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Biomedical subjects

Howard J Federoff

Publications and source records attributed to Howard J Federoff.

At least 19 recordsLinked to original sources
PubMed2006

Effects of ex vivo transduction of mesencephalic reaggregates with bcl-2 on grafted dopamine neuron survival.

Survival rates of dopamine (DA) neurons grafted to the denervated striatum are extremely poor (5-20%). Gene transfer of survival promoting factors, such as the anti-apoptotic protein bcl-2, to mesencephalic DA neurons prior to transplantation (ex vivo transduction) offers a novel approach to increase graft survival. However, specific criteria to assess the efficacy of various vectors must be adhered to in order to reasonably predict successful gene transfer with appropriate timing and levels of protein expression. Cell culture results utilizing three different herpes simplex virus (HSV) vectors to deliver the reporter beta-galactosidase gene (lacZ) indicate that transduction of mesencephalic cells with a helper virus-free HSV amplicon (HF HSV-TH9lac) that harbors the 9-kb tyrosine hydroxylase (TH) promoter to drive lacZ gene expression elicits the transduction of the highest percentage (approximately 50%) of TH-immunoreactive (THir) neurons without significant cytotoxic effects. This transduction efficiency and limited cytotoxicity was superior to that observed following transduction with helper virus-containing HSV (HC HSVlac) and helper virus-free HSV amplicons (HF HSVlac) expressing lacZ under the transcriptional control of the HSV immediate-early 4/5 gene promoter. Subsequently, we assessed the ability of HSV-TH9lac and the bcl-2 expressing HSV-TH9bcl-2 amplicon to transduce mesencephalic reaggregates. Although an increase in bcl-2 and beta-galactosidase protein was induced by transduction, amplicon-mediated overexpression of bcl-2 did not lead to an increase in grafted THir neuron number. Even with highly efficient viral vector-mediated transduction, our results demonstrate that ex vivo gene transfer of bcl-2 to mesencephalic reaggregates is ineffective in increasing grafted DA neuron survival.

Animals↗
PubMed2006

Effect of promoter strength on protein expression and immunogenicity of an HSV-1 amplicon vector encoding HIV-1 Gag.

Helper-free herpes simplex virus type-1 (HSV-1) amplicon vectors elicit robust immune responses to encoded proteins, including human immunodeficiency virus type-1 (HIV-1) antigens. To improve this vaccine delivery system, seven amplicon vectors were constructed, each encoding HIV-1 Gag under the control of a different promoter. Gag expression levels were analyzed in murine and human cell lines, as well as in biopsied tissue samples from injected mice; these data were then compared with Gag-specific T cell responses in BALB/c mice. The magnitude of the amplicon-induced immune response was found to correlate strongly with the level of Gag production both in vitro and in vivo. Interestingly, the best correlation of the strength of the amplicon-induced immune response was with antigen expression in cultured DC rather than expression at the tissue site of injection or in cultured cell lines. These findings may have implications for the generation of improved HSV-1 amplicon vectors for HIV-1 vaccine delivery.

3T3 Cells↗
PubMed2006

VIP is a transcriptional target of Nurr1 in dopaminergic cells.

The orphan nuclear receptor Nurr1 is required for the development of the ventral mesencephalic dopaminergic neurons. These are the same neurons that are invariantly lost in patients with Parkinson's disease. Nurr1 mRNA expression is not confined to the developing midbrain, and yet Nurr1 appears to be essential for either the maturation of progenitors into fully post-mitotic dopaminergic neurons and/or once formed, their survival. The function of Nurr1 in the transactivation of gene(s) important for neuronal development and/or maintenance is uncharacterized. To characterize potential downstream target genes of Nurr1, we sought to identify mRNAs that are differentially affected by Nurr1 expression. Using a dopaminergic cell line in which Nurr1 content was tightly regulated, differential display analysis identified transcripts altered by Nurr1 expression, including the mRNA encoding vasoactive intestinal peptide (VIP). Herein, we demonstrate that Nurr1 regulates VIP mRNA and protein levels, and transactivates the VIP promoter through Nurr1-responsive cis elements. In addition, dopaminergic cells release and utilize VIP to mediate survival when challenged with paraquat. Nurr1 regulation of VIP is also demonstrated in vivo as loss of Nurr1 function results in diminished VIP mRNA levels within the developing midbrain.

