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1.
Gene Ther ; 26(10-11): 432-440, 2019 11.
Article in English | MEDLINE | ID: mdl-30770896

ABSTRACT

Biological rejuvenation by partial cell reprogramming is an emerging avenue of research. In this context, regulatable pluripotency gene expression systems are the most widely used at present. We have constructed a regulatable bidirectional adenovector expressing the humanized green fluorescent protein (GFP) and oct4, sox2, klf4, and c-myc genes (known as the Yamanaka genes or OSKM). The OSKM genes are arranged as a bicistronic tandem (hSTEMCCA tandem), which is under the control of a Tet-Off bidirectional promoter that also controls the expression of the gFP gene. Separately, a constitutive cassette expresses the regulatory protein tTA. Vector DNA was transfected in HEK293 Cre cells, which were additionally infected with the helper adenovector H14, unable to package its DNA due to the Cre recombinase produced by the HEK293 Cre cells. The newly generated vector was expanded by six iterated coinfections of the above cells which were lysed at the end of the process and the adenovector purified by ultracentrifugation in a CsCl gradient. The titer of the initial preparation was 1.2 × 1012 physical viral particles/ml. As expected, GFP fluorescence in vector-transduced rat fibroblast cultures declined with the dose of doxycycline (DOX) present in the medium. Immunocytochemical analysis of transduced cells confirmed the expression of the four Yamanaka genes. Additionally, 3 days after vector injection in the hypothalamus of rats, a significant level of fluorescence was observed in the region. Addition of 2 mg/ml DOX to the drinking water reduced the GFP expression. This adenovector constitutes a promising tool for implementing nonintegrative partial cell reprogramming.


Subject(s)
Brain/physiology , Genetic Therapy/methods , Kruppel-Like Transcription Factors/genetics , Octamer Transcription Factor-3/genetics , Proto-Oncogene Proteins c-myc/genetics , Regeneration , SOXB1 Transcription Factors/genetics , Adenoviridae/genetics , Animals , Cells, Cultured , Female , Gene Transfer Techniques , Genetic Vectors/genetics , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , HEK293 Cells , Humans , Kruppel-Like Factor 4 , Kruppel-Like Transcription Factors/metabolism , Octamer Transcription Factor-3/metabolism , Proto-Oncogene Proteins c-myc/metabolism , Rats , Rats, Sprague-Dawley , SOXB1 Transcription Factors/metabolism
2.
J Neural Transm (Vienna) ; 125(12): 1787-1803, 2018 12.
Article in English | MEDLINE | ID: mdl-30244292

ABSTRACT

Sporadic Alzheimer's disease (SAD) is the most common form of dementia; therefore, there is an urgent need for a model that recapitulates the main pathologic hallmarks of this disease. The intracerebroventricular (icv) injection of streptozotocin (icv-STZ) in rats constitutes a promising model, and thus, icv-STZ rats develop insulin-resistant brain state and cognitive impairments. Even though a great piece of studies has hitherto described this system as a model for SAD, further behavioral and morphometric studies are still needed to fully characterize it. In this study, using Sprague Dawley rats, we evaluated short-term effects on behavior and hippocampus morphometry of the icv-STZ injection at two doses: 1 (STZ1) and 3 mg/kg (STZ3). We found that, following icv-STZ injection, STZ3 animals, but not STZ1, exhibited impairments in spatial reference learning and memory (Barnes maze test) and in recognition memory (object recognition test). Furthermore, the results from behavioral and morpho-histochemical data are compatible. STZ3 rats displayed Stratum Radiatum volume reduction and a decreased NeuN immunoreactivity (neuron loss) in hippocampal CA1 region, together with an increased immunoreactivity for microglial (Iba1) and astroglial (GFAP) markers (neuroinflammation). Sholl analysis revealed the vulnerability of hippocampal astrocytes to STZ in CA1 and CA3. Thus, both doses induced a reduction in process length and in the number of main processes, accompanied by a frank decrease in branching complexity. The present study provides important knowledge of this AD rat model. Overall, we found that the only high STZ dose induced severe and acute neurodegenerative lesions, associated with an inflammation process.


Subject(s)
Astrocytes/drug effects , CA1 Region, Hippocampal/drug effects , CA3 Region, Hippocampal/drug effects , Spatial Memory/drug effects , Streptozocin/pharmacology , Animals , Astrocytes/cytology , CA1 Region, Hippocampal/cytology , CA3 Region, Hippocampal/cytology , Cell Shape/drug effects , Injections, Intraventricular , Male , Maze Learning/drug effects , Rats , Rats, Sprague-Dawley
3.
Hippocampus ; 27(4): 435-449, 2017 04.
Article in English | MEDLINE | ID: mdl-28085212

