Activated protein C (APC) is a protease with anticoagulant and cell-signaling activities. In the central nervous system, APC and its analogs with reduced anticoagulant activity but preserved cell signaling activities, such as 3K3A-APC, exert neuroprotective, vasculoprotective and anti-inflammatory effects. Murine APC promotes subependymal neurogenesis in rodents in vivo after ischemic and traumatic brain injury. Whether human APC can influence neuronal production from resident progenitor cells in humans is unknown. Here we show that 3K3A-APC, but not S360A-APC (an enzymatically inactive analog of APC), stimulated neuronal mitogenesis and differentiation from fetal human neural stem and progenitor cells (NPCs). 3K3A-APC’s effects on proliferation and differentiation were comparable to those respectively obtained with fibroblast growth factor and brain-derived growth factor. Its promoting effect on neuronal differentiation was accompanied by inhibition of astroglial differentiation. In addition, 3K3A-APC exerted modest anti-apoptotic effects during neuronal production. These effects appeared mediated through specific protease activated (PAR) and sphingosine-1-phosphate (S1PR) receptors, in that siRNA-mediated inhibition of PARs 1–4 and S1PRs 1–5 revealed that PAR1, PAR3 and S1PR1 are required for the neurogenic effects of 3K3A-APC. 3K3A-APC activated Akt, a downstream target of S1PR1, which was inhibited by S1PR1, PAR1 and PAR3 silencing. Adenoviral transduction of NPCs with a kinase-defective Akt mutant abolished the effects of 3K3A-APC on NPCs, confirming a key role of Akt activation in 3K3A-APC-mediated neurogenesis. Thus, APC and its pharmacological analogues, by influencing PAR and S1PR signals in resident neural progenitor cells, may be potent modulators of both development and repair in the human CNS.
Cardiac troponin levels help risk-stratify patients presenting with an acute coronary syndrome (ACS). Although they may be elevated in patients presenting with Non-ACS conditions, specific diagnoses and long-term outcomes within that cohort are unclear.
Methods and Results
Using the Veterans Affairs (VA) centralized databases, we identified all hospitalized patients in 2006 who had a troponin assay obtained during their initial reference hospitalization. Based on ICD-9 diagnostic codes, primary diagnoses were categorized as either ACS or Non-ACS conditions. Of a total of 21,668 patients with an elevated troponin level who were discharged from the hospital, 12,400 (57.2%) had a Non-ACS condition. Among that cohort, the most common diagnostic category involved the cardiovascular system and congestive heart failure (N=1661) and chronic coronary artery disease (N=1648) accounted for the major classifications. At one-year following hospital discharge, mortality in patients with a Non-ACS condition was 22.8% and was higher than the ACS cohort (Odds Ratio=1.39; 95%CI: 1.30–1.49). Despite the high prevalence of cardiovascular diseases in patients with a Non-ACS diagnosis, utilization of cardiac imaging within 90 days of hospitalization was low compared with ACS patients (Odds Ratio=0.25; 95%CI: 0.23–0.27).
Hospitalized patients with an elevated troponin level most often have a primary diagnosis that is not an acute coronary syndrome. Their long-term survival is poor and justifies novel diagnostic or therapeutic strategy-based studies to target the highest risk subsets prior to hospital discharge.
outcomes; troponins; non-ACS diagnosis; cardiac imaging; coronary artery disease
Because it lacks a lymphatic circulation, the brain must clear extracellular proteins by an alternative mechanism. The cerebrospinal fluid (CSF) functions as a sink for brain extracellular solutes, but it is not clear how solutes from the brain interstitium move from the parenchyma to the CSF. We demonstrate that a substantial portion of subarachnoid CSF cycles through the brain interstitial space. On the basis of in vivo two-photon imaging of small fluorescent tracers, we showed that CSF enters the parenchyma along paravascular spaces that surround penetrating arteries and that brain interstitial fluid is cleared along paravenous drainage pathways. Animals lacking the water channel aquaporin-4 (AQP4) in astrocytes exhibit slowed CSF influx through this system and a ~70% reduction in interstitial solute clearance, suggesting that the bulk fluid flow between these anatomical influx and efflux routes is supported by astrocytic water transport. Fluorescent-tagged amyloid β, a peptide thought to be pathogenic in Alzheimer’s disease, was transported along this route, and deletion of the Aqp4 gene suppressed the clearance of soluble amyloid β, suggesting that this pathway may remove amyloid β from the central nervous system. Clearance through paravenous flow may also regulate extracellular levels of proteins involved with neurodegenerative conditions, its impairment perhaps contributing to the mis-accumulation of soluble proteins.
