Cystic Fibrosis Transport Regulator and its mRNA are Expressed in Human Epidermis

Cystic fibrosis transport regulator is a cAMP-dependent chloride channel protein. Normal (non cystic fibrosis) human epidermis stained positive for cystic fibrosis transport regulator as densely as did the eccrine sweat gland when three monoclonal antibodies for R (regulatory) and C (C-terminus) domains of cystic fibrosis transport regulator were used. All the layers of the epidermis took up staining uniformly. A peptide for C-epitope completely blocked the staining with monoclonal antibodies for C. Nested reverse transcription polymerase chain reaction of freshly isolated human epidermal fragments and the eccrine sweat glands amplified the cystic fibrosis transport regulator mRNA sequence derived from exons 13 and 14 to comparable extents. The 526 base pair antisense, but not sense, RNA probe derived from exons 10-13 stained cystic fibrosis transport regulator mRNA in both the epidermis and the sweat gland to a similar extent. In the epidermis, the cytoplasm of basal cells, stratum spinosum cells, and granular layer cells were all stained uniformly, but not corneocytes in the stratum corneum. In the sweat secretory coils, both clear and dark cells were stained but not the myoepithelium, with the dark cells staining more densely than the clear cells as in a previous study. In the duct, both luminal and basal ductal cells took up cystic fibrosis transport regulator staining uniformly but luminal cytoplasm of luminal ductal cells was devoid of cystic fibrosis transport regulator mRNA. Although the function of cystic fibrosis transport regulator in the epidermis is totally unknown, its recently proposed role as a universal regulator of a variety of cellular and membrane functions necessitates further studies on its regulation and function in health and disease

Cystic Fibrosis Transport Regulator and its mRNA are Expressed in Human Epidermis

Cystic fibrosis transport regulator is a cAMP-dependent chloride channel protein. Normal (non cystic fibrosis) human epidermis stained positive for cystic fibrosis transport regulator as densely as did the eccrine sweat gland when three monoclonal antibodies for R (regulatory) and C (C-terminus) domains of cystic fibrosis transport regulator were used. All the layers of the epidermis took up staining uniformly. A peptide for C-epitope completely blocked the staining with monoclonal antibodies for C. Nested reverse transcription polymerase chain reaction of freshly isolated human epidermal fragments and the eccrine sweat glands amplified the cystic fibrosis transport regulator mRNA sequence derived from exons 13 and 14 to comparable extents. The 526 base pair antisense, but not sense, RNA probe derived from exons 10-13 stained cystic fibrosis transport regulator mRNA in both the epidermis and the sweat gland to a similar extent. In the epidermis, the cytoplasm of basal cells, stratum spinosum cells, and granular layer cells were all stained uniformly, but not corneocytes in the stratum corneum. In the sweat secretory coils, both clear and dark cells were stained but not the myoepithelium, with the dark cells staining more densely than the clear cells as in a previous study. In the duct, both luminal and basal ductal cells took up cystic fibrosis transport regulator staining uniformly but luminal cytoplasm of luminal ductal cells was devoid of cystic fibrosis transport regulator mRNA. Although the function of cystic fibrosis transport regulator in the epidermis is totally unknown, its recently proposed role as a universal regulator of a variety of cellular and membrane functions necessitates further studies on its regulation and function in health and disease

The Effect of Seat Back Inclination on Spinal Alignment in Automotive Seating Postures

