Neurologic complications after allogeneic hematopoietic stem cell transplantation: risk factors and impact.

Neurologic complications (NCs) may be a significant source of morbidity and mortality after hematopoietic cell transplantation (HCT). We performed a retrospective study of 263 consecutive patients undergoing allogeneic HCT for hematological malignancies to determine the incidence, risk factors and clinical impact of NCs in the first 5 years after HCT. We determined the incidence of central nervous system (CNS) infection, intracranial hemorrhage, ischemic stroke, metabolic encephalopathy, posterior reversal encephalopathy syndrome, seizure and peripheral neuropathy. In all, 50 patients experienced 63 NCs-37 early (⩽day +100), 21 late (day +101 to 2 years) and 5 very late (2 to 5 years). The 1- and 5-year cumulative incidences of all NCs were 15.6% and 19.2%, respectively, and of CNS complication (CNSC; all of the above complications except peripheral neuropathy) were 12.2 and 14.5%. Risk factors for CNSC were age (hazard ratio (HR)=1.06 per year, P=0.0034), development of acute GvHD grade III-IV (HR=2.78, P=0.041), transfusion-dependent thrombocytopenia (HR=3.07, P=0.025) and delayed platelet engraftment (>90th centile; HR=2.77, P=0.043). CNSCs negatively impacted progression-free survival (HR=2.29, P=0.0001), overall survival (HR=2.63, P<0.0001) and non-relapse mortality (HR=8.51, P<0.0001). NCs after HCT are associated with poor outcomes, and usually occur early after HCT.

Regulation of platelet lifespan in the presence and absence of thrombopoietin signaling.

UNLABELLED: Essentials We examined platelet survival in models of absent or enhanced thrombopoietin (TPO) signaling. Platelet lifespan is normal in transgenic mice with chronically enhanced TPO signaling. Mpl deficiency does not negatively affect platelet lifespan in the absence of thrombocytopenia. We conclude that TPO and its receptor Mpl are dispensable for platelet survival in adult mice. SUMMARY: Background It is well established that thrombopoietin (TPO), acting via its receptor Mpl, is the major cytokine regulator of platelet biogenesis. The primary mechanism by which TPO signaling stimulates thrombopoiesis is via stimulation of Mpl-expressing hematopoietic progenitors; Mpl on megakaryocytes and platelets acts to control the amount of TPO available. TPO could potentially reduce platelet and/or megakaryocyte apoptosis, and therefore increase the platelet count. However, the effect of TPO receptor signaling on platelet survival is unresolved. Methods and results Here, we investigated platelet survival in mouse models of absent or enhanced TPO signaling. In the absence of thrombocytopenia, Mpl deficiency did not negatively influence platelet lifespan, and nor was platelet survival affected in transgenic mice with chronically increased TPO signaling. Conclusions We conclude that TPO and its receptor Mpl are dispensable for platelet survival in adult mice.

Quantal and graded stimulation of B lymphocytes as alternative strategies for regulating adaptive immune responses.

Lymphocytes undergo a typical response pattern following stimulation in vivo: they proliferate, differentiate to effector cells, cease dividing and predominantly die, leaving a small proportion of long-lived memory and effector cells. This pattern results from cell-intrinsic processes following activation and the influence of external regulation. Here we apply quantitative methods to study B-cell responses in vitro. Our results reveal that B cells stimulated through two Toll-like receptors (TLRs) require minimal external direction to undergo the basic pattern typical of immunity. Altering the stimulus strength regulates the outcome in a quantal manner by varying the number of cells that participate in the response. In contrast, the T-cell-dependent CD40 activation signal induces a response where division times and differentiation rates vary in relation to stimulus strength. These studies offer insight into how the adaptive antibody response may have evolved from simple autonomous response patterns to the highly regulable state that is now observed in mammals.

Reassessment of the pharmacology of Sphingosine-1-phosphate S1P3 receptor ligands using the DiscoveRx PathHunter™ and Ca2+ release functional assays.

BACKGROUND AND PURPOSE: DiscoverRx's PathHunter™ assay measures GPCR agonist potency, via the recruitment of ß-arrestin, independent of the subtype of G(α) protein activated. This assay is frequently used in drug discovery although little is known about the agonist pharmacology generated. Here we have compared agonist potency, efficacy and affinity values obtained in PathHunter™ assays with those from more established radioligand binding and functional techniques. EXPERIMENTAL APPROACH: Using cells expressing the human sphingosine-1-phosphate S1P(3) receptor at four different densities, we compared pharmacological affinity and efficacy values of four structurally distinct ligands - FTY720-P, VPC24191, CYM5442 and the endogenous agonist S1P - obtained from competition binding, functional Ca(2+) release and PathHunter™ assays. KEY RESULTS: The pK(i) values for S1P were significantly different (9.34 ± 0.10 and 8.92 ± 0.15) in clones expressing different receptor levels using the binding assay. In the PathHunter™ and Ca(2+) assays, S1P and CYM5442 were full agonists, FTY720-P was a partial agonist, while the efficacy of VPC24191 could not be detected in PathHunter™ assays. VPC23019, previously described as a S1P(1/3) receptor antagonist, behaved as an S1P(3) receptor partial agonist in the Ca(2+) release assay. CONCLUSIONS AND IMPLICATIONS: Comparison of data from the PathHunter™ assay with binding and functional Ca(2+) assays suggest that PathHunter™ assays measured a different agonist-bound receptor conformation. While this assay has great utility in drug discovery, care must be taken as high-efficacy, low-affinity agonist compounds would not be detected. Therefore highly amplified, more traditional assays are necessary to identify agonists with low efficacy.

Automated and semi-automated cell tracking: addressing portability challenges.

Cell tracking is a key task in the high-throughput quantitative study of important biological processes, such as immune system regulation and neurogenesis. Variability in cell density and dynamics in different videos, hampers portability of existing trackers across videos. We address these potability challenges in order to develop a portable cell tracking algorithm. Our algorithm can handle noise in cell segmentation as well as divisions and deaths of cells. We also propose a parameter-free variation of our tracker. In the tracker, we employ a novel method for recovering the distribution of cell displacements. Further, we present a mathematically justified procedure for determining the gating distance in relation to tracking performance. For the range of real videos tested, our tracker correctly recovers on average 96% of cell moves, and outperforms an advanced probabilistic tracker when the cell detection quality is high. The scalability of our tracker was tested on synthetic videos with up to 200 cells per frame. For more challenging tracking conditions, we propose a novel semi-automated framework that can increase the ratio of correctly recovered tracks by 12%, through selective manual inspection of only 10% of all frames in a video.

A model of immune regulation as a consequence of randomized lymphocyte division and death times.

The magnitude of an adaptive immune response is controlled by the interplay of lymphocyte quiescence, proliferation, and apoptosis. How lymphocytes integrate receptor-mediated signals influencing these cell fates is a fundamental question for understanding this complex system. We examined how lymphocytes interleave times to divide and die to develop a mathematical model of lymphocyte growth regulation. This model provides a powerful method for fitting and analyzing fluorescent division tracking data and reveals how summing receptor-mediated kinetic changes can modify the immune response progressively from rapid tolerance induction to strong immunity. An important consequence of our results is that intrinsic variability in otherwise identical cells, usually dismissed as noise, may have evolved to be an essential feature of immune regulation.