Oligodendrocyte progenitor cells (OPCs) persist in human white matter, yet the mechanisms by which they are maintained in an undifferentiated state are unknown. Human OPCs differentially express protein tyrosine phosphatase receptor β/ζ (PTPRZ1), and its inhibitory ligand, pleiotrophin, suggesting the maintenance of an autocrine loop by which PTPRZ1 activity is tonically suppressed. PTPRZ1 constitutively promotes the tyrosine dephosphorylation of β-catenin, and thus β-catenin participation in TCF-mediated transcription. Using CD140a/PDGFRα-based FACS to isolate fetal OPCs from the fetal brain at gestational ages 16-22 weeks, we asked if pleiotrophin modulated the expansion of OPCs, and if so, whether this was effected through the serial engagement of PTPRZ1 and β-catenin-dependent signals, such as TCF-mediated transcription. Lentiviral shRNAi knockdown of PTPRZ1 induced TCF-mediated transcription, and substantially augmented GSK3β inhibition-induced TCF-reporter luciferase expression, suggesting dual regulation of β-catenin and the importance of PTPRZ1 as a tonic brake upon TCF-dependent transcription. Pharmacological inhibition of GSK3β triggered substrate detachment and initiated sphere formation, yet had no effect on either proliferation or net cell number. In contrast, pleiotrophin strongly potentiated the proliferation of CD140a+-sorted OPCs, as did PTPRZ1 knockdown, which significantly increased the total number of population doublings exhibited by OPCs before mitotic senescence. These observations suggest that pleiotrophin inhibition of PTPRZ1 contributes to the homeostatic self-renewal of OPCs, and that this process is mediated by the tonic activation of β-catenin/TCF-dependent transcription.
progenitor cells; β-catenin; Wnt signaling; pleiotrophin; PTPRZ1; PDGFRA; RPTPβ/ζ; CD140a; human; oligodendrocyte progenitor
The perivascular niche for neurogenesis was first reported as the co-association of newly generated neurons and their progenitors with both dividing and mitotically quiescent endothelial cells in restricted regions of the brain in adult birds and mammals alike. This review attempts to summarize our present understanding of the interaction of blood vessels with neural stem and progenitor cells, addressing both glial and neuronal progenitor cell interactions in the perivascular niche. We review the molecular interactions that are most critical to the endothelial control of stem and progenitor cell mobilization and differentiation. The focus throughout will be on defining those perivascular ligand-receptor interactions shared among these systems, as well as those that clearly differ as a function of cell type and setting, by which specificity may be achieved in the development of targeted therapeutics.
Oligodendrocyte progenitor cells (OPCs) persist in human white matter, yet the mechanisms by which they are maintained in an undifferentiated state are unknown. Human OPCs differentially express protein tyrosine phosphatase receptor β/ζ (PTPRZ1) and its inhibitory ligand, pleiotrophin, suggesting the maintenance of an autocrine loop by which PTPRZ1 activity is tonically suppressed. PTPRZ1 constitutively promotes the tyrosine dephosphorylation of β-catenin and, thus, β-catenin participation in T cell factor (TCF)-mediated transcription. Using CD140a/PDGFRα-based fluorescence-activated cell sorting to isolate fetal OPCs from the fetal brain at gestational ages 16–22 weeks, we asked whether pleiotrophin modulated the expansion of OPCs and, if so, whether this was effected through the serial engagement of PTPRZ1 and β-catenin-dependent signals, such as TCF-mediated transcription. Lentiviral shRNAi knockdown of PTPRZ1 induced TCF-mediated transcription and substantially augmented GSK3β inhibition-induced TCF-reporter luciferase expression, suggesting dual regulation of β-catenin and the importance of PTPRZ1 as a tonic brake upon TCF-dependent transcription. Pharmacological inhibition of GSK3β triggered substrate detachment and initiated sphere formation, yet had no effect on either proliferation or net cell number. In contrast, pleiotrophin strongly potentiated the proliferation of CD140a+-sorted OPCs, as did PTPRZ1 knockdown, which significantly increased the total number of population doublings exhibited by OPCs before mitotic senescence. These observations suggest that pleiotrophin inhibition of PTPRZ1 contributes to the homeostatic self-renewal of OPCs and that this process is mediated by the tonic activation of β-catenin/TCF-dependent transcription.
