Next generation physiologically based kinetic (NG-PBK) models in support of regulatory decision making.

The fields of toxicology and chemical risk assessment seek to reduce, and eventually replace, the use of animals for the prediction of toxicity in humans. In this context, physiologically based kinetic (PBK) modelling based on in vitro and in silico kinetic data has the potential to a play significant role in reducing animal testing, by providing a methodology capable of incorporating in vitro human data to facilitate the development of in vitro to in vivo extrapolation of hazard information. In the present article, we discuss the challenges in: 1) applying PBK modelling to support regulatory decision making under the toxicology and risk-assessment paradigm shift towards animal replacement; 2) constructing PBK models without in vivo animal kinetic data, while relying solely on in vitro or in silico methods for model parameterization; and 3) assessing the validity and credibility of PBK models built largely using non-animal data. The strengths, uncertainties, and limitations of PBK models developed using in vitro or in silico data are discussed in an effort to establish a higher degree of confidence in the application of such models in a regulatory context. The article summarises the outcome of an expert workshop hosted by the European Commission Joint Research Centre (EC-JRC) - European Union Reference Laboratory for Alternatives to Animal Testing (EURL ECVAM), on "Physiologically-Based Kinetic modelling in risk assessment - reaching a whole new level in regulatory decision-making" held in Ispra, Italy, in November 2016, along with results from an international survey conducted in 2017 and recently reported activities occurring within the PBK modelling field. The discussions presented herein highlight the potential applications of next generation (NG)-PBK modelling, based on new data streams.

Intergenerational and early life influences on the well-being of Australian Aboriginal and Torres Strait Islander children: overview and selected findings from Footprints in Time, the Longitudinal Study of Indigenous Children.

Footprints in Time: The Longitudinal Study of Indigenous Children (LSIC) is a national study of 1759 Australian Aboriginal and Torres Strait Islander children living across urban, regional and remote areas of Australia. The study is in its 11th wave of annual data collection, having collected extensive data on topics including birth and early life influences, parental health and well-being, identity, cultural engagement, language use, housing, racism, school engagement and academic achievement, and social and emotional well-being. The current paper reviews a selection of major findings from Footprints in Time relating to the developmental origins of health and disease for Australian Aboriginal and Torres Strait Islander peoples. Opportunities for new researchers to conduct further research utilizing the LSIC data set are also presented.

Cerenkov emission of quasiparallel whistlers by fast electron phase-space holes during magnetic reconnection.

Kinetic simulations of magnetotail reconnection have revealed electromagnetic whistlers originating near the exhaust boundary and propagating into the inflow region. The whistler production mechanism is not a linear instability, but rather is Cerenkov emission of almost parallel whistlers from localized moving clumps of charge (finite-size quasiparticles) associated with nonlinear coherent electron phase space holes. Whistlers are strongly excited by holes without ever growing exponentially. In the simulation the whistlers are emitted in the source region from holes that accelerate down the magnetic separatrix towards the x line. The phase velocity of the whistlers vφ in the source region is everywhere well matched to the hole velocity vH as required by the Cerenkov condition. The simulation shows emission is most efficient near the theoretical maximum vφ=half the electron Alfven speed, consistent with the new theoretical prediction that faster holes radiate more efficiently. While transferring energy to whistlers the holes lose coherence and dissipate over a few local ion inertial lengths. The whistlers, however, propagate to the x line and out over many 10's of ion inertial lengths into the inflow region of reconnection. As the whistlers pass near the x line they modulate the rate at which magnetic field lines reconnect.

Paget-Schroetter syndrome treated with cutting-balloon angioplasty.

Here, a case of Paget-Schroetter Syndrome in a 25-year-old guitar player is reported. After thrombolysis, conventional angioplasty failed to dilate the underlying subclavian stenosis both before and after first-rib excision with scalenus anterior and medius division. For the third attempt at angioplasty, a cutting balloon was used, which immediately produced a good result. Venography at 4-year follow-up showed no restenosis and no functional deficit. This case report demonstrates that cutting-balloon angioplasty may be considered when conventional balloon fails and may have greater durability than conventional balloon angioplasty in the treatment of Paget-Schroetter syndrome.

