Infection of Mouse Neural Progenitor Cells by Toxoplasma gondii Reduces Proliferation, Migration, and Neuronal Differentiation in Vitro.

Congenital toxoplasmosis constitutes a major cause of pre- and postnatal complications. Fetal infection with Toxoplasma gondii influences development and can lead to microcephaly, encephalitis, and neurologic abnormalities. Systematic studies concerning the effects of neural progenitor cell infection with T. gondii are unavailable. Cortical intermediate progenitor cells cultivated as neurospheres obtained from E16.5 Swiss Webster mice were infected with T. gondii (ME49 strain) tachyzoites to mimic the developing mouse cerebral cortex in vitro. Infection was associated with decreased cell proliferation, detected by Ki-67 staining at 48 and 72 hours after infection in floating neurospheres, and reduced cellularity at 96 hours. Transient decreases in the expression of the neurogenesis-related transcription factors T-box brain protein 1, mouse atonal homolog protein 1, and hairy and enhancer of split protein 1 were found in infected cultures, while the level of transcription factor SOX-2 remained unaltered. Neurogenic potential, assessed in plated neurospheres, was impaired in infected cultures, as indicated by decreased late neuronal marker neurofilament heavy chain immunoreactivity. Infected cultures exhibited decreased overall migration rates at 48 and 120 hours. These findings indicate that T. gondii infection of neural progenitor cells may lead to reduced neurogenesis due to an imbalance in cell proliferation alongside an altered migratory profile. If translated to the in vivo situation, these data could explain, in part, cortical malformations in congenitally infected individuals.

The innate and adaptive immunological aspects in neurodegenerative diseases.

Neurodegenerative diseases affect a considerable percentage of the elderly population. New therapeutic approaches are warranted, aiming to at least delay and possibly reverse disease progression. Strategies to elaborate such approaches require knowledge of specific immune system involvement in disease pathogenesis. In this review, innate and adaptive immunological aspects of neurodegenerative disorders, in particular Parkinson's disease, Alzheimer's disease, and amyotrophic lateral sclerosis (ALS), are discussed. Initiating disease factors, as well as common mechanistic pathways, are detailed and potential immunological therapeutic targets are identified.

Blood-brain barrier alterations provide evidence of subacute diaschisis in an ischemic stroke rat model.

BACKGROUND: Comprehensive stroke studies reveal diaschisis, a loss of function due to pathological deficits in brain areas remote from initial ischemic lesion. However, blood-brain barrier (BBB) competence in subacute diaschisis is uncertain. The present study investigated subacute diaschisis in a focal ischemic stroke rat model. Specific focuses were BBB integrity and related pathogenic processes in contralateral brain areas. METHODOLOGY/PRINCIPAL FINDINGS: In ipsilateral hemisphere 7 days after transient middle cerebral artery occlusion (tMCAO), significant BBB alterations characterized by large Evans Blue (EB) parenchymal extravasation, autophagosome accumulation, increased reactive astrocytes and activated microglia, demyelinization, and neuronal damage were detected in the striatum, motor and somatosensory cortices. Vascular damage identified by ultrastuctural and immunohistochemical analyses also occurred in the contralateral hemisphere. In contralateral striatum and motor cortex, major ultrastructural BBB changes included: swollen and vacuolated endothelial cells containing numerous autophagosomes, pericyte degeneration, and perivascular edema. Additionally, prominent EB extravasation, increased endothelial autophagosome formation, rampant astrogliosis, activated microglia, widespread neuronal pyknosis and decreased myelin were observed in contralateral striatum, and motor and somatosensory cortices. CONCLUSIONS/SIGNIFICANCE: These results demonstrate focal ischemic stroke-induced pathological disturbances in ipsilateral, as well as in contralateral brain areas, which were shown to be closely associated with BBB breakdown in remote brain microvessels and endothelial autophagosome accumulation. This microvascular damage in subacute phase likely revealed ischemic diaschisis and should be considered in development of treatment strategies for stroke.

Nestin overexpression precedes caspase-3 upregulation in rats exposed to controlled cortical impact traumatic brain injury.

