RESUMEN
Opioid use in HIV infection has been associated with an increased frequency of neurological disease and cognitive impairment and vitamin A deficiency has been linked to progressive HIV disease in drug users. In this report the potential effects of these factors, alone and in combination, on gamma amino butyric acid (GABA)-expression interneurons in hippocampus in the HIV-1 transgenic rat (TG) model were studied. TG and wild-type (WT) F344 Fisher rats deficient in vitamin A from birth were implanted either with a 37.5 mg morphine tablet or with a matching placebo and total numbers of neurons and of parvalbumin+ neurons were quantitated and parvalbumin expression was quantitated in the CA1 hippocampal region of the rats. These studies showed that total neuronal numbers were decreased in the TG versus WT Fisher rats and that this decrease was enhanced by the vitamin A deficient diet and by treatment with morphine. In contrast, there was no significant change noted in numbers of parvalbumin+ neurons. However, levels of parvalbumin expression were decreased for vitamin A deficient and morphine-treated WT rats as compared to WT rats on the normal diet and placebo-treated WT rats. For TG rats, parvalbumin expression was higher for vitamin A deficient TG rats treated with either placebo or morphine than for WT vitamin A deficient rats treated with placebo, and placebo treated vitamin A deficient TG rats showed higher expression than morphine treated vitamin A deficient rats. Expression was also higher for vitamin A deficient morphine-treated rats than for the corresponding WT rat groups and for vitamin A deficient TG rats treated with placebo. For the remaining groups, parvalbumin was similar for the TG and WT rats. These findings suggest that in hippocampus vitamin A deficiency and morphine can increase parvalbumin expression, perhaps as a manifestation of a stress response. Parvalbumin-expressing GABA-ergic interneurons regulate the primary neuronal output from hippocampus that is important for memory and behavior. Therefore, these studies suggest that vitamin A deficiency and morphine might have effects that may impact such outputs and thereby have lasting effects on cognitive status.
Asunto(s)
Región CA1 Hipocampal/patología , VIH-1/metabolismo , Interneuronas/metabolismo , Morfina/farmacología , Narcóticos/farmacología , Parvalbúminas/biosíntesis , Deficiencia de Vitamina A/patología , Animales , Región CA1 Hipocampal/efectos de los fármacos , Femenino , Regulación Viral de la Expresión Génica/efectos de los fármacos , VIH-1/efectos de los fármacos , Inmunohistoquímica , Interneuronas/efectos de los fármacos , Masculino , Embarazo , Ratas , Ratas Transgénicas , Receptores Opioides mu/biosíntesis , Proteínas Virales/biosíntesisRESUMEN
Vitamin A (VA) deficiency in human immunodeficiency virus (HIV) infection has been associated with more progressive HIV disease, which may be enhanced by opioid use. In these studies, we examined the effects of VA deficiency and morphine on frontal cortex neuronal numbers in the HIV-1 transgenic (Tg) rat. These studies showed that total numbers of neurons were similar for rats on the VA-deficient diet as for rats on the normal diet and these numbers were not affected by treatment with morphine. In contrast, numbers of neurons that expressed the calcium-binding protein parvalbumin, which is a marker interneurons that express the inhibitory neurotransmitter gamma-aminobutyric acid (GABAergic neurons) were decreased for wild-type (Wt) rats on the VA-deficient diet and for Wt rats treated with morphine. In addition, parvalbumin+ neurons were also decreased for Tg rats on a normal diet but increased to normal levels when these animals were placed on the VA-deficient diet and treated with morphine. Analysis of expression of the genes that code for the HIV regulatory proteins vif, vpr, nef, and tat in frontal cortex and adjacent subcortical white matter showed that tat expression was increased in the morphine-treated Tg rat on the VA-deficient diet as compared to untreated Tg rats on the normal diet and untreated VA-deficient rats. These studies therefore suggest that VA deficiency, opioid exposure, and HIV infection alone and in combination may potentially alter neuronal metabolic activity and induce cellular stress, resulting in the observed changes in levels of parvalbumin expression. The specific mechanisms that underlie these effects require further study.