ABSTRACT

There have been a few descriptive studies in aged rodents about transcriptome changes in the hippocampus, most of them in males. Here, we assessed the age changes in spatial memory performance and hippocampal morphology in female rats and compared those changes with changes in the hippocampal transcriptome. Old rats displayed significant deficits in spatial memory. In both age groups, hole exploration frequency showed a clear peak at hole 0 (escape hole), but the amplitude of the peak was significantly higher in the young than in the old animals. In the hippocampus, there was a dramatic reduction in neurogenesis, whereas reactive microglial infiltrates revealed an inflammatory hippocampal state in the senile rats. Hippocampal RNA-sequencing showed that 210 genes are differentially expressed in the senile rats, most of them being downregulated. Our RNA-Seq data showed that various genes involved in the immune response, including TYROBP, CD11b, C3, CD18, CD4, and CD74, are overexpressed in the hippocampus of aged female rats. Enrichment analysis showed that the pathways overrepresented in the senile rats matched those of an exacerbated inflammatory environment, reinforcing our morphologic findings. After correlating our results with public data of human and mouse hippocampal gene expression, we found an 11-gene signature of overexpressed genes related to inflammatory processes that was conserved across species. We conclude that age-related hippocampal deficits in female rats share commonalities between human and rodents. Interestingly, the 11-gene signature that we identified may represent a cluster of immune and regulatory genes that are deregulated in the hippocampus and possibly other brain regions during aging as well as in some neurodegenerative diseases and low-grade brain tumors. Our study further supports neuroinflammation as a promising target to treat cognitive dysfunction in old individuals and some brain tumors. © 2017 Wiley Periodicals, Inc.


Subject(s)
Aging/immunology , Aging/pathology , Hippocampus/immunology , Hippocampus/pathology , Spatial Memory/physiology , Adult , Aged , Aged, 80 and over , Aging/genetics , Aging/psychology , Animals , Dementia/immunology , Dementia/pathology , Female , Gene Expression , Humans , Male , Maze Learning/physiology , Microglia/metabolism , Microglia/pathology , Middle Aged , Neurogenesis/physiology , Neurons/immunology , Neurons/pathology , Rats, Sprague-Dawley , Species Specificity , Transcriptome , Young Adult
4.
Eur J Neurosci ; 44(4): 2120-8, 2016 08.
Article in English | MEDLINE | ID: mdl-27188415

ABSTRACT

In rats, learning and memory performance decline during aging, which makes this rodent species a suitable model to evaluate therapeutic strategies of potential value for correcting age-related cognitive deficits. Some of these strategies involve neurotrophic factors like insulin-like growth factor-I (IGF-I), a powerful neuroprotective molecule in the brain. Here, we implemented 18-day long intracerebroventricular (ICV) IGF-I gene therapy in 28 months old Sprague-Dawley female rats, and assessed spatial memory performance in the Barnes maze. We also studied hippocampal morphology using an unbiased stereological approach. Adenovectors expressing the gene for rat IGF-I or the reporter DsRed were used. Cerebrospinal fluid (CSF) samples were taken and IGF-I levels determined by radioimmunoassay. At the end of the study, IGF-I levels in the CSF were significantly higher in the experimental group than in the DsRed controls. After treatment, the IGF-I group showed a significant improvement in spatial memory accuracy as compared with DsRed counterparts. In the dentate gyrus (DG) of the hippocampus, the IGF-I group showed a higher number of immature neurons than the DsRed controls. The treatment increased hippocampal astrocyte branching and reduced their number in the hippocampal stratum radiatum. We conclude that the ependymal route is an effective approach to increase CSF levels of IGF-I and that this strategy improves the accuracy of spatial memory in aging rats. The favorable effect of the treatment on DG neurogenesis and astrocyte branching in the stratum radiatum may contribute to improving memory performance in aging rats.


Subject(s)
Astrocytes/metabolism , Hippocampus/metabolism , Insulin-Like Growth Factor I/metabolism , Neurogenesis/physiology , Spatial Memory/physiology , Animals , Cognition/physiology , Cognition Disorders/genetics , Cognition Disorders/metabolism , Cognition Disorders/therapy , Female , Genetic Therapy , Hippocampus/cytology , Insulin-Like Growth Factor I/genetics , Memory Disorders/genetics , Memory Disorders/therapy , Rats, Sprague-Dawley
5.
Ageing Res Rev ; 99: 102408, 2024 Aug.
Article in English | MEDLINE | ID: mdl-38969142

ABSTRACT

Alzheimer's disease (AD) and osteoporosis are two diseases that mainly affect elderly people, with increases in the occurrence of cases due to a longer life expectancy. Several epidemiological studies have shown a reciprocal association between both diseases, finding an increase in incidence of osteoporosis in patients with AD, and a higher burden of AD in osteoporotic patients. This epidemiological relationship has motivated the search for molecules, genes, signaling pathways and mechanisms that are related to both pathologies. The mechanisms found in these studies can serve to improve treatments and establish better patient care protocols.


