Assessment of on-treatment platelet reactivity at high and low shear stress and platelet activation status after the addition of dipyridamole to aspirin in the early and late phases after TIA and ischaemic stroke.

BACKGROUND: Data are limited on the ability of dipyridamole to additionally inhibit platelet function/reactivity in ischaemic cerebrovascular disease (CVD) patients on aspirin. AIMS: To assess inhibition of platelet function/reactivity and platelet activation with dipyridamole in CVD. METHODS: This prospective, observational study assessed TIA/ischaemic stroke patients before (baseline; N = 60), at 14 ±7 days (14d, N = 39) and ≥ 90 days (90d, N = 31) after adding dipyridamole to aspirin. Platelet function/reactivity at high shear stress (PFA-100® C-ADP) and low shear stress (VerifyNow® P2Y12 and Multiplate® ADP assays), and platelet activation status (% expression of CD62P, CD63 and leucocyte-platelet complexes on whole blood flow cytometry) were quantified. 'Dipyridamole-high on-treatment platelet reactivity (HTPR)' was defined as failure to inhibit ADP-induced platelet aggregation +/- adhesion compared with the patient's baseline on aspirin monotherapy by more than twice the coefficient-of-variation of the assay after adding dipyridamole to aspirin. RESULTS: Dipyridamole-HTPR was identified in 71.4-75% of patients on PFA-100 C-ADP, 83.9-86.8% of patients on VerifyNow P2Y12, and 81.5-83.3% of patients on Multiplate ADP assays. There were no changes in CD62P/CD63 expression (P ≥ 0.18), or consistent changes in leucocyte-platelet complexes in CVD patients overall at 14d or 90d vs. baseline after commencing dipyridamole. Monocyte-platelet complexes increased in the patient subgroup with dipyridamole-HTPR at 14d and 90d on PFA-100, and at 14d on VerifyNow (P ≤ 0.04), but not in those without dipyridamole-HTPR. DISCUSSION: Additional antiplatelet effects of dipyridamole are detectable under high and low shear stress conditions with user-friendly platelet function/reactivity tests ex vivo. Increasing circulating monocyte-platelet complexes over time are associated with dipyridamole-HTPR.

A longitudinal, observational study of women who persisted in smoking in successive pregnancies.

BACKGROUND: This longitudinal study examined the profile and pregnancy-related behaviours of women who reported smoking in two successive pregnancies when they presented for prenatal care in a large maternity hospital. METHODS: Using the hospital electronic medical records, women who delivered two successive singleton pregnancies during the years 2011-15 were analyzed. Standardized data were computerized by a midwife at the first prenatal visit, following delivery and before discharge. RESULTS: Over the 5 years, 6647 women delivered twice. Overall 5754 (86.6%) were persistent non-smokers in both pregnancies, 609 (9.2%) were persistent smokers in both pregnancies and between pregnancies 202 (3.0%) quit and 82 (1.2%) started smoking. Compared with persistent non-smokers, persistent smokers had higher rates of reported illicit drug use, alcohol consumption and psychological problems and lower rates of planned pregnancy, folic acid supplementation and breastfeeding in both pregnancies (all P < 0.001). In persistent smokers, folic acid supplementation practices deteriorated and illicit drug use increased in the subsequent pregnancy. CONCLUSIONS: We found that approximately one in 10 women smoked in two consecutive pregnancies. Furthermore, compared with non-smokers, persistent smokers were more likely to report other health behaviours associated with adverse pregnancy outcomes and may require additional multidisciplinary support.

A National Audit of Smoking Cessation Services in Irish Maternity Units.

There is international consensus that smoking cessation in the first half of pregnancy improves foetal outcomes. We surveyed all 19 maternity units nationally about their antenatal smoking cessation practices. All units recorded details on maternal smoking at the first antenatal visit. Only one unit validated the self-reported smoking status of pregnant women using a carbon monoxide breath test. Twelve units (63%) recorded timing of smoking cessation. In all units women who reported smoking were given verbal cessation advice. This was supported by written advice in 12 units (63%), but only six units (32%) had all midwives trained to provide this advice. Only five units (26%) reported routinely revisiting smoking status later in pregnancy. Although smoking is an important modifiable risk factor for adverse pregnancy outcomes, smoking cessation services are inadequate in the Irish maternity services and there are variations in practices between hospitals.

Profile of von Willebrand factor antigen and von Willebrand factor propeptide in an overall TIA and ischaemic stroke population and amongst subtypes.