Experimental studies have demonstrated a relationship between spinal injury severity and vertebral kinematics, influenced by the initial spinal alignment of automotive occupants. Spinal alignment has been considered one of the possible causes of gender differences in the risk of sustaining spinal injuries. To predict vertebral kinematics and investigate spinal injury mechanisms, including gender-related mechanisms, under different seat back inclinations, it is needed to investigate the effect of the seat back inclination on initial spinal alignment in automotive seating postures for both men and women. The purpose of this study was to investigate the effect of the seat back inclination on spinal alignments, comparing spinal alignments of automotive seating postures in the 20\ub0 and 25\ub0 seat back angle and standing and supine postures. The spinal columns of 11 female and 12 male volunteers in automotive seating, standing, and supine postures were scanned in an upright open magnetic resonance imaging system. Patterns of their spinal alignments were analyzed using Multidimensional Scaling presented in a distribution map. Spinal segmental angles (cervical curvature, T1 slope, total thoracic kyphosis, upper thoracic kyphosis, lower thoracic kyphosis, lumbar lordosis, and sacral slope) were also measured using the imaging data. In the maximum individual variances in spinal alignment, a relationship between the cervical and thoracic spinal alignment was found in multidimensional scaling analyses. Subjects with a more lordotic cervical spine had a pronounced kyphotic thoracic spine, whereas subjects with a straighter to kyphotic cervical spine had a less kyphotic thoracic spine. When categorizing spinal alignments into two groups based on the spinal segmental angle of cervical curvature, spinal alignments with a lordotic cervical spine showed significantly greater absolute average values of T1 slope, total thoracic kyphosis, and lower thoracic kyphosis for both the 20\ub0 and 25\ub0 seat back angles. For automotive seating postures, the gender difference in spinal alignment was almost straight cervical and less-kyphotic thoracic spine for the female subjects and lordotic cervical and more pronounced kyphotic thoracic spine for the male subjects. The most prominent influence of seatback inclination appeared in Total thoracic kyphosis, with increased angles for 25\ub0 seat back, 8.0\ub0 greater in spinal alignments with a lordotic cervical spine, 3.2\ub0 greater in spinal alignments with a kyphotic cervical spine. The difference in total thoracic kyphosis between the two seatback angles and between the seating posture with the 20\ub0 seat back angle and the standing posture was greater for spinal alignments with a lordotic cervical spine than for spinal alignments with a kyphotic cervical spine. The female subjects in this study had a tendency toward the kyphotic cervical spine. Some of the differences between average gender-specific spinal alignments may be explained by the findings observed in the differences between spinal alignments with a lordotic and kyphotic cervical spine

Female and Male Whole Spinal Alignment and Cervical Kinematic Responses in Rear Impacts

The susceptibility of women to Whiplash Associated Disorders (WADs) has been the focus of numerous epidemiologic studies. Summarising the epidemiologic WAD studies, women were found to be at three times higher risk of sustaining WADs than men. Analysis of insurance claims records indicate that certain whiplash protection seats have reduced the risk of sustaining WADs more effectively for men than for women. However, many aspects of WADs are still unknown, including what role gender differences play in the risk of sustaining WADs. In order to obtain fundamental knowledge to understand the gender difference involved in the risk of sustaining WADs, this thesis reanalysed previous rear impact sled test series comprising female and male volunteers to clarify the dynamic characteristics of inertia-induced cervical vertebral kinematics during rear impacts for women and men. Furthermore, cervical spinal alignment has been suggested as one of several possible causes of the gender differences seen in the risk of sustaining WADs. In addition, it has been reported that the initial position of the thoracolumbar spine against a seatback affects vertebral kinematics as well as the cervical spine. Therefore, this study also investigated whole spinal alignments in one automotive seated posture using an upright open MRI system for both genders, and estimated average gender specific spinal alignment patterns. During rear impacts, the female subjects presented with a more pronounced S-shape in the cervical spine than the male subjects, beyond the voluntary muscle-induced cervical kinematics range for female subjects. In contrast, for the male subjects, the peak S-shape appeared within the voluntary muscle-induced cervical kinematics range. The estimated average spinal alignment pattern in the automotive seated posture was slight kyphotic, or almost straight cervical spine with less-kyphotic thoracic spine for the female subjects, and lordotic cervical spine with more pronounced kyphotic thoracic spine for the male subjects. The findings support previous studies which have indicated influences of cervical spinal alignment on cervical vertebral kinematics. Potential impacts of any gender differences in whole spinal alignment on cervical vertebral kinematics can be investigated with a whole-body human finite element model in future work based on this thesis

Does spinal alignment influence car occupant responses? -The influence of variation in whole spinal alignment patterns on vertebral kinematics under rear impact conditions-