Asunto(s)
Analgésicos Opioides/efectos adversos , Regulación Viral de la Expresión Génica , Infecciones por VIH/patología , Infecciones por VIH/virología , VIH-1 , Morfina/efectos adversos , Neuronas/efectos de los fármacos , Neuronas/patología , Parvalbúminas/biosíntesis , Deficiencia de Vitamina A/patología , Animales , Recuento de Células , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Corteza Cerebral/virología , Regulación Viral de la Expresión Génica/efectos de los fármacos , Regulación Viral de la Expresión Génica/fisiología , Infecciones por VIH/complicaciones , Infecciones por VIH/metabolismo , VIH-1/genética , VIH-1/metabolismo , Humanos , Neuronas/metabolismo , Neuronas/virología , Ratas , Deficiencia de Vitamina A/etiología , Deficiencia de Vitamina A/metabolismo , Productos del Gen nef del Virus de la Inmunodeficiencia Humana/biosíntesis , Productos del Gen nef del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen rev del Virus de la Inmunodeficiencia Humana/biosíntesis , Productos del Gen rev del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/biosíntesis , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen vif del Virus de la Inmunodeficiencia Humana/biosíntesis , Productos del Gen vif del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen vpr del Virus de la Inmunodeficiencia Humana/biosíntesis , Productos del Gen vpr del Virus de la Inmunodeficiencia Humana/genéticaRESUMEN
Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, is neuroprotective in animal models of neurodegenerative diseases. However, BDNF has a short half-life and its efficacy in the central nervous system (CNS), when delivered peripherally, is limited due to the blood-brain barrier (BBB). We have developed a means of delivering BDNF into the CNS using genetically engineered bone marrow stem cells (BMSCs) as a vehicle, and have explored the clinical effects of BDNF on outcomes in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). BDNF-engineered-BMSCs were transplanted (i.v.) into irradiated 2-week-old SJL/J female mice. Eight weeks after transplantation, mice were immunized with a peptide of proteolipid protein (PLP(139-151)). Mice, which had received BDNFengineered BMSCs, showed a significant delay in EAE onset and a reduction in overall clinical severity compared to mice receiving BMSC transfected with an empty vector lacking the BDNF gene. In addition, pathological examination showed that BDNF delivery reduced demyelination and increased remyelination. Inhibition of pro-inflammatory cytokines TNF-alpha and IFN-gamma and enhanced expression of the antiinflammatory cytokines IL-4, IL-10, and IL-11 were found in the CNS tissues of the BDNF transplanted group. These results support the use of BMSCs as vehicles to deliver BDNF into the CNS of EAE animals. This is a potentially novel therapeutic approach that might be used to deliver BDNF gene or genes for other therapeutic proteins into the CNS in MS or in other diseases of the CNS in which accessibility of therapeutic proteins is limited due to the BBB.
