Neuromuscular reinnervation efficacy using a YFP model.

INTRODUCTION: The gold standard reconstruction for facial reanimation is the functional muscle transfer. The reinnervation of a muscle is never complete, and clinical results are variable with 20% not achieving a satisfactory outcome. We hypothesise that this may be due to a mismatch between the characteristics of the donor nerve and transferred muscle. METHOD: 81 YFP-16 and 14 YFP-H mice were studied in three intervention groups over three time periods. Two parameters were investigated: the number and surface area of reinnervated neuromuscular junctions and regenerating axons. An assessment was made of motor unit proportions. RESULTS: All cases of nerve repair and nerve graft, the neuromuscular junctions (NMJ) were completely reinnervated by regenerating axons. The number and calibre of the regenerating axons were significantly different from controls for both intervention groups. The motor units were smaller in both intervention groups. DISCUSSION: Reinnervation occurs after nerve repair or graft; however, the arbour was reinnervated by large numbers of much smaller axons. These axons showed some evidence of remodelling in the repair group, but not in the graft group. Neither group achieved the parameters of the control group. There were persistent qualitative changes to the morphology of both axons and junctions. Imaging documented both synkinesis and alterations that resemble those seen in ageing. CONCLUSION: Overall, the efficacy of reinnervation is very high with all NMJ reoccupied by regenerating axons. The way small axons are remodelled is different in the nerve repairs compared with the nerve grafts.

High-resolution, high-throughput imaging with a multibeam scanning electron microscope.

Electron-electron interactions and detector bandwidth limit the maximal imaging speed of single-beam scanning electron microscopes. We use multiple electron beams in a single column and detect secondary electrons in parallel to increase the imaging speed by close to two orders of magnitude and demonstrate imaging for a variety of samples ranging from biological brain tissue to semiconductor wafers.

Correlative light and electron microscopy using cathodoluminescence from nanoparticles with distinguishable colours.

Correlative light and electron microscopy promises to combine molecular specificity with nanoscale imaging resolution. However, there are substantial technical challenges including reliable co-registration of optical and electron images, and rapid optical signal degradation under electron beam irradiation. Here, we introduce a new approach to solve these problems: imaging of stable optical cathodoluminescence emitted in a scanning electron microscope by nanoparticles with controllable surface chemistry. We demonstrate well-correlated cathodoluminescence and secondary electron images using three species of semiconductor nanoparticles that contain defects providing stable, spectrally-distinguishable cathodoluminescence. We also demonstrate reliable surface functionalization of the particles. The results pave the way for the use of such nanoparticles for targeted labeling of surfaces to provide nanoscale mapping of molecular composition, indicated by cathodoluminescence colour, simultaneously acquired with structural electron images in a single instrument.

Outcomes after ischemic stroke for hospitals with and without Joint Commission-certified primary stroke centers.

BACKGROUND: The Joint Commission (JC) began certifying primary stroke centers (PSCs) in the United States in 2003. We assessed whether 30-day risk-standardized mortality (RSMR) and readmission (RSRR) rates differed between hospitals with and without JC-certified PSCs in 2006. METHODS: The study cohort included all fee-for-service Medicare beneficiaries ≥65 years old discharged with a primary diagnosis of ischemic stroke (International Classification of Diseases, ninth revision, Clinical Modification 433, 434, 436) in 2006. Hierarchical linear regression models calculated hospital-level RSMRs and RSRRs, adjusting for patient demographics, comorbid conditions, and hospital referral region. Hospitals were categorized as being higher than, no different from, or lower than the national average. RESULTS: There were 310,381 ischemic stroke discharges from 315 JC-certified PSC and 4,231 noncertified hospitals. Mean overall 30-day RSMR and RSRR were 10.9% ± 1.7% and 12.5% ± 1.4%, respectively. The RSMRs of hospitals with JC-certified PSCs were lower than in noncertified hospitals (10.7% ± 1.7% vs 11.0% ± 1.7%), but the RSRRs were comparable (12.5% ± 1.3% vs 12.4% ± 1.7%). Almost half of JC-certified PSC hospitals had RSMRs lower than the national average compared with 19% of noncertified hospitals, but 13% of JC-certified PSC hospitals had lower RSRRs vs 15% of noncertified hospitals. CONCLUSIONS: Hospitals with JC-certified PSCs had lower RSMRs compared with noncertified hospitals in 2006; however, differences were small. Readmission rates were similar between the 2 groups. PSC certification generally identified better-performing hospitals for mortality outcomes, but some hospitals with certified PSCs may have high RSMRs and RSRRs whereas some hospitals without PSCs have low rates. Unmeasured factors may contribute to this heterogeneity.

