Addition of lateral bending range of motion measurement to standard sagittal measurement to improve diagnosis sensitivity of ligamentous injury in the human lower cervical spine.

PURPOSE: This study examined the cervical spine range of motion (ROM) resulting from whiplash-type hyperextension and hyperflexion type ligamentous injuries, and sought to improve the accuracy of specific diagnosis of these injuries. METHODS: The study was accomplished by measurement of ROM throughout axial rotation, lateral bending, and flexion and extension, using a validated finite element model of the cervical spine that was modified to simulate hyperextension and/or hyperflexion injuries. RESULTS: It was found that the kinematic difference between hyperextension and hyperflexion injuries was minimal throughout the combined flexion and extension ROM measurement that is commonly used for clinical diagnosis of cervical ligamentous injury. However, the two injuries demonstrated substantially different ROM under axial rotation and lateral bending. CONCLUSIONS: It is recommended that other bending axes beyond flexion and extension are incorporated into clinical diagnosis of cervical ligamentous injury.

Similar response patterns to topical minoxidil foam 5% in frontal and vertex scalp of men with androgenetic alopecia: a microarray analysis.

BACKGROUND: There are regional variations in the scalp hair miniaturization seen in androgenetic alopecia (AGA). Use of topical minoxidil can lead to reversal of miniaturization in the vertex scalp. However, its effects on other scalp regions have been less well studied. OBJECTIVES: To determine whether scalp biopsies from men with AGA show variable gene expression before and after 8 weeks of treatment with minoxidil topical foam 5% (MTF) vs. placebo. METHODS: A placebo-controlled double-blinded prospective pilot study of MTF vs. placebo was conducted in 16 healthy men aged 18-49 years with Hamilton-Norwood type IV-V thinning. The subjects were asked to apply the treatment (active drug or placebo) to the scalp twice daily for 8 weeks. Stereotactic scalp photographs were taken at the baseline and final visits, to monitor global hair growth. Scalp biopsies were taken at the leading edge of hair loss from the frontal and vertex scalp before and after treatment with MTF and placebo, and microarray analysis was performed using the Affymetrix GeneChip HG U133 Plus 2.0. RESULTS: Global stereotactic photographs showed that MTF induced hair growth in both the frontal and vertex scalp of patients with AGA. Regional differences in gene expression profiles were observed before treatment. However, MTF treatment induced the expression of hair keratin-associated genes and decreased the expression of epidermal differentiation complex and inflammatory genes in both scalp regions. CONCLUSIONS: These data suggest that MTF is effective in the treatment of both the frontal and vertex scalp of patients with AGA.

The effects of ligamentous injury in the human lower cervical spine.

Damage is often sustained by the anterior longitudinal ligament (ALL) and ligamentum flavum (LF) in the cervical spine subsequent to whiplash or other cervical trauma. These ligaments afford substantial cervical stability when healthy, but the ability of the ALL and LF to stabilize the spine when injured is not as conclusively studied. In order to address this issue, the current study excised ALL and LF tissues from cadaveric spines and experimentally simulated whiplash-type damage to the isolated ligaments. Stiffnesses and toe region lengths were measured for both the uninjured and damaged states. These ligamentous mechanical properties were then inputted into a previously-validated finite element (FE) model of the cervical spine and the kinematic effects of various clinically relevant combinations of ligamentous injury were predicted. The data indicated three and five-fold increases in toe region length for the LF and ALL injury variants, respectively. These toe length distensions resulted in FE predictions of supra-physiologic ranges of motion, and these motions were comparable to spines with no ligamentous support. Finally, a set of cadaveric cervical spine ligament-sectioning experiments confirmed the FE predictions and supported the finding that partial injury to the relevant ligaments produces equivalent cervical kinematic signatures to spines that have completely compromised ALL and LF tissues.

Finite element modeling of kinematic and load transmission alterations due to cervical intervertebral disc replacement.