Our understanding of biological mechanisms and treatment options for traumatic brain injury (TBI) is limited. Here, we employed quantitative real-time PCR (QRT-PCR) and immunohistochemical analyses to determine the dynamic expression of cell proliferation and apoptosis in an effort to provide insights into the therapeutic window for developing regenerative strategies for TBI. For this purpose, young adult Sprague-Dawley rats were subjected to experimental TBI using a controlled cortical impactor, then euthanized 1-48 hours after TBI for QRT-PCR and immunohistochemistry. QRT-PCR revealed that brains from TBI exposed rats initially displayed nestin mRNA expression that modestly increased as early as 1-hour post-TBI, then significantly peaked at 8 hours, but thereafter reverted to pre-TBI levels. On the other hand, caspase-3 mRNA expression was slightly elevated at 8 hours post-TBI, which did not become significantly upregulated until 48 hours. Immunofluorescent microscopy revealed a significant surge in nestin immunoreactive cells in the cortex, corpus callosum, and subventricular zone at 24 hours post-TBI, whereas a significant increase in the number of active caspase-3 immunoreactive cells was only found in the cortex and not until 48 hours. These results suggest that the injured brain attempts to repair itself via cell proliferation immediately after TBI, but that this endogenous regenerative mechanism is not sufficient to abrogate the secondary apoptotic cell death. Treatment strategies designed to amplify cell proliferation and to prevent apoptosis are likely to exert maximal benefits when initiated at the acute phase of TBI.

Impaired blood-brain/spinal cord barrier in ALS patients.

Vascular pathology, including blood-brain/spinal cord barrier (BBB/BSCB) alterations, has recently been recognized as a key factor possibly aggravating motor neuron damage, identifying a neurovascular disease signature for ALS. However, BBB/BSCB competence in sporadic ALS (SALS) is still undetermined. In this study, BBB/BSCB integrity in postmortem gray and white matter of medulla and spinal cord tissue from SALS patients and controls was investigated. Major findings include (1) endothelial cell damage and pericyte degeneration, (2) severe intra- and extracellular edema, (3) reduced CD31 and CD105 expressions in endothelium, (4) significant accumulation of perivascular collagen IV, and fibrin deposits (5) significantly increased microvascular density in lumbar spinal cord, (6) IgG microvascular leakage, (7) reduced tight junction and adhesion protein expressions. Microvascular barrier abnormalities determined in gray and white matter of the medulla, cervical, and lumbar spinal cord of SALS patients are novel findings. Pervasive barrier damage discovered in ALS may have implications for disease pathogenesis and progression, as well as for uncovering novel therapeutic targets.

Multiple intravenous administrations of human umbilical cord blood cells benefit in a mouse model of ALS.

BACKGROUND: A promising therapeutic strategy for amyotrophic lateral sclerosis (ALS) is the use of cell-based therapies that can protect motor neurons and thereby retard disease progression. We recently showed that a single large dose (25 × 106 cells) of mononuclear cells from human umbilical cord blood (MNC hUCB) administered intravenously to pre-symptomatic G93A SOD1 mice is optimal in delaying disease progression and increasing lifespan. However, this single high cell dose is impractical for clinical use. The aim of the present pre-clinical translation study was therefore to evaluate the effects of multiple low dose systemic injections of MNC hUCB cell into G93A SOD1 mice at different disease stages. METHODOLOGY/PRINCIPAL FINDINGS: Mice received weekly intravenous injections of MNC hUCB or media. Symptomatic mice received 106 or 2.5 × 106 cells from 13 weeks of age. A third, pre-symptomatic, group received 106 cells from 9 weeks of age. Control groups were media-injected G93A and mice carrying the normal hSOD1 gene. Motor function tests and various assays determined cell effects. Administered cell distribution, motor neuron counts, and glial cell densities were analyzed in mouse spinal cords. Results showed that mice receiving 106 cells pre-symptomatically or 2.5 × 106 cells symptomatically significantly delayed functional deterioration, increased lifespan and had higher motor neuron counts than media mice. Astrocytes and microglia were significantly reduced in all cell-treated groups. CONCLUSIONS/SIGNIFICANCE: These results demonstrate that multiple injections of MNC hUCB cells, even beginning at the symptomatic disease stage, could benefit disease outcomes by protecting motor neurons from inflammatory effectors. This multiple cell infusion approach may promote future clinical studies.

Stem cell therapies for type 1 diabetes mellitus.

The present review discusses the use of autologous hematopoietic stem cell transplantation (HSCT) for the treatment of diabetes mellitus type 1 (DM 1). It has been observed that high dose immunosuppression followed by HSCT shows better results among other immunotherapeutic treatments for the disease as the patients with adequate beta cell reserve achieve insulin independence. However, this response is not maintained and reoccurrence of the disease is major a major challenge to use HSCT in future to prevent or control relapse of DM 1.