The joint contribution of sex, age and type of myocardial infarction on hospital mortality following acute myocardial infarction.

OBJECTIVE: Younger, but not older, women have a higher mortality than men of similar age after a myocardial infarction (MI). We sought to determine whether this relationship is true for both ST elevation MI (STEMI) and non-ST elevation MI (NSTEMI). DESIGN: Retrospective cohort study. SETTING: 1057 USA hospitals participant in the National Registry of Myocardial Infarction between 2000 and 2006. PATIENTS: 126 172 STEMI and 235 257 NSTEMI patients. MAIN OUTCOME MEASURE: Hospital death. RESULTS: For both STEMI and NSTEMI, the younger the patient's age, the greater the excess mortality risk for women compared with men, while older women fared similarly (STEMI) or better (NSTEMI) than men (p<0.0001 for the age-sex interaction). In STEMI, the unadjusted women-to-men RR was 1.68 (95% CI 1.41 to 2.01), 1.78 (1.59 to 1.99), 1.45 (1.34 to 1.57), 1.08 (1.02 to 1.14) and 1.03 (0.98 to 1.07) for age <50 years, age 50-59, age 60-69, age 70-79 and age 80-89, respectively. For NSTEMI, corresponding unadjusted RRs were 1.56 (1.31 to 1.85), 1.42 (1.27 to 1.58), 1.17 (1.09 to 1.25), 0.92 (0.88 to 0.96) and 0.86 (0.83 to 0.89). After adjusting for risk status, the excess risk for younger women compared with men decreased to approximately 15-20%, while a better survival of older NSTEMI women compared with men persisted. CONCLUSIONS: Sex-related differences in short-term mortality are age-dependent in both STEMI and NSTEMI patients.

Knowledge of blood pressure levels and targets in patients with coronary artery disease in the USA.

Little is known about patient awareness of nationally recommended blood pressure targets, especially among patients with cardiac disease. To examine this issue, we interviewed 738 patients hospitalized with coronary artery disease to assess their knowledge of their systolic and diastolic blood pressure levels as well as corresponding national targets. We used bivariate and multivariate analyses to determine if any patient demographic or clinical characteristics were associated with blood pressure knowledge. Only 66.1% of patients could recall their own systolic and diastolic blood pressure levels. Only 48.9% of all patients could correctly name targets for these values. Knowledge of target blood pressure levels was particularly poor among patients who were female (odds ratio (OR) 0.69; 95% confidence interval (CI) 0.49-0.98), aged > or =60 years (OR 0.70, CI 0.51-0.97), without any college education (OR 0.48, CI 0.35-0.65), without a documented history of hypertension (OR 0.57, CI 0.39-0.84), and with known diabetes (OR 0.46, CI 0.33-0.66). Patients in the highest risk group, according to Joint National Committee guidelines stratification, were no more knowledgeable about their blood pressure levels and targets than lower risk patients. A significant proportion of patients hospitalized with coronary artery disease do not know their own blood pressure levels or targets. Current blood pressure education efforts appear inadequate, particularly for certain patient subgroups in which hypertension is an important modifiable risk factor.

