Myelin Water Fraction Is Transiently Reduced after a Single Mild Traumatic Brain Injury – A Prospective Cohort Study in Collegiate Hockey Players

Impact-related mild traumatic brain injuries (mTBI) are a major public health concern, and remain as one of the most poorly understood injuries in the field of neuroscience. Currently, the diagnosis and management of such injuries are based largely on patient-reported symptoms. An improved understanding of the underlying pathophysiology of mTBI is urgently needed in order to develop better diagnostic and management protocols. Specifically, dynamic post-injury changes to the myelin sheath in the human brain have not been examined, despite ‘compromised white matter integrity’ often being described as a consequence of mTBI. In this preliminary cohort study, myelin water imaging was used to prospectively evaluate changes in myelin water fraction, derived from the T2 decay signal, in two varsity hockey teams (45 players) over one season of athletic competition. 11 players sustained a concussion during competition, and were scanned at 72 hours, 2 weeks, and 2 months post-injury. Results demonstrated a reduction in myelin water fraction at 2 weeks post-injury in several brain areas relative to preseason scans, including the splenium of the corpus callosum, right posterior thalamic radiation, left superior corona radiata, left superior longitudinal fasciculus, and left posterior limb of the internal capsule. Myelin water fraction recovered to pre-season values by 2 months post-injury. These results may indicate transient myelin disruption following a single mTBI, with subsequent remyelination of affected neurons. Myelin disruption was not apparent in the athletes who did not experience a concussion, despite exposure to repetitive subconcussive trauma over a season of collegiate hockey. These findings may help to explain many of the metabolic and neurological deficits observed clinically following mTBI.     

A limit on the extent to which increased egg size can compensate for a poor postnatal environment revealed experimentally in the burying beetle,Nicrophorus vespilloides

It is often assumed that there is a positive relationship between egg size and offspring fitness. However, recent studies have suggested that egg size has a greater effect on offspring fitness in low‐quality environments than in high‐quality environments. Such observations suggest that mothers may compensate for poor posthatching environments by increasing egg size. In this paper we test whether there is a limit on the extent to which increased egg size can compensate for the removal of posthatching parental care in the burying beetle, Nicrophorus vespilloides. Previous experiments with N. vespilloides suggest that an increased egg size can compensate for a relatively poor environment after hatching. Here, we phenotypically engineered female N. vespilloides to produce large or small eggs by varying the amount of time they were allowed to feed on the carcass as larvae. We then tested whether differences between these groups in egg size translated into differences in larval performance in a harsh postnatal environment that excluded parental care. We found that females engineered to produce large eggs did not have higher breeding success, and nor did they produce larger larvae than females engineered to produce small eggs. These results suggest that there is a limit on the extent to which increased maternal investment in egg size can compensate for a poor posthatching environment. We discuss the implication of our results for a recent study showing that experimental N. vespilloides populations can adapt rapidly to the absence of posthatching parental care.     

Response of three enzymes to oleic acid, trypsin, and calmodulin chemically modified with a reactive phenothiazine

Calmodulin was covalently modified with 10-(1-propionyloxysuccinimide)-2-trifluoromethylphenothiazine++ + to stoichiometries between 0 and 2 mol/mol in the presence of Ca2+. The modified calmodulins, oleic acid, and trypsin were assayed for their ability to activate pea plant NAD kinase, bovine brain 3',5'-cAMP phosphodiesterase, and human erythrocyte Ca2+-ATPase. All modified calmodulins activated both phosphodiesterase and Ca2+-ATPase; at the highest concentration assayed, calmodulin modified with 2 mol of reagent/mol activated phosphodiesterase and Ca2+-ATPase to 53% and 100%, respectively, of the activation obtained with unmodified calmodulin. However, higher concentrations of the modified calmodulins were required to observe the same activation; at least 900-fold and 100-fold higher concentrations were required for the two enzymes, respectively. NAD kinase was not activated by any calmodulin labeled to a stoichiometry greater than 1 mol/mol even when a concentration equal to 17,000 times the apparent dissociation constant of calmodulin for NAD kinase was assayed. Therefore, the modified protein (and not some fraction resistant to labeling) is active toward the mammalian enzymes but inactive toward plant NAD kinase. The different response of the three enzymes to the chemical modification suggests that the enzymes may utilize different binding domains on calmodulin. NAD kinase also was not activated by other known activators of the two mammalian enzymes, namely lipids and limited proteolysis. In parallel experiments using the same agents on each enzyme, NAD kinase was the only enzyme of the three that was not activated by oleic acid and several other lipids or by limited trypsin digestion. These results show that NAD kinase possesses several attributes which would not be predicted by current models of the mechanism of activation of enzymes by calmodulin.     

Regulation of rat ovarian cell growth and steroid secretion

A cultured rat ovarian cell line (31 A-F(2)) was used to study the effect of growth factors (epidermal growth factor [EGF] and fibroblast growth factor [FGF]), a survival factor (ovarian growth factor [OGF]), a hormone (insulin), and an iron-binding protein (transferring) on cell proliferation and steroid production under defined culture conditions. EGF and insulin were shown to be mitogenic (half-maximal response at 0.12 nM and 0.11 muM, respectively) for 31A-F(2) cells incubated in serum-free medium. EGF induced up to three doublings in the cell population, whereas insulin induced an average of one cell population doubling. FGF, OGF, and transferrin were found not to have any prominent effect on cell division when incubated individually with 31A-F(2) cells in serum-free medium. However, a combination of EGF, OGF, insulin, and transferrin stimulated cell division to the same approximate extent as cells incubated in the presence of 5 percent fetal calf serum. EGF or insulin did not significantly affect total cell cholesterol levels (relative to cells incubated in serum-free medium) when incubated individually with 31A-F(2) cells. However, cell cholesterol levels were increased by the addition of OGF (250 percent), FGF (370 percent), or a combination of insulin and EGF (320 percent). Progesterone secretion from 31A-F(2) cells was enhanced by EGF (25 percent), FGF (80 percent), and insulin (115 percent). However, the addition of a mitogenic mixture of EGF, OGF, insulin, and transferrin suppressed progesterone secretion 150 percent) below that of control cultures. These studies have permitted us to determine that EGF and insulin are mitogenic factors that are required for the growth of 31A-F(2) cells and that OGF and transferrin are positive cofactors that enhance growth. Also, additional data suggest that cholesterol and progesterone production in 31A-F(2) cells can be regulated by peptide growth factors and the hormone insulin.     