The diseases of myelin are among the most prevalent and disabling conditions in neurology. These diseases include both the vascular and inflammatory demyelinating disorders of adulthood, as well as the childhood leukodystrophies and cerebral palsy. These fundamentally glial disorders may be amenable to treatment by glial progenitor cells (GPCs), which give rise to astroglia and myelin-producing oligodendrocytes. Given the development of new methods for generating and isolating human GPCs, the myelin disorders may now be compelling targets for cell-based therapy. In addition, the efficient engraftment and expansion of human GPCs in murine hosts has led to the development of human glial chimeric mouse brains, which provides new opportunities for studying the species-specific roles of human glia in cognition, as well as in disease pathogenesis.
Astrocytes participate in all essential CNS functions, including blood flow regulation, energy metabolism, ion and water homeostasis, immune defence, neurotransmission, and adult neurogenesis. It is thus not surprising that astrocytic morphology and function differ between regions, and that different subclasses of astrocytes exist within the same brain region. Recent lines of work also show that the complexity of protoplasmic astrocytes increases during evolution. Human astrocytes are structurally more complex, larger, and propagate calcium signals significantly faster than rodent astrocytes. In this chapter, we review the diversity of astrocytic form and function, while considering the markedly expanded roles of astrocytes with phylogenetic evolution. We also define major challenges for the future, which include determining how astrocytic functions are locally specified, defining the molecular controls upon astrocytic fate and physiology and establishing how evolutionary changes in astrocytes contribute to higher cognitive functions.
Astrocyte; NG2 cell; Glia; Glia progenitor; Potassium buffering; Epilepsy; Calcium signaling; Purinergic receptors
Huntington’s disease (HD) is an inherited, relentlessly progressive neurodegenerative disease with an invariably fatal outcome. HD is inherited in an autosomal dominant fashion, and is characterized pathologically by the loss of cortical and striatal neurons, and clinically by involuntary choreiform movements accompanied by progressive cognitive impairment and emotional lability. The disorder is caused by an expanded cystosine adenine guanine (CAG) tri-nucleotide repeat encoding polyglutamine (polyQ) in the first exon of the Huntingtin gene. There is a correlation between the number of CAG repeats and disease onset, such that in patients with CAG repeat lengths of 36 to 60, disease symptoms typically manifest after 35 years of age, whereas CAG repeat lengths >60 yield the more severe juvenile form of the disease. Even though mutant huntingtin is expressed throughout the brain, it is characterized by the selective degeneration of medium spiny neurons of the caudate and putamen, which heralds more widespread neuronal degeneration with disease progression. The mechanisms of cell dysfunction and death in HD have been the subjects of a number of studies, which have led to therapeutic strategies largely based on the amelioration of mutant huntingtin-related metabolic impairment and cellular toxicity. Each of these approaches has aimed to delay or stop the preferential degeneration of medium spiny neurons early in the disease course. Yet, in later stages of the disease, after cell death has become prominent, cell replacement therapy (whether by direct cell transplantation or by the mobilization of endogenous progenitors) may comprise a stronger potential avenue for therapy. In this review, we will consider recent progress in the transplantation of fetal striatal cells to the HD brain, as well as emerging alternative sources for human striatal progenitor cells. We will then consider the potential application of gene therapy toward the induction of striatal neurogenesis and neuronal recruitment, with an eye toward its potential therapeutic use in HD.
Electronic supplementary material
The online version of this article (doi:10.1007/s13311-011-0075-8) contains supplementary material, which is available to authorized users.