Animals↗
PubMed2006

Identification of human alpha-synuclein specific single chain antibodies.

Parkinson's disease (PD) is a common neurodegenerative disease of unknown etiology. Evidence suggests a role for protein misfolding in disease pathogenesis. One pathologic feature observed in dopaminergic neurons is the intracytoplasmic eosinophilic inclusions known as Lewy bodies. One component of Lewy bodies, the presynaptic protein, alpha-synuclein forms oligomers and higher order aggregates and is proposed to be involved in dopaminergic neuronal death. In an effort to discriminate between alpha-synuclein conformational forms as well as design potential disruptors of pathogenic misfolding we panned a human phage antibody library for anti-synuclein single chain antibodies (scFvs). We identified six scFvs which recognize different conformers of alpha-synuclein in both an ELISA and Western blot analysis. These scFvs may further our understanding of alpha-synuclein's role in PD.

Humans↗
PubMed2006

Visual deficits in a mouse model of Batten disease are the result of optic nerve degeneration and loss of dorsal lateral geniculate thalamic neurons.

Juvenile neuronal ceroid lipofuscinosis (JNCL) is an autosomal recessive disorder of childhood caused by mutations in CLN3. Although visual deterioration is typically the first clinical sign to manifest in affected children, loss of Cln3 in a mouse model of JNCL does not recapitulate this retinal deterioration. This suggests that either the loss of CLN3 does not directly affect retinal cell survival or that nuclei involved in visual processing are affected prior to retinal degeneration. Having previously demonstrated that Cln3(-/-) mice have decreased optic nerve axonal density, we now demonstrate a decrease in nerve conduction. Examination of retino-recipient regions revealed a decreased number of neurons within the dorsal lateral geniculate nucleus (LGNd). We demonstrate decreased transport of amino acids from the retina to the LGN, suggesting an impediment in communication between the retina and projection nuclei. This study defines a novel path of degeneration within the LGNd, providing a mechanism for causation of JNCL visual deficits.

Animals↗
PubMed2006

Neuronal precursor-restricted transduction via in utero CNS gene delivery of a novel bipartite HSV amplicon/transposase hybrid vector.

The ability to modify genetically in utero the precursors of neuronal lineage contributing to multiple postmitotic cell types in the adult central nervous system would provide a means to evaluate strategies to ameliorate conditions affecting cellular patterning, metabolism, or survival. The herpes simplex virus (HSV)-derived amplicon, a vector devoid of viral genes and with the largest known payload capacity, normally exists episomally within nuclei of transduced cells, thus precluding conveyance during mitosis. Herein, we modify the Tc1-like Sleeping Beauty (SB) transposon system to create an integrating amplicon vector platform wherein provision of transposase in trans effectively catalyzes integration of a transgenomic segment. Cotransduction with a Rous sarcoma virus promoter-driven beta-galactosidase-neomycin (betageo) fusion flanked by SB terminal repeats (HSVT-betageo) and a second expressing the SB transposase gene under HSV immediate-early 4/5 gene promoter control (HSVsb) resulted in integration and extension of expression duration. Most notably, in utero intraventricular application led to extensive transgene expression within neuronal precursors and their derivatives without attendant adverse consequences, suggesting this new platform could be used to evaluate prenatally the function of gene products in neuronal lineages and evaluate therapeutic strategies for correction of genetic abnormalities affecting the developing CNS.

Animals↗
PubMed2006

Microarrays in Parkinson's disease: a systematic approach.

Neurological disease (ND) is one of the greatest challenges facing our population, from medical, financial, and social perspectives. The application of new research approaches to understand the underlying pathogenesis of ND is critical. In this article, we review the use of microarray analysis in Parkinson's disease (PD). Microarrays have tremendous power, simultaneously querying the expression of tens of thousands of genes from a given biological sample. Coupled with impressive advances in statistical tools for analyzing large, complex data sets, well-designed microarray experiments are poised to make a big impact in the field of ND. Parkinson's disease is a devastating neurodegenerative disease well suited to a systems-based microarray analysis. Genetic and environmental rodent models of PD emulate many of the cardinal features of human PD, providing the unique opportunity to compare gene expression profiles from different etiologies of the same disease. The elucidation of important gene expression patterns during disease will make possible identification of genetic susceptibility markers, biomarkers of disease progression, and new therapeutic targets.

Animals↗
PubMed2006

HSV amplicons: neuro applications.