STUDY DESIGN: A parametric finite element investigation of the cervical spine. OBJECTIVE: To determine what effect, if any, cervical disc replacement has on kinematics, facet contact parameters, and anterior column loading. SUMMARY OF BACKGROUND DATA: Anterior cervical discectomy and fusion has been a standard treatment for certain spinal degenerative disorders, but evidence suggests that fusion contributes to adjacent-segment degeneration. Motion-sparing disc replacement implants are believed to reduce adjacent-segment degeneration by preserving kinematics at the treated level. Such implants have been shown to maintain the mobility of the intact spine, but the effects on load transfer between the anterior and posterior elements remain poorly understood. METHODS: To investigate the effects of disc replacement on load transfer in the lower cervical spine, a finite element model was generated using cadaver-based computed tomography imagery. Mesh resolution was varied to establish model convergence, and cadaveric testing was undertaken to validate model predictions. The validated model was altered to include disc replacement prosthesis at the C4/C5 level. The effect of disc-replacement on range of motion, anteroposterior load distribution, contact forces in the facets, as well as the distribution of contact pressure on the facets were examined. Three sizes of implants were examined. RESULTS: Model predictions indicate that the properly sized implant retains the mobility, load sharing, and contact force magnitude and distribution of the intact case. Mobility, load sharing, nuclear pressures, and contact pressures at the adjacent motion segments were not strongly affected by the presence of the properly sized implant, indicating that disc replacement may not be a significant cause of postoperative adjacent-level degeneration. Implant size affected certain mechanical parameters, such as anteroposterior load sharing, and did not affect compliance or range of motion. CONCLUSION: The results of this work support the continued use of motion sparing implants in the lower cervical spine. Load sharing data indicate that implant size may be an important factor that merits further study; although, the deleterious effects of improper size selection may be less significant than those of fusion.

Correlation of mechanical properties within the equine third metacarpal with trabecular bending and multi-density micro-computed tomography data.

Computed tomography (CT) data can be employed with respect to determining mechanical properties and has been used to predict parameters such as elastic modulus, yield strength, and ultimate strength of intact bone. Micro-computed tomography (muCT) possesses the resolution capable of detecting apparent bone density in extremely local regions and can characterize the trabecular structure. It has been asserted that this micro-structure is susceptible to micro-buckling and bending, which has a controversial role in predicting the global mechanical properties of bone. The current study measured the mechanical properties of relatively high apparent density bone from the equine distal third metacarpal. The mechanical properties were correlated with trabecular morphology parameters and apparent densities of localized regions obtained with muCT. These data were used to test two hypotheses: (1) accounting for trabecular bending using trabecular morphology parameters would provide better global mechanical property predictions than using only apparent density, and (2) regions of low apparent density dominate the overall mechanical behavior and provide greater correlation to the measured mechanical properties than regions of high apparent density. The data indicated that accounting for trabecular bending with morphological parameters resulted in stronger correlations to mechanical properties than correlations that relied only on apparent density (r2= 0.91 versus r2= 0.81). Low apparent density regions were more strongly correlated with mechanical properties than high apparent density regions (r2= 0.85 versus r2= 0.77), demonstrating the importance of selecting appropriate regions when attempting to predict mechanical properties from CT data.

Extraocular muscle is defined by a fundamentally distinct gene expression profile.

Skeletal muscle fibers are defined by patterned covariation of key traits that determine contractile and metabolic characteristics. Although the functional properties of most skeletal muscles result from their proportional content of a few conserved muscle fiber types, some, typically craniofacial, muscles exhibit fiber types that appear to lie outside the common phenotypic range. We analyzed gene expression profiles of three putative muscle classes, limb, masticatory, and extraocular muscle (EOM), in adult mice by high-density oligonucleotide arrays. Pairwise comparisons using conservative acceptance criteria identified expression differences in 287 genes between EOM and limb and/or masticatory muscles. Use of significance analysis of microarrays methodology identified up to 400 genes as having an EOM-specific expression pattern. Genes differentially expressed in EOM reflect key aspects of muscle biology, including transcriptional regulation, sarcomeric organization, excitation-contraction coupling, intermediary metabolism, and immune response. These patterned differences in gene expression define EOM as a distinct muscle class and may explain the unique response of these muscles in neuromuscular diseases.

Sterol regulatory element-binding protein-1c mimics the negative effect of insulin on phosphoenolpyruvate carboxykinase (GTP) gene transcription.

We have assessed the potential role of sterol regulatory element-binding protein-1c (SREBP-1c) on the transcription of the gene for the cytosolic form of phosphoenolpyruvate carboxykinase (GTP) (EC ) (PEPCK-C). SREBP-1c introduced into primary hepatocytes with an adenovirus vector caused a total loss of PEPCK-C mRNA and a marked induction of fatty acid synthase mRNA that directly coincided with the appearance of SREBP-1c in the hepatocytes. It also blocked the induction of PEPCK-C mRNA by cAMP and dexamethasone in these cells. In contrast, a dominant negative form of SREBP-1c (dnSREBP-1c) stimulated the accumulation of PEPCK-C mRNA in these cells. SREBP-1c completely blocked the induction of PEPCK-C gene transcription by the catalytic subunit of protein kinase A (PKA), and increasing concentrations of dnSREBP-1c reversed the negative effect of insulin on transcription from the PEPCK-C gene promoter in WT-IR cells. The more than 10-fold induction of PKA-stimulated PEPCK-C gene transcription caused by the co-activator CBP, was also blocked by SREBP-1c. In addition, dnSREBP-1c reversed the strong negative effect of E1A and NF1 on PKA-stimulated transcription from the PEPCK-C gene promoter. An analysis of the possible site of action of SREBP-1c using stepwise truncations of the PEPCK-C gene promoter indicated that the negative effect of SREBP-1c on transcription is exerted at a site between -355 and -277. We conclude that SREBP-1c is an intermediate in the action of insulin on PEPCK-C gene transcription in the liver and acts by blocking the stimulatory effect cAMP that is mediated via an interaction with cAMP-binding protein.