Subject(s)
Alzheimer Disease , Osteoporosis , Humans , Alzheimer Disease/epidemiology , Osteoporosis/epidemiology , Incidence
6.
Neural Regen Res ; 17(3): 469-476, 2022 Mar.
Article in English | MEDLINE | ID: mdl-34380873

ABSTRACT

Hippocampus serves as a pivotal role in cognitive and emotional processes, as well as in the regulation of the hypothalamus-pituitary axis. It is known to undergo mild neurodegenerative changes during normal aging and severe atrophy in Alzheimer's disease. Furthermore, dysregulation in the hippocampal function leads to epilepsy and mood disorders. In the first section, we summarized the most salient knowledge on the role of glial cell-line-derived neurotrophic factor and its receptors focused on aging, cognition and neurodegenerative and hippocampal-related neurological diseases mentioned above. In the second section, we reviewed the therapeutic approaches, particularly gene therapy, using glial cell-line-derived neurotrophic factor or its gene, as a key molecule in the development of neurological disorders. In the third section, we pointed at the potential of regenerative medicine, as an emerging and less explored strategy for the treatment of hippocampal disorders. We briefly reviewed the use of partial reprogramming to restore brain functions, non-neuronal cell reprogramming to generate neural stem cells, and neural progenitor cells as source-specific neuronal types to be implanted in animal models of specific neurodegenerative disorders.

7.
J Gerontol A Biol Sci Med Sci ; 77(12): 2387-2394, 2022 12 29.
Article in English | MEDLINE | ID: mdl-35917578

ABSTRACT

In humans and rats, aging is associated with a progressive deterioration of spatial learning and memory. These functional alterations are correlated with morphological and molecular changes in the hippocampus. Here, we assessed age-related changes in DNA methylation (DNAm) landscape in the rat hippocampus and the correlation of spatial memory with hippocampal DNAm age in 2.6- and 26.6-month-old rats. Spatial memory performance was assessed with the Barnes maze test. To evaluate learning ability and spatial memory retention, we assessed the time spent by animals in goal sector 1 (GS1) and 3 (GS3) when the escape box was removed. The rat pan-tissue clock was applied to DNAm data from hippocampal tissue. An enrichment pathway analysis revealed that neuron fate commitment, brain development, and central nervous system development were processes whose underlying genes were enriched in hypermethylated CpGs in the old rats. In the old rat hippocampi, the methylation levels of CpG proximal to transcription factors associated with genes Pax5, Lbx1, Nr2f2, Hnf1b, Zic1, Zic4, Hoxd9; Hoxd10, Gli3, Gsx1 and Lmx1b, and Nipbl showed a significant regression with spatial memory performance. Regression analysis of different memory performance indices with hippocampal DNAm age was significant. These results suggest that age-related hypermethylation of transcription factors related to certain gene families, such as Zic and Gli, may play a causal role in the decline in spatial memory in old rats. Hippocampal DNAm age seems to be a reliable index of spatial memory performance in young and old rats.


Subject(s)
DNA Methylation , Spatial Memory , Animals , Rats , Aging/genetics , Cell Cycle Proteins/genetics , Epigenesis, Genetic , Hippocampus , Maze Learning/physiology , Spatial Memory/physiology , Transcription Factors/genetics
8.
Neuropeptides ; 83: 102072, 2020 Oct.
Article in English | MEDLINE | ID: mdl-32690313

ABSTRACT

Spatial memory performance declines in both normal aging and Alzheimer's disease. This cognitive deficit is related to hippocampus dysfunction. Gene therapy using neurotrophic factors like Glial cell line-derived neurotrophic factor (GDNF) emerges as a promising approach to ameliorate age-related cognitive deficits. We constructed a two vector regulatable system (2VRS) which consists of a recombinant adenoviral vector (RAd) harboring a Tet-Off bidirectional promoter flanked by GDNF and Green Fluorescent Protein (GFP) genes. A second adenovector, RAd-tTA, constitutively expresses the regulatory protein tTA. When cells are cotransduced by the 2VRS, tTA activates the bidirectional promoter and both transgenes are expressed. In the presence of the antibiotic doxycycline (DOX) transgene expression is silenced. We tested the 2VRS in CHO-K1 cells where we observed a dose-dependent GFP expression that was completely inhibited by DOX (1 mg/ml). The 2VRS injected in the hippocampal CA1 region transduced both neurons and astrocytes and was efficiently inhibited by DOX added to the drinking water. In order to assess GDNF biological activity we injected 2VRS and its Control (CTRL) vector in the hypothalamus and monitored body weight for one month. The results showed that GDNF retards weight recovery 6 days more than CTRL. In conclusion, our 2VRS demonstrated optimal GFP expression and showed a bioactive effect of transgenic GDNF in the brain.


