Effects of erythropoietin and methylprednisolone on AQP4 expression in astrocytes

Abstract

Methylprednisolone sodium succinate (MPSS) has been suggested as a treatment for spinal cord injury (SCI), but its use has been limited due to its adverse effects. Erythropoietin (EPO) has been suggested as a promising candidate for limiting SCI in mammals. The aim of the present study was to investigate the effects of EPO in combination with MPSS on astrocytes following ischemic injury in vitro. Astrocytes were isolated from the cerebral cortex of postnatal day 3 Sprague-Dawley rats and cultured in vitro. Astrocyte ischemic injury was induced by oxygen and glucose deprivation for 4 h, and reperfusion was simulated by subsequent culture under normoxic conditions. The effects of EPO and MPSS on the expression of aquaporin-4 (AQP4) were investigated. Ischemic astrocytes were treated with EPO (10 U/ml), MPSS (10 µg/ml), or EPO (10 U/ml) in combination with MPSS (10 µg/ml) during reperfusion. The cell viability of astrocytes was assessed using an MTT assay. The mRNA and protein expression levels of AQP4 were determined using reverse transcription-quantitative polymerase chain reaction and western blot analysis, respectively. The role of the protein kinase C (PKC) signaling pathway in the molecular mechanisms underlying the effects of EPO and MPSS was also investigated. The present results demonstrated that following treatment with EPO and MPSS, the mRNA expression levels of AQP4 were upregulated and cell viability was enhanced. EPO and MPSS effectively inhibited the oxygen and glucose deprivation-mediated downregulation of AQP4 following reperfusion. In addition, the combined treatment with EPO and MPSS exhibited higher AQP4 expression levels and cell viability compared with each treatment alone. Finally, the effects of EPO and MPSS on AQP4 expression were partially reversed by pretreatment with the PKC inhibitor Ro 31–8220. The present study indicated that EPO and MPSS had a synergistic effect on AQP4 expression following reperfusion, and suggest that they may be combined in the treatment of SCI.

Multivalent Display of Erythropoietin on Quantum Dots Enhances Aquaporin-4 Expression and Water Transport in Human Astrocytes In Vitro

“…Notably, at a ratio of 5 EPO/QD, we observed a 1.8-fold increase in APQN-4 expression over the level of induction mediated by free EPO at the same concentration. While it is established that EPO induces the upregulation and expression of AQPN-4 in astrocytes both in vitro 35 and in vivo, 16 our results here demonstrating the greater level of JAK/ STAT signaling and overall induction of protein expression by EPO-QD versus free EPO are, to our knowledge, the first demonstration of the augmented expression of AQPN-4 mediated by an NP-displayed form of EPO.…”

The Efficacy of Combined Medication With Methylprednisolone and Erythropoietin in the Treatment of Ischemia-Reperfusion Injury to the Spinal Cord in Patients With Cervical Spondylotic Myelopathy

“…Methylprednisolone is an effective drug in the treatment of spinal ischemia-reperfusion. However, it has been observed that high doses of methylprednisolone can cause complications such as gastrointestinal bleeding and bone necrosis [ 9 , 10 ]. Erythropoietin largely determines the oxygenation state of tissues.…”