Preventive Triple Gene Therapy Reduces the Negative Consequences of Ischemia-Induced Brain Injury after Modelling Stroke in a Rat.

Vage Markosyan,Farid Bashirov,Rustem Islamov,Andrei Izmailov,Ilnur Salafutdinov,Maksim Kuznetsov,Dmitry Trofimov,Leniz Nurullin,Evgeny Kim,Mikhail Sokolov,Grayr Kundakchyan,Filip Fadeev,Airat Gibadullin,Zufar Safiullov

doi:10.3390/ijms21186858

Abstract

Currently, the main fundamental and clinical interest for stroke therapy is focused on developing a neuroprotective treatment of a penumbra region within the therapeutic window. The development of treatments for ischemic stroke in at-risk patients is of particular interest. Preventive gene therapy may significantly reduce the negative consequences of ischemia-induced brain injury. In the present study, we suggest the approach of preventive gene therapy for stroke. Adenoviral vectors carrying genes encoding vascular endothelial growth factor (VEGF), glial cell-derived neurotrophic factor (GDNF) and neural cell adhesion molecule (NCAM) or gene engineered umbilical cord blood mononuclear cells (UCB-MC) overexpressing recombinant VEGF, GDNF, and NCAM were intrathecally injected before distal occlusion of the middle cerebral artery in rats. Post-ischemic brain recovery was investigated 21 days after stroke modelling. Morphometric and immunofluorescent analysis revealed a reduction of infarction volume accompanied with a lower number of apoptotic cells and decreased expression of Hsp70 in the peri-infarct region in gene-treated animals. The lower immunopositive areas for astrocytes and microglial cells markers, higher number of oligodendrocytes and increased expression of synaptic proteins suggest the inhibition of astrogliosis, supporting the corresponding myelination and functional recovery of neurons in animals receiving preventive gene therapy. In this study, for the first time, we provide evidence of the beneficial effects of preventive triple gene therapy by an adenoviral- or UCB-MC-mediated intrathecal simultaneous delivery combination of vegf165, gdnf, and ncam1 on the preservation and recovery of the brain in rats with subsequent modelling of stroke.

Highlights

The current options for ischemic stroke treatment are extremely limited and are aimed at restoring blood flow in the ischemic area by intravenous infusion of recombinant tissue plasminogen activator and/or physical removal of the clots [1]
We showed the positive effect of gene-modified umbilical cord blood mononuclear cells (UCB-MC), simultaneously producing three recombinant molecules—vascular endothelial growth factor (VEGF), glial cell-derived neurotrophic factor (GDNF) and neural cell adhesion molecule (NCAM)—in animal models of amyotrophic lateral sclerosis [23], spinal cord injury [24], and stroke [25]
In the model of middle cerebral artery occlusion (MCAO) in rats, we demonstrated that intrathecal injection of genetically-engineered UCB-MC over-expressing VEGF, GDNF, and NCAM, four hours after MCAO results in a reduction of infarct volume, the positive reaction of neuroglial cells and an increase in synaptic protein expression

Summary

Introduction

The current options for ischemic stroke treatment are extremely limited and are aimed at restoring blood flow in the ischemic area by intravenous infusion of recombinant tissue plasminogen activator and/or physical removal of the clots [1]. The strategy of cell-, gene-, and gene-cell therapy for neuroprotection in stroke treatment has been proven by numerous experiments in animal models [2,3,4]. Besides brain-specific cell types, umbilical cord blood (UCB) is widely used for neuroprotection in the central nervous system (CNS) for different pathological conditions [5]. UCB cells are considered a valuable source of stem cells, growth and neurotrophic factors for cell therapy. The mononuclear fraction of UCB contains populations of different immature cells that are capable of differentiating into many cell types [6] and, represent an alternative to embryonic stem cells for transplantation to patients with post-ischemic, post-traumatic and degenerative diseases [7,8]. The following have been discovered in UCB: Hematopoietic stem cells (HSCs), endothelial progenitor cells, mesenchymal stem cells (MSCs), unrestricted somatic stem cells (USSCs), and side population cells (SP) [9,10,11,12]

Methods

Discussion

Conclusion