Cell therapy; Neural transplants; Stem cells; Neurogenesis; Medium spiny neuron
The value of Single Photon Emission Computed Tomography stress myocardial perfusion imaging (SPECT-MPI) for detecting graft disease after coronary artery bypass surgery (CABG) has not been studied prospectively in an unselected cohort.
Radial Artery Versus Saphenous Vein Graft Study is a Veterans Affairs Cooperative Study to determine graft patency rates after CABG surgery. Seventy-nine participants agreed to SPECT-MPI within 24 hours of their coronary angiogram, one-year after CABG. The choice of the stress protocol was made at the discretion of the nuclear radiologist and was either a symptom-limited exercise test (n = 68) or an adenosine infusion (n = 11). The SPECT-MPI results were interpreted independent of the angiographic results and estimates of sensitivity, specificity and accuracy were based on the prediction of a graft stenosis of ≥70% on coronary angiogram.
A significant stenosis was present in 38 (48%) of 79 patients and 56 (22%) of 251 grafts. In those stress tests with an optimal exercise heart rate response (>80% maximum predicted heart rate) (n = 26) sensitivity, specificity and accuracy of SPECT-MPI for predicting the graft stenosis was 77%, 69% and 73% respectively. With adenosine (n = 11) it was 75%, 57% and 64%, respectively. Among participants with a suboptimal exercise heart rate response, the sensitivity of SPECT-MPI for predicting a graft stenosis was <50%. The accuracy of SPECT-MPI for detecting graft disease did not vary significantly with ischemic territory.
Under optimal stress conditions, SPECT-MPI has a good sensitivity and accuracy for detecting graft disease in an unselected patient population 1 year post-CABG.
Coronary artery bypass grafts; CABG; Coronary artery imaging; Cardiac catheterization/intervention
Malignant gliomas are aggressive brain tumors with limited therapeutic options, and improvements in treatment require a deeper molecular understanding of this disease. As in other cancers, recent studies have identified highly tumorigenic subpopulations within malignant gliomas, known generally as cancer stem cells. Here we demonstrate that glioma stem cells (GSCs) produce nitric oxide via elevated nitric oxide synthase-2 (NOS2) expression. GSCs depend on NOS2 activity for growth and tumorigenicity, distinguishing them from non-GSCs and normal neural progenitors. Gene expression profiling identified many NOS2-regulated genes, including the cell cycle inhibitor cell division autoantigen-1 (CDA1). Further, high NOS2 expression correlates with decreased survival in human glioma patients, and NOS2 inhibition slows glioma growth in a murine intracranial model. These data provide insight into how GSCs are mechanistically distinct from their less tumorigenic counterparts, and suggest that NOS2 inhibition may be an efficacious approach to treating this devastating disease.
Testosterone-induced singing in songbirds is thought to involve testosterone-dependent morphological changes that include angiogenesis and neuronal recruitment into the HVC, a central part of the song control circuit. Previous work showed that testosterone induces the production of vascular endothelial growth factor (VEGF) and its receptor (VEGFR2 tyrosine kinase), which in turn leads to an upregulation of brain-derived neurotrophic factor (BDNF) production in HVC endothelial cells. Here we report for the first time that systemic inhibition of the VEGFR2 tyrosine kinase is sufficient to block testosterone-induced song in adult female canaries, despite sustained androgen exposure and the persistence of the effects of testosterone on HVC morphology. Expression of exogenous BDNF in HVC, induced locally by in situ transfection, reversed the VEGFR2 inhibition-mediated blockade of song development, thereby restoring the behavioral phenotype associated with androgen-induced song. The VEGFR2-inhibited, BDNF-treated females developed elaborate male-like song that included large syllable repertoires and high syllable repetition rates, features known to attract females. Importantly, although functionally competent new neurons were recruited to HVC after testosterone treatment, the time course of neuronal addition appeared to follow BDNF-induced song development. These findings indicate that testosterone-associated VEGFR2 activity is required for androgen-induced song in adult songbirds and that the behavioral effects of VEGFR2 inhibition can be rescued by BDNF within the adult HVC.
