Ferumoxytol nanoparticle uptake in brain during acute neuroinflammation is cell-specific.

Ferumoxytol ultrasmall superparamagnetic iron oxide nanoparticles can enhance contrast between neuroinflamed and normal-appearing brain tissue when used as a contrast agent for high-sensitivity magnetic resonance imaging (MRI). Here we used an anti-dextran antibody (Dx1) that binds the nanoparticle's carboxymethyldextran coating to differentiate ferumoxytol from endogenous iron and localize it unequivocally in brain tissue. Intravenous injection of ferumoxytol into immune-competent rats that harbored human tumor xenograft-induced inflammatory brain lesions resulted in heterogeneous and lesion-specific signal enhancement on MRI scans in vivo. We used Dx1 immunolocalization and electron microscopy to identify ferumoxytol in affected tissue post-MRI. We found that ferumoxytol nanoparticles were taken up by astrocyte endfeet surrounding cerebral vessels, astrocyte processes, and CD163(+)/CD68(+) macrophages, but not by tumor cells. These results provide a biological basis for the delayed imaging changes seen with ferumoxytol and indicate that ferumoxytol-MRI can be used to assess the inflammatory component of brain lesions in the clinic.

Pallidal neuronal apolipoprotein E in pantothenate kinase-associated neurodegeneration recapitulates ischemic injury to the globus pallidus.

Pantothenate kinase-associated neurodegeneration (PKAN) is a progressive movement disorder that is due to mutations in PANK2. Pathologically, it is a member of a class of diseases known as neurodegeneration with brain iron accumulation (NBIA) and features increased tissue iron and ubiquitinated proteinaceous aggregates in the globus pallidus. We have previously determined that these aggregates represent condensed residue derived from degenerated pallidal neurons. However, the protein content, other than ubiquitin, of these aggregates remains unknown. In the present study, we performed biochemical and immunohistochemical studies to characterize these aggregates and found them to be enriched in apolipoprotein E that is poorly soluble in detergent solutions. However, we did not determine a significant association between APOE genotype and the clinical phenotype of disease in our database of 81 cases. Rather, we frequently identified similar ubiquitin- and apolipoprotein E-enriched lesions in these neurons in non-PKAN patients in the penumbrae of remote infarcts that involve the globus pallidus, and occasionally in other brain sites that contain large γ-aminobutyric acid (GABA)ergic neurons. Our findings, taken together, suggest that tissue or cellular hypoxic/ischemic injury within the globus pallidus may underlie the pathogenesis of PKAN.

Surgery versus physical therapy for a meniscal tear and osteoarthritis.

BACKGROUND: Whether arthroscopic partial meniscectomy for symptomatic patients with a meniscal tear and knee osteoarthritis results in better functional outcomes than nonoperative therapy is uncertain. METHODS: We conducted a multicenter, randomized, controlled trial involving symptomatic patients 45 years of age or older with a meniscal tear and evidence of mild-to-moderate osteoarthritis on imaging. We randomly assigned 351 patients to surgery and postoperative physical therapy or to a standardized physical-therapy regimen (with the option to cross over to surgery at the discretion of the patient and surgeon). The patients were evaluated at 6 and 12 months. The primary outcome was the difference between the groups with respect to the change in the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) physical-function score (ranging from 0 to 100, with higher scores indicating more severe symptoms) 6 months after randomization. RESULTS: In the intention-to-treat analysis, the mean improvement in the WOMAC score after 6 months was 20.9 points (95% confidence interval [CI], 17.9 to 23.9) in the surgical group and 18.5 (95% CI, 15.6 to 21.5) in the physical-therapy group (mean difference, 2.4 points; 95% CI, -1.8 to 6.5). At 6 months, 51 active participants in the study who were assigned to physical therapy alone (30%) had undergone surgery, and 9 patients assigned to surgery (6%) had not undergone surgery. The results at 12 months were similar to those at 6 months. The frequency of adverse events did not differ significantly between the groups. CONCLUSIONS: In the intention-to-treat analysis, we did not find significant differences between the study groups in functional improvement 6 months after randomization; however, 30% of the patients who were assigned to physical therapy alone underwent surgery within 6 months. (Funded by the National Institute of Arthritis and Musculoskeletal and Skin Diseases; METEOR ClinicalTrials.gov number, NCT00597012.).