The neuronal naturalist: watching neurons in their native habitat.

Dynamic processes of neural development, such as migrations of precursor cells, growth of axons and dendrites, and formation and modification of synapses, can be fully analyzed only with techniques that monitor changes over time. Although there has been long-standing motivation for following cellular and synaptic events in vivo (intravital microscopy), until recently few preparations have been studied, and then often only with great effort. Innovations in low-light and laser-scanning microscopies, coupled with developments of new dyes and of genetically encoded indicators, have increased both the breadth and depth of in situ imaging approaches. Here we present the motivations and challenges for dynamic imaging methods, offer some illustrative examples and point to future opportunities with emerging technologies.

Utilization of intravenous tissue-type plasminogen activator for ischemic stroke at academic medical centers: the influence of ethnicity.

BACKGROUND AND PURPOSE: We sought to measure the overall rate of usage of tissue-type plasminogen activator (tPA) for ischemic stroke at academic medical centers, and to determine whether ethnicity was associated with usage. METHODS: Between June and December 1999, 42 academic medical centers in the United States each identified 30 consecutive ischemic stroke cases. Medical records were reviewed and information on demographics, medical history, and treatment were abstracted. Rates of tPA use were compared for African Americans and whites in univariate analysis and after adjustment for age, gender, stroke severity, and type of medical insurance with multivariable logistic regression. RESULTS: Complete information was available for 1195 ischemic stroke patients; 788 were whites and 285 were African Americans: Overall, 49 patients (4.1%) received tPA. In the subgroup of 189 patients without a documented contraindication to therapy, 39 (20.6%) received tPA. Ten (20%) of those receiving tPA had documented contraindication. African Americans were one fifth as likely to receive tPA as whites (1.1% African Americans versus 5.3%; P=0.001), and the difference persisted after adjustment (OR 0.21, 95% CI 0.06 to 0.68; P=0.01). When comparison was restricted to those without a documented contraindication to tPA, the difference remained significant (OR 0.24, 95% CI 0.06 to 0.93; P=0.04). Medical insurance type was independently associated with tPA treatment. After adjustment for ethnicity and other demographic characteristics, those with Medicaid or no insurance were one ninth as likely to receive tPA as those with private medical insurance (OR 0.11, 95% CI 0.02 to 0.17; P=0.003). CONCLUSIONS: tPA is used infrequently for ischemic stroke at US academic medical centers, even among qualifying candidates. African Americans are significantly less likely to receive tPA for ischemic stroke. Contraindications to treatment do not appear to account for the difference.

Asynchronous synapse elimination in neonatal motor units: studies using GFP transgenic mice.

In developing muscle, synapse elimination reduces the number of motor axons that innervate each postsynaptic cell. This loss of connections is thought to be a consequence of axon branch trimming. However, branch retraction has not been observed directly, and many questions remain, such as: do all motor axons retract branches, are eliminated branches withdrawn synchronously, and are withdrawing branches localized to particular regions? To address these questions, we used transgenic mice that express fluorescent proteins in small subsets of motor axons, providing a unique opportunity to reconstruct complete axonal arbors and identify all the postsynaptic targets. We found that, during early postnatal development, each motor axon loses terminal branches, but retracting branches withdraw asynchronously and without obvious spatial bias, suggesting that local interactions at each neuromuscular junction regulate synapse elimination.

Glial cell line-derived neurotrophic factor administration in postnatal life results in motor unit enlargement and continuous synaptic remodeling at the neuromuscular junction.