Strategies that employ HSV amplicon vectors in the prevention and/or amelioration of pathogenic states afflicting the central nervous system (CNS) have been extensively documented in preclinical disease models. The versatility of the HSV amplicon platform allows for the implementation of therapeutic approaches that require expression of genes exhibiting neuroprotective or neuroplastic activities, or even applications that necessitate the elaboration of antigen-specific immune responses to pathogenic proteins/structures harbored within the CNS. This discourse highlights the successes and challenges encountered using HSV amplicon vectors as tools for the dissection of neural network function and as therapeutics directed against a variety of neurologic disorders.

Animals↗
PubMed2006

CNS gene therapy and a nexus of complexity: systems and biology at a crossroads.

Gene therapy is a potentially promising new treatment for neurodegenerative disorders such as Alzheimer's disease (AD), which has been difficult to treat with conventional therapeutics. Viral vector-mediated somatic gene therapy is a rapidly developing methodology for providing never before achieved capability to deliver specific genes to the CNS in a highly localized and controlled manner. With the advent and refinements of this technology one focus is directed to which genes are the most appropriate to select for specific disease indications. Nerve growth factor (NGF), a potent survival factor for critical cell populations that degenerate in AD, has been chosen already for clinical gene therapy trials in human AD patients. Much knowledge about the pathophysiological underpinnings of AD is still lacking to make clear which patients may benefit from a gene therapy approach. Moreover, a detailed understanding of sustained NGF action in the normal and diseased CNS needs to be resolved before conclusions can be drawn regarding the utility of NGF gene therapy. Systematic efforts to acquire this new knowledge should compel clinically and biologically sophisticated efforts to advance gene therapy for neurodegenerative diseases.

Alzheimer Disease↗
PubMed2005

Immune shaping and the development of Alzheimer's disease vaccines.

Given the emotional, social, and financial devastation wrought by Alzheimer's disease (AD), it is imperative that effective therapeutics be devised to ameliorate this presently incurable disorder. Vaccine-based approaches have been developed to target and eliminate amyloid beta (Abeta), a key peptide implicated in AD pathogenesis. Preclinical successes in AD mouse models created excitement and impetus for the clinical application of an Abeta-based vaccine. Eliciting immune responses against a self-peptide (that is, a peptide produced by the organism itself), such as Abeta, carries with it the potential to induce autoimmune and inflammatory conditions in the vaccinated individual, a caveat borne out in multiple patients enrolled as part of a recent clinical trial. These clinical adverse events seemingly overshadowed interesting behavioral stabilization and alterations of Abeta burden in these and other vaccinated patients, thus speaking to the potential of immunotherapy for AD. Understanding the mechanisms by which vaccines reduce Abeta burden in AD brain and the types of immune responses raised, as well as developing new modalities of vaccine delivery that facilitate the modulation of elicited immune responses, will undoubtedly lead to a new generation of efficacious Abeta immunotherapeutics with improved safety profiles.

Age Factors↗
PubMed2005

Combined delivery of neurotrophin-3 and NMDA receptors 2D subunit strengthens synaptic transmission in contused and staggered double hemisected spinal cord of neonatal rat.

We investigated whether administration of neurotrophin-3 (NT-3) and NMDA-2D-expressing units, found previously to enhance transmission in neonatal rat spinal cord, strengthens synaptic connections in the injured neonatal cord. We employed electrophysiological methods to evaluate the strength of synaptic transmission to individual motoneurons in the contusion and staggered double hemisection spinal cord injury (SCI) models. SCI at caudal thoracic levels (T11-T12) was carried out at postnatal day 2 (P2). Plugs containing NT-3- secreting fibroblasts and NR2D-expressing HSV-1 amplicons (HSVnr2d) were implanted above the lesion. Control animals were treated with an amplicon-expressing beta-galactosidase (HSVlac). After 8-10 days of treatment, the rats were sacrificed and spinal cords were removed for intracellular recording. Untreated contused cords preserved a fraction of white matter and weak monosynaptic responses were observed through the injury region. However, no synaptic connections were observed in control cords receiving double hemisection injury. Combined treatment with NT-3 and HSVnr2d strengthened monosynaptic connections in contused cords and induced the appearance of weak but functional multisynaptic connections in double hemisected cords. In contrast, treatment with either NT-3 or HSVnr2d alone failed to induce appearance of synaptic responses through the hemisected region. These results suggest that chronic treatment with NT-3 secreting fibroblasts combined with facilitated function of NMDA receptors by HSVnr2d treatment strengthens connections that survive incomplete SCI and therefore that such combined treatment might facilitate recovery of function following SCI.