Conservation of the sequence and temporal expression of let-7 heterochronic regulatory RNA.

Two small RNAs regulate the timing of Caenorhabditis elegans development. Transition from the first to the second larval stage fates requires the 22-nucleotide lin-4 RNA, and transition from late larval to adult cell fates requires the 21-nucleotide let-7 RNA. The lin-4 and let-7 RNA genes are not homologous to each other, but are each complementary to sequences in the 3' untranslated regions of a set of protein-coding target genes that are normally negatively regulated by the RNAs. Here we have detected let-7 RNAs of approximately 21 nucleotides in samples from a wide range of animal species, including vertebrate, ascidian, hemichordate, mollusc, annelid and arthropod, but not in RNAs from several cnidarian and poriferan species, Saccharomyces cerevisiae, Escherichia coli or Arabidopsis. We did not detect lin-4 RNA in these species. We found that let-7 temporal regulation is also conserved: let-7 RNA expression is first detected at late larval stages in C. elegans and Drosophila, at 48 hours after fertilization in zebrafish, and in adult stages of annelids and molluscs. The let-7 regulatory RNA may control late temporal transitions during development across animal phylogeny.

Molecular and cellular adaptations to carbohydrate and fat intake.

Adaptation to carbohydrate and fat intake involves changes in a number of biochemical parameters at the cellular level. A change in the concentration of fat or carbohydrate in the blood acts directly to influence metabolic pathways by altering the flux of intermediates into cells. This in turn alters the concentration of hormones and other signaling molecules and changes the rate of expression of genes coding for key regulatory proteins or enzymes in metabolic pathways. These effects occur at different rates and in a tissue-specific manner in response to diet. A key metabolic adaptation involves changes in the level of expression of genes coding for proteins of critical importance in energy metabolism; this is largely due to an altered rate of transcription of selected genes under the control of hormones and/or carbohydrate and lipid. The mediators of this effect are transcription factors, that is, nuclear proteins which integrate the effects of hormones and substrates with the transcription process by binding to response elements in the promoters of regulated genes and interacting with the transcription machinery at the TATA box, thereby altering the activity of RNA polymerase II. In this review we will outline the hierarchy of cellular adaptations to diet and will emphasize the latest concepts of gene regulation in response to metabolites and hormones. In particular, we will review the role of the various transcription factors involved in the regulated expression of the gene for the cytosolic form of phosphoenolpyruvate carboxykinase (GTP) (EC 4.1.132), a key gluconeogenic enzyme.

Microsatellite loci in wild-type and inbred Strongylocentrotus purpuratus.

Strongylocentrotus purpuratus, a major research model in developmental molecular biology, has been inbred through six generations of sibling matings. Though viability initially decreased, as described earlier, the inbred line now consists of healthy, fertile animals. These are intended to serve as a genomic resource in which the level of polymorphism is decreased with respect to wild S. purpuratus. To genotype the inbred animals eight simple sequence genomic repeats were isolated, in context, and PCR primers were generated against the flanking single-copy sequences. Distribution and polymorphism of these regions of the genome were studied in the genomes of 27 wild individuals and in a sample of the inbred animals at F2 and F3 generations. All eight regions were polymorphic, though to different extents, and their homozygosity was increased by inbreeding as expected. The eight markers suffice to identify unambiguously the cellular DNA of any wild or F3 S. purpuratus individual.

CREB binding protein coordinates the function of multiple transcription factors including nuclear factor I to regulate phosphoenolpyruvate carboxykinase (GTP) gene transcription.