Experimental models of myelin disorders can be treated by the transplantation of oligodendrocyte progenitor cells (OPCs) into the affected brain or spinal cord. OPCs express gangliosides recognized by MAb A2B5, but this marker also identifies lineage-restricted astrocytes and immature neurons. To establish a more efficient means of isolating myelinogenic OPCs, we asked if FACS could be used to sort PDGFα receptor+ cells from fetal human forebrain, based on expression of the PDGFRα epitope CD140a. CD140a+ isolates were maintained as mitotic bipotential progenitors that could be instructed to either oligodendrocyte or astrocyte fate. Transplanted CD140a+ cells were highly migratory, and rapidly and robustly myelinated the hypomyelinated shiverer mouse brain, more efficiently than did A2B5-sorted cells. Microarray analysis of CD140a+ cells revealed their differential expression of CD9, as well as of PTN-PTPRZ1, wnt, notch and BMP pathway components, indicating the dynamic interaction of self-renewal and fate-restricting pathways in these cells.
oligodendrocyte progenitor; PDGF receptor; myelin; remyelination
The childhood leukodystrophies are characterized by neonatal or childhood deficiencies in myelin production or maintenance; these may be due to hereditary defects in genes for myelin maintenance, as in Pelizaeus-Merzbacher disease, or to enzymatic deficiencies resulting in substrate misaccumulation or misprocessing, as in the lysosomal storage disorders. Regardless of their respective etiologies, these disorders are essentially all manifested by a profound deterioration in neurological function with age. A congenital deficit in forebrain myelination is also noted in children with the periventricular leukomalacia of cerebral palsy, which yields a more static morbidity. In light of the wide range of disorders to which congenital hypomyelination or postnatal demyelination may contribute, and the relative homogeneity of oligodendrocytes and their progenitors, the leukodystrophies may be especially attractive targets for cell-based therapeutic strategies. As a result, glial progenitor cells, which can give rise to new myelinogenic oligodendrocytes, have become of great interest as potential vectors for the restoration of myelin to the dysmyelinated brain and spinal cord. In addition, by distributing throughout the neuraxis after perinatal graft, and giving rise to astrocytes as well as oligodendrocytes, glial progenitor cells may be of great utility in rectifying the dysmyelination-associated enzymatic deficiencies of the lysosomal storage disorders.
Congenitally hypomyelinated shiverer mice fail to generate compact myelin, and die by 18–21 weeks of age. Using multifocal anterior and posterior fossa delivery of sorted fetal human glial progenitor cells into neonatal shiverer x rag2−/− immunodeficient and myelin-deficient mice, we achieved whole neuraxis myelination of the engrafted hosts, which in a significant fraction of cases rescued this otherwise lethal phenotype. The transplanted mice exhibited greatly prolonged survival, some surviving beyond a year, with progressive resolution of their neurological deficits. Accordingly, they exhibited substantial myelination of the brain, brainstem and cerebellum, as well as the spinal cord, optic nerves, and cranial ganglia. This was accompanied by the acquisition of normal nodes of Ranvier and transcallosal conduction velocities, ultrastructurally normal and complete myelination of most axons, and a restoration of a substantially normal neurological phenotype. Notably, the resultant mice were cerebral chimeras, with murine gray matter but a predominantly human white matter glial composition. These data demonstrate that the neonatal transplantation of human glial progenitor cells can effectively treat disorders of congenital and perinatal hypomyelination.
oligodendrocyte progenitor; neural stem cell; transplantation; myelin; remyelination; leukodystrophy
Sox2 is expressed by neural stem and progenitor cells, and a sox2 enhancer identifies these cells in the forebrains of both fetal and adult transgenic mouse reporters. We found that an adenovirus encoding EGFP placed under the regulatory control of a 0.4 kb sox2 core enhancer selectively identified multipotential and self-renewing neural progenitor cells in dissociates of human fetal forebrain. Upon EGFP-based fluorescence activated cell sorting (FACS), the E/sox2:EGFP+ isolates were propagable for up to a year in vitro, and remained multilineage competent throughout. E/sox2:EGFP+ cells expressed more telomerase enzymatic activity than matched sox2-depleted populations, and maintained their telomeric lengths with successive passage. Gene expression analysis of E/sox2:EGFP-sorted neural progenitor cells, normalized to the unsorted forebrain dissociates from which they derived, revealed marked over-expression of genes within the notch and wnt pathways, and identified multiple elements of each pathway that appear selective to human neural progenitors. Sox2 enhancer-based FACS thus permits the prospective identification and direct isolation of a telomerase-active population of neural stem cells from the human fetal forebrain, and the elucidation of both the transcriptome and dominant signaling pathways of these critically important cells.