Animals↗
PubMed2005

Herpes simplex virus amplicon delivery of a hypoxia-inducible soluble vascular endothelial growth factor receptor (sFlk-1) inhibits angiogenesis and tumor growth in pancreatic adenocarcinoma.

BACKGROUND: Tumor hypoxia induces vascular endothelial growth factor (VEGF) expression, which stimulates angiogenesis and tumor proliferation. The VEGF signaling pathway is inhibited by soluble VEGF receptors (soluble fetal liver kinase 1; sFlk-1), which bind VEGF and block its interaction with endothelial cells. Herpes simplex virus (HSV) amplicons are replication-incompetent viruses used for gene delivery. We attempted to attenuate angiogenesis and inhibit pancreatic tumor growth through HSV amplicon-mediated expression of sFlk-1 under hypoxic control. METHODS: A multimerized hypoxia-responsive enhancer (10 x HRE) was cloned upstream of the sFlk-1 gene (10 x HRE/sFlk-1). A novel HSV amplicon expressing 10 x HRE/sFlk-1 was genetically engineered (HSV10 x HRE/sFlk-1).Human pancreatic adenocarcinoma cells (AsPC1) were transduced with HSV10 x HRE/sFlk-1 and incubated in normoxia (21% oxygen) or hypoxia (1% oxygen). Capillary inhibition was evaluated by human umbilical vein endothelial cell assay. Western blot assessed sFlk-1 expression. AsPC1 flank tumor xenografts (n = 24) were transduced with HSV10 x HRE/sFlk-1. RESULTS: Media from normoxic AsPC1 transduced with HSV10 x HRE/sFlk-1 yielded a 36% reduction in capillary formation versus controls (P < .05), whereas hypoxic AsPC1 yielded a 76% reduction (P < .005). Western blot of AsPC1 transduced with HSV10 x HRE/sFlk-1 demonstrated greater sFlk-1 expression in hypoxia versus normoxia. AsPC1 flank tumors treated with HSV10 x HRE/sFlk-1 exhibited a 59% reduction in volume versus controls (P < .000001). CONCLUSIONS: HSV amplicon delivery of a hypoxia-inducible soluble VEGF receptor significantly reduces new vessel formation and tumor growth. Tumor hypoxia can thus be used to direct antiangiogenic therapy to pancreatic adenocarcinoma.

Adenocarcinoma↗
PubMed2005

Early correlation of microglial activation with enhanced tumor necrosis factor-alpha and monocyte chemoattractant protein-1 expression specifically within the entorhinal cortex of triple transgenic Alzheimer's disease mice.

BACKGROUND: Alzheimer's disease is a complex neurodegenerative disorder characterized pathologically by a temporal and spatial progression of beta-amyloid (Abeta) deposition, neurofibrillary tangle formation, and synaptic degeneration. Inflammatory processes have been implicated in initiating and/or propagating AD-associated pathology within the brain, as inflammatory cytokine expression and other markers of inflammation are pronounced in individuals with AD pathology. The current study examines whether inflammatory processes are evident early in the disease process in the 3xTg-AD mouse model and if regional differences in inflammatory profiles exist. METHODS: Coronal brain sections were used to identify Abeta in 2, 3, and 6-month 3xTg-AD and non-transgenic control mice. Quantitative real-time RT-PCR was performed on microdissected entorhinal cortex and hippocampus tissue of 2, 3, and 6-month 3xTg-AD and non-transgenic mice. Microglial/macrophage cell numbers were quantified using unbiased stereology in 3xTg-AD and non-transgenic entorhinal cortex and hippocampus containing sections. RESULTS: We observed human Abeta deposition at 3 months in 3xTg-AD mice which is enhanced by 6 months of age. Interestingly, we observed a 14.8-fold up-regulation of TNF-alpha and 10.8-fold up-regulation of MCP-1 in the entorhinal cortex of 3xTg-AD mice but no change was detected over time in the hippocampus or in either region of non-transgenic mice. Additionally, this increase correlated with a specific increase in F4/80-positive microglia and macrophages in 3xTg-AD entorhinal cortex. CONCLUSION: Our data provide evidence for early induction of inflammatory processes in a model that develops amyloid and neurofibrillary tangle pathology. Additionally, our results link inflammatory processes within the entorhinal cortex, which represents one of the earliest AD-affected brain regions.