INTRODUCTION: Von Willebrand factor propeptide (VWF:Ag II) is proposed to be a more sensitive marker of acute endothelial activation than von Willebrand factor antigen (VWF:Ag). Simultaneous data on VWF:Ag and VWF:Ag II profiles are very limited following TIA and ischaemic stroke. METHODS: In this prospective, observational, case-control study, plasma VWF:Ag and VWF:Ag II levels were quantified in 164 patients≤4weeks of TIA or ischaemic stroke (baseline), and then ≥14days (14d) and ≥90days (90d) later, and compared with those from 27 healthy controls. TIA and stroke subtyping was performed according to the TOAST classification. The relationship between VWF:Ag and VWF:Ag II levels and platelet activation status was assessed. RESULTS: 'Unadjusted' VWF:Ag and VWF:Ag II levels were higher in patients at baseline, 14d and 90d than in controls (p≤0.03). VWF:Ag levels remained higher in patients than controls at baseline (p≤0.03), but not at 14d or 90d after controlling for differences in age or hypertension, and were higher in patients at baseline and 90d after controlling for smoking status (p≤0.04). 'Adjusted' VWF:Ag II levels were not higher in patients than controls after controlling for age, hypertension or smoking (p≥0.1). Patients with symptomatic carotid stenosis (N=46) had higher VWF:Ag and VWF:Ag II levels than controls at all time-points (p≤0.002). There was no significant correlation between platelet activation status and VWF:Ag or VWF:Ag II levels. CONCLUSIONS: VWF:Ag and VWF:Ag II levels are increased in an overall TIA and ischaemic stroke population, especially in patients with recently symptomatic carotid stenosis. VWF:Ag II was not superior to VWF:Ag at detecting acute endothelial activation in this cohort and might reflect timing of blood sampling in our study.

Five year trends in maternal smoking behaviour reported at the first prenatal appointment.

BACKGROUND: Maternal smoking is a key modifiable risk factor in preventing adverse pregnancy outcomes such as intrauterine growth restriction, preterm birth and stillbirth. AIM: This observational study examined annual trends of maternal smoking reported at the first prenatal visit in women who delivered in a large university maternity hospital for the 5 years 2011-2015. METHODS: We examined clinical and sociodemographic data computerised routinely for women who presented for prenatal care at the hospital between 2011 and 2015. Multinomial logistic regression was used to determine the maternal characteristics, health behaviours and psychiatric history associated with smoking behaviours. RESULTS: Of the 42,509 women the mean age was 31.4 ± 5.5 years, mean Body Mass Index (BMI) was 25.6 ± 5.1 kg/m2, and 39.5% were nulliparas. Overall, 52.6% reported they had never smoked, 34.9% were ex-smokers, 10.5% smoked ≤10 cigarettes per day, 1.9% smoked ≥11 cigarettes per day and 0.1% smoked e-cigarettes. Between 2011 and 2015 the prevalence of maternal cigarette smoking decreased from 14.3 to 10.9% (P < 0.001). Smoking during pregnancy was most strongly associated with younger age, multiparity, unemployment, unplanned pregnancy, a history of psychiatric problems, alcohol intake and illicit drug usage. CONCLUSIONS: The number of women who reported smoking at the first prenatal visit decreased annually. Amongst women who continue to smoke during pregnancy, there is a clustering of adverse lifestyle behaviour and psychological problems that may need to be addressed if smoking cessation interventions are going to succeed in improving fetal programming.

Increased platelet activation in early symptomatic vs. asymptomatic carotid stenosis and relationship with microembolic status: results from the Platelets and Carotid Stenosis Study.