Nuclear factor I (NFI) binds to a region of the phosphoenolpyruvate carboxykinase (GTP) (PEPCK) gene promoter adjacent to the cAMP regulatory element (CRE) and inhibits the induction of transcription from the gene promoter caused by the catalytic subunit of protein kinase A. In vivo footprinting studies demonstrated that both the CRE and the NFI-binding site are occupied by transcription factors, regardless of the presence of factors that stimulate (dibutyryl cAMP or dexamethasone) or inhibit (insulin) transcription from the PEPCK gene promoter. The NFI effects on transcription from the PEPCK gene promoter were observed even in the absence of the NFI binding site, suggesting the possibility of other weaker binding sites on the promoter or an interaction of NFI with a transcriptional co-activator. A mammalian two-hybrid system was used to demonstrate direct interaction between the transactivation domain of NFI-C and the CREB binding domain of the CREB-binding protein (CBP). Overexpression of a gene fragment encoding the CREB binding domain of CBP stimulates transcription from the PEPCK gene promoter. The inhibitory effect of NFI on transcription of the PEPCK gene induced by the catalytic subunit of protein kinase A appears to be the result of an interaction between NFI and the CREB-binding protein in which NFI competes with CREB for binding to the CREB-binding site on CBP. In contrast, glucocorticoids and thyroid hormone use the steroid hormone receptor binding domain of CBP to stimulate transcription from the PEPCK gene promoter. NFI-A combines with dexamethasone or thyroid hormone in an additive manner to stimulate PEPCK gene transcription. We conclude that CBP coordinates the action of the multiple factors known to control transcription of the PEPCK gene.

Embryonic and post-embryonic utilization and subcellular localization of the nuclear receptor SpSHR2 in the sea urchin.

SpSHR2 (Strongylocentrotus purpuratus steroid hormone receptor 2) is a nuclear receptor, encoded by a maternal RNA in the sea urchin embryo. These maternal SpSHR2 transcripts, which are present in all cells, persist until the blastula stage and then are rapidly turned over. A small fraction of the embryonic SpSHR2 protein is maternal, but the majority of this nuclear receptor in the embryo is the product of new synthesis, presumably from the maternal RNA after fertilization. In agreement with the mRNA distribution, the SpSHR2 protein is also detected in all embryonic cells. Contrary to the RNA though, the SpSHR2 protein persists throughout embryonic development to the pluteus stage, long after the mRNA is depleted. Following fertilization and as soon as the 2-cell stage, the cytoplasmic SpSHR2 protein enters rapidly into the embryonic nuclei where it appears in the form of speckles. During subsequent stages (from fourth cleavage onward), SpSHR2 resides in speckled form in both the nucleus and the cytoplasm of the embryonic cells. The cytoplasmic localization of SpSHR2 differs between polarized and non-polarized cells, maintaining an apical position in the ectoderm and endoderm versus a uniform distribution in mesenchyme cells. Following the end of embryonic development (pluteus stage), the SpSHR2 protein is depleted from all tissues. During the ensuing four weeks of larval development, the SpSHR2 is not detected in either the larval or the rudiment cells which will give rise to the adult. Just prior to metamorphosis, at about 35 days post-fertilization, the protein is detected again but in contrast to the uniform distribution in the early embryo, the larval SpSHR2 is specifically expressed in cells of the mouth epithelium and the epaulettes. In adult ovaries and testes, SpSHR2 is specifically detected in the myoepithelial cells surrounding the ovarioles and the testicular acini. Nuclear SpSHR2 in blastula extracts binds to the C1R hormone response element in the upstream promoter region of the CyIIIb actin gene indicating that the latter may be a target of this nuclear receptor in the sea urchin embryo.

Nuclear factor I regulates expression of the gene for phosphoenolpyruvate carboxykinase (GTP).

Nuclear factor-I (NFI) binds to the phosphoenolpyruvate carboxykinase (GTP) (PEPCK) gene promoter immediately 5' to the cAMP regulatory element (CRE). This suggests an interaction between NFI and factors that bind the CRE. Of the four NFI isoforms expressed in mammalian tissues, NFI-A and -B stimulate basal transcription from the PEPCK gene promoter in HepG2 cells, while NFI-C and -X are slightly inhibitory. All four NFI isoforms abrogate the 20-fold protein kinase Ac (PKAc)-mediated induction of transcription from the PEPCK gene promoter. Normal PKAc-mediated induction was noted when the CRE was moved 10 base pairs 3' of its original location. However if the CRE was moved 5 base pairs 3', placing it out of phase with the other elements in the promoter, or moved 5' to -285 (the P3(I) site in the promoter), some PKA-mediated stimulation was lost. The NFI-C isoform effectively inhibited PKAc induction regardless of the relative positions of the CRE and the NFI binding sites. NFI-C also abrogated cAMP regulatory element-binding protein (CREB)-induced activity of wild type and mutant PEPCK promoters. There was some cooperativity in the binding of CREB and NFI to their respective binding sites but this did not appear to be physiologically important.

