The composition and dynamics of cell-substratum adhesions in locomoting fish keratocytes.
Close contacts are the predominant type of cell-substratum adhesion in rapidly moving cells yet little is known about their composition and dynamics. To address these issues we have attempted to identify the molecular components of close contacts formed in rapidly moving fish epithelial keratocytes. In addition we have utilized the simple shape of keratocytes to explore the relationship between close contact formation and rapid locomotion. Beta1 integrin and talin molecules were found to be localized within a narrow rim of very close contact along the leading edge. These molecules together with vinculin were also found within small foci distributed evenly throughout the lamella, corresponding to regions of variable close contact. Alpha-actinin was found in foci within older, more posteriorly located regions of the lamella and along stress fibers. In addition to close contacts, small focal adhesion-like structures which stained positively for all antibodies tested were found at the tips of stress fibers within retracting cell margins. Interference reflection and total internal reflection microscopy of moving keratocytes showed cell-substratum contacts to be organized into distinct patterns that appear to move forwards, in concert with the leading edge. A feature common to all cells is a rim of very close contact at the leading edge. This region is specialized for the formation of new cell-substratum adhesions and is the site where patterns of close contact are generated. We have found that cell locomotion is most rapid when a uniform contact pattern is present but cell speed is progressively reduced as the contact pattern becomes more irregular. Furthermore, the local rate of lamellar extension is most rapid when underlain by regions of intermediate closeness to the substratum, but is reduced or ceases if the underlying contact is either very close or more distant, respectively. Our results suggest that close contacts and focal adhesions are related structures formed from a common hierarchy of molecular interactions. In addition the relationship between close contact formation and lamellar extension indicates a direct coupling between these two processes at the leading edge. Furthermore, we can explain the dynamic behaviour of close contacts in terms of the relative rates of trapping and release of component molecules that is initiated at the leading edge. We suggest that regulation of the molecular dynamics involved in leading edge specialization determines both the pattern of cell-substratum contacts and the net rate of actin filament assembly.

title: Principles of locomotion for simple-shaped cells
abstract: MOVING cells display a variety of shapes and modes of locomotion1, but it is not clear how motility at the molecular level relates to the locomotion of a whole cell, a problem compounded in studies of cells with complex shapes2–5. A striking feature of fish epidermal keratocyte locomotion is its apparent simplicity6. Here we present a kinematic description of locomotion which is consistent with the semicircular shape and persistent 'gliding'; motion of fish epidermal keratocytes. We propose that extension of the front and retraction of the rear of these cells occurs perpendicularly to the cell edge, and that a graded distribution of extension and retraction rates along the cell margin maintains cell shape and size during locomotion. Evidence for this description is provided by the predicted circumferential motion of lamellar features and the curvature of 'photo-marked' lines within specific molecular components of moving keratocytes. Our description relates the dynamics of molecular assemblies to the movement of a whole cell.

title: Akt mediates self-renewal division of mouse spermatogonial stem cells
abstract: Spermatogonial stem cells have unique properties to self-renew and support spermatogenesis throughout their lifespan. Although glial cell line-derived neurotrophic factor (GDNF) has recently been identified as a self-renewal factor for spermatogonial stem cells, the molecular mechanism of spermatogonial stem cell self-renewal remains unclear. In the present study, we assessed the role of the phosphoinositide-3 kinase (PI3K)-Akt pathway using a germline stem (GS) cell culture system that allows in vitro expansion of spermatogonial stem cells. Akt was rapidly phosphorylated when GDNF was added to the GS cell culture, and the addition of a chemical inhibitor of PI3K prevented GS cell self-renewal. Furthermore, conditional activation of the myristoylated form of Akt-Mer (myr-Akt-Mer) by 4-hydroxy-tamoxifen induced logarithmic proliferation of GS cells in the absence of GDNF for at least 5 months. The myr-Akt-Mer GS cells expressed spermatogonial markers and retained androgenetic imprinting patterns. In addition, they supported spermatogenesis and generated offspring following spermatogonial transplantation into the testes of infertile recipient mice, indicating that they are functionally normal. These results demonstrate that activation of the PI3K-Akt pathway plays a central role in the self-renewal division of spermatogonial stem cells.

