Insulin-like growth factor-I-mediated neurite outgrowth in vitro requires mitogen-activated protein kinase activation

Insulin-like growth factor-I (IGF-I) induces neuronal differentiation in vitro. In the present study, we examined the signaling pathway underlying IGF-I-mediated neurite outgrowth. In SH-SY5Y human neuroblastoma cells, treatment with IGF-I induced concentration- and time-dependent tyrosine phosphorylation of the type I IGF receptor (IGF-IR) and extracellular signal-regulated protein kinases (ERK) 1 and 2. These effects of IGF-I were blocked by a neutralizing antibody against IGF-IR. Whereas IGF-IR phosphorylation was observed within 1 min, maximal phosphorylation of ERKs was not reached for 30 min. Both IGF-IR and ERK phosphorylation were maintained for at least 24 h. Also, the concentration dependence of IGF-I-stimulated IGF-IR and ERK tyrosine phosphorylation paralleled that of IGF-I-mediated neurite outgrowth. We further examined the role of mitogen-activated protein kinase activation in IGF-I-stimulated neuronal differentiation using the mitogen-activated protein kinase/ERK kinase inhibitor PD98059. Whereas PD98059 had no effect on IGF-IR phosphorylation, PD98059 reduced IGF-I-mediated ERK tyrosine phosphorylation and ERK phosphorylation of the substrate Elk-1. PD98059 also produced a parallel reduction of IGF-I-stimulated neurite outgrowth. Finally, consistent with its ability to block neuronal differentiation, PD98059 inhibited IGF-I-dependent changes of GAP-43 and c-myc gene expression. Together these results suggest that activation of ERKs is essential for IGF-I-stimulated neuronal differentiation.     

Reconstruction of the scapholunate ligament in a cadaver model using a bone-ligament-bone autograft from the foot

This study is an investigation of a new procedure in which the scapholunate interosseous ligament (SLIL) is reconstructed using a bone-ligament-bone autograft from the foot. After investigation, the dorsal medial portion of the navicular-first cuneiform ligament (NFCL) was chosen for testing as a potential donor since it is similar in length and thickness to the SLIL and it is easily harvested with minimal potential donor site morbidity. Eight SLILs and NFCLs were harvested from fresh-frozen cadavers. Biomechanical extensometry testing was performed using an Instron 1000 machine. A 5-mm-wide central portion of the NFCL was tested since this width was compatible with the technical aspects of reconstructing the SLIL. Both ligaments were tested for elastic properties, including stiffness, load to failure, and deformation to failure. Mean length of the NFCL was 7.6 mm (range, 5.5-8.5 mm). Stiffness of the NFCL was 10.6 x 10(5) Nm (range, 8.0-13.0 Nm) compared with 14.4 x 10(5) Nm for the SLIL (range, 10.0-19.5 Nm). Peak load to failure for the NFCL was 1,980 N (range, 1,530-2,940 N) compared with 2,940 N for the SLIL (range, 1,780-4,050 N). Total elongation to failure for the NFCL was 2.50 mm (range, 1.7-3.2 mm) compared with 3.2 mm for the SLIL (range, 2.1-5.2 mm). Thus, the biomechanical characteristics of the NFCL were found to be very similar to those of the SLIL. Having established the biomechanical similarities of the 2 ligaments, we are currently using the NFCL to reconstruct the sectioned SLIL in a fresh-frozen cadaver model. Early results suggest that this procedure is feasible for restoration of normal kinematics of the wrist.     

Double-strand break repair in the absence of RAD51 in yeast: a possible role for break-induced DNA replication

In wild-type diploid cells of Saccharomyces cerevisiae, an HO endonuclease-induced double-strand break (DSB) at the MAT locus can be efficiently repaired by gene conversion using the homologous chromosome sequences. Repair of the broken chromosome was nearly eliminated in rad52delta diploids; 99% lost the broken chromosome. However, in rad51delta diploids, the broken chromosomes were repaired approximately 35% of the time. None of these repair events were simple gene conversions or gene conversions with an associated crossover, instead, they created diploids homozygous for the MAT locus and all markers in the 100-kb region distal to the site of the DSB. In rad51delta diploids, the broken chromosome can apparently be inherited for several generations, as many of these repair events are found as sectored colonies, with one part being repaired and the other part being lost the broken chromosome. Similar events occur in about 2% of wild-type cells. We propose that a broken chromosome end can invade a homologous template in the absence of RAD51 and initiate DNA replication that may extend to the telomere, 100 or more kb away. Such break-induced replication appears to be similar to recombination-initiated replication in bacteria.     

Differential thymic selection outcomes stimulated by focal structural alteration in peptide/major histocompatibility complex ligands

The T lineage repertoire is shaped by T cell receptor (TCR)-dependent positive and negative thymic selection processes. Using TCR-transgenic (N15tg) beta2-microglobulin-deficient (beta2m-/-) RAG-2(-/-) H-2(b) mice specific for the VSV8 (RGYVYQGL) octapeptide bound to Kb, we identified a single weak agonist peptide variant V4L (L4) inducing phenotypic and functional T cell maturation. The cognate VSV8 peptide, in contrast, triggers negative selection. The crystal structure of L4/Kb was determined and refined to 2.1 A for comparison with the VSV8/Kb structure at similar resolution. Aside from changes on the p4 side chain of L4 and the resulting alteration of the exposed Kb Lys-66 side chain, these two structures are essentially identical. Hence, a given TCR recognizes subtle distinctions between highly related ligands, resulting in dramatically different selection outcomes. Based on these finding and the recent structural elucidation of the N15-VSV8/Kb complex, moreover, it appears that the germ-line Valpha repertoire contributes in a significant way to positive selection.     