Subject(s)
Glial Cell Line-Derived Neurotrophic Factor/administration & dosage , Green Fluorescent Proteins/administration & dosage , Hippocampus/drug effects , Neurons/drug effects , Adenoviridae , Animals , CHO Cells , Cricetinae , Cricetulus , Genetic Vectors , Glial Cell Line-Derived Neurotrophic Factor/genetics , Glial Cell Line-Derived Neurotrophic Factor/metabolism , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Hippocampus/metabolism , Neurons/metabolism , Rats
9.
Rejuvenation Res ; 23(6): 516-525, 2020 Dec.
Article in English | MEDLINE | ID: mdl-32340558

ABSTRACT

Several countries have established self-help cryonics groups whose mission is to cryopreserve human bodies or brains after legal death and ship them to cryonics organizations. The objective of this study was to report the first case of human brain cryopreservation in Argentina and complementary experiments in rats. After legal death, the body of a 78-year-old Caucasian woman was transported to a funeral home where her head was submitted to intracarotid perfusion with 5 L cold physiologic saline followed by the same volume of cold saline containing 13% dimethyl sulfoxide and 13% glycerol. The brain was removed, temporarily frozen at -80°C, and shipped to a U.S. cryostasis facility. Three groups of rats were intracardially perfused with fixative but not frozen (Reference group), vitrification solution VM1 (Control group), or the cryoprotection solution used in the patient (Experimental group). Control and Experimental brains were stored at -80°C and subsequently assessed by immunohistochemistry for the adult neuron marker (NeuN), the immature neuron marker doublecortin (DCX), the dopaminergic neuron marker tyrosine hydroxylase, and the presynaptic marker synaptophysin (SYN). The number of NeuN-positive neurons remained unchanged in the experimental brain cortex, whereas the number of immature DCX neurons in the hippocampus fell markedly in the cryoprotected brains. The results were highly variable for hypothalamic dopaminergic neurons. Confocal microscopy for SYN revealed that cryopreservation did not affect the synaptic network in the hippocampus. To our knowledge, this is the first report correlating a human cryoprotection procedure with results in complementary experiments in laboratory animals.


Subject(s)
Brain , Cryopreservation , Models, Animal , Aged , Animals , Brain/anatomy & histology , Cadaver , Cryopreservation/methods , Cryoprotective Agents , Doublecortin Protein , Female , Freezing , Hippocampus , Humans , Rats , Tissue Fixation , Vitrification
10.
Mol Neurobiol ; 57(2): 600-615, 2020 Feb.
Article in English | MEDLINE | ID: mdl-31399955

ABSTRACT

Sporadic Alzheimer's disease (sAD) is the most prevalent neurodegenerative pathology with no effective therapy until date. This disease promotes hippocampal degeneration, which in turn affects multiple cognitive domains and daily life activities. In this study, we hypothesized that long-lasting therapy with mesenchymal stem cells (MSC) would have a restorative effect on the behavioral alterations and cognitive decline typical of sAD, as they have shown neurogenic and immunomodulatory activities. To test this, we chronically injected intravenous human MSC in a sAD rat model induced by the intracerebroventricular injection of streptozotocin (STZ). During the last 2 weeks, we performed open field, Barnes maze, and marble burying tests. STZ-treated rats displayed a poor performance in all behavioral tests. Cell therapy increased exploratory behavior, decreased anxiety, and improved spatial memory and marble burying behavior, the latter being representative of daily life activities. On the hippocampus, we found that STZ promotes neuronal loss in the Cornus Ammoni (CA1) field and decreased neurogenesis in the dentate gyrus. Also, STZ induced a reduction in hippocampal volume and presynaptic protein levels and an exacerbated microgliosis, relevant AD features. The therapy rescued CA1 neurodegeneration but did not reverse the decrease of immature neurons, suggesting that the therapy effect varied among hippocampal neuronal populations. Importantly, cell therapy ameliorated microgliosis and restored hippocampal atrophy and some presynaptic protein levels in the sAD model. These findings, by showing that intravenous injection of human MSC restores behavioral and hippocampal alterations in experimental sAD, support the potential use of MSC therapy for the treatment of neurodegenerative diseases.


Subject(s)
Behavior, Animal , Hippocampus/pathology , Mesenchymal Stem Cell Transplantation , Mesenchymal Stem Cells/cytology , Animals , Anxiety/complications , Anxiety/pathology , Anxiety/physiopathology , Exploratory Behavior , Glial Fibrillary Acidic Protein/metabolism , Gliosis/complications , Gliosis/pathology , Male , Maze Learning , Memory , Nerve Tissue Proteins/metabolism , Neurogenesis , Neurons/pathology , Organ Size , Rats, Sprague-Dawley , Spatial Learning , Streptozocin , Synapses/metabolism
11.
Stem Cell Rev Rep ; 15(4): 612-617, 2019 08.
Article in English | MEDLINE | ID: mdl-31119513

