Combined Adipose Tissue-Derived Mesenchymal Stem Cell Therapy and Rehabilitation in Experimental Stroke

ORIGINAL RESEARCH published: 26 March 2019 doi: 10.3389/fneur.2019.00235 Combined Adipose Tissue-Derived Mesenchymal Stem Cell Therapy and Rehabilitation in Experimental Stroke Jingwei Mu 1,2† , Abdulhameed Bakreen 2† , Miia Juntunen 3,4† , Paula Korhonen 5 , Ella Oinonen 2 , Lili Cui 2 , Mikko Myllyniemi 2 , Shanshan Zhao 2 , Susanna Miettinen 3,4 and Jukka Jolkkonen 2,5,6* 1 Department of Neurology, The People’s Hospital of China Medical University, Shenyang, China, 2 Department of Neurology, University of Eastern Finland, Kuopio, Finland, 3 Faculty of Medicine and Health Technology, Tampere University, Tampere, Finland, 4 Research, Development and Innovation Centre, Tampere University Hospital, Tampere, Finland, 5 A. I. Virtanen Institute for Molecular Sciences, University of Eastern Finland, Kuopio, Finland, 6 Neurocenter, Kuopio University Hospital, Kuopio, Finland Edited by: Paulo Henrique Rosado-de-Castro, Instituto D’Or de Pesquisa e Ensino (IDOR), Brazil Reviewed by: Aurel Popa-Wagner, University Hospital Essen, Germany Fivos Panetsos, Complutense University of Madrid, Spain *Correspondence: Jukka Jolkkonen † These authors have contributed equally to this work Specialty section: This article was submitted to Stroke, a section of the journal Frontiers in Neurology Received: 21 December 2018 Accepted: 22 February 2019 Published: 26 March 2019 Citation: Mu J, Bakreen A, Juntunen M, Korhonen P, Oinonen E, Cui L, Myllyniemi M, Zhao S, Miettinen S and Jolkkonen J (2019) Combined Adipose Tissue-Derived Mesenchymal Stem Cell Therapy and Rehabilitation in Experimental Stroke. Front. Neurol. 10:235. doi: 10.3389/fneur.2019.00235 Frontiers in Neurology | www.frontiersin.org Background/Objective: Stroke is a leading global cause of adult disability. As the population ages as well as suffers co-morbidities, it is expected that the stroke burden will increase further. There are no established safe and effective restorative treatments to facilitate a good functional outcome in stroke patients. Cell-based therapies, which have a wide therapeutic window, might benefit a large percentage of patients, especially if combined with different restorative strategies. In this study, we tested whether the therapeutic effect of human adipose tissue-derived mesenchymal stem cells (ADMSCs) could be further enhanced by rehabilitation in an experimental model of stroke. Methods: Focal cerebral ischemia was induced in adult male Sprague Dawley rats by permanently occluding the distal middle cerebral artery (MCAO). After the intravenous infusion of vehicle (n = 46) or ADMSCs (2 × 106 ) either at 2 (n = 37) or 7 (n = 7) days after the operation, half of the animals were housed in an enriched environment mimicking rehabilitation. Subsequently, their behavioral recovery was assessed by a neurological score, and performance in the cylinder and sticky label tests during a 42-day behavioral follow-up. At the end of the follow-up, rats were perfused for histology to assess the extent of angiogenesis (RECA-1), gliosis (GFAP), and glial scar formation. Results: No adverse effects were observed during the follow-up. Combined ADMSC therapy and rehabilitation improved forelimb use in the cylinder test in comparison to MCAO controls on post-operative days 21 and 42 (P < 0.01). In the sticky label test, ADMSCs and rehabilitation alone or together, significantly decreased the removal time as compared to MCAO controls on post-operative days 21 and 42. An early initiation of combined therapy seemed to be more effective. Infarct size, measured by MRI on post-operative days 1 and 43, did not differ between the experimental groups. Stereological counting revealed an ischemia-induced increase both in the density of blood vessels and the numbers of glial cells in the perilesional cortex, but there were no differences among MCAO groups. Glial scar volume was also similar in MCAO groups. 1 March 2019 | Volume 10 | Article 235 Mu et al. Combined Cell Therapy and Rehabilitation in Stroke Conclusion: Early delivery of ADMSCs and combined rehabilitation enhanced behavioral recovery in an experimental stroke model. The mechanisms underlying these treatment effects remain unknown. Keywords: stroke, cell therapy, rehabilitation, combination therapy, functional outcome, mechanisms, translational research INTRODUCTION skilled training (32–34), and special rehabilitative training devices (35, 36) have all been employed. In addition, housing the experimental animals in an enriched environment (EE) has also been used to provide multiple sensory, motor, social, and visual stimuli (37). Although very non-specific, housing in EE is one of the most promising approaches for improving an animal’s sensorimotor functions after an experimental stroke (38, 39). EE has also been shown to improve spatial learning and memory in ischemia-reperfusion models (40). The combination of different restorative approaches represents an intriguing approach to maximize treatment effects (41). Furthermore, cell-based therapies offer the possibility of combining different neurorestorative strategies to achieve an additive or even a synergistic therapeutic effect. However, only a few studies have been published (12, 13, 37, 42–45), and thus, more research is required in this regard to examine not only the stand-alone effects of each therapy, but also their potential combined effect (46, 47). Here, we hypothesized that the combination of an enriched environment with the IV infusion of ADMSCs after permanent middle cerebral artery occlusion (MCAO) would result in an improved behavioral recovery, perhaps even a maximal therapeutic effect. In order to explore the therapeutic window, we infused ADMSCs at either 2 or 7 d post-MCAO. Angiogenesis was evaluated as a possible repair mechanism related to treatment effect. In addition, glial cell staining was used to assess the extent of gliosis since the presence of a glial scar is considered to impede neuronal plasticity and prevent the functional recovery. Stroke is one of the leading global causes of death and long-term disability, with about 5 million survivors becoming permanently disabled annually (1–3). Despite advances in acute stroke care (4), the narrow therapeutic time windows for early thrombolysis and thrombectomy make them available to only about 10% of stroke patients (5, 6). Safe and effective treatments beyond the acute phase are urgently needed. Cell therapy represents a potential breakthrough in the treatment of stroke. In particular, mesenchymal stem cells (MSCs) are of major interest due to their advantages over other cell types, including their abundance and good availability (7), their relatively low immunogenicity (8) and tumorigenicity (9), and the lack of ethical concerns (10, 11). The non-invasive intravenous (IV) route has been most commonly used for delivery of MSCs in both preclinical and clinical studies (Cui et al. in press). More importantly, preclinical studies have (...truncated)