Overexpression of glial cell line-derived neurotrophic factor (GDNF) in embryonic muscle fibers causes dramatic hyperinnervation of neuromuscular junctions. However, it is not known whether GDNF induces the extra innervation by regulation of axonal branching and/or synaptic maintenance. To address this issue, high levels of circulating GDNF were established by administering subcutaneous injections starting either at birth or later and continuing for up to 40 d. Treatment with exogenous GDNF beginning in the first week, but not later, increased the number of axons converging at neuromuscular junctions. The effect of GDNF on the branching pattern of individual motor axons was determined by reconstructing labeled axonal arbors from transgenic mice expressing yellow fluorescent protein in subsets of motor neurons. Whereas, at postnatal day 8 (P8) individual axons in control animals branched to sporadically innervate junctions within circumscribed regions of the muscle, motor units from GDNF injected animals had significantly more axonal branches and exhibited a high degree of localized arborization such that adjacent muscle fibers were often innervated by the same axon. Administration beginning at P0 and continuing through P40 prolonged multiple innervation of most fibers throughout the period of injection. Between P30 and P40 there was no net change in multiple innervation, although there was evidence of retraction bulbs, suggesting that axon extension and retraction were in equilibrium. We conclude that GDNF has a developmentally regulated effect on presynaptic branching and that sustained administration of GDNF induces a state of continuous synaptic remodeling.

Getting a bead on receptor movements.

Studies of populations of receptor proteins suggest that their number and location are highly regulated. Single-particle tracking of glycine receptors now reveals the direct movement of receptors between different clusters of the anchoring protein gephyrin.

Can practice guidelines be transported effectively to different settings? Results from a multicenter interventional study.

RATIONALE: Although clinical guidelines have become increasingly popular as a means to reduce variation in care, increase efficiency, and improve patient outcomes, little is known about their effectiveness when they are transported outside their original setting, or about the factors that influence their successful translation into clinical practice. This study assessed whether a clinical guideline for low-risk chest pain patients, implemented with a standardized protocol, could be effectively transported to five hospital settings. METHODS: In a prospective, interventional trial, a standardized protocol for low-risk chest pain was implemented at each site. A total of 553 consecutively hospitalized low-risk patients with chest pain were enrolled during a 3-month baseline period followed by a standardized 6-month intervention period. During the intervention period, each patient's physician was contacted about eligibility for discharge within the specified 2-day guideline period. Guideline adherence (discharged within 48 hours) and postdischarge patient outcomes were measured. Local guideline champions were interviewed about their implementation experience. RESULTS: Guideline adherence during the intervention period ranged from 61% to 100%, with only two sites achieving significant increases of > or = 10% from the baseline values. Guideline implementation did not affect clinical outcomes or patient satisfaction. Implementation factors such as preexisting hospital environment, implementation team staffing, and the rapid identification and resolution of barriers may influence the successful translation of guidelines into practice. CONCLUSIONS: Even with a standardized implementation protocol, consistent results across institutions were not obtained when a clinical guideline for chest pain was implemented beyond its original setting. These findings demonstrate the importance of understanding the local factors that influence guideline implementation.

The effect of fibrin sealant on the healing of laser-resurfaced skin.

BACKGROUND: Fibrin sealant is an effective hemostatic agent and a useful tissue sealant. Studies have also suggested that fibrin sealant may accelerate the normal wound-healing process. OBJECTIVE: This study was designed to ascertain whether fibrin sealant would enhance wound healing after CO(2) laser resurfacing in a guinea pig model. METHODS: The CO(2) laser was used to create equal areas of skin resurfacing on both sides of 14 Dunkin Hartley guinea pigs. Fibrin sealant was applied to the treatment side, whereas bacitracin was applied to the control side. Biopsies of these areas were performed on days 1, 3, 7, and 10. A histologic evaluation was performed with the use of a grading scale that compared acute and chronic inflammation, granulation tissue, collagen deposition, and epidermal regeneration. RESULTS: The wounds treated with fibrin sealant demonstrated a statistically significant reduction in the degree of acute and chronic inflammation as well as collagen deposition. At day 7, fibrin sealant was noted to enhance neovascularization and result in a slight delay in reepithelialization. All wounds were completely reepithelialized at day 10. No wound infections or other complications were noted as a result of the application of fibrin sealant. CONCLUSIONS: Although wound healing was not accelerated, the application of fibrin sealant after CO(2) laser resurfacing diminished the acute and chronic inflammatory response, enhanced neovascularization, and reduced collagen accumulation. Further research is needed to assess whether the effects of fibrin sealant noted in this study result in improved cosmetic healing after CO(2) laser resurfacing. (Aesthetic Surg J 2001;21:509-517.).