Variations in the quality of consent for open mesh repair of inguinal hernia.

INTRODUCTION: Correctly performed informed consent acts as a shield against complaints by patients and claims of malpractice against doctors. Inguinal hernia repair accounts for 10% of the general surgical workload and shows a marked variation in the consenting of individuals. AIM: To compare the variations in consenting practice among various grades of surgeons for the primary repair of open inguinal hernia with specific reference to the documentation of significant risk of surgery. METHOD: A proforma was devised which included the grade of the surgeon and significant and/or commonly recognised complications of the open repair of inguinal hernia. This was then cross-referenced with the consent forms for the 47 patients and the documented risks explained in each case were noted. RESULT: A considerable variation was noted in the documentation of complications by various grades of surgeons. Of the 47 consent forms assessed, only five were completed by the consultant who was performing the operation, 26 were completed by Specialist Registrars (SpRs), 14 by Senior House Officers (SHOs), one by a staff grade and one by an intern. Consultants are less likely to mention recurrence (60%) than juniors (SpR 88% and SHO 93%). They are, however, more likely to mention the complication of chronic pain (60% vs. 35% for SpRs and 7% for SHOs). CONCLUSION: Patients are not provided with consistent information to make informed consent. There is a need for standardised consent forms to achieve consistency and effectiveness of the consenting practice of inguinal hernia surgery.

A magnetic reconnection X-line extending more than 390 Earth radii in the solar wind.

Magnetic reconnection in a current sheet converts magnetic energy into particle energy, a process that is important in many laboratory, space and astrophysical contexts. It is not known at present whether reconnection is fundamentally a process that can occur over an extended region in space or whether it is patchy and unpredictable in nature. Frequent reports of small-scale flux ropes and flow channels associated with reconnection in the Earth's magnetosphere raise the possibility that reconnection is intrinsically patchy, with each reconnection X-line (the line along which oppositely directed magnetic field lines reconnect) extending at most a few Earth radii (R(E)), even though the associated current sheets span many tens or hundreds of R(E). Here we report three-spacecraft observations of accelerated flow associated with reconnection in a current sheet embedded in the solar wind flow, where the reconnection X-line extended at least 390R(E) (or 2.5 x 10(6) km). Observations of this and 27 similar events imply that reconnection is fundamentally a large-scale process. Patchy reconnection observed in the Earth's magnetosphere is therefore likely to be a geophysical effect associated with fluctuating boundary conditions, rather than a fundamental property of reconnection. Our observations also reveal, surprisingly, that reconnection can operate in a quasi-steady-state manner even when undriven by the external flow.

Composition and dynamics of plasma in Saturn's magnetosphere.

During Cassini's initial orbit, we observed a dynamic magnetosphere composed primarily of a complex mixture of water-derived atomic and molecular ions. We have identified four distinct regions characterized by differences in both bulk plasma properties and ion composition. Protons are the dominant species outside about 9 RS (where RS is the radial distance from the center of Saturn), whereas inside, the plasma consists primarily of a corotating comet-like mix of water-derived ions with approximately 3% N+. Over the A and B rings, we found an ionosphere in which O2+ and O+ are dominant, which suggests the possible existence of a layer of O2 gas similar to the atmospheres of Europa and Ganymede.

Autonomic innervation of the human cardiac conduction system: changes from infancy to senility--an immunohistochemical and histochemical analysis.

