Resolving the Role of Actoymyosin Contractility in Cell Microrheology

Einstein's original description of Brownian motion established a direct relationship between thermally-excited random forces and the transport properties of a submicron particle in a viscous liquid. Recent work based on reconstituted actin filament networks suggests that nonthermal forces driven by the motor protein myosin II can induce large non-equilibrium fluctuations that dominate the motion of particles in cytoskeletal networks. Here, using high-resolution particle tracking, we find that thermal forces, not myosin-induced fluctuating forces, drive the motion of submicron particles embedded in the cytoskeleton of living cells. These results resolve the roles of myosin II and contractile actomyosin structures in the motion of nanoparticles lodged in the cytoplasm, reveal the biphasic mechanical architecture of adherent cells—stiff contractile stress fibers interdigitating in a network at the cell cortex and a soft actin meshwork in the body of the cell, validate the method of particle tracking-microrheology, and reconcile seemingly disparate atomic force microscopy (AFM) and particle-tracking microrheology measurements of living cells

A Distinct Macrophage Population Mediates Metastatic Breast Cancer Cell Extravasation, Establishment and Growth

Background: The stromal microenvironment and particularly the macrophage component of primary tumors influence their malignant potential. However, at the metastatic site the role of these cells and their mechanism of actions for establishment and growth of metastases remain largely unknown. Methodology/Principal Findings: Using animal models of breast cancer metastasis, we show that a population of host macrophages displaying a distinct phenotype is recruited to extravasating pulmonary metastatic cells regardless of species of origin. Ablation of this macrophage population through three independent means (genetic and chemical) showed that these macrophages are required for efficient metastatic seeding and growth. Importantly, even after metastatic growth is established, ablation of this macrophage population inhibited subsequent growth. Furthermore, imaging of intact lungs revealed that macrophages are required for efficient tumor cell extravasation. Conclusion/Significance: These data indicate a direct enhancement of metastatic growth by macrophages through their effects on tumor cell extravasation, survival and subsequent growth and identifies these cells as a new therapeutic target fo

Outcome of primary resurfacing hip replacement: evaluation of risk factors for early revision: 12,093 replacements from the Australian Joint Registry

BACKGROUND AND PURPOSE: The outcome of modern resurfacing remains to be determined. The Australian Orthopaedic Association National Joint Replacement Registry (AOANJRR) started collection of data on hip resurfacing at a time when modern resurfacing was started in Australia. The rate of resurfacing has been higher in Australia than in many other countries. As a result, the AOANJRR has one of the largest series of resurfacing procedures. This study was undertaken to determine the results of this series and the risk factors associated with revision. PATIENTS AND METHODS: Data from the AOANJRR were used to analyze the survivorship of 12,093 primary resurfacing hip replacements reported to the Joint Replacement Registry between September 1999 and December 2008. This was compared to the results of primary conventional total hip replacement reported during the same period. The Kaplan-Meier method and proportional hazards models were used to determine risk factors such as age, sex, femoral component size, primary diagnosis, and implant design. RESULTS: Female patients had a higher revision rate than males; however, after adjusting for head size, the revision rates were similar. Prostheses with head sizes of less than 50 mm had a higher revision rate than those with head sizes of 50 mm or more. At 8 years, the cumulative per cent revision of hip resurfacing was 5.3 (4.6-6.2), as compared to 4.0 (3.8-4.2) for total hip replacement. However, in osteoarthritis patients aged less than 55 years with head sizes of 50 mm or more, the 7-year cumulative per cent revision for hip resurfacing was 3.0 (2.2-4.2). Also, hips with dysplasia and some implant designs had an increased risk of revision. INTERPRETATION: Risk factors for revision of resurfacing were older patients, smaller femoral head size, patients with developmental dysplasia, and certain implant designs. These results highlight the importance of patient and prosthesis selection in optimizing the outcome of hip resurfacing

Bistability in the actin cortex

Multi-color fluorescence imaging experiments of wave forming Dictyostelium cells have revealed that actin waves separate two domains of the cell cortex that differ in their actin structure and phosphoinositide composition. We propose a bistable model of actin dynamics to account for these experimental observation. The model is based on the simplifying assumption that the actin cytoskeleton is composed of two distinct network types, a dendritic and a bundled network. The two structurally different states that were observed in experiments correspond to the stable fixed points in the bistable regime of this model. Each fixed point is dominated by one of the two network types. The experimentally observed actin waves can be considered as trigger waves that propagate transitions between the two stable fixed points

Inferior outcome after hip resurfacing arthroplasty than after conventional arthroplasty: Evidence from the Nordic Arthroplasty Register Association (NARA) database, 1995 to 2007