MAP and kinesin-dependent nuclear positioning is required for skeletal muscle function.

The basic unit of skeletal muscle in all metazoans is the multinucleate myofibre, within which individual nuclei are regularly positioned. The molecular machinery responsible for myonuclear positioning is not known. Improperly positioned nuclei are a hallmark of numerous diseases of muscle, including centronuclear myopathies, but it is unclear whether correct nuclear positioning is necessary for muscle function. Here we identify the microtubule-associated protein ensconsin (Ens)/microtubule-associated protein 7 (MAP7) and kinesin heavy chain (Khc)/Kif5b as essential, evolutionarily conserved regulators of myonuclear positioning in Drosophila and cultured mammalian myotubes. We find that these proteins interact physically and that expression of the Kif5b motor domain fused to the MAP7 microtubule-binding domain rescues nuclear positioning defects in MAP7-depleted cells. This suggests that MAP7 links Kif5b to the microtubule cytoskeleton to promote nuclear positioning. Finally, we show that myonuclear positioning is physiologically important. Drosophila ens mutant larvae have decreased locomotion and incorrect myonuclear positioning, and these phenotypes are rescued by muscle-specific expression of Ens. We conclude that improper nuclear positioning contributes to muscle dysfunction in a cell-autonomous fashion.

Mechanisms guiding primordial germ cell migration: strategies from different organisms.

The regulated migration of cells is essential for development and tissue homeostasis, and aberrant cell migration can lead to an impaired immune response and the progression of cancer. Primordial germ cells (PGCs), precursors to sperm and eggs, have to migrate across the embryo to reach somatic gonadal precursors, where they carry out their function. Studies of model organisms have revealed that, despite important differences, several features of PGC migration are conserved. PGCs require an intrinsic motility programme and external guidance cues to survive and successfully migrate. Proper guidance involves both attractive and repulsive cues and is mediated by protein and lipid signalling.

Live imaging of Drosophila myoblast fusion.

Myoblast fusion requires a number of cellular behaviors, including cell migration, recognition, and adhesion, as well as a series of subcellular behaviors, such as cytoskeletal rearrangements, vesicle trafficking, and membrane dynamics, leading to two cells becoming one. With the discovery of fluorescent proteins that can be introduced and studied within living cells, the possibility of monitoring these complex processes within the living embryo is now a reality. Live imaging, unlike imaging techniques for fixed embryos, allows the opportunity to visualize and measure the dynamics of these processes in vivo. This chapter describes the development and use of live imaging techniques to study myoblast fusion in Drosophila.

Myoblast fusion in fly and vertebrates: new genes, new processes and new perspectives.

Muscle formation and repair depends critically on the fusion of myoblasts. Despite the importance of this process, little is known about the cellular and molecular mechanisms regulating fusion. Forward genetic screens in Drosophila melanogaster have uncovered genes that, when mutated, prevent myoblast fusion. Analyses of these gene products have indicated that the actin cytoskeleton and its regulation play a central role in the fusion process. In this review, we discuss recent advances in the field, including new imaging approaches to analyze fusion as well as a description of novel genes required for fusion. In particular, we highlight what has been learned about the requirement of a specific actin structure at the site of fusion. We also place these findings from Drosophila within the context of myoblast fusion in vertebrates.

SCAR/WAVE and Arp2/3 are crucial for cytoskeletal remodeling at the site of myoblast fusion.

Myoblast fusion is crucial for formation and repair of skeletal muscle. Here we show that active remodeling of the actin cytoskeleton is essential for fusion in Drosophila. Using live imaging, we have identified a dynamic F-actin accumulation (actin focus) at the site of fusion. Dissolution of the actin focus directly precedes a fusion event. Whereas several known fusion components regulate these actin foci, others target additional behaviors required for fusion. Mutations in kette/Nap1, an actin polymerization regulator, lead to enlarged foci that do not dissolve, consistent with the observed block in fusion. Kette is required to positively regulate SCAR/WAVE, which in turn activates the Arp2/3 complex. Mutants in SCAR and Arp2/3 have a fusion block and foci phenotype, suggesting that Kette-SCAR-Arp2/3 participate in an actin polymerization event required for focus dissolution. Our data identify a new paradigm for understanding the mechanisms underlying fusion in myoblasts and other tissues.