Amyotrophic lateral sclerosis: a neurovascular disease.

Amyotrophic lateral sclerosis (ALS) is a severe neurodegenerative disease with a complicated pathogenesis. Compelling evidence indicates impairment of all neurovascular unit components including the blood-brain and blood-spinal cord barriers (BBB/BSCB) in both patients and animal models, leading to classification of ALS as a neurovascular disease. The present review provides an updated analysis of the normal and impaired BBB/BSCB, focusing on the ALS-altered barrier. Here we describe the roles of cellular components, tight junctions, transport systems, cell interactions, cytokines, matrix metalloproteinases, and free radicals in the BBB/BSCB disruption, along with recent evidence from experimental and clinical ALS studies. The BBB/BSCB is a promising research area in ALS and this review will reveal some aspects of microvascular pathology in ALS and hopefully provide ideas for the development of new therapeutic strategies.

Thyroid hormone receptor binding to DNA and T3-dependent transcriptional activation are inhibited by uremic toxins.

BACKGROUND: There is a substantial clinical overlap between chronic renal failure (CRF) and hypothyroidism, suggesting the presence of hypothyroidism in uremic patients. Although CRF patients have low T3 and T4 levels with normal thyroid-stimulating hormone (TSH), they show a higher prevalence of goiter and evidence for blunted tissue responsiveness to T3 action. However, there are no studies examining whether thyroid hormone receptors (TRs) play a role in thyroid hormone dysfunction in CRF patients. To evaluate the effects of an uremic environment on TR function, we investigated the effect of uremic plasma on TRbeta1 binding to DNA as heterodimers with the retinoid X receptor alpha (RXRalpha) and on T3-dependent transcriptional activity. RESULTS: We demonstrated that uremic plasma collected prior to hemodialysis (Pre-HD) significantly reduced TRbeta1-RXRalpha binding to DNA. Such inhibition was also observed with a vitamin D receptor (VDR) but not with a peroxisome proliferator-activated receptor gamma (PPARgamma). A cell-based assay confirmed this effect where uremic pre-HD ultrafiltrate inhibited the transcriptional activation induced by T3 in U937 cells. In both cases, the inhibitory effects were reversed when the uremic plasma and the uremic ultrafiltrate were collected and used after hemodialysis (Post-HD). CONCLUSION: These results suggest that dialyzable toxins in uremic plasma selectively block the binding of TRbeta1-RXRalpha to DNA and impair T3 transcriptional activity. These findings may explain some features of hypothyroidism and thyroid hormone resistance observed in CRF patients.

Thyroid hormone transport is disturbed in erythrocytes from patients with chronic renal failure on hemodialysis.

AIMS: To now, there are no studies reporting whether thyroid hormones (THs) transport play a role in thyroid hormone dysfunction observed in chronic renal failure (CRF). Therefore, the aim of this study was to investigate the transport of THs in erythrocytes from patients with CRF on hemodialysis (HD). METHODS: [125I]-L-triiodothyronine ([125I]T3) and [125I]-L-thyroxine ([125I]T4) erythrocytes uptake was measured at 1 min and 5 min. To study L-triiodothyronine (LT3) and L-thyroxine (LT4) efflux from erythrocytes, we preloaded the cells during 180 min with [125I]T3 or [125I]T4 and measured their [125I]T3 or [125I]T4 efflux during 60 min. RESULTS: [125I]T3 uptake in erythrocytes from uremic patients pre-HD was higher than control subjects by 50% at 1 min and by 55% at 5 min. However, [125I]T4 uptake in erythrocytes from uremic patients was significantly lower at 1min (88%) and at 5 min (63%). LT3 efflux rate was lower and LT4 efflux was significantly higher than in control subjects. After 60-min of efflux, LT3 remained in erythrocytes was 80% higher and LT4 was 57% lower than in normal individuals. Neither [125I]T3 and [125I]T4 uptake, nor efflux rates were changed by hemodialysis. CONCLUSION: Despite the fact that uremic patients on hemodialysis show low serum levels of LT3, changes in LT3 influx and efflux could act as a compensatory mechanism that neutralize thyroid hormone dysfunction in order to maintain the euthyroid state.