In order to study the changes in the pattern of autonomic innervation of the human cardiac conduction system in relation to age, the innervation of the conduction system of 24 human hearts (the age of the individuals ranged from newborn to 80 years), freshly obtained at autopsy, was evaluated by a combination of immunofluorescence and histochemical techniques. The pattern of distribution and density of nerves exhibiting immunoreactivity against protein gene product 9.5 (PGP), a general neural marker, dopamine beta-hydroxylase (DBH) and tyrosine hydroxylase (TH), indicators for presumptive sympathetic neural tissue, and those demonstrating positive acetylcholinesterase (AChE) activity, were studied. All these nerves showed a similar pattern of distribution and developmental changes. The density of innervation, assessed semiquantitatively, was highest in the sinus node, and exhibited a decreasing gradient through the atrioventricular node, penetrating and branching bundle, to the bundle branches. Other than a paucity of those showing AChE activity, nerves were present in substantial quantities in infancy. They then increased in density to a maximum in childhood, at which time the adult pattern was achieved and then gradually decreased in density in the elders to a level similar to or slightly less than that in infancy. In contrast, only scattered AChE-positive nerves were found in the sinus and atrioventricular nodes, but were absent from the bundle branches of the infant heart, whereas these conduction tissues themselves possessing a substantial amount of pseudocholinesterase. During maturation into adulthood, however, the conduction tissues gradually lost their content of pseudocholinesterase but acquired a rich supply of AChE-positive nerves, comparable in density to those of DBH and TH nerves. The decline in density of AChE-positive nerves in the conduction tissues in the elders was also similar to those of DBH and TH nerves. Our findings of initial sympathetic dominance in the neural supply to the human cardiac conduction system in infancy, and its gradual transition into a sympathetic and parasympathetic codominance in adulthood, correlate well with the physiologic alterations known to occur in cardiac rate during postnatal development. The finding of reduction in density of innervation of the conduction tissue with ageing is also in agreement with clinical and electrophysiological findings such as age-associated reduction in cardiac response to parasympathetic stimulation. Finally, our findings also support the hypothesis that, in addition to the para-arterial route, the parafascicular route of extension along the conduction tissue constitutes another pathway for the innervation of the conduction system of the human heart during development.

Enhanced pulmonary allergic responses to Aspergillus in CCR2-/- mice.

Allergic responses to Aspergillus species exacerbate asthma and cystic fibrosis. The natural defense against live Aspergillus fumigatus spores or conidia depends on the recruitment and activation of mononuclear and polymorphonuclear leukocytes, events that are dependent on chemotactic cytokines. In this study, we explored the relative contribution of the monocyte chemoattractant protein-1 receptor, CCR2, in the pulmonary response to A. fumigatus conidia. Following sensitization to soluble A. fumigatus Ags, mice lacking CCR2 due to targeted deletion were markedly more susceptible to the injurious effects of an intrapulmonary challenge with live conidia compared with mice that expressed CCR2 or CCR2+/+. CCR2-/- mice exhibited a major defect in the recruitment of polymorphonuclear cells, but these mice also had significantly more eosinophils and lymphocytes in bronchoalveolar lavage samples. CCR2-/- mice also had significant increases in serum levels of total IgE and whole lung levels of IL-5, IL-13, eotaxin, and RANTES compared with CCR2+/+ mice. Airway inflammation, hyper-responsiveness to spasmogens, and subepithelial fibrosis were significantly enhanced in CCR2-/- mice compared with CCR2+/+ mice after the conidia challenge. Thus, these findings demonstrate that CCR2 plays an important role in the immune response against A. fumigatus, thereby limiting the allergic airway inflammatory and remodeling responses to this fungus.

Exaggerated hepatic injury due to acetaminophen challenge in mice lacking C-C chemokine receptor 2.