Journal Article↗
PubMed2005

Blockade of gap junctions in vivo provides neuroprotection after perinatal global ischemia.

BACKGROUND AND PURPOSE: We investigated the contribution of gap junctions to brain damage and delayed neuronal death produced by oxygen-glucose deprivation (OGD). METHODS: Histopathology, molecular biology, and electrophysiological and fluorescence cell death assays in slice cultures after OGD and in developing rats after intrauterine hypoxia-ischemia (HI). RESULTS: OGD persistently increased gap junction coupling and strongly activated the apoptosis marker caspase-3 in slice cultures. The gap junction blocker carbenoxolone applied to hippocampal slice cultures before, during, or 60 minutes after OGD markedly reduced delayed neuronal death. Administration of carbenoxolone to ischemic pups immediately after intrauterine HI prevented caspase-3 activation and dramatically reduced long-term neuronal damage. CONCLUSIONS: Gap junction blockade may be a useful therapeutic tool to minimize brain damage produced by perinatal and early postnatal HI.

Animals↗
PubMed2005

Robust dysregulation of gene expression in substantia nigra and striatum in Parkinson's disease.

Large-scale genomics approaches are now widely utilized to study a myriad of human diseases. These powerful techniques, when combined with data analysis tools, detect changes in transcript abundance in diseased tissue relative to control. We hypothesize that specific differential gene expression underlies important pathogenic processes in Parkinson's disease, which is characterized by the gradual loss of dopaminergic neurons in the substantia nigra and consequent loss of dopamine in the striatum. We have therefore examined gene expression levels in the human parkinsonian nigrostriatal pathway, and compared them with those of neurologically normal controls. Using unsupervised clustering methods, we demonstrate that relatively few genes' expression levels can effectively distinguish between disease and control brains. Further, we identify several interesting patterns of gene expression that illuminate pathogenic cascades in Parkinson's disease. In particular is the robust loss of synaptic gene expression in diseased substantia nigra and striatum.

Aged↗
PubMed2005

Synuclein, dopamine and oxidative stress: co-conspirators in Parkinson's disease?

The etiology of Parkinson's disease (PD) is presently unknown. The unifying hallmark of disease is depletion of dopamine and loss of nigrostriatal dopamine neurons. Familial and sporadic forms of the disease are described. The familial mutations occur within alpha-synuclein and molecules involved in protein degradation and mitochondrial function. Sporadic PD is thought to involve the interplay of genetic and environmental factors. Despite disparate initiating triggers, a convergent pathobiologic model for this common neurodegenerative disease has been proposed. Likely players have emerged that may form the basis for this common pathway model of disease. In this review, we examine the role of three most implicated PD pathogenic conspirators: synuclein, dopamine and oxidative stress.

Animals↗
PubMed2005

beta-hexosaminidase lentiviral vectors: transfer into the CNS via systemic administration.

Brain inflammation in GM2 gangliosidosis has been recently realized as a key factor in disease development. The aim of this study was to investigate the effects of a FIV beta-hexosaminidase vector in the brain of HexB-deficient (Sandhoff disease) mice following intraperitoneal administration to pups of neonatal age. Since brain inflammation, lysosomal storage and neuromuscular dysfunction are characteristics of HexB deficiency, these parameters were employed as experimental outcomes in our study. The ability of the lentiviral vector FIV(HEX) to infect murine cells was initially demonstrated with success in normal mouse fibroblasts and human Tay-Sachs cells in vitro. Furthermore, systemic transfer of FIV(HEX) to P2 HexB-/- knockout pups lead to transduction of peripheral and central nervous system tissues. Specifically, beta-hexosaminidase expressing cells were immunolocalized in periventricular areas of the cerebrum as well as in the cerebellar cortex. FIV(HEX) neonatal treatment resulted in reduction of GM2 storage along with attenuation of the brain inflammation and amelioration of the attendant neuromuscular deterioration. In conclusion, these results demonstrate the effective transfer of a beta-hexosaminidase lentiviral vector to the brain of Sandhoff mice and resolution of the GM2 gangliosidosis after neonatal intraperitoneal administration.

Animals↗