Shoulder joint kinetics during the push phase of wheelchair propulsion

The purpose of this investigation was to quantify the forces and moments at the shoulder joint during free, level wheelchair propulsion and to document changes imposed by increased speed, inclined terrain, and 15 minutes of continuous propulsion. Data were collected using a six-camera VICON motion analysis system, a strain gauge instrumented wheel, and a wheelchair ergometer. Seventeen men with low level paraplegia participated in this study. Shoulder joint forces and moments were calculated using a three-dimensional model applying the inverse dynamics approach. During free propulsion, peak shoulder joint forces were in the posterior (46 N) and superior directions (14 N), producing a peak resultant force of 51 N at an angle of 185 degrees (180 degrees = posterior). Peak shoulder joint moments were greatest in extension (14 Newton-meters [Nm]), followed by abduction (10 Nm), and internal rotation (6 Nm). With fast and inclined propulsion, peak vertical force increased by greater than 360%, and the increase in posterior force and shoulder moments ranged from 107% to 167%. At the end of 15 minutes of continuous free propulsion, there were no significant changes compared with short duration free propulsion. The increased joint loads documented during fast and inclined propulsion could lead to compression of subacromial structures against the overlying acromion.     

Antagonistic activity of Lactobacillus plantarum C11: two new two-peptide bacteriocins, plantaricins EF and JK, and the induction factor plantaricin A

Six bacteriocinlike peptides (plantaricin A [PlnA], PlnE, PlnF, PlnJ, PlnK, and PlnN) produced by Lactobacillus plantarum C11 were detected by amino acid sequencing and mass spectrometry. Since purification to homogeneity was problematic, all six peptides were obtained by solid-phase peptide synthesis and were tested for bacteriocin activity. It was found that L. plantarum C11 produces two two-peptide bacteriocins (PlnEF and PlnJK); a strain-specific antagonistic activity was detected at nanomolar concentrations when PlnE and PlnF were combined and when PlnJ and PlnK were combined. Complementary peptides were at least 10(3) times more active when they were combined than when they were present individually, and optimal activity was obtained when the complementary peptides were present in approximately equal amounts. The interaction between complementary peptides was specific, since neither PlnE nor PlnF could complement PlnJ or PlnK, and none of these peptides could complement the peptides constituting the two-peptide bacteriocin lactococcin G. Interestingly, PlnA, which acts as an extracellular signal (pheromone) that triggers bacteriocin production, also possessed a strain-specific antagonistic activity. No bacteriocin activity could be detected for PlnN.     

Induction of immunologic refractoriness in adults by meningococcal C polysaccharide vaccination

Thirty-four adults were vaccinated with 1/50 of the usual dose of meningococcal polysaccharide vaccine (1 microg of A, C, Y, and W135 polysaccharides, given intramuscularly). This dose was selected as a probe to assess B cell memory. The probe elicited meningococcal C bactericidal antibody responses in all 18 adults who had been vaccinated 4 years earlier with an investigational meningococcal A and C oligosaccharide-protein conjugate vaccine and in the majority of the 11 subjects vaccinated for the first time. In contrast, the responses of the 5 adults given a full dose of licensed polysaccharide vaccine 4 years earlier were <1/10 of those of the other 2 groups. Thus, adults previously given a full dose of meningococcal polysaccharide vaccine show evidence of immunologic refractoriness to group C polysaccharide, whereas refractoriness is not observed after conjugate vaccination. These findings have implications for the use of meningococcal polysaccharide vaccine when the risk of disease is low.     

Analyses of coronary graft patency after aprotinin use: results from the International Multicenter Aprotinin Graft Patency Experience (IMAGE) trial

OBJECTIVE: We examined the effects of aprotinin on graft patency, prevalence of myocardial infarction, and blood loss in patients undergoing primary coronary surgery with cardiopulmonary bypass. METHODS: Patients from 13 international sites were randomized to receive intraoperative aprotinin (n = 436) or placebo (n = 434). Graft angiography was obtained a mean of 10.8 days after the operation. Electrocardiograms, cardiac enzymes, and blood loss and replacement were evaluated. RESULTS: In 796 assessable patients, aprotinin reduced thoracic drainage volume by 43% (P < .0001) and requirement for red blood cell administration by 49% (P < .0001). Among 703 patients with assessable saphenous vein grafts, occlusions occurred in 15.4% of aprotinin-treated patients and 10.9% of patients receiving placebo (P = .03). After we had adjusted for risk factors associated with vein graft occlusion, the aprotinin versus placebo risk ratio decreased from 1.7 to 1.05 (90% confidence interval, 0.6 to 1.8). These factors included female gender, lack of prior aspirin therapy, small and poor distal vessel quality, and possibly use of aprotinin-treated blood as excised vein perfusate. At United States sites, patients had characteristics more favorable for graft patency, and occlusions occurred in 9.4% of the aprotinin group and 9.5% of the placebo group (P = .72). At Danish and Israeli sites, where patients had more adverse characteristics, occlusions occurred in 23.0% of aprotinin- and 12.4% of placebo-treated patients (P = .01). Aprotinin did not affect the occurrence of myocardial infarction (aprotinin: 2.9%; placebo: 3.8%) or mortality (aprotinin: 1.4%; placebo: 1.6%). CONCLUSIONS: In this study, the probability of early vein graft occlusion was increased by aprotinin, but this outcome was promoted by multiple risk factors for graft occlusion.