Neural stem cells; telomerase; gene expression; cell sorting
A history of severe allergic reaction to iodine contrast leading to anaphylactic shock presents a dilemma in patients requiring cardiac catheterization. As an alternative, gadolinium has been an interesting and potentially useful agent. However, gadolinium produces poor image quality and has been associated with significant arrhythmias in small case series. Furthermore, there is no consensus about the maximal allowable dose that can be administered to a patient. In the present report, a successful combination of gadolinium contrast with a power injector that produced adequate image quality in a patient with severe allergy to iodine contrast is described. The case was complicated by the occurrence of ventricular fibrillation when damping occurred during injection of contrast into the right coronary artery. This complication has been reported previously with intracoronary gadolinium injection. The report is followed by a brief literature review.
Angiography; Complication; Gadolinium use; Iodine contrast allergy; PCI complication; Percutaneous coronary intervention; Stenting; Ventricular fibrillation
Recent in vivo studies have established astrocytes as a major target for locus coeruleus activation (Bekar et al., Cereb. Cortex 18, 2789–2795), renewing interest in cell culture studies on noradrenergic effects on astrocytes in primary cultures and calling for additional information about the expression of adrenoceptor subtypes on different types of brain cells. In the present communication, mRNA expression of α1-, α2- and β-adrenergic receptors and their subtypes was determined in freshly-isolated, cell marker-defined populations of astrocytes, NG2-positive cells, microglia, endothelial cells, and Thy1-positive neurons (mainly glutamatergic projection neurons) in murine cerebral cortex. Immediately after dissection of frontal, parietal and occipital cortex of 10–12-week-old transgenic mice, which combined each cell-type marker with a specific fluorescent signal, the tissue was digested, triturated and centrifuged, yielding a solution of dissociated cells of all types, which were separated by fluorescence-activated cell sorting (FACS). mRNA expression in each cell fraction was determined by microarray analysis. α1A-Receptors were unequivocally expressed in astrocytes and NG2-positive cells, but absent in other cell types, and α1B-receptors were not expressed in any cell population. Among α2-receptors only α2A-receptors were expressed, unequivocally in astrocytes and NG-positive cells, tentatively in microglia and questionably in Thy1-positive neurons and endothelial cells. β1-Receptors were unequivocally expressed in astrocytes, tentatively in microglia, and questionably in neurons and endothelial cells, whereas β2-adrenergic receptors showed tentative expression in neurons and astrocytes and unequivocal expression in other cell types. This distribution was supported by immunochemical data and its relevance established by previous studies in well-differentiated primary cultures of mouse astrocytes, showing that stimulation of α2-adrenoceptors increases glycogen formation and oxidative metabolism, the latter by a mechanism depending on intramitochondrial Ca2+, whereas α1-adrenoceptor stimulation enhances glutamate uptake, and β-adrenoceptor activation causes glycogenolysis and increased Na+,K+-ATPase activity. The Ca2+- and cAMP-mediated association between energy-consuming and energy-yielding processes is emphasized.
alpha2A receptor; astrocytes; fluorescence-activated cell sorting; free cytosolic Ca2+ concentration; NG2-positive cells; oxidative metabolism
Cell-based therapy for myocardial infarction (MI) holds great promise; however, the ideal cell type and delivery system have not been established. Obstacles in the field are the massive cell death after direct injection and the small percentage of surviving cells differentiating into cardiomyocytes. To overcome these challenges we designed a novel study to deliver cardiac progenitor cells as a cell sheet.
