Núria Cabezas-Llobet1, Sandra Camprubí2, Beatriz García2, Jordi Alberch3, Xavier Xifró4. 1. New Therapeutic Targets Group (TargetsLab), Departament de Ciències Mèdiques, Facultat de Medicina, Universitat de Girona, E-17071 Girona, Spain. 2. Grifols Bioscience Research Group, Barcelona, Spain. 3. Departament de Biomedicina, Institut de Neurociències, Facultat de Medicina, Universitat de Barcelona, E-08036 Barcelona, Spain; Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), E-08036 Barcelona, Spain; Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Spain. 4. New Therapeutic Targets Group (TargetsLab), Departament de Ciències Mèdiques, Facultat de Medicina, Universitat de Girona, E-17071 Girona, Spain; Departament de Biomedicina, Institut de Neurociències, Facultat de Medicina, Universitat de Barcelona, E-08036 Barcelona, Spain; Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), E-08036 Barcelona, Spain; Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Spain. Electronic address: xavier.xifro@udg.edu.
Abstract
BACKGROUND: Death due to cerebral stroke afflicts a large number of neuronal populations, including glial cells depending on the brain region affected. Drugs with a wide cellular range of protection are needed to develop effective therapies for stroke. Human alpha 1-antitrypsin (hAAT) is a serine proteinase inhibitor with potent anti-inflammatory, anti-apoptotic and immunoregulatory activities. This study aimed to test whether hAAT can protect different kind of neurons and glial cells after the oxygen and glucose deprivation (OGD). METHODS: Addition of hAAT to mouse neuronal cortical, hippocampal and striatal cultures, as well as glial cultures, was performed 30 min after OGD induction and cell viability was assessed 24 h later. The expression of different apoptotic markers and several inflammatory parameters were assessed by immunoblotting and RT-PCR. RESULTS: hAAT had a concentration-dependent survival effect in all neuronal cultures exposed to OGD, with a maximal effect at 1-2 mg/mL. The addition of hAAT at 1 mg/mL reduced the OGD-mediated necrotic and apoptotic death in all neuronal cultures. This neuroprotective activity of hAAT was associated with a decrease of cleaved caspase-3 and an increase of MAP2 levels. It was also associated with a reduction of pro-inflammatory cytokines protein levels and expression, increase of IL-10 protein levels and decrease of nuclear localization of nuclear factor-kappaB. Similar to neurons, addition of hAAT protected astrocytes and oligodendrocytes against OGD-induced cell death. CONCLUSIONS: Human AAT protects neuronal and glial cells against OGD through interaction with cytokines. GENERAL SIGNIFICANCE: Human AAT could be a good therapeutic neuroprotective candidate to treat ischemic stroke.
BACKGROUND: Death due to cerebral stroke afflicts a large number of neuronal populations, including glial cells depending on the brain region affected. Drugs with a wide cellular range of protection are needed to develop effective therapies for stroke. Humanalpha 1-antitrypsin (hAAT) is a serine proteinase inhibitor with potent anti-inflammatory, anti-apoptotic and immunoregulatory activities. This study aimed to test whether hAAT can protect different kind of neurons and glial cells after the oxygen and glucose deprivation (OGD). METHODS: Addition of hAAT to mouse neuronal cortical, hippocampal and striatal cultures, as well as glial cultures, was performed 30 min after OGD induction and cell viability was assessed 24 h later. The expression of different apoptotic markers and several inflammatory parameters were assessed by immunoblotting and RT-PCR. RESULTS:hAAT had a concentration-dependent survival effect in all neuronal cultures exposed to OGD, with a maximal effect at 1-2 mg/mL. The addition of hAAT at 1 mg/mL reduced the OGD-mediated necrotic and apoptotic death in all neuronal cultures. This neuroprotective activity of hAAT was associated with a decrease of cleaved caspase-3 and an increase of MAP2 levels. It was also associated with a reduction of pro-inflammatory cytokines protein levels and expression, increase of IL-10 protein levels and decrease of nuclear localization of nuclear factor-kappaB. Similar to neurons, addition of hAAT protected astrocytes and oligodendrocytes against OGD-induced cell death. CONCLUSIONS:HumanAAT protects neuronal and glial cells against OGD through interaction with cytokines. GENERAL SIGNIFICANCE: HumanAAT could be a good therapeutic neuroprotective candidate to treat ischemic stroke.