BACKGROUND: Cerebral microembolic signals (MES) may predict increased stroke risk in carotid stenosis. However, the relationship between platelet counts or platelet activation status and MES in symptomatic vs. asymptomatic carotid stenosis has not been comprehensively assessed. SETTING: University teaching hospitals. METHODS: This prospective, pilot observational study assessed platelet counts and platelet activation status, and the relationship between platelet activation and MES in asymptomatic vs. early (≤ 4 weeks after TIA/stroke) and late phase (≥ 3 months) symptomatic moderate or severe (≥ 50%) carotid stenosis patients. Full blood count measurements were performed, and whole blood flow cytometry was used to quantify platelet surface activation marker expression (CD62P and CD63) and circulating leucocyte-platelet complexes. Bilateral simultaneous transcranial Doppler ultrasound monitoring of the middle cerebral arteries was performed for 1 h to classify patients as MES positive or MES negative. RESULTS: Data from 31 asymptomatic patients were compared with 46 symptomatic patients in the early phase, and 35 of these patients were followed up to the late phase after symptom onset. The median platelet count (211 vs. 200 × 10(9)  L(-1) ; P = 0.03) and the median percentage of lymphocyte-platelet complexes was higher in early symptomatic than asymptomatic patients (2.8 vs. 2.4%; P = 0.001). The percentage of lymphocyte-platelet complexes was higher in early symptomatic than in asymptomatic patients with ≥ 70% carotid stenosis (P = 0.0005) and symptomatic patients recruited within 7 days of symptom onset (P = 0.028). Complete TCD data were available in 25 asymptomatic, 31 early phase symptomatic and 27 late phase symptomatic patients. Twelve per cent of asymptomatic vs. 32% of early phase symptomatic (P = 0.02) and 19% of late phase symptomatic patients (P = 0.2) were MES positive. Early symptomatic MES-negative patients had a higher percentage of lymphocyte-platelet complexes than asymptomatic MES-negative patients (2.8 vs. 2.3%; P = 0.0085). DISCUSSION: Recently, symptomatic carotid stenosis patients have had higher platelet counts (potentially reflecting increased platelet production, mobilization or reduced clearance) and platelet activation status than asymptomatic patients. MES were more frequently detected in early symptomatic than asymptomatic patients, but the differences between late symptomatic and asymptomatic groups were not significant. Increased lymphocyte-platelet complex formation in recently symptomatic vs. asymptomatic MES-negative patients indicates enhanced platelet activation in this early symptomatic subgroup. Platelet biomarkers, in combination with TCD, have the potential to aid risk-stratification in asymptomatic and symptomatic carotid stenosis patients.

The cost utility of a multi-disciplinary foot protection clinic (MDFPC) in an Irish hospital setting.

BACKGROUND: Foot ulceration which may result in lower limb amputation is one of the most feared complications among patients with diabetes and the prevention of both ulceration and amputation is a major challenge facing the health service. Many studies have proposed dedicated diabetic foot teams as the future of diabetic foot care. AIMS: We aimed to quantify the cost benefit and sustainability of a multi-disciplinary foot protection clinic (MDFPC) in an Irish university hospital setting. METHODS: A dedicated bi-weekly consultant-led MDFPC including Vascular Surgery, Endocrinology, Orthopaedic Surgery, Podiatry, Orthotics and Tissue Viability was established in June 2008. RESULTS: Between 2006 and 2010, a total of 221 lower limb procedures (major/minor amputations and debridement) were performed. The number of major amputations decreased from 12 during the control period (2 years before the clinic) to 7 in the study period (2 years after the clinic). After costing all activity associated with the clinic, there was an overall saving of €114,063 per year associated with the introduction of the MDFPC. CONCLUSION: This is the first study in an Irish context, and one of few international studies, to demonstrate that an aggressive-coordinated approach to diabetic foot care is both cost effective and clinically efficient in reducing the burden of foot-related complications in a diabetic population.

High on-treatment platelet reactivity on commonly prescribed antiplatelet agents following transient ischaemic attack or ischaemic stroke: results from the Trinity Antiplatelet Responsiveness (TRAP) study.

BACKGROUND AND PURPOSE: The prevalence of ex vivo 'high on-treatment platelet reactivity' (HTPR) to antiplatelet regimens in patients with ischaemic cerebrovascular disease (CVD) is uncertain. METHODS: HTPR was assessed with PFA-100 collagen-epinephrine (C-EPI) and collagen-ADP (C-ADP) cartridges. Platelet activation (CD62P, CD63 and leucocyte-platelet complex formation) was assessed with whole-blood flow cytometry. Patients were assessed at baseline [≤ 4 weeks of transient ischaemic attack (TIA) or ischaemic stroke], and at 14 days and ≥ 90 days after changing treatment from (i) no medication to aspirin monotherapy (N = 26) or (ii) aspirin to clopidogrel monotherapy (N = 22). HTPR was defined in a novel, 'longitudinal fashion' as failure to prolong relevant closure times compared with the patient's 'baseline value' before he/she underwent an antiplatelet change by more than twice the coefficient of variation of the assay. RESULTS: (i) C-EPI closure times increased at 14 days and 90 days after commencing aspirin (P = 0.002); 24% at 14 days and 18% at 90 days demonstrated HTPR on aspirin. (ii) C-ADP closure times increased at 14 days (P = 0.001) but not 90 days (P = 0.09) after changing from aspirin to clopidogrel; 41% at 14 days, and 35% at 90 days demonstrated HTPR on clopidogrel. Platelet activation was unaffected by aspirin (P = 0.09). The percentage neutrophil-platelet complexes decreased at 14 days (P = 0.02), but this reduction was not maintained 90 days after changing to clopidogrel (P = 0.3). No patient had a recurrent vascular event during prospective follow-up. CONCLUSIONS: Longitudinal definitions of HTPR in patients with ischaemic CVD who are undergoing a change in antiplatelet therapy have the potential to provide more clinically meaningful information than traditional 'cross-sectional definitions' of HTPR which are usually based on the comparison of patients' values with those in healthy controls. Using our novel, longitudinal definition of HTPR, the PFA-100 could be used to monitor ex vivo responsiveness to aspirin, and larger, prospective studies are warranted to assess the clinical predictive value of this and other platelet function tests in patients with ischaemic CVD.