Monocyte chemoattractant protein-1 is one of the major C-C chemokines that has been implicated in liver injury. The C-C chemokine receptor, CCR2, has been identified as the primary receptor that mediates monocyte chemoattractant protein-1 (MCP-1) responses in the mouse. Accordingly, the present study addressed the role of CCR2 in mice acutely challenged with acetaminophen (APAP). Mice genetically deficient in CCR2 (CCR2(-/-)) and their wild-type counterparts (CCR2(+/+)) were fasted for 10 hours before receiving an intraperitoneal injection of APAP (300 mg/kg). Liver and serum samples were removed from both groups of mice before and at 24 and 48 hours post APAP. Significantly elevated levels of MCP-1 were detected in liver samples from CCR2(+/+) and CCR2(-/-) mice at 24 hours post-APAP. Although CCR2(+/+) mice exhibited no liver injury at any time after receiving APAP, CCR2(-/-) mice exhibited marked evidence of necrotic and TUNEL-positive cells in the liver, particularly at 24 hours post-APAP. Enzyme-linked immunosorbent assay analysis of liver homogenates from both groups of mice at the 24 hours time point revealed that liver tissue from CCR2(-/-) mice contained significantly greater amounts of immunoreactive IFN-gamma and TNF-alpha. The in vivo immunoneutralization of IFN-gamma or TNF-alpha significantly attenuated APAP-induced liver injury in CCR2(-/-) mice and increased hepatic IL-13 levels. Taken together, these findings demonstrate that CCR2 expression in the liver provides a hepatoprotective effect through its regulation of cytokine generation during APAP challenge.

Correlation between the structure and function of the rabbit urinary bladder following partial outlet obstruction.

PURPOSE: To understand the relationship between contractile and structural changes in the obstructed bladder, rabbit bladder was partially obstructed for up to 70 days and alterations in tension response to field stimulation and carbachol were compared with alterations in ultrastructure and innervation of detrusor smooth muscle (SM). The effect of partial outlet obstruction on the physiological responses to field stimulation (FS) (nerve mediated contraction) and carbachol (receptor mediated contraction) were correlated with the structure and innervation of the detrusor smooth muscle (SM) of the same animal during a 70 day period. MATERIALS AND METHODS: 28 rabbits were subjected to 1 to 70 days of mild partial outlet obstruction. Sham operated rabbits were euthanized at 7, 14, 28, and 70 days post-obstruction. At each time period, isolated strips of bladder body were mounted in individual baths and the contractile response to FS and carbachol determined. Three additional strips from each bladder were fixed for electron microscopy. RESULTS: Bladder mass increased rapidly during the first 7 days after obstruction, was constant for the next 7 days, and then continued to increase gradually. Dysfunction of the contractile response to FS was noted as early as 3 days and progressively increased over the 70-day study period. The decrease in the response to FS increased at a significantly faster rate than the decrease in the contractile response to carbachol. In ultrastructure studies, at 3 and 7 days post-obstruction the majority of SM cells displayed the characteristics of hypertrophy. At 28 days some SM cells displayed loosely packed myofilaments and an irregular distribution of sarcoplasmic dense bodies. At 70 days swollen mitochondria were present in all cell types of the bladder wall. Evidence of axonal degeneration was first observed at 7 days post-obstruction and became more extensive thereafter. No evidence of mitotic figures, nerve growth cones or regenerating SM cells was observed. CONCLUSIONS: Prolonged partial bladder outflow obstruction is accompanied by a progressive decrease in contractility of SM. The present study describes the structural damage that occurs in the bladder wall in response to partial outlet obstruction and correlates these observations with the contractile dysfunction with which it is associated. Furthermore, mitochondrial damage in vessels and fibroblasts is suggestive of bladder wall ischemia.

Cutting edge: identification of a novel chemokine receptor that binds dendritic cell- and T cell-active chemokines including ELC, SLC, and TECK.