Methods and results
Cell sheets composed of rat or human cardiac progenitor cells (cardiospheres), and cardiac stromal cells were transplanted onto the infarcted myocardium after coronary artery ligation in rats. Three weeks later, transplanted cells survived, proliferated, and differentiated into cardiomyocytes (14.6 ± 4.7%). Cell sheet transplantation suppressed cardiac wall thinning and increased capillary density (194 ± 20 vs. 97 ± 24 per mm2, P < 0.05) compared with the untreated MI. Cell migration from the sheet was observed along the necrotic trails within the infarcted area. The migrated cells were located in the vicinity of stromal-derived factor (SDF-1) released from the injured myocardium, and about 20% of these cells expressed CXCR4, suggesting that the SDF-1/CXCR4 axis plays, at least, a role in cell migration. Transplantation of cell sheets resulted in a preservation of cardiac contractile function after MI, as was shown by a greater ejection fraction and lower left ventricular end diastolic pressure compared with untreated MI.
The scaffold-free cardiosphere-derived cell sheet approach seeks to efficiently deliver cells and increase cell survival. These transplanted cells effectively rescue myocardium function after infarction by promoting not only neovascularization but also inducing a significant level of cardiomyogenesis.
Myocardial infarction; Cardiac progenitor cells; Cardiospheres; Cardiac regeneration; Contractility
Cerebral lactic acid, a product of ischemic anaerobic glycolysis, may directly contribute to ischemic brain damage in vivo. In this study we evaluated the effects of extracellular acid exposure on 7-day-old cultures of embryonic rat forebrain. Mixed neuronal and glial cultures were exposed to either lactic or hydrochloric acid to compare the toxicities of relatively permeable and impermeable acids. Neurons were relatively resistant to extracellular HCl acidosis, often surviving 10-min exposures to pH 3.8. In the same cultures, immunochemically defined astrocytes survived 10-min HCl exposures to a maximum acidity of pH 4.2. Similarly, axonal bundles defasciculated in HCl-titrated media below pH 4.4, although their constituent fibers often survived pH 3.8. Cell death occurred at higher pH in cultures subjected to lactic acidosis than in those exposed to HCl. Over half of forebrain neurons and glia subjected for 10 min to lactic acidification failed to survive exposure to pH 4.9. Longer 1-h lactic acid incubations resulted in cell death below pH 5.2. The potent cytotoxicity of lactic acid may be a direct result of the relatively rapid transfer of its neutral protonated form across cell membranes. This process would rapidly deplete intracellular buffer stores, resulting in unchecked cytosolic acidification. Neuronal and glial death from extracellular acidosis may therefore be a function of both the degree and the rapidity of intracellular acidification.
Acidosis; Astrocyte; Cell death; Hypoxia; Ischemia; Neurons; Stroke
We have examined primary human neuronal precursors (HNPs) from 18–22-week-old fetuses. We showed that E-NCAM/MAP2/β-III tubulin-immunoreactive neuronal precursors divide in vitro and could be induced to differentiate into mature neurons in 2 weeks. HNPs did not express nestin and differentiated slowly compared to rodent neuronal restricted precursors (NRPs, 5 days). Immunocytochemical and physiological analyses showed that HNPs could generate a heterogeneous population of neurons that expressed neurofilament-associated protein and various neurotransmitters, neurotransmitter synthesizing enzymes, voltage-gated ion channels, and ligand-gated neurotransmitter receptors and could fire action potentials. Undifferentiated and differentiated HNPs did not coexpress glial markers. Only a subset of cells that expressed GFP under the control of the Tα1 tubulin promoter was E-NCAM/β-III tubulin-immunoreactive, indicating nonexclusive overlap between these two HNP cell populations. Overall, HNPs resemble NRPs isolated from rodent tissue and appear to be a neuronal precursor population.
stem cells; differentiation; electrophysiology
This study investigated the hypothesis that baseline calcified coronary atherosclerosis may determine cardiovascular disease events in response to intensive glycemic control within the Veterans Affairs Diabetes Trial (VADT).
RESEARCH DESIGN AND METHODS
At baseline, 301 type 2 diabetic participants in the VADT, a randomized trial comparing the effects of intensive versus standard glucose lowering on cardiovascular events, had baseline coronary atherosclerosis assessed by coronary artery calcium (CAC) measured by computed tomography. Participants were followed over the 7.5-year study for development of cardiovascular end points.
