Andrew G. Clark

Diagnosis and management of acute graft-versus-host disease

A joint working group established by the Haemato‐oncology subgroup of the British Committee for Standards in Haematology (BCSH) and the British Society for Bone Marrow Transplantation (BSBMT) has reviewed the available literature and made recommendations for the diagnosis and management of acute graft‐versus‐host disease. This guideline includes recommendations for the diagnosis and grading of acute graft‐versus‐host disease as well as primary treatment and options for patients with steroid‐refractory disease. The goal of treatment should be effective control of graft‐versus‐host disease while minimizing risk of toxicity and relapse.

Peripheral blood and bone marrow abnormalities in patients with HIV related disease

Between February 1983 and April 1986 we studied peripheral blood and bone marrow samples from 20 patients with human immunodeficiency virus (HIV) related disease. 14 patients had AIDS, three had ARC, two had PGL and one had ITP as a sole manifestation of HIV related disease.

Monitoring Actin Cortex Thickness in Live Cells

Animal cell shape is controlled primarily by the actomyosin cortex, a thin cytoskeletal network that lies directly beneath the plasma membrane. The cortex regulates cell morphology by controlling cellular mechanical properties, which are determined by network structure and geometry. In particular, cortex thickness is expected to influence cell mechanics. However, cortex thickness is near the resolution limit of the light microscope, making studies relating cortex thickness and cell shape challenging. To overcome this, we developed an assay to measure cortex thickness in live cells, combining confocal imaging and subresolution image analysis. We labeled the actin cortex and plasma membrane with chromatically different fluorophores and measured the distance between the resulting intensity peaks. Using a theoretical description of cortex geometry and microscopic imaging, we extracted an average cortex thickness of ∼190 nm in mitotic HeLa cells and tested the validity of our assay using cell images generated in silico. We found that thickness increased after experimental treatments preventing F-actin disassembly. Finally, we monitored physiological changes in cortex thickness in real-time during actin cortex regrowth in cellular blebs. Our investigation paves the way to understanding how molecular processes modulate cortex structure, which in turn drives cell morphogenesis.

Integration of Single and Multicellular Wound Responses

Single cells and multicellular tissues rapidly heal wounds. These processes are considered distinct, but one mode of healing--Rho GTPase-dependent formation and closure of a purse string of actin filaments (F-actin) and myosin-2 around wounds--occurs in single cells and in epithelia. Here, we show that wounding of one cell in Xenopus embryos elicits Rho GTPase activation around the wound and at the nearest cell-cell junctions in the neighbor cells. F-actin and myosin-2 accumulate at the junctions and around the wound itself, and as the resultant actomyosin array closes over the wound site, junctional F-actin and myosin-2 become mechanically integrated with the actin and myosin-2 around the wound, forming a hybrid purse string. When cells are ablated rather than wounded, Rho GTPase activation and F-actin accumulation occur at cell-cell junctions surrounding the ablated cell, and the purse string closes the hole in the epithelium. Elevation of intracellular free calcium, an essential upstream signal for the single-cell wound response, also occurs at the cell-cell contacts and in neighbor cells. Thus, the single and multicellular purse string wound responses represent points on a signaling and mechanical continuum that are integrated by cell-cell junctions.

Diagnosis and management of chronic graft-versus-host disease.

A joint working group established by the Haemato‐oncology subgroup of the British Committee for Standards in Haematology (BCSH) and the British Society for Bone Marrow Transplantation (BSBMT) has reviewed the available literature and made recommendations for the diagnosis and management of chronic graft‐versus‐host disease (GvHD). This guideline includes recommendations for the diagnosis and staging of chronic GvHD as well as primary treatment and options for patients with steroid‐refractory disease. The goal of treatment should be the effective control of GvHD while minimizing the risk of toxicity and relapse.

Stresses at the Cell Surface during Animal Cell Morphogenesis

Cell shape is determined by cellular mechanics. Cell deformations in animal cells, such as those required for cell migration, division or epithelial morphogenesis, are largely controlled by changes in mechanical stress and tension at the cell surface. The plasma membrane and the actomyosin cortex control surface mechanics and determine cell surface tension. Tension in the actomyosin cortex primarily arises from myosin-generated stresses and depends strongly on the ultrastructural architecture of the network. Plasma membrane tension is controlled mainly by the surface area of the membrane relative to cell volume and can be modulated by changing membrane composition, shape and the organization of membrane-associated proteins. We review here our current understanding of the control of cortex and membrane tension by molecular processes. We particularly highlight the need for studies that bridge the scales between microscopic events and emergent properties at the cellular level. Finally, we discuss how the mechanical interplay between membrane dynamics and cortex contractility is key to understanding the biomechanical control of cell morphogenesis.