Amyloid fibril formation requires a chemically discriminating nucleation event: studies of an amyloidogenic sequence from the bacterial protein OsmB.

The sequence of the Escherichia coli OsmB protein was found to resemble that of the C-terminal region of the beta amyloid protein of Alzheimer's disease, which seems to be the major determinant of its unusual structural and solubility properties. A peptide corresponding to residues 28-44 of the OsmB protein was synthesized, and its conformational properties and aggregation behavior were analyzed. The peptide OsmB(28-44) was shown to form amyloid fibrils, as did two sequence analogs designed to test the sequence specificity of fibril formation. These fibrils bound Congo red, and two of the peptides showed birefringence. The peptide fibrils were analyzed by electron microscopy and Fourier transform infrared spectroscopy. Subtle differences were observed which were not interpretable at the molecular level. The rate of fibril formation by each peptide was followed by monitoring the turbidity of supersaturated aqueous solutions. The kinetics of aggregation were characterized by a delay period during which the solution remained clear, followed by a nucleation event which led to a growth phase, during which the solution became viscous and turbid due to the presence of insoluble fibrils. The observation of a kinetic barrier to aggregation is typical of a crystallization event. The delay period could be eliminated by seeding the supersaturated solution with previously formed fibrils. Each peptide could be nucleated by fibrils formed from that same peptide, but not by fibrils from closely related sequences, suggesting that fibril growth requires specific hydrophobic interactions. It appears likely that this repeated sequence motif, which comprises most of the OsmB protein sequence, dictates the structure and possibly the function of that protein.(ABSTRACT TRUNCATED AT 250 WORDS)     

Some viral and protozool diseases in the European wildcat (Felis silvestris).

Ten European wildcats (Felis silvestris) were examined at necropsy and an additional 23 were examined clinically for evidence of viral diseases in Scotland. Two plasma samples taken from live free-living wildcats showed positive ELISA reactions to feline leukemia antigen. A feline leukemia virus of subgroup A was isolated from one of these samples, taken from a wildcat in north-western Scotland. No antibodies to feline coronavirus or feline immunodeficiency virus were detected in any sample. Three of the live wildcats and one of the dead had chronic mucopurulent rhinotracheitis suggestive of "cat flu." One other dead wildcat had diffuse enlargement of anterior lymph nodes. The findings indicated that feline leukemia virus infection can occur in free-living Felis silvestris. It is possible that the disease exists as a sustained infection in some wildcat populations, although the close interaction between wildcat and the domestic cat means that the latter could act as a continual source of infection.     

The effects of deletions in the central helix of calmodulin on enzyme activation and peptide binding

Using site-directed mutagenesis we have expressed in Escherichia coli three engineered calmodulins (CaM) containing deletions in the solvent-exposed region of the central helix. These are CaM delta 84, Glu-84 removed; CaM delta 83-84, Glu-83 and Glu-84 removed; and CaM delta 81-84, Ser-81 through Glu-84 removed. The abilities of these proteins to activate skeletal muscle myosin light chain kinase, plant NAD kinase, and bovine brain calcineurin activities were determined, as were their abilities to bind a synthetic peptide based on the calmodulin-binding domain of skeletal muscle myosin light chain kinase. Similar results were obtained with all three deletion proteins. Vm values for enzymes activated by the deletion proteins are all within 10-20% of those values obtained with bacterial control calmodulin. Relative to bacterial control values, changes in Kact or Kd values associated with the deletions are all less than an order of magnitude: Kact values for NAD kinase and myosin light chain kinase are increased 5-7-fold, Kd values for binding of the synthetic peptide are increased 4-7-fold, and Kact values for calcineurin are increased only 1-3-fold. In assays of NAD kinase and myosin light chain kinase activation some differences between bovine calmodulin and bacterial control calmodulin were observed. With NAD kinase, Kact values for the bacterial control protein are increased 4-fold relative to values for bovine calmodulin, and Vm values are increased by 50%; with myosin light chain kinase, Kact values are increased 2-fold and Vm values are decreased 10-15% relative to those values obtained with bovine calmodulin. These differences between bacterial control and bovine calmodulins probably can be attributed to known differences in postranslational processing of calmodulin in bacterial and eucaryotic cells. No differences between bovine and control calmodulins were observed in assays of calcineurin activation or peptide binding. Our observations indicate that contacts with the deleted residues, Ser-81 through Glu-84, are not critical in the calmodulin-target complexes we have evaluated. Formation of these calmodulin-target complexes also does not appear to be greatly affected by the global alterations in the structure of calmodulin that are associated with the deletions. In models in which the central helix is maintained in the altered calmodulins, each deleted residue causes the two lobes of calmodulin to be twisted 100 degrees relative to one another and brought 1.5 A closer together.(ABSTRACT TRUNCATED AT 400 WORDS)