Asunto(s)
Trasplante de Médula Ósea/métodos , Factor Neurotrófico Derivado del Encéfalo/genética , Factor Neurotrófico Derivado del Encéfalo/uso terapéutico , Encefalomielitis Autoinmune Experimental/terapia , Técnicas de Transferencia de Gen , Terapia Genética/métodos , Animales , Células Cultivadas , Citocinas/metabolismo , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/metabolismo , Femenino , Regulación de la Expresión Génica/genética , Vectores Genéticos/farmacología , Vectores Genéticos/uso terapéutico , Ratones , Proteína Proteolipídica de la Mielina/inmunología , Vaina de Mielina/inmunología , Vaina de Mielina/metabolismo , Vaina de Mielina/patología , Fragmentos de Péptidos/inmunología , Resultado del TratamientoRESUMEN
Interferon-beta (IFN-beta), an approved treatment of multiple sclerosis (MS), produces only partial clinical responses. IFN-beta therapy has been limited by its short serum half-life and limited ability to cross the blood brain barrier. We have developed a means of delivering the IFN-beta gene both systemically and into the central nervous system (CNS) using bone marrow stem cells (BMSCs) as a vehicle and examined the therapeutic efficacy of this approach in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. A retroviral expression vector (pLXSN-IFNbeta) was used to stably transfect virus producer PA317 cells to generate retrovirus containing the IFN-beta gene which then was used to transduce BMSCs. IFN-beta engineered BMSCs were transplanted (i.v.) into mice that then were immunized with proteolipoprotein (PLP) to initiate EAE. IFN-beta-engineered BMSCs transplanted mice showed a significant inhibition of EAE onset, and the overall clinical severity was less compared to control groups. IFN-beta delivery strongly reduced infiltration of mononuclear cells possibly by inhibiting cell adhesion molecules. Reduced demyelination and increased remyelination were also observed in the IFN-beta treated group. Furthermore, inhibition of the pro-inflammatory cytokines TNF-alpha, IFN-gamma and IL-12 and enhanced expression of the anti-inflammatory cytokines IL-10, IL-4 and TGF-beta was observed in CNS tissue. In addition, mice receiving IFN-beta had reduced apoptosis and increases in growth promoting factors including BDNF, CNTF, PDGF and VEGF. These results suggest that BMSCs can be used as vehicles to deliver the IFN-beta into the CNS. This is a potentially novel therapeutic approach which might be used in MS and other diseases of the CNS in which drug access is limited.
Asunto(s)
Células de la Médula Ósea/fisiología , Trasplante de Médula Ósea/métodos , Encefalomielitis Autoinmune Experimental/prevención & control , Interferón beta/uso terapéutico , Animales , Antígenos CD34/metabolismo , Células de la Médula Ósea/metabolismo , Caspasa 3/genética , Caspasa 3/metabolismo , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/inducido químicamente , Ensayo de Inmunoadsorción Enzimática/métodos , Femenino , Técnicas de Transferencia de Gen , Etiquetado Corte-Fin in Situ , Molécula 1 de Adhesión Intercelular/genética , Molécula 1 de Adhesión Intercelular/metabolismo , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Interferón beta/biosíntesis , Interferón beta/genética , Ratones , Proteína Básica de Mielina/genética , Proteína Básica de Mielina/metabolismo , Proteína Proteolipídica de la Mielina , Fragmentos de Péptidos , Prevención Secundaria , Molécula 1 de Adhesión Celular Vascular/genética , Molécula 1 de Adhesión Celular Vascular/metabolismoRESUMEN
Multiple sclerosis is an inflammatory disease of the central nervous system (CNS) which includes a neurodegenerative component. Brain derived neurotrophic factor (BDNF) is a neuroprotective agent which might be useful in preventing neurodegeneration but its application has been limited because the blood brain barrier restricts its access to the CNS. We have developed a novel delivery system for BDNF using transformed bone marrow stem cells (BMSC) and undertook studies of EAE to determine whether the delivery of BDNF could reduce inflammation and apoptosis. Mice receiving BDNF producing BMSC had reduced clinical impairment compared to control mice receiving BMSC that did not produce BDNF. Pathological examination of brain and spinal cord showed a reduction in inflammatory infiltrating cells in treated compared to control mice. Apoptosis was reduced in brain and spinal cord based on TUNEL and cleaved Caspase-3 staining. Consistent with the known mechanism of action of BDNF on apoptosis, Bcl-2 and Akt were increased in treated mice. Further studies suggested that these increases could be mediated by inhibition of both caspase dependent and caspase independent pathways. These results suggest that the BDNF delivered by the transformed bone marrow stem cells reduced clinical severity, inflammation and apoptosis in this model.