Management of cytomegalovirus infection in haemopoietic stem cell transplantation

● Cytomegalovirus (CMV) infection and CMV diseaseshould be diagnosed according to established, interna-tionally accepted, standardized criteria (Grade 1C).● Risk-adapted patient assessment should inform clinicalmanagement (Grade 1B).● All potential haemopoietic stem cell transplantation(HSCT) recipients should be tested for the presence ofCMV IgG antibody at diagnosis (Grade 1C).● Once optimum human leucocyte antigen (HLA) match-ing has been performed, a CMV IgG-negative donorshould be chosen for a CMV IgG-negative recipient anda CMV IgG-positive donor should be chosen for a CMVIgG-positive recipient when possible (Grade 1A).● Donors or recipients who are initially found to be CMVIgG-negative should be retested pre-transplant toexclude primary CMV infection (Grade 1C).● Apparent CMV seroconversion in potential allograftrecipients who have received unscreened blood productsshould be actively investigated to exclude passive acqui-sition of antibody (Grade 1C)● Any CMV IgG-negative HSCT recipient transplantedfrom a CMV IgG-negative donor who develops CMVinfection post-transplant must be reported to the SeriousHazards of Transfusion (SHOT) scheme (Grade 1C).● Primary prophylaxis with ganciclovir is not generallyrecommended as toxicity outweighs efficacy in HSCTpatients (Grade 1B).● Primary prophylaxis with aciclovir or valaciclovir canbe deployed but only in conjunction with appropriatemonitoring of CMV in blood (Grade 1B).● Valaciclovir or valganciclovir are valid treatmentoptions for secondary prophylaxis with appropriatemonitoring of CMV in blood (Grade 1C).● Intravenous immunoglobulin is not recommended forprophylaxis of CMV infection (Grade 1A).● Real time quantitative polymerase chain reaction (PCR)is the preferred choice for monitoring CMV DNA levelsin HSCT patients (Grade 1B).● All diagnostic laboratories should deploy the CMVinternational standard to allow viral loads to becompared between centres (Grade 1C).● Monitoring of CMV load should be undertaken atleast weekly for the first 3 months post-HSCT (Grade2C).● CMV viral load monitoring should continue for 6-12 months if the patient has chronic graft-versus-hostdisease (GvHD) or prolonged T-cell immunodeficiency(Grade 1B).● Each transplant centre should have a risk-adapted policydetailing threshold values for treatment of CMV infec-tion, taking into account patient factors and PCR meth-odology (Grade 2C).● Ganciclovir is recommended as first line pre-emptivetherapy for CMV in HSCT patients (Grade 1A).● Oral valganciclovir is a valid alternative when gastroin-testinal absorption is normal or only minimallyimpaired (Grade 1A).● Foscarnet is recommended as an alternative first-lineagent if neutropenia is present or for ganciclovir treat-ment failure (Grade 1A).

Organ-specific management and supportive care in chronic graft-versus-host disease

A joint working group established by the Haemato‐oncology subgroup of the British Committee for Standards in Haematology and the British Society for Bone Marrow Transplantation has reviewed the available literature and made recommendations for the supportive care and management of organ‐specific complications of chronic graft‐versus‐host disease (cGvHD). This guideline includes recommendations for the specific therapy of skin, oral, liver, gut, lung, ocular and genital manifestations of cGvHD and for the supportive care of these patients, including vaccinations and prophylaxis against infection. The goal of treatment should be effective control of GvHD while minimizing the risk of toxicity and relapse.

Mechanics and Regulation of Cell Shape During the Cell Cycle

Many cell types undergo dramatic changes in shape throughout the cell cycle. For individual cells, a tight control of cell shape is crucial during cell division, but also in interphase, for example during cell migration. Moreover, cell cycle-related cell shape changes have been shown to be important for tissue morphogenesis in a number of developmental contexts. Cell shape is the physical result of cellular mechanical properties and of the forces exerted on the cell. An understanding of the causes and repercussions of cell shape changes thus requires knowledge of both the molecular regulation of cellular mechanics and how specific changes in cell mechanics in turn effect global shape changes. In this chapter, we provide an overview of the current knowledge on the control of cell morphology, both in terms of general cell mechanics and specifically during the cell cycle.

Actin cortex architecture regulates cell surface tension

Animal cell shape is largely determined by the cortex, a thin actin network underlying the plasma membrane in which myosin-driven stresses generate contractile tension. Tension gradients result in local contractions and drive cell deformations. Previous cortical tension regulation studies have focused on myosin motors. Here, we show that cortical actin network architecture is equally important. First, we observe that actin cortex thickness and tension are inversely correlated during cell-cycle progression. We then show that the actin filament length regulators CFL1, CAPZB and DIAPH1 regulate mitotic cortex thickness and find that both increasing and decreasing thickness decreases tension in mitosis. This suggests that the mitotic cortex is poised close to a tension maximum. Finally, using a computational model, we identify a physical mechanism by which maximum tension is achieved at intermediate actin filament lengths. Our results indicate that actin network architecture, alongside myosin activity, is key to cell surface tension regulation.