During a median follow-up duration of 5.2 years, 89 cardiovascular events occurred. Although intensive glucose-lowering therapy did not significantly reduce cardiovascular events in the substudy cohort as a whole, there was evidence that the response was modified by baseline CAC, as indicated by significant P values for treatment by log(CAC + 1) interaction terms in unadjusted and multivariable-adjusted models (0.01 and 0.03, respectively). Multivariable-adjusted hazard ratios (HRs) for the effect of treatment indicated a progressive diminution of benefit with increasing CAC. Subgroup analyses were also conducted for clinically relevant CAC categories: those above and below an Agatston score of 100. Among those randomized to intensive treatment, for the subgroup with CAC >100, 11 of 62 individuals had events, while only 1 of 52 individuals with CAC ≤100 had an event. The multivariable HR for intensive treatment for those with CAC >100 was 0.74 (95% CI 0.46–1.20; P = 0.21), while for the subgroup with CAC ≤100, the corresponding HR was 0.08 (0.008–0.77; P = 0.03), with event rates of 39 and 4 per 1,000 person-years, respectively.
These data indicate that intensive glucose lowering reduces cardiovascular events in those with less extensive calcified coronary atherosclerosis.
Defining the microanatomic differences between the human brain and that of other mammals is key to understanding its unique computational power. While much effort has been devoted to comparative studies of neurons, astrocytes have received far less attention. We report here that protoplasmic astrocytes in human neocortex are 2.6 fold larger in diameter and extend 10-fold more GFAP+ primary processes than their rodent counterparts. In cortical slices prepared from acutely resected surgical tissue, protoplasmic astrocytes propagate Ca2+ waves with a speed of 36 µm/second, approximately 4-fold faster than rodent. Human astrocytes also transiently increase cystosolic Ca2+ in response to glutamatergic and purinergic receptor agonists. The human neocortex also harbors several anatomically-defined subclasses of astrocytes not represented in rodents. These include a population of astrocytes that reside in layers 5–6 and extend long fibers characterized by regularly spaced varicosities. Another specialized type of astrocyte, the interlaminar astrocyte, abundantly populates the superficial cortical layers and extends long processes without varicosities to cortical layers 3 and 4. Human fibrous astrocytes resemble their rodent counterpart, but are larger in diameter. Thus, human cortical astrocytes are both larger, and structurally both more complex and more diverse, than those of rodents. On this basis, we posit that this astrocytic complexity has permitted the increased functional competence of the adult human brain.
astrocyte; cortex; evolution; Gfa [Glial Fibrillary Acidic Protein]; Glutamate receptor; Histochemistry; Human; Purinergic; Structure
This study was designed to determine if a viable biodegradable three-dimensional fibroblast construct (3DFC) patch implanted on the left ventricle after myocardial infarction (MI) improves left ventricular (LV) function and blood flow. We ligated the left coronary artery of adult male Sprague-Dawley rats and implanted the 3DFC at the time of the infarct. Three weeks after MI, the 3DFC improved LV systolic function by increasing (p < 0.05) ejection fraction (37 ± 3% to 62 ± 5%), increasing regional systolic displacement of the infarcted wall (0.04 ± 0.02 to 0.11 ± 0.03 cm), and shifting the passive LV diastolic pressure volume relationship toward the pressure axis. The 3FDC improved LV remodeling by decreasing (p < 0.05) LV end-systolic and end-diastolic diameters with no change in LV systolic pressure. The 3DFC did not change LV end-diastolic pressure (LV EDP; 25 ± 2 vs. 23 ± 2 mmHg) but the addition of captopril (2mg/L drinking water) lowered (p < 0.05) LV EDP to 12.9 ± 2.5 mmHg and shifted the pressure–volume relationship toward the pressure axis and decreased (p < 0.05) the LV operating end-diastolic volume from 0.49 ± 0.02 to 0.34 ± 0.03 ml. The 3DFC increased myocardial blood flow to the infarcted anterior wall after MI over threefold (p < 0.05). This biodegradable 3DFC patch improves LV function and myocardial blood flow 3 weeks after MI. This is a potentially new approach to cell-based therapy for heart failure after MI.
Acute myocardial infarct; Fibroblasts; Growth factors; Angiogenesis; Extracellular matrix; Bioabsorbable scaffold