ABSTRACT

There is a growing interest in the potential of adult stem cells for implementing regenerative medicine in the brain. We assessed the effect of intracerebroventricular (icv) administration of human umbilical cord perivascular cells (HUCPVCs) on spatial memory of senile (27 mo) female rats, using intact senile counterparts as controls. Approximately one third of the animals were injected in the lateral ventricles with a suspension containing 4.8 X 105 HUCPVC in 8 µl per side. The other third received 4.8 X 105 transgenic HUCPVC overexpressing Insulin-like growth factor-1 (IGF-1) and the last third of the rats received no treatment. Spatial memory performance was evaluated using a modified version of the Barnes maze test. In order to evaluate learning ability as well as spatial memory retention, we assessed the time spent (permanence) by animals in goal sector 1 (GS1) and 3 (GS3) when the escape box was removed. Fluorescence microscopy revealed the prescence of Dil-labeled HUCPVC in coronal sections of treated brains. The HUCPVC were located in close contact with the ependymal cells with only a few labeled cells migrating into the brain parenchyma. After treatment with naïve or IGF-1 transgenic HUCPVC, permanence in GS1 and GS3 increased significantly whereas there were no changes in the intact animals. We conclude that HUCPVC injected icv are effective to improve some components of spatial memory in senile rats. The ready accessibility of HUCPVC constitutes a significant incentive to continue the exploration of their therapeutic potential on neurodegenerative diseases.


Subject(s)
Aging , Brain/physiopathology , Cell Transplantation , Memory Disorders/therapy , Spatial Memory , Umbilical Cord , Animals , Female , Humans , Memory Disorders/pathology , Memory Disorders/physiopathology , Rats , Rats, Sprague-Dawley
12.
Behav Brain Res ; 374: 111887, 2019 11 18.
Article in English | MEDLINE | ID: mdl-30951751

ABSTRACT

There is a growing interest in the potential of mesenchymal stem cells (MSCs) for implementing regenerative medicine in the brain as they have shown neurogenic and immunomodulatory activities. We assessed the effect of intracerebroventricular (icv) administration of human bone marrow-derived MSCs (hBM-MSCs) on spatial memory and hippocampal morphology of senile (27 months) female rats, using 3-months-old counterparts as young controls. Half of the animals were injected in the lateral ventricles (LV) with a suspension containing 5 × 105hBM-MSCs in 8 µl per side. The other half received no treatment (senile controls). Spatial memory performance was assessed with a modified version of the Barnes maze test. We employed one probe trial, one day after training in order to evaluate learning ability as well as spatial memory retention. Neuroblast (DCX) and microglial (Iba-1 immunoreactive) markers were also immunohistochemically quantitated in the animals by means of an unbiased stereological approach. In addition, hippocampal presynaptic protein expression was assessed by immunoblotting analysis. After treatment, the senile MSC-treated group showed a significant improvement in spatial memory accuracy and extended permanence in a one- and 3-hole goal sectors as compared with senile controls. The MSC treatment increased the number of neuroblasts in the hippocampal dentate gyrus, reduced the number of reactive microglial cells, and restored presynaptic protein levels as compared to senile controls. We conclude that icv injected hBM-MSCs are effective in improving spatial memory in senile rats and that the strategy improves some functional and morphologic brain features typically altered in aging rats.


Subject(s)
Aging/drug effects , Mesenchymal Stem Cell Transplantation/methods , Spatial Memory/drug effects , Aging/metabolism , Animals , Dentate Gyrus/drug effects , Dentate Gyrus/metabolism , Doublecortin Protein , Female , Hippocampus/drug effects , Hippocampus/metabolism , Humans , Mesenchymal Stem Cells/metabolism , Microglia/drug effects , Neural Stem Cells/drug effects , Neurogenesis/drug effects , Neurons/drug effects , Rats , Rats, Sprague-Dawley , Spatial Memory/physiology , Temporal Lobe/drug effects , Temporal Lobe/metabolism
13.
Mol Cancer ; 7: 13, 2008 Jan 25.
Article in English | MEDLINE | ID: mdl-18218140

ABSTRACT

BACKGROUND: The implementation of gene therapy for the treatment of pituitary tumors emerges as a promising complement to surgery and may have distinct advantages over radiotherapy for this type of tumors. Up to now, suicide gene therapy has been the main experimental approach explored to treat experimental pituitary tumors. In the present study we assessed the effectiveness of insulin-like growth factor I (IGF-I) gene therapy for the treatment of estrogen-induced prolactinomas in rats. RESULTS: Female Sprague Dawley rats were subcutaneously implanted with silastic capsules filled with 17-beta estradiol (E2) in order to induce pituitary prolactinomas. Blood samples were taken at regular intervals in order to measure serum prolactin (PRL). As expected, serum PRL increased progressively and 23 days after implanting the E2 capsules (Experimental day 0), circulating PRL had undergone a 3-4 fold increase. On Experimental day 0 part of the E2-implanted animals received a bilateral intrapituitary injection of either an adenoviral vector expressing the gene for rat IGF-I (RAd-IGFI), or a vector (RAd-GFP) expressing the gene for green fluorescent protein (GFP). Seven days post vector injection all animals were sacrificed and their pituitaries morphometrically analyzed to evaluate changes in the lactotroph population. RAd-IGFI but not RAd-GFP, induced a significant fall in serum PRL. Furthermore, RAd-IGFI but not RAd-GFP significantly reversed the increase in lactotroph size (CS) and volume density (VD) induced by E2 treatment. CONCLUSION: We conclude that IGF-I gene therapy constitutes a potentially useful intervention for the treatment of prolactinomas and that bioactive peptide gene delivery may open novel therapeutic avenues for the treatment of pituitary tumors.