Imaging neuronal subsets in transgenic mice expressing multiple spectral variants of GFP.

We generated transgenic mice in which red, green, yellow, or cyan fluorescent proteins (together termed XFPs) were selectively expressed in neurons. All four XFPs labeled neurons in their entirety, including axons, nerve terminals, dendrites, and dendritic spines. Remarkably, each of 25 independently generated transgenic lines expressed XFP in a unique pattern, even though all incorporated identical regulatory elements (from the thyl gene). For example, all retinal ganglion cells or many cortical neurons were XFP positive in some lines, whereas only a few ganglion cells or only layer 5 cortical pyramids were labeled in others. In some lines, intense labeling of small neuronal subsets provided a Golgi-like vital stain. In double transgenic mice expressing two different XFPs, it was possible to differentially label 3 neuronal subsets in a single animal.

Multicolor "DiOlistic" labeling of the nervous system using lipophilic dye combinations.

We describe a technique for rapid labeling of a large number of cells in the nervous system with many different colors. By delivering lipophilic dye-coated particles to neuronal preparations with a "gene gun," individual neurons and glia whose membranes are contacted by the particles are quickly labeled. Using particles that are each coated with different combinations of various lipophilic dyes, many cells within a complex neuronal network can be simultaneously labeled with a wide variety of colors. This approach is most effective in living material but also labels previously fixed material. In living material, labeled neurons continue to show normal synaptic responses and undergo dendritic remodeling. This technique is thus useful for studying structural plasticity of neuronal circuits in living preparations. In addition, the Golgi-like labeling of neurons with many different colors provides a novel way to study neuronal connectivity.

Intracranial hemorrhage associated with thrombolytic therapy for elderly patients with acute myocardial infarction: results from the Cooperative Cardiovascular Project.

BACKGROUND AND PURPOSE: Intracranial hemorrhage is a serious complication of thrombolytic therapy for acute myocardial infarction, especially among the elderly, but little information exists on estimating risk. Better estimation of risk in individual patients may allow for withholding or using alternate therapies among those at highest risk. METHODS: To quantify the risk and identify predictors of intracranial hemorrhage associated with thrombolytic therapy, we performed a retrospective cohort study using data from medical charts. The study involved nearly all acute-care hospitals in the United States. All Medicare patients discharged with a principal diagnosis of acute myocardial infarction during a 9-month period in 1994 to 1995 were included. The main outcome measure was intracranial hemorrhage among those treated with thrombolytic therapy. RESULTS: The rate of intracranial hemorrhage was 1.43% (455 of 31 732). In a logistic model, age > or =75 years, female, black race, prior stroke, blood pressure > or =160 mm Hg, tissue plasminogen activator (versus other thrombolytic agent), excessive anticoagulation (international normalized ratio > or =4 or prothrombin time > or =24), and below median weight (< or =65 kg for women; < or =80 kg for men) were independent predictors. A risk stratification scale was developed on the basis of these factors: with none or 1 of the factors (n=6651), the rate of intracranial hemorrhage was 0.69%; with 2 factors (n=10 509), 1.02%; with 3 factors (n=9074), 1.63%; with 4 factors (n=4298), 2.49%; and with > or =5 factors (n=1071), 4. 11% (Mantel-Haenszel; P<0.001). CONCLUSIONS: The rate of intracranial hemorrhage in older patients after treatment with thrombolytic therapy exceeds 1%. Readily available factors can identify elderly patients with acute myocardial infarction at high and low risk for intracranial hemorrhage associated with thrombolytic therapy.