Glycemia-lowering effect of cobalt chloride in the diabetic rat: role of decreased gluconeogenesis.

Results of previous studies indicated that treatment of diabetic rats (induced by streptozotocin) with cobalt chloride (CoCl2) resulted in a significant decrement in serum glucose concentration. The present study was designed to determine the potential role of enhanced glucose uptake vs. decreased glucose production in the above response. The rate of systemic appearance of glucose, measured under fasting conditions using [3-3H]glucose tracer, was reduced from 35.5 +/- 2.5 to 17.5 +/- 1.8 micromol . kg-1 . min-1 in diabetic rats treated with 2 mM CoCl2 added to the drinking water for 10-14 days (P < 0.01). Tissue accumulation of intravenously administered 2-deoxy-[14C]glucose was significantly reduced in kidney and eye of diabetic rats treated with CoCl2, whereas the uptake remained unchanged in several other tissues including cerebrum, red and white skeletal muscle, heart, and liver. The relative content of phosphoenolpyruvate carboxykinase (PEPCK) mRNA was increased 3.1-fold in livers of diabetic compared with normal rats (P < 0.001), and treatment of diabetic rats with CoCl2 decreased hepatic PEPCK mRNA levels to normal. The content of PEPCK mRNA in the liver was decreased by 33% in CoCl2-treated normal rats (P < 0.05). Treatment with CoCl2 resulted in no change in cAMP levels in the livers of either diabetic or normal rats. These results suggest that the glycemia-lowering effect of CoCl2 is mediated by reductions in the rate of systemic appearance of glucose and hepatic gluconeogenesis.

Reliability of measurements obtained with a modified functional reach test in subjects with spinal cord injury.

BACKGROUND AND PURPOSE: The primary purpose of this study was to determine whether the Functional Reach Test (FRT) could be modified to provide reliable measurements of sitting balance. A secondary purpose was to determine whether the test could be used to measure differences among levels of spinal cord injury. SUBJECTS: Thirty male subjects with spinal cord injuries were divided into into three groups based on injury type. Group 1 consisted of subjects with C5-6 tetraplegia, group 2 consisted of subjects with T1-4 paraplegia, and group 3 consisted of subjects with T10-12 paraplegia. METHODS: Subjects sat on similar mat tables (tables varied based on what was available at a given clinic) against the same backboard, set at 80 degrees. During two sessions, forward reach was measured with a yardstick, with a 10-minute break between sessions. RESULTS: Intraclass correlation coefficients (3,2) were high and varied from .85 to .94. Post hoc testing revealed that differences occurred between groups 1 and 3 and groups 2 and 3, but not between groups 1 and 2. CONCLUSION AND DISCUSSION: Test-retest reliability was high with modification of the FRT with a single rater. The measurements reflected differences among levels of lesion. Further study is needed to determine normal values for all levels of lesion, relationships to functional outcomes, and effects of equipment on sitting balance. The modified FRT appears to provide reliable measurements of sitting balance in nonstanding persons with spinal cord injuries.

Transcription from plasmid expression vectors is increased up to 14-fold when plasmids are transfected as concatemers.

A protocol for increasing transcription from plasmid expression vectors is presented. A vector containing chloramphenicol acetyltransferase (CAT) gene was digested leaving the transcription cassette intact. Heat inactivation of restriction enzymes followed by ligation of the digestion products yielded concatemers which migrated as a single band in agarose gel electrophoresis. Mouse fibroblasts transfected with the concatemers gave a CAT activity that was 14-fold greater than that of cells transfected with a similar mass (equimolar gene number) of the native plasmid. The effect was independent of promoter type, restriction enzyme, number of restriction sites and with a noted exception, cell line.

The development of a comprehensive stroke programme in the acute hospital.

Stroke is the third leading cause of death and an important cause of hospital admissions and long term disability. The incidence of stroke is estimated at 200 per 100,000 of the population, Dublin has approximately 2,400 new strokes per annum. In 1989 a comprehensive stroke programme was introduced in St Vincent's Hospital. The aim was to provide patients with a coordinated approach to rehabilitation. A study of the programme was conducted in 1993 by detailed chart review. 129 patients with a clinical diagnosis of stroke were referred to the programme during a 12 month period. 27.13% were under age 65 and 72.87% were over 65 years, 18% died and 44.2% were discharged home, independently mobile. The study findings suggest that the comprehensive stroke programme in St Vincent's Hospital represents a feasible model of care in the acute general hospital setting.