Searching for new receptors of dendritic cell- and T cell-active chemokines, we used a combination of techniques to interrogate orphan chemokine receptors. We report here on human CCX CKR, previously represented only by noncontiguous expressed sequence tags homologous to bovine PPR1, a putative gustatory receptor. We employed a two-tiered process of ligand assignment, where immobilized chemokines constructed on stalks (stalkokines) were used as bait for adhesion of cells expressing CCX CKR. These cells adhered to stalkokines representing ELC, a chemokine previously thought to bind only CCR7. Adhesion was abolished in the presence of soluble ELC, SLC (CCR7 ligands), and TECK (a CCR9 ligand). Complete ligand profiles were further determined by radiolabeled ligand binding and competition with >80 chemokines. ELC, SLC, and TECK comprised high affinity ligands (IC50 <15 nM); lower affinity ligands include BLC and vMIP-II (IC50 <150 nM). With its high affinity for CC chemokines and homology to CC receptors, we provisionally designate this new receptor CCR10.

The distribution of vesicular acetylcholine transporter in the human male genitourinary organs and its co-localization with neuropeptide Y and nitric oxide synthase.

Because doubt still remains concerning the distribution of nerves that are unequivocally cholinergic in the human genitourinary organs, we have used a specific marker, namely, an antibody to vesicular acetylcholine transporter (VAChT), to immunolabel cholinergic axons and cell bodies in specimens of urinary bladder, seminal vesicle, vas deferens, and prostate gland obtained from neonates and children post mortem. In addition some sections were double-immunolabeled with VAChT and either neuropeptide Y (NPY) or nitric oxide synthase (NOS). The results demonstrated a rich cholinergic innervation to the muscle coat of the bladder body with a much less prominent, but nonetheless significant, cholinergic innervation to the smooth muscle components of the seminal vesicle, vas deferens, and prostate. Small ganglia were scattered throughout the detrusor muscle of the urinary bladder, approximately 75% of the intramural neurons being VAChT immunoreactive, whereas approximately 95% contained NPY and approximately 40% contained NOS. VAChT immunoreactivity was observed in 40% of neurons in ganglia scattered throughout the pelvic plexus. Almost all these cholinergic neurons contained NPY and approximately 65% contained NOS. Almost all the cholinergic nerve fibers throughout the genitourinary organs also contained NPY. Although NOS was sparse in the cholinergic nerves of the bladder body, it occurred in the majority of cholinergic nerves at the bladder neck and was also present in a proportion of the cholinergic nerves in the other organs examined. VAChT-immunoreactive nerves were also observed in a sub-epithelial location in all the organs examined, the majority containing NPY, whereas a small proportion contained NOS. Although doubt remains about the function of sub-epithelial cholinergic nerves in the urinary bladder, the majority of similar nerves in the seminal vesicle, vas deferens, and prostate gland are considered to be secretomotor. Collectively these findings demonstrate that the cholinergic innervation of the male genitourinary system is well established in the neonate and child. Neurourol. Urodynam. 19:185-194, 2000.

Fractalkine receptor expression by T lymphocyte subpopulations and in vivo production of fractalkine in human.

Expression and function of the fractalkine receptor CX3CR1 by T lymphocyte subpopulations was evaluated in healthy individuals. In CD8(+) T lymphocytes, CX3CR1 was expressed by and functional in both CD45RO(-) and CD45RO(+) cells. In CD4(+) T lymphocytes, CX3CR1 was expressed mainly by CD45RO(+) cells, and almost exclusively by activated HLA-DR(+) T lymphocytes. This receptor was functional in CD45RO(+) cells, but not in CD45RO(-) cells. Expression of fractalkine was detected by in situ hybridization and immunohistochemistry in endothelial cells of normal lung and thymus. In hyperplastic lymph nodes, fractalkine was expressed by endothelial cells of high endothelial venules and of subcapsular vessels, by follicular dendritic cells (FDC) and by some follicle lymphocytes. Fractalkine mRNA was constitutively present in the HK FDC-like cell line, and it was induced in vitro in B lymphocytes stimulated by an anti-micro or by a CD40 mAb. These findings indicate that fractalkine may contribute to the recruitment of effector T helper lymphocytes, either in peripheral tissues or in lymphoid organs. In these tissues, fractalkine and its receptor may favor contact within follicles between activated T helper lymphocytes, activated B lymphocytes and FDC, thus contributing to the maturation of the B lymphocyte response.