Subject(s)
Genetic Therapy , Hyperprolactinemia/pathology , Hyperprolactinemia/therapy , Insulin-Like Growth Factor I/genetics , Insulin-Like Growth Factor I/therapeutic use , Prolactinoma/pathology , Prolactinoma/therapy , Animals , Cell Size , Female , Green Fluorescent Proteins/metabolism , Hyperprolactinemia/genetics , Lactotrophs/pathology , Prolactin/blood , Prolactinoma/genetics , Rats , Rats, Sprague-Dawley , Thymidine Kinase/metabolism , Transgenes
14.
Curr Gene Ther ; 8(1): 49-53, 2008 Feb.
Article in English | MEDLINE | ID: mdl-18336249

ABSTRACT

The aim of the present article is to discuss the potential of gene therapy for thymic hormones as a novel therapeutic strategy to treat dyshomeostatic conditions associated with congenital athymia or hypofunction of the endocrine thymus. Recent studies using an adenoviral vector harboring a synthetic gene for the thymic peptide thymulin are reviewed. This adenoviral vector was injected intramuscularly in thymectomized and nude mice as well as in thymectomized rats. Transduced myocytes acted as an ectopic source of thymulin thus restoring circulating thymulin levels to normal values. This restorative effect was long lasting (several months) even though an adenoviral vector was used. In the rat brain, adenovirally-mediated delivery of the synthetic gene for thymulin achieved longer expression than in the case of adenovirally-delivered reporter genes, which is consistent with the reported antiinflammatory activity of thymulin in the brain. Furthermore, neonatal thymulin gene therapy in nude female mice was able to prevent the pituitary and ovarian alterations that typically occur in this mutant after puberty. Neonatal thymulin gene therapy in nude mice was able to prevent some of the alterations in lipid metabolism that develop during adult life in congenitally athymic mice. We conclude that the availability of the above biotechnological tools should boost basic studies on the molecular biology of thymulin and should also allow an assessment of the potential of gene therapy to restore circulating thymulin levels in thymodeficient animal models and eventually, in humans.


Subject(s)
Aging/physiology , Genetic Therapy/methods , Immunity, Innate/genetics , Thymic Factor, Circulating/genetics , Thymus Gland/abnormalities , Animals , Base Sequence , Models, Animal , Molecular Sequence Data , Peptide Fragments/genetics , Sequence Homology, Nucleic Acid , Thymus Gland/physiology
15.
J Gerontol A Biol Sci Med Sci ; 73(4): 459-467, 2018 03 14.
Article in English | MEDLINE | ID: mdl-28645186

ABSTRACT

In rats, learning and memory performance decline during normal aging, which makes this rodent species a suitable model to evaluate therapeutic strategies. In aging rats, insulin-like growth factor-I (IGF-I), is known to significantly improve spatial memory accuracy as compared to control counterparts. A constellation of gene expression changes underlie the hippocampal phenotype of aging but no studies on the effects of IGF-I on the hippocampal transcriptome of old rodents have been documented. Here, we assessed the effects of IGF-I gene therapy on spatial memory performance in old female rats and compared them with changes in the hippocampal transcriptome. In the Barnes maze test, experimental rats showed a significantly higher exploratory frequency of the goal hole than controls. Hippocampal RNA-sequencing showed that 219 genes are differentially expressed in 28-month-old rats intracerebroventricularly injected with an adenovector expressing rat IGF-I as compared with placebo adenovector-injected counterparts. From the differentially expressed genes, 81 were down and 138 upregulated. From those genes, a list of functionally relevant genes, concerning hippocampal IGF-I expression, synaptic plasticity as well as neuronal function was identified. Our results provide an initial glimpse at the molecular mechanisms underlying the neuroprotective actions of IGF-I in the aging brain.