The susceptibility of women to Whiplash Associated Disorders (WADs) has been the focus of numerous epidemiologic studies. Summarising the epidemiologic WAD studies, women were found to be subject to up to three times higher risk of sustaining WADs than men. To improve occupant safety for both men and women, the overall aim of this thesis was to provide fundamental knowledge to understand how gender differences influence the injury mechanisms and risk of sustaining WADs. WADs are generally considered to be derived from cervical soft tissue damage caused by excessive cervical vertebral kinematics. This thesis has focused on the whole spinal alignment in automotive seated postures as one of the gender differences, and investigated its potential impact on cervical vertebral kinematics during a rear impact, clarifying gender specific dynamic characteristics of cervical vertebral kinematics.Previous rear impact sled test series comprising female and male volunteers were reanalysed to determine the dynamic characteristics of inertia-induced cervical vertebral kinematics during rear impacts. For spinal alignment, image data of the spinal column in automotive seated postures, acquired with an upright open Magnetic Resonance Imaging (MRI) system, were analysed. Typical patterns of the whole spinal alignment, including average gender specific spinal alignment patterns, were obtained through Multi-Dimensional Scaling (MDS). Implementing these typical spinal alignment patterns in a whole-body occupant FE model, the potential impact of whole spinal alignment on cervical vertebral kinematicswere investigated in reconstruction simulations of previous rear impact sled tests.In the sled tests, the female subjects were subjected to a more pronounced cervical S-shape than the male subjects, beyond the voluntary muscle-induced cervical kinematics range for female subjects. The average gender specific spinal alignment patterns of the automotive seated posture included a slight kyphotic, or almost straight cervical spine, with a less-kyphotic thoracic spine for the female subjects, and a lordotic cervical spine with a more pronounced kyphotic thoracic spine for the male subjects. In the reconstructed simulations, the average female spinal alignment pattern demonstrated greater intervertebral displacements from the lower cervical spine to the upper thoracic spine with a more pronounced cervical S-shape, compared to the average male spinal alignment pattern. Greater elongation of the cervical ligaments occurred at intervertebral levels where greater intervertebral displacement was found.Rear impact reconstruction simulations performed in this thesis demonstrated a potential impact of gender differences in whole spinal alignment on cervical vertebral kinematics and ligament elongation. The female spinal alignment trend may make women exposed to more significant deformation of the cervical soft tissues due to greater cervical vertebral kinematics during a rear impact. The findings may partially contribute to a greater understanding of the increased injury risk of women sustaining WADs

Finite element human body models with active reflexive muscles suitable for sex based whiplash injury prediction

Previous research has not produced a satisfactory resource to study reflexive muscle activity for investigating potentially injurious whiplash motions. Various experimental and computational studies are available, but none provided a comprehensive biomechanical representation of human response during rear impacts. Three objectives were addressed in the current study to develop female and male finite element human body models with active reflexive neck muscles: 1) eliminate the buckling in the lower cervical spine of the model observed in earlier active muscle controller implementations, 2) evaluate and quantify the influence of the individual features of muscle activity, and 3) evaluate and select the best model configuration that can beused for whiplash injury predictions. The current study used an open-source finite element model of the human body for injury assessment representing an average 50th percentile female anthropometry, together with the derivative 50th percentile male morphed model. Based on the head-neck kinematics and CORelation and Analyis (CORA) tool for evaluation, models with active muscle controller and parallel damping elements showed improved head-neckkinematics agreement with the volunteers over the passive models. It was concluded that this model configuration would be the most suitable for gender-based whiplash injury prediction when different impact severities are to be studied

Comparison of control strategies for the cervical muscles of an average female head-neck finite element model

Objective: ViVA OpenHBM is the first open-source Human Body Model (HBM) for crash safety assessment. It represents an average size (50th percentile) female and was created to assess whiplash protection systems in a car. To increase the biofidelity of the current model, further enhancements are being made by implementing muscle reflex response capabilities as cervical muscles alter the head and neck kinematics of the occupant during low-speed rear crashes. The objective of this study was to assess how different neck muscle activation control strategies affect head-neck kinematics in low-speed rear impacts.Methods: The VIVA OpenHBM head-neck model, previously validated to PMHS data, was used for this study. To represent the 34 cervical muscles, 129 beam elements with Hill-type material models were used. Two different muscle activation control strategies were implemented: a control strategy to mimic neural feedback from the vestibular system and a control strategy to represent displacement feedback from muscle spindles. To identify control gain values for these controller strategies, parameter calibrations were conducted using optimization. The objective of these optimizations was to match the head linear and angular displacements measured in volunteer tests.Results: Muscle activation changed the head kinematics by reducing the peak linear displacements, as compared to the model without muscle activation. For the muscle activation model mimicking the human vestibular system, a good agreement was observed for the horizontal head translation.However, in the vertical direction, there was a discrepancy of head kinematic response caused by buckling of the cervical spine. In the model with a control strategy that represents muscle spindle feedback, improvements in translational head kinematics were observed and less cervical spinebuckling was observed. Although, the overall kinematic responses were better in the first strategy.Conclusions: Both muscle control strategies improved the head kinematics compared to the passive model and comparable to the volunteer kinematics responses with overall better agreement achieved by the model with active muscles mimicking the human vestibular syste