Increased expression of IRF4 and ETS1 in CD4+ cells from patients with intermittent allergic rhinitis.

BACKGROUND: The transcription factor (TF) IRF4 is involved in the regulation of Th1, Th2, Th9, and Th17 cells, and animal studies have indicated an important role in allergy. However, IRF4 and its target genes have not been examined in human allergy. METHODS: IRF4 and its target genes were examined in allergen-challenged CD4(+) cells from patients with IAR, using combined gene expression microarrays and chromatin immunoprecipitation chips (ChIP-chips), computational target prediction, and RNAi knockdowns. RESULTS: IRF4 increased in allergen-challenged CD4(+) cells from patients with IAR, and functional studies supported its role in Th2 cell activation. IRF4 ChIP-chip showed that IRF4 regulated a large number of genes relevant to Th cell differentiation. However, neither Th1 nor Th2 cytokines were the direct targets of IRF4. To examine whether IRF4 induced Th2 cytokines via one or more downstream TFs, we combined gene expression microarrays, ChIP-chips, and computational target prediction and found a putative intermediary TF, namely ETS1 in allergen-challenged CD4(+) cells from allergic patients. ETS1 increased significantly in allergen-challenged CD4(+) cells from patients compared to controls. Gene expression microarrays before and after ETS1 RNAi knockdown showed that ETS1 induced Th2 cytokines as well as disease-related pathways. CONCLUSIONS: Increased expression of IRF4 in allergen-challenged CD4(+) cells from patients with intermittent allergic rhinitis leads to activation of a complex transcriptional program, including Th2 cytokines.

Popliteo-pedal bypass surgery for critical limb ischemia.

BACKGROUND: Critical limb ischaemia due to distal arterial disease represents a significant challenge. Randomised controlled evidence suggests that open surgery may be superior to endovascular intervention but there is limited data on the specific clinical cohort with exclusively infra-popliteal disease. AIM: We analysed indications for, and outcome from all, popliteo-pedal bypass procedures performed between July 1998 to November 2008. PATIENTS AND METHODS: Twenty-eight bypass procedures were performed in 24 patients. Autologous vein was used exclusively. The proximal anastomosis was to the below-knee popliteal artery in all the patients; the distal anastomosis was to plantar artery (n = 15) or dorsalis pedis artery (n = 13). Mean patient age was 63.Eight years of age (range 37-92 years). Indications for surgery were tissue loss (n = 21) and rest pain (n = 7). Ultrasound graft surveillance was performed every 6-months. RESULTS: Using life table analysis, primary graft patency was 63.3% at 1-, 3- and 5-years and secondary patency (after three interventions) was 74.6% at 1-, 3- and 5-years. Limb salvage rate was 81.8% after 1-, 3- and 5-years as all five limb amputations were performed in the first 3-months following the surgery. Overall survival was 75, 75 and 47.1% at 1-, 3- and 5-years, respectively. The major amputation free survival rate was 54.2, 54.2 and 21.3% at 1-, 3- and 5-years, respectively. Seventy-nine percent (n = 19) patients were diabetic. CONCLUSION: Our data supports popliteo-pedal bypass as an effective treatment for distal vascular disease. Comparison with endovascular treatment in a randomised trial needs to be performed.

Changing trends in surgical treatment of carotid disease in Ireland (1996-2003).

