile ring formation. Although we examined the differentiation of these cells, the multinucleated cells could not be categorized into any type of cell in vivo known to exhibit multinuclei.

Changes in optical properties of rat cerebral cortical slices during oxygen glucose deprivation.

We simultaneously measured the diffuse reflectance spectra and transmittance spectra of in vitro rat cerebral cortical tissue slices perfused with artificial cerebrospinal fluid (aCSF) in the wavelength range from 500 to 900 nm. An ischemia-like condition in the cortical tissue was induced by oxygen/glucose deprivation (OGD) of the aCSF. Diffuse reflectance and transmittance of the cortical slices were decreased and increased, respectively, during OGD. Spectral data of reduced scattering coefficients and absorption coefficients were estimated by the inverse Monte Carlo simulation for light transport in tissue. As with OGD, significant decrease of the reduced scattering coefficients and alteration of the absorption coefficient spectrum were observed over the measured wavelength range. The mean maximum amplitudes of change in the absorption coefficient at 520, 550, 605, and 830 nm were 0.33 ± 0.14, 0.30 ± 0.12, 0.30 ± 0.14, and -0.04 ± 0.16, respectively, whereas those in the reduced scattering coefficient at 520, 550, 605, and 830 nm were -0.37 ± 0.08, -0.38 ± 0.08, -0.38 ± 0.08, and -0.39 ± 0.08. Variations in the reduced scattering coefficients implied cell deformation mainly due to cell swelling, whereas those in the absorption spectra indicated reductions in heme aa(3) and CuA in cytochrome c oxidase and cytochrome c.

Oligopotent mesenchymal stem cell-like clone becomes multinucleated following phorbol ester, TPA stimulation.

We established a mesenchymal stem cell clone, 5F9A, from rat bone marrow substrate adherent cells by repeated limiting dilutions. The cells have a fibroblastic shape and form intimate contacts with adjacent cells with interdigitations and junctions similar to adherence and tight junctions in a semi-confluent culture. Analysis of the phenotypes of these cells by RT-PCR and FACS demonstrated that they resembled mesenchymal stem cells, and the cells could differentiate into adiopocytes and osteoblasts under appropriate conditions in vitro showing their oligopotency. Furthermore, the cells were induced to become multinuclear cells by TPA (12-o-tetradecanoylphorbol 13-acetate) stimulation.

TPA-induced multinucleation of a mesenchymal stem cell-like clone is mediated primarily by karyokinesis without cytokinesis, although cell-cell fusion also occurs.

The 5F9A cell, which is a mesenchymal stem cell-like clone established from rat bone marrow substrate adherent cells, can differentiate into adipocytes and osteoblasts in vitro under the appropriate conditions. Multinucleated cells could be also induced by 12-O-tetradecanoylphorbol 13-acetate (TPA) in 5F9A cells. This effect was mediated by protein kinase C. Possible mechanisms of multinucleation by TPA were hypothesized to be either karyokinesis without cytokinesis or cell-cell fusion. By observation using time-lapse phase-contrast microscopy, we determined that the multinucleated cells were generated mainly by karyokinesis without cytokinesis. Cell fusion was studied using time-lapse photography, and confocal laser scanning microscopy using two differentially labeled cells. These techniques demonstrated that multinucleated 5F9A cells could be produced by cell fusion, albeit at a low frequency. We conclude that multinucleated 5F9A cells are formed primarily by karyokinesis without cytokinesis, although some cells are also formed by cell-cell fusion.

Nicotinic environment affects the differentiation and functional maturation of monocytes derived dendritic cells (DCs).

Differentiation of tissue monocytes into DCs is a critical phase in the development of a competent immune system. We show that in a nicotinic environment, while human monocytes differentiate into DCs (henceforth called nicDCs) with a typical morphology, they display unique phenotype and cytokine profile that adversely affect their function. Despite an increased capacity for receptor-dependent antigen uptake, nicDCs do not express CD1a and fail to fully up-regulate MHCs, molecules essential for their antigen-presenting function. Additionally, in response to bacterial antigen LPS, maturing nicDCs hardly express the chemotactic cytokine receptor 7 required for their entry into lymphatic vessels. Furthermore, in parallel with their differential expression of costimulatory molecules CD80 and CD86 and lack of IL-12, nicDCs display profoundly reduced Th1 promoting capacity. These findings thus indicate that nicotine impedes the development of cell-mediated immunity by skewing DC differentiation. These effects of nicotinic environment on DC differentiation may contribute to the increased risks of respiratory tract infection and various cancers in smokers.