Optimization of Female Head–Neck Model with Active Reflexive Cervical Muscles in Low Severity Rear Impact Collisions

ViVA Open Human Body Model (HBM) is an open-source human body model that was developed to fill the gap of currently available models that lacked the average female size. In this study, the head–neck model of ViVA OpenHBM was further developed by adding active muscle controllers for the cervical muscles to represent the human neck muscle reflex system as studies have shown that cervical muscles influence head–neck kinematics during impacts. The muscle controller was calibrated by conducting optimizationbased parameter identification of published-volunteer data. The effects of different calibration objectives to head–neck kinematics were analyzed and compared. In general, a model with active neck muscles improved the head–neck kinematics agreement with volunteer responses. The current study highlights the importance of including active muscle response to mimic the volunteer’s kinematics. A simple PD controller has found to be able to represent the behavior of the neck muscle reflex system. The optimum gains that defined the muscle controllers in the present study were able to be identified using optimizations. The present study provides a basis for describing an active muscle controller that can be used in future studies to investigate whiplash injuries in rear impacts

Host range and receptor utilization of canine distemper virus analyzed by recombinant viruses: Involvement of heparin-like molecule in CDV infection

AbstractWe constructed recombinant viruses expressing enhanced green fluorescent protein (EGFP) or firefly luciferase from cDNA clones of the canine distemper virus (CDV) (a Japanese field isolate, Yanaka strain). Using these viruses, we examined susceptibilities of different cell lines to CDV infection. The results revealed that the recombinant CDVs can infect a broad range of cell lines. Infectivity inhibition assay using a monoclonal antibody specific to the human SLAM molecule indicated that the infection of B95a cells with these recombinant CDVs is mainly mediated by SLAM but the infection of 293 cell lines with CDV is not, implying the presence of one or more alternative receptors for CDV in non-lymphoid tissue. Infection of 293 cells with the recombinant CDV was inhibited by soluble heparin, and the recombinant virus bound to immobilized heparin. Both F and H proteins of CDV could bind to immobilized heparin. These results suggest that heparin-like molecules are involved in CDV infection

Determination of a phosphorylation site in Nipah virus nucleoprotein and its involvement in virus transcription

Many viruses use their host’s cellular machinery to regulate the functions of viral proteins. The phosphorylation of viral proteins is known to play a role in genome transcription and replication in paramyxoviruses. The paramyxovirus nucleoprotein (N), the most abundant protein in infected cells, is a component of the N–RNA complex and supports the transcription and replication of virus mRNA and genomic RNA. Recently, we reported that the phosphorylation of measles virus N is involved in the regulation of viral RNA synthesis. In this study, we report a rapid turnover of phosphorylation in the Nipah virus N (NiV-N). The phosphorylated NiV-N was hardly detectable in steady-state cells, but was detected after inhibition of cellular protein phosphatases. We identified a phosphorylated serine residue at Ser451 of NiV-N by peptide mass fingerprinting by electrospray ionization–quadrupole time-of-flight mass spectrometry. In the NiV minigenome assay, using luciferase as a reporter gene, the substitution of Ser451 for alanine in NiV-N resulted in a reduction in luciferase activity of approximately 45 % compared with the wild-type protein. Furthermore, the substitution of Ser451 for glutamic acid, which mimics a phosphoserine, led to a more significant decrease in luciferase activity – approximately 81 %. Northern blot analysis showed that both virus transcription and replication were reduced by these mutations. These results suggest that a rapid turnover of the phosphorylation of NiV-N plays an important role in virus transcription and replication