Subject(s)
Aging/genetics , Genetic Therapy/methods , Hippocampus/metabolism , Insulin-Like Growth Factor I/genetics , Insulin-Like Growth Factor I/pharmacology , Memory Disorders/genetics , Spatial Memory/physiology , Age Factors , Animals , Female , Maze Learning/physiology , Neuronal Plasticity/physiology , Neurons/metabolism , Radioimmunoassay , Rats , Rats, Sprague-Dawley , Real-Time Polymerase Chain Reaction , Transcriptome
16.
Rejuvenation Res ; 21(2): 102-108, 2018 Apr.
Article in English | MEDLINE | ID: mdl-28673122

ABSTRACT

The aging female rat constitutes an interesting model of spontaneous and progressive age-related dopaminergic dysfunction as it allows assessing new therapeutic strategies for Parkinson's disease. Insulin-like growth factor I (IGF-I) is emerging as a powerful neuroprotective molecule, strongly induced in the central nervous system after different insults. We constructed a helper-dependent recombinant adenoviral vector (HDRAd-IGFI) harboring the gene for rat IGF-I. This was used to implement long-term IGF-I gene therapy in the hypothalamus of aged female rats, which display hypothalamic dopaminergic (DA) dysfunction and, as a consequence, chronic hyperprolactinemia. Rejuvenating long-term IGF-I gene therapy was implemented in young (3 months) and aged (24 months) female rats, which received a single intrahypothalamic injection of 4 × 109 viral particles of either HD-RAd-IGFI or HD-RAd-DsRed (control vector) and were sacrificed 119 days postinjection. In the young animals, neither vector modified serum prolactin (PRL) levels, but in the RAd-IGFI-injected aged rats a nearly full reversion of their hyperprolactinemic status was recorded. Morphometric analysis revealed a significant increase in the total number of tyrosine hydroxylase (TH)-positive cells in the hypothalamus of experimental compared with control aged animals (5874 ± 486 and 3390 ± 498, respectively). Our results indicate that IGF-I gene therapy in aged female rats is highly effective in rejuvenating the hypothalamic DA neuron groups.


Subject(s)
Dopamine/metabolism , Genetic Therapy/methods , Genetic Vectors/administration & dosage , Hyperprolactinemia/therapy , Insulin-Like Growth Factor I/genetics , Rejuvenation , Adenoviridae/genetics , Animals , Female , Hyperprolactinemia/genetics , Hyperprolactinemia/pathology , Hypothalamus/cytology , Hypothalamus/metabolism , Rats , Rats, Sprague-Dawley
17.
Neurogenesis (Austin) ; 4(1): e1259709, 2017.
Article in English | MEDLINE | ID: mdl-28405590

ABSTRACT

In rats, learning and memory performance decline during normal aging, which is paralleled by a severe reduction of the levels of neurogenesis in the hippocampal dentate gyrus (DG). A promising therapeutic strategy to restore neurogenesis in the hippocampus of old rats and their spatial memory involves the use of insulin-like growth factor-I (IGF-I). The peptide exerts pleiotropic effects in the brain, regulating multiple cellular processes. Thus, 4-week intracerebroventricular (ICV) perfusion of IGF-I significantly restored spatial memory and hippocampal neurogenesis in old male rats. Similar results were achieved by ICV IGF-I gene therapy in aging female rats. Thus, the treatment seemed to increase the number of immature neurons in the DG of 28 mo old rats, which was paralleled by an increase in the accuracy of the animals to remember specific patterns, which is known as pattern separation memory. The DG is thought to be the main hippocampal structure involved in pattern separation memory and there is evidence that the level of neurogenesis in the DG is directly related to pattern separation performance in rodents. Summing up, IGF-I emerges as a promising restorative molecule for increasing hippocampal neurogenesis and memory accuracy in aged individuals and possibly, in neurodegenerative pathologies.

18.
Curr Gene Ther ; 6(1): 125-9, 2006 Feb.
Article in English | MEDLINE | ID: mdl-16475950

ABSTRACT

The implementation of experimental gene therapy in animal models of neurological diseases is an area of growing interest. Although the neuroendocrine system offers unique advantages for the assessment of in vivo gene therapy, little work has been done in this model. Here we review the core of documented studies in which in vivo gene therapy has been implemented in the neuroendocrine system of rodent models. In the hypothalamus, restorative gene therapy has been successfully implemented in Brattleboro rats, an arginine vasopressin (AVP) mutant which suffers from diabetes insipidus, in Koletsky (fa(k)/fa(k)) and in Zucker (fa/fa) rats which have leptin receptor mutations that render them obese, hyperphagic and hyperinsulinemic. In the above models, viral vectors expressing AVP, leptin receptor b and proopiomelanocortin, respectively were stereotaxically injected in the relevant hypothalamic regions. In rats, aging brings about a progressive degeneration and loss of hypothalamic tuberoinfundibular dopaminergic neurons, which are involved in the tonic inhibitory control of prolactin secretion and lactotrophic cell proliferation. Stereotaxic injection of an adenoviral vector expressing Insulin-like Growth Factor-I (IGF-I) was able to correct their chronic hyperprolactinemia and restore tuberoinfundibular dopaminergic (TIDA) neuron numbers. In young and old F-344 male rats, Glial Cell Line-derived Neurotrophic Factor (GDNF) gene delivery in the hypothalamus induced body weight loss. These results suggest that further implementation of gene therapy strategies in neuroendocrine models may be highly rewarding.