BACKGROUND: Carotid endarterectomy (CEA) is a well-established method of stroke prevention in patients with symptomatic, high-grade internal carotid artery stenosis. AIMS: To assess the change in carotid surgery practice in health board regions in Ireland over two different time periods in the past 11 years (1996-1998/2001-2003). METHODS: Numbers of discharges of patients with a procedure code CEA (38.12) between 1996-1998 and 2001-2003 were obtained from the Hospital In-Patient Enquiry (HIPE) database maintained by the Economic and Social Research Institute (ESRI). Population data was obtained from national censuses. RESULTS: There has been considerable change in the level of carotid surgery activity in specific health board regions between the two periods. CONCLUSIONS: Still no region comes close to achieving levels of CEA recommended by population studies. This is important in the context of the MRC asymptomatic carotid surgery trial (ACST), as the numbers suitable for CEA will probably increase further in the future.

Transfusion requirements and outcomes in patients undergoing abdominal aortic surgery using intra-operative cell salvage.

BACKGROUND: Intraoperative cell salvage (ICS) is the recovery, anticoagulation, filtration and reinfusion of blood lost during surgery. The aim of this study is to determine the safety and efficacy of ICS in emergency and elective abdominal aortic surgery. METHODS: This study reviews volumes of blood loss, blood salvaged with ICS, allogenic blood requirements, and clinical outcomes in patients undergoing abdominal aortic surgery using ICS. RESULTS: Seventy-nine patients undergoing abdominal aortic surgery are included. Supplemental allogenic blood was not required in 45/79 (57%) of all patients. Transfusion with allogenic blood was not necessary in 41/63 (66%) of elective abdominal aortic aneurysm repairs. ICS was associated with no major complications. CONCLUSION: ICS is a safe procedure and substantially reduces the need for blood transfusion in patients undergoing abdominal aortic surgery. It may substantially alleviate shortages of allogenic blood and should be part of the armamentarium of vascular units.

Genetic Mechanisms and Aberrant Gene Expression during the Development of Gastric Intestinal Metaplasia and Adenocarcinoma.

Gastric adenocarcinoma occurs via a sequence of molecular events known as the Correa's Cascade which often progresses over many years. Gastritis, typically caused by infection with the bacterium H. pylori, is the first step of the cascade that results in gastric cancer; however, not all cases of gastritis progress along this carcinogenic route. Despite recent antibiotic intervention of H. pylori infections, gastric adenocarcinoma remains the second most common cause of cancer deaths worldwide. Intestinal metaplasia is the next step along the carcinogenic sequence after gastritis and is considered to be a precursor lesion for gastric cancer; however, not all patients with intestinal metaplasia develop adenocarcinoma and little is known about the molecular and genetic events that trigger the progression of intestinal metaplasia into adenocarcinoma. This review aims to highlight the progress to date in the genetic events involved in intestinal-type gastric adenocarcinoma and its precursor lesion, intestinal metaplasia. The use of technologies such as whole genome microarray analysis, immunohistochemical analysis and DNA methylation analysis has allowed an insight into some of the events which occur in intestinal metaplasia and may be involved in carcinogenesis. There is still much that is yet to be discovered surrounding the development of this lesion and how, in many cases, it develops into a state of malignancy.

Effect of intravenous taurine on endotoxin-induced acute lung injury in sheep.

OBJECTIVE: To find out if pretreatment with taurine would reduce the severity of endotoxin-induced acute lung injury in a large animal model. DESIGN: Randomised controlled study under licence from the Department of Health. SETTING: Department of Surgical Research, Ireland. ANIMALS: 15 male Suffolk sheep. INTERVENTIONS: Vascular catheters were placed in the femoral artery and vein and a Swan-Ganz catheter in the external jugular vein under general anaesthetic. Animals were randomized into three groups: control with measurements taken at baseline and half hourly up to 90 minutes; endotoxin, given Escherichia coli endotoxin intravenously after baseline measurements and taurine given 300 mg/kg 1 hour before endotoxin was given. MAIN OUTCOME MEASURES: Mean systemic arterial pressure, mean pulmonary arterial pressure, arterial oxygen tension (PO2), pulmonary myeloperoxidase activity, and neutrophil respiratory burst activity. RESULTS: Endotoxin induced a severe lung injury characterised by a decrease in mean systemic blood pressure and an increase in pulmonary artery pressure, hypoxia, and an increase in pulmonary myeloperoxidase activity. Pretreatment with intravenous taurine significantly reduced these haemodynamic changes. It reduced pulmonary myeloperoxidase activity and peripheral neutropenia and increased neutrophil respiratory burst activity. CONCLUSIONS: This data suggest that taurine may have a therapeutic role in preventing the lung injury seen in endotoxaemia.