Rearranged marriages: marital relationships after head injury.

An in depth study of 18 heterosexual couples investigated the quality of the marital and sexual relationships 1-7 years after the male partner had suffered a severe head injury. Both quantitative and qualitative methods were employed and the focus was on the perspective of the uninjured female partner. The female partners reported both marital and sexual satisfaction as lower following injury. They rated their current marital satisfaction as significantly less than their brain injured partners. The quantitative part of the study revealed major role changes experienced by the women, with many comparing their new role to that of a parent with total decision making responsibility. The incompatibility of this role with that of sexual partner was mentioned by many. A tendency for the males to express gratitude but not to communicate their feelings was described by many women. Most women were resigned to the expectation that there would be little change in the future and, for most, the only positive aspect of the relationship was a sense of commitment and continuing companionship. The implications of the findings for rehabilitation and couple therapy are discussed.

Monocyte chemoattractant protein-1 mediates cockroach allergen-induced bronchial hyperreactivity in normal but not CCR2-/- mice: the role of mast cells.

Bronchial eosinophil and mononuclear cell infiltrates are a hallmark of the asthmatic lung and are associated with the induction of reversible airway hyperreactivity. In these studies, we have found that monocyte chemotactic protein-1 (MCP-1), a CC (beta) chemokine, mediates airway hyperreactivity in normal and allergic mice. Using a murine model of cockroach Ag-induced allergic airway inflammation, we have demonstrated that anti-MCP-1 Abs inhibit changes in airway resistance and attenuate histamine release into the bronchoalveolar lavage, suggesting a role for MCP-1 in mast cell degranulation. In normal mice, instillation of MCP-1 induced prolonged airway hyperreactivity and histamine release. In addition, MCP-1 directly induced pulmonary mast cell degranulation in vitro. These latter effects would appear to be selective because no changes were observed when macrophage-inflammatory protein-1alpha, eotaxin, or MCP-3 were instilled into the airways of normal mice or when mast cells were treated in vitro. Airway hyperreactivity was mediated by MCP-1 through CCR2 because allergen-induced as well as direct MCP-1 instilled-induced changes in airway hyperreactivity were significantly attenuated in CCR2 -/- mice. The neutralization of MCP-1 in allergic animals and instillation of MCP-1 in normal animals was related to leukotriene C4 levels in the bronchoalveolar lavage and was directly induced in pulmonary mast cells by MCP-1. Thus, these data identify MCP-1 and CCR2 as potentially important therapeutic targets for the treatment of hyperreactive airway disease.

The distribution of noradrenergic nerves in the human lower urinary tract. A review.

The purpose of this presentation is to describe the distribution of noradrenergic nerves in the human genitourinary system. The techniques which have been employed include formaldehyde-induced fluorescence and immunocytochemical methods to demonstrate dopamine beta-hydroxylase and tyrosine hydroxylase. These methods have been applied to human fetal, neonatal, infant, child and adult tissues removed either at post mortem examination or by surgical excision. The innervation of the fetal urinary bladder is well established by 13 weeks and, as in older specimens, the detrusor receives a sparse noradrenergic nerve supply. In contrast the smooth muscle of the terminal ureter is well supplied by this type of autonomic nerve. An additional incomplete muscle layer has been identified as a nomal component of the terminal ureter which is richly innervated by noradrenergic nerves. In some cases this muscle forms a complete collar which may be responsible for ureteric obstruction. By comparison with the detrusor, bladder neck smooth muscle receives a dense noradrenergic nerve supply particularly in the male. Unlike the detrusor, the structure and innervation of the vas deferens, seminal vesicle and prostate are poorly differentiated in the fetus. In the infant and child, the structure of the intramural smooth muscle of these organs remains immature although a rich noradrenergic nerve supply resembing the adult has been established in the fetus by 30 weeks. In the fetus, autonomic ganglia occur in association with noradrenaline rich paraganglia and surprisingly, with sensory nerve endings resembling pacinian corpuscles. Shortly after birth paraganglia are no longer associated with the autonomic ganglia of the genitourinary system. On the basis of size at least two types of autonomic neuron populate these autonomic ganglia. One type is relatively large and devoid of catecholamines but is closely associated with pericellular noradrenergic nerve fibres. The second type of neuron is small, contains noradrenaline and is arranged in clusters closely related to the capsule of the prostate gland. The significance of these observations will be considered with respect to the neurological control of the genitourinary system.