Subject(s)
Genetic Therapy , Neurosecretory Systems , Animals , Disease Models, Animal , Female , Genetic Therapy/methods , Glial Cell Line-Derived Neurotrophic Factor/administration & dosage , Glial Cell Line-Derived Neurotrophic Factor/genetics , Humans , Male , Mice , Rats , Receptors, Cell Surface/administration & dosage , Receptors, Cell Surface/genetics , Receptors, Leptin
19.
Brain Res Bull ; 69(6): 647-51, 2006 May 31.
Article in English | MEDLINE | ID: mdl-16716832

ABSTRACT

Thymulin is a thymic peptide with antiinflammatory activity in the brain. We constructed a recombinant adenoviral vector, RAd-FTS, expressing a synthetic DNA sequence encoding met-FTS, a biologically active analog of thymulin and used it for peripheral and central gene transfer in rats. Thymulin concentration in serum and brain tissue was determined by bioassay. Reporter gene expression in the substantia nigra (SN) was quantitated by enzymohistochemistry or fluorescence microscopy using an appropriate image analysis software. A single intramuscular injection (10(8) plaque forming units (pfu)/animal) of RAd-FTS in thymectomized rats (nondetectable serum thymulin) induced supraphysiologic serum thymulin levels for at least 110 days (123+/-22 fg/ml versus 598+/-144 fg/ml in intact and vector-injected rats, respectively). Stereotaxic intranigral injection of RAd-FTS induced steady expression levels of met-FTS for at least 90 days, whereas expression of adenovirally transferred reporter genes coding for green fluorescent protein fused to HSV thymidine kinase (GFP-TK)(fus) or E.coli beta-galactosidase (beta-gal), declined drastically within a month (% transgene expression in the SN on post-injection day 30 relative to day 2 was: 18, <1 and 125%, for beta-gal, (GFP-TK)(fus) and met-FTS, respectively). We conclude that RAd-FTS constitutes a suitable biotechnological tool for the assessment of peripheral and central thymulin gene therapy in animal models of nigral dopaminergic neurodegeneration induced by pro-inflammatory agents.


Subject(s)
Gene Transfer Techniques , Substantia Nigra/metabolism , Thymic Factor, Circulating/genetics , Adenoviridae/genetics , Animals , Diagnostic Imaging/methods , Female , Gene Expression Regulation/physiology , Genes, Reporter/physiology , Genes, Synthetic/physiology , Genetic Vectors/physiology , Immunohistochemistry/methods , Male , Rats , Rats, Sprague-Dawley , Thymectomy/methods , Thymic Factor, Circulating/metabolism , Time Factors
20.
Curr Gene Ther ; 14(1): 24-34, 2014 Feb.
Article in English | MEDLINE | ID: mdl-24450294

ABSTRACT

In the central nervous system, cholinergic and dopaminergic (DA) neurons are among the cells most susceptible to the deleterious effects of age. Thus, the basal forebrain cholinergic system is known to undergo moderate neurodegenerative changes during normal aging as well as severe atrophy in Alzheimer's disease (AD). Parkinson's disease (PD), a degeneration of nigro-striatal DA neurons is the most conspicuous reflection of the vulnerability of DA neurons to age. Overall, there is growing evidence that a progressive decline in cognitive function and central DA activity represents basic features of normal aging both in humans and laboratory rodents. Spontaneous or environmental neurotoxin-mediated exacerbation of these processes contributes to the symptoms of AD and PD, respectively. In this context, neurotrophic factors that can prevent or delay the decline in cognitive function and central DA activity are of clinical interest. Among them, Insulin-like Growth Factor I and Glial cell line-Derived Neurotrophic Factor are emerging as powerful neuroprotective molecules. This article discusses the experimental evidence supporting the neuroprotective relevance of these and related factors in the aging brain. The availability of induced pluripotent stem cells offers a new promise for the treatment of pathologies associated with the loss of specific cell types as for instance, nigral DA neurons (in PD) or basal forebrain cholinergic neurons (BFCN) in the early stages of AD. Recent studies documenting the use of cell reprogramming for the generation of multipotent neuronal precursors as well as functional BFCN and DA neurons are reviewed.


Subject(s)
Alzheimer Disease/genetics , Genetic Therapy , Glial Cell Line-Derived Neurotrophic Factor/genetics , Parkinson Disease/genetics , Alzheimer Disease/pathology , Alzheimer Disease/therapy , Brain/metabolism , Brain/pathology , Cellular Reprogramming , Cholinergic Neurons/metabolism , Cholinergic Neurons/pathology , Dopaminergic Neurons/metabolism , Dopaminergic Neurons/pathology , Glial Cell Line-Derived Neurotrophic Factor/metabolism , Humans , Insulin-Like Growth Factor I/genetics , Insulin-Like Growth Factor I/metabolism , Parkinson Disease/pathology , Parkinson Disease/therapy
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