title: Targeted cell immobilization by ultrasound microbeam.
abstract: Various techniques exerting mechanical stress on cells have been developed to investigate cellular responses to externally controlled stimuli. Fundamental mechanotransduction processes about how applied physical forces are converted into biochemical signals have often been examined by transmitting such forces through cells and probing its pathway at cellular levels. In fact, many cellular biomechanics studies have been performed by trapping (or immobilizing) individual cells, either attached to solid substrates or suspended in liquid media. In that context, we demonstrated two-dimensional acoustic trapping, where a lipid droplet of 125 µm in diameter was directed transversely toward the focus (or the trap center) similar to that of optical tweezers. Under the influence of restoring forces created by a 30 MHz focused ultrasound beam, the trapped droplet behaved as if tethered to the focus by a linear spring. In order to apply this method to cellular manipulation in the Mie regime (cell diameter > wavelength), the availability of sound beams with its beamwidth approaching cell size is crucial. This can only be achieved at a frequency higher than 100 MHz. We define ultrasound beams in the frequency range from 100 MHz to a few GHz as ultrasound microbeams because the lateral beamwidth at the focus would be in the micron range. Hence a zinc oxide (ZnO) transducer that was designed and fabricated to transmit a 200 MHz focused sound beam was employed to immobilize a 10 µm human leukemia cell (K-562) within the trap. The cell was laterally displaced with respect to the trap center by mechanically translating the transducer over the focal plane. Both lateral displacement and position trajectory of the trapped cell were probed in a two-dimensional space, indicating that the retracting motion of these cells was similar to that of the lipid droplets at 30 MHz. The potential of this tool for studying cellular adhesion between white blood cells and endothelial cells was discussed, suggesting its capability as a single cell manipulator.

title: Multidrug-resistant tuberculosis in a lung transplant recipient.
abstract: Tuberculosis infection has been a relatively rare complication after lung transplantation. However, as more countries in which Mycobacterium tuberculosis infection remains endemic embark on lung transplant programs, the occurrence of multidrug-resistant tuberculosis after transplantation is a genuine threat. We report the first case of multidrug-resistant tuberculosis in a double-lung transplant recipient who probably acquired the disease from the donor. We discuss the problems in clinical management of post-transplant tuberculosis infection and of drug-resistance.

title: Respiration-induced B0 field fluctuation compensation in balanced SSFP: real-time approach for transition-band SSFP fMRI.
abstract: In functional MRI (fMRI) the resonance frequency shift induced from respiration is a major source of physiological noise. In transition-band SSFP fMRI the respiration-induced resonance offset not only increases noise interference, it also shifts the activation band. This leads to a reduction in the contrast-to-noise ratio (CNR) and the potential for varying contrast levels during the experiment. A novel real-time method that compensates for the respiration-induced resonance offset frequency is presented. This method utilizes free induction decay (FID) phase information to measure the resonance offset. For compensation, one can update the resonant frequency in real time by changing the transmit RF pulse and receiver phases to track the measured offset. The results show decreased signal power in the respiration frequency band and increased numbers of activated voxels with higher Z-scores compared to uncompensated experiments.

title: Quantitative measures of facial expression for patients with head and neck cancer
abstract: In addition to changes in facial morphology, head and neck cancer treatment can impact the facial expression. Quantification of changes in facial expression, particularly of smiling, would enable a deeper understanding of the relationship between physical changes and psychosocial adjustment in patients being treated for facial cancer. This study proposes 48 quantitative facial expression measures, which consist of the length normalized distances and slopes between 27 manually annotated fiducial points on standard 2D clinical photographs of patients with head and neck cancer. Using the proposed measures, the maximum intensity of smiling for patients with head and neck cancer prior to their treatments was compared to that of a sample from a healthy population. A total of 7 facial expression measures captured statistically significant differences between patients with head and neck cancer and healthy individuals. These measures suggest that patients with head and neck cancer are less expressive than people without head and neck cancer. This study is the first attempt to quantify the facial expression of patients with head and neck cancer and to build a foundation for studying how surgical interventions may affect their facial expressions. Moreover, this study lays the groundwork for future investigation of the relationship between facial expression and psychosocial adjustment in cancer patients.