MCP-1 deficiency reduces susceptibility to atherosclerosis in mice that overexpress human apolipoprotein B.

The earliest recognizable atherosclerotic lesions are fatty streaks composed of lipid-laden macrophages (foam cells). Circulating monocytes are the precursors of these foam cells, but the molecular mechanisms that govern macrophage trafficking through the vessel wall are poorly understood. Monocyte chemoattractant protein-1 (MCP-1), a member of the chemokine (chemotactic cytokine) family, is a potent monocyte agonist that is upregulated by oxidized lipids. Recent studies in hypercholesterolemic mice lacking apo E or the low-density lipoprotein receptor have suggested a role for MCP-1 in monocyte recruitment to early atherosclerotic lesions. To determine if MCP-1 is critically involved in atherogenesis in the setting of elevated physiological plasma cholesterol levels, we deleted the MCP-1 gene in transgenic mice expressing human apo B. Here we report that the absence of MCP-1 provides dramatic protection from macrophage recruitment and atherosclerotic lesion formation in apo B transgenic mice, without altering lipoprotein metabolism. Taken together with the results of earlier studies, these data provide compelling evidence that MCP-1 plays a critical role in the initiation of atherosclerosis.

Enhanced myeloid progenitor cell cycling and apoptosis in mice lacking the chemokine receptor, CCR2.

Chemokines regulate hematopoiesis in part by influencing the proliferative status of myeloid progenitor cells (MPC). Human MCP-1/murine JE, a myelosuppressive chemokine, specifically binds C-C chemokine receptor 2 (CCR2). Transgenic mice containing a targeted disruption in CCR2 that prevents expression of CCR2 mRNA and protein and have MPC that are insensitive to inhibition by MCP-1 and JE in vitro were assessed for potential abnormalities in growth of bone marrow (BM) and spleen MPC. MPC in both unseparated and c-kit+lin- populations of BM from CCR2-deficient (-/-) mice were in a greatly increased proliferation state compared with CCR2 littermate control (+/+) mice, an effect not apparent with progenitors from spleens of CCR2 (-/-) mice. Increased cycling status of CCR2 (-/-) BM MPC did not result in increased numbers of nucleated cells or MPC in BM or spleens of CCR2 (-/-) mice. Possible reasons for this apparent discrepancy were highlighted by flow cytometric analysis of c-kit+lin- BM cells and colony formation by MPC subjected to delayed addition of growth factors. The c-kit+lin- population of BM cells from CCR2 (-/-) mice had a significantly higher percentage of apoptotic cells than those from CCR2 (+/+) BM. However, elevated apoptosis was not associated with decreased numbers of c-kit+lin- cells. The increased percentage of apoptotic c-kit+lin- cells was due to elevated apoptosis within the c-kitdimlin-, but not the c-kitbrightlin-, subpopulations of cells. Consistent with enhanced apoptosis of phenotypically defined cells, MPC from CCR2 (-/-) BM and purified c-kit+lin- cells demonstrated decreased cell survival in vitro upon delayed addition of growth factors. The data suggest that signals received by CCR2 limit proliferation of progenitor cells in the BM, but also enhance survival of these cells.