Today, total hip arthroplasty (THA) is one of the safest and most efficient surgical treatments. New materials, surgical techniques and design concepts intended to improve THA have not always been successful. Thorough preclinical and early clinical investigations can detect some aspects of under-performing, while continuing surveillance is recommended to detect and analyze reasons for any later appearing flaws. In this thesis, several ways to monitor and assess THA performance are explored and carried out, using survival analysis in registry studies, radiostereometry (RSA), radiology and clinical outcome. In Paper I, a study using the Nordic Arthroplasty Register Association (NARA) registry shows that HRA had an almost 3-fold increased early non-septic revision risk and that risk factors were found to be female sex, certain HRA designs and units having performed few HRA procedures. Papers II and III contain comparisons of highly cross-linked polyethylene (XLPE) and conventional polyethylene (PE). XLPE had a considerably lower wear rate up to 10 years but showed no obvious improvements regarding implant fixation, BMD or clinical outcome. In the NARA registry, in 2 of 4 studied cup designs the XLPE version had a lower risk of revision for aseptic loosening compared to the PE version. Paper IV describes that stem subsidence and retrotorsion measured with RSA at 2 years predicted later aseptic stem failure in an unfavorably altered, previously well-functioning cemented femoral stem. In Paper V and VI, a novel approach to measure articulation wear with RSA in radiodense hip arthroplasty articulations was presented and evaluated. Subsequently, a comparison between ceramic-on-ceramic (COC) and metal-on-conventional PE uncemented THA displayed a considerably lower wear rate, smaller periacetabular bone lesions and a relatively high squeaking rate, the latter with unknown long-term consequences, in the COC hips. Implant fixation, heterotopic ossification and clinical outcome did not differ between articulation types. In conclusion, it was confirmed that implant surveillance can be done with RSA, also in radiodense THA. Early migration predicts later aseptic implant failure. Prolonged surveillance can confirm long-term material and design performance, verify or contradict anticipated advantages as well as detect unanticipated long-term complications

An Experimental and Computational Study of the Effect of ActA Polarity on the Speed of Listeria monocytogenes Actin-based Motility

Listeria monocytogenes is a pathogenic bacterium that moves within infected cells and spreads directly between cells by harnessing the cell's dendritic actin machinery. This motility is dependent on expression of a single bacterial surface protein, ActA, a constitutively active Arp2,3 activator, and has been widely studied as a biochemical and biophysical model system for actin-based motility. Dendritic actin network dynamics are important for cell processes including eukaryotic cell motility, cytokinesis, and endocytosis. Here we experimentally altered the degree of ActA polarity on a population of bacteria and made use of an ActA-RFP fusion to determine the relationship between ActA distribution and speed of bacterial motion. We found a positive linear relationship for both ActA intensity and polarity with speed. We explored the underlying mechanisms of this dependence with two distinctly different quantitative models: a detailed agent-based model in which each actin filament and branched network is explicitly simulated, and a three-state continuum model that describes a simplified relationship between bacterial speed and barbed-end actin populations. In silico bacterial motility required a cooperative restraining mechanism to reconstitute our observed speed-polarity relationship, suggesting that kinetic friction between actin filaments and the bacterial surface, a restraining force previously neglected in motility models, is important in determining the effect of ActA polarity on bacterial motility. The continuum model was less restrictive, requiring only a filament number-dependent restraining mechanism to reproduce our experimental observations. However, seemingly rational assumptions in the continuum model, e.g. an average propulsive force per filament, were invalidated by further analysis with the agent-based model. We found that the average contribution to motility from side-interacting filaments was actually a function of the ActA distribution. This ActA-dependence would be difficult to intuit but emerges naturally from the nanoscale interactions in the agent-based representation

Arp2/3 complex interactions and actin network turnover in lamellipodia

Cell migration is initiated by lamellipodia—membrane-enclosed sheets of cytoplasm containing densely packed actin filament networks. Although the molecular details of network turnover remain obscure, recent work points towards key roles in filament nucleation for Arp2/3 complex and its activator WAVE complex. Here, we combine fluorescence recovery after photobleaching (FRAP) of different lamellipodial components with a new method of data analysis to shed light on the dynamics of actin assembly/disassembly. We show that Arp2/3 complex is incorporated into the network exclusively at the lamellipodium tip, like actin, at sites coincident with WAVE complex accumulation. Capping protein likewise showed a turnover similar to actin and Arp2/3 complex, but was confined to the tip. In contrast, cortactin—another prominent Arp2/3 complex regulator—and ADF/cofilin—previously implicated in driving both filament nucleation and disassembly—were rapidly exchanged throughout the lamellipodium. These results suggest that Arp2/3- and WAVE complex-driven actin filament nucleation at the lamellipodium tip is uncoupled from the activities of both cortactin and cofilin. Network turnover is additionally regulated by the spatially segregated activities of capping protein at the tip and cofilin throughout the mesh