Bedford v. Canada: a paradigmatic case toward ensuring the human and health rights of sex workers.

The Criminal Code of Canada prohibits certain aspects of sex work: the keeping of a common bawdy-house, living off the avails of prostitution and communicating for the purposes of prostitution in a public place. These legal constraints impede sex workers' ability to practise their profession safely and without risk to their bodily integrity; they also impair their personal autonomy and can lead to their stigmatization. Bedford v. Canada is a groundbreaking case, since the applicants and intervening organizations seek to overturn aspects of Canadian law that specifically put the health and human rights of sex workers at risk.

Paxillin associates with the microtubule cytoskeleton and the immunological synapse of CTL through its leucine-aspartic acid domains and contributes to microtubule organizing center reorientation.

The cytoskeletal adaptor protein paxillin localizes to the microtubule organizing center (MTOC) in T cells and, upon target cell binding, is recruited to the supramolecular activation complex (SMAC). We mapped the region of paxillin that associates with both the MTOC and SMAC to the leucine-aspartic acid (LD) domains and showed that a protein segment containing LD2-4 was sufficient for MTOC and SMAC recruitment. Examination of the localization of paxillin at the SMAC revealed that paxillin localizes to the peripheral area of the SMAC along with LFA-1, suggesting that LFA-1 may contribute to its recruitment. LFA-1 or CD3 engagement alone was insufficient for paxillin recruitment because there was no paxillin accumulation at the site of CTL contact with anti-LFA-1- or anti-CD3-coated beads. In contrast, paxillin accumulation was detected when beads coated with both anti-CD3 and anti-LFA-1 were bound to CTL, suggesting that signals from both the TCR and LFA-1 are required for paxillin accumulation. Paxillin was shown to be phosphorylated downstream of ERK, but when we generated a mutation (S83A/S130A) that abolished the mobility shift as a result of phosphorylation, we found that paxillin still bound to the MTOC and was recruited to the SMAC. Furthermore, ERK was not absolutely required for MTOC reorientation in CTL that require ERK for killing. Finally, expression of the LD2-4 region of paxillin substantially reduced MTOC reorientation. These studies demonstrated that paxillin is recruited, through its LD domains, to sites of integrin engagement and may contribute to MTOC reorientation required for directional degranulation.

Sequence of physical changes to the cell membrane during glucocorticoid-induced apoptosis in S49 lymphoma cells.

During apoptosis, physical changes in the plasma membrane prepare the cell for clearance by phagocytes and hydrolysis by secretory phospholipase A(2) (sPLA(2)). The relationships among these changes have not been adequately established, especially for hormone-stimulated apoptosis. This study addresses these issues for glucocorticoid-induced apoptosis in S49 lymphoma cells. Flow cytometry, microscopy, and fluorescence spectroscopy were used to assess merocyanine 540 emission, laurdan generalized polarization, phosphatidylserine exposure, caspase activation, and membrane permeability to propidium iodide in the absence and presence of sPLA(2). The earliest event observed was activation of cellular caspases. Results with membrane probes suggest that interlipid spacing also increases early during apoptosis and precedes transbilayer migration of phosphatidylserine, DNA fragmentation, and a general increase in lipid order associated with blebbing and dissolution of the cells. The activity of sPLA(2) appeared to be linked more to lipid spacing than to loss of membrane asymmetry. The early nature of some of these events and their ability to promote activity of a proinflammatory enzyme suggests the possibility of an inflammatory response during T-lymphocyte apoptosis.

Relationship between membrane physical properties and secretory phospholipase A2 hydrolysis kinetics in S49 cells during ionophore-induced apoptosis.

During apoptosis, changes occur in lymphocyte membranes that render them susceptible to hydrolysis by secretory phospholipase A(2) (sPLA(2)). To study the relevant mechanisms, a simplified model of apoptosis using a calcium ionophore was applied. Kinetic and flow cytometry experiments provided key observations regarding ionophore treatment: the initial rate of hydrolysis was elevated at all enzyme concentrations, the total amount of reaction product was increased fourfold, and adsorption of the enzyme to the membrane surface was unaltered. Analysis of these results suggested that susceptibility during calcium-induced apoptosis is limited by availability of substrate rather than adsorption of enzyme. Fluorescence experiments identified three membrane alterations during apoptosis that might affect substrate access to the sPLA(2) active site. First, intercalation of merocyanine 540 into the membrane was improved, suggesting an increase in lipid spacing. Second, laurdan detected increased solvation of the lower headgroup region of the membrane. Third, the rate at which fluorescent lipids could be removed from the membrane by albumin was enhanced, implying greater vertical mobility of phospholipids. Thus, it is proposed that the membranes of apoptotic cells become susceptible to sPLA(2) through a reduction in lipid-neighbor interactions that facilitates migration of phospholipids into the enzyme active site.

Risk, citizenship, and public discourse: coeval dialogues on war and health in Vancouver's Downtown Eastside.

This article is about September 11, 2001, and its narrated effects on the lives of nine street-involved women in Vancouver's Downtown Eastside. I outline the locations from which they spoke about war and health: as consumers and economic agents whose bodies are linked to transnational economic processes; as residents in a local community of shared knowledge and practices; and as marginalized citizens of a nation-state. I hope to emphasize the value of engaging research subjects in coeval dialogues that work against essentializing, state-sanctioned discourses narrated in the context of armed conflict and a public health crisis. To women drug users in Vancouver's Downtown Eastside, the "War against Terror" evokes particular sites of knowledge: the body, the local community, and transnational processes. Their repertoires of war stimulate questions about citizenship and perceptions of risk, challenging dominating medical and political discourses that tend to temporally and spatially localize their engagement with the world.

A role for phosphatidylinositol 3-kinase in TCR-stimulated ERK activation leading to paxillin phosphorylation and CTL degranulation.

PI3K is an important regulator of a number of cellular processes. We examined the contribution of PI3K to mouse CTL signaling, leading to degranulation. We show that TCR-triggered, but not phorbol ester and calcium ionophore-induced, CTL degranulation is dependent on PI3K activity. Although PI3K activity is required for optimal LFA-1-mediated adhesion and cell spreading, this most likely does not account for its full contribution to degranulation. We demonstrate that PI3K is required for TCR-stimulated ERK activation in CTL, which we have shown previously to be required for CTL degranulation. We thus define a pathway through which PI3K most likely regulates degranulation and in which ERK appears to be a key signaling molecule. Furthermore, we identified the cytoskeletal adaptor paxillin as a target of ERK downstream of TCR stimulation. Consistent with a role in degranulation, we demonstrate that paxillin is localized to the microtubule organizing center in resting cells and upon target cell binding is recruited to the contact point with the target cell. These studies demonstrate that PI3K regulates ERK activity leading to CTL degranulation, and identify paxillin as a target of ERK downstream of the TCR. That paxillin is independently phosphorylated by both tyrosine kinase(s) and ERK downstream of the TCR and localized both at the microtubule organizing center and at the target cell contact point suggests an important role for paxillin in CTL-mediated killing.