Signaling events mediating the additive effects of oleic acid and angiotensin II on vascular smooth muscle cell migration.

Obese hypertensive patients with cardiovascular risk factor clustering and increased risk for atherosclerotic disease have increased plasma nonesterified fatty acid levels, including oleic acid (OA), and a more active renin-angiotensin-aldosterone system. Vascular smooth muscle cell (VSMC) migration and proliferation participate in the development of atherosclerotic plaque. OA and angiotensin (Ang) II induce synergistic mitogenic responses in VSMCs through sequential signaling pathways dependent on the activation of protein kinase C (PKC), oxidants (reactive oxygen species, ROS), and extracellular signal-regulated kinase (ERK) activation. We tested the hypotheses that (1) OA and Ang II have additive or synergistic effects on VSMC migration and (2) PKC, ROS, and mitogen-activated protein kinase are critical signaling molecules. OA at 100 micromol/L increases VSMC migration 60+/-10% over control (P:<0.001). Ang II (10(-)(9) mol/L) increases VSMC migration by 62+/-13% and 73% over control, respectively (P:<0.01). Coincubation of cells with OA and Ang II produces a nearly additive increase in VSMC cell migration at 107+/-20% (P:<0.01). Increases in VSMC migration induced by OA alone and combined with Ang II were reduced by PKC inhibition and downregulation. VSMC migration in response to OA alone and with Ang II was also inhibited by N:-acetyl-cysteine, MEK inhibition, and ERK antisense. VSMC migration in response to OA alone or combined with Ang II is dependent on activation of PKC, ROS, and ERK activation, further raising the possibility that increased plasma nonesterified fatty acids and an activated renin-angiotensin-aldosterone system in subjects with the risk factor cluster contribute to accelerated atherosclerosis through a PKC, ROS, and ERK-dependent signaling pathway.

Bone marrow mobilized with granulocyte colony-stimulating factor in related allogeneic transplant recipients: a study of 29 patients.

We studied whether a short course of granulocyte colony-stimulating factor (G-CSF) administered to normal donors immediately before bone marrow (BM) harvest would shorten time to neutrophil and platelet engraftment in matched related allogeneic BM recipients. Twenty-nine normal donors received 4 consecutive daily subcutaneous injections of G-CSF (median dose, 12.1 microg/kg per day; range, 9.6-15.7 microg/kg per day) immediately before BM harvest. Donors tolerated G-CSF well, with only mild myalgias and arthralgias, and BM was easy to aspirate. The BM harvest contained a median of 5.3 x 10(8) white blood cells (WBCs)/kg (range, 3.1-11.1 x 10(8) WBCs/kg) and 2.5 x 10(6) CD34+ cells per kg (range, 1.5-7.3 x 10(6) CD34+ cells per kg). Median times to neutrophil (18 days [range, 11-30 days] versus 22 days [range, 16-36 days]; P = .05) and platelet (22 days [range, 15-55 days] versus 27 days [range, 18-46 days]; P = .04) engraftment were statistically shorter than those of historical control subjects whose donors had not received G-CSF before BM harvest. However, secondary engraftment-dependent outcomes including red blood cell and platelet transfusions, febrile days, days on antibiotics, days from transplant to hospital discharge, and days in hospital during the first 60 days after transplant were not statistically different from historical control subjects. We conclude that G-CSF administered to normal donors immediately before harvest facilitates BM aspiration, increases the WBC content of the harvest, and hastens neutrophil and platelet engraftment compared with historical control subjects.

Targeting of tail-anchored proteins to yeast mitochondria in vivo.

Tail-anchored proteins are inserted into intracellular membranes via a C-terminal transmembrane domain. The topology of the protein is such that insertion must occur post-translationally, since the insertion sequence is not available for membrane insertion until after translation of the tail-anchored polypeptide is completed. Here, we show that the targeting information in one such tail-anchored protein, translocase in the outer mitochondrial membrane 22, is contained in a short region flanking the transmembrane domain. An equivalent region is sufficient to specify the localisation of Bcl2 and SNARE proteins to the secretory membranes. We discuss the targeting process for directing members of this protein family to the secretory and mitochondrial membranes in vivo.