NSC-derived exosomes enhance therapeutic effects of NSC transplantation on cerebral ischemia in mice
Curation statements for this article:-
Curated by eLife
eLife assessment
The current study employed NSCs derived from human induced pluripotent stem cells (iPSCs) together with NSC-derived exosomes extracted from NSCs to treat cerebral ischemia, and they made an important observation. Remarkably, NSC-derived exosomes could promote NSCs differentiation, reduce the oxidative stress and inflammation, and alleviate the formation of glial scars after ischemia and reperfusion, and as a result, could enhance the therapeutic effects of NSC transplantation, which is compelling. The solid experimental evidence strongly supports their major claims.
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (eLife)
- Neuroscience (eLife)
Abstract
Transplantation of neural stem cells (NSCs) has been proved to promote functional rehabilitation of brain lesions including ischemic stroke. However, the therapeutic effects of NSC transplantation are limited by the low survival and differentiation rates of NSCs due to the harsh environment in the brain after ischemic stroke. Here, we employed NSCs derived from human induced pluripotent stem cells together with exosomes extracted from NSCs to treat cerebral ischemia induced by middle cerebral artery occlusion/reperfusion in mice. The results showed that NSC-derived exosomes significantly reduced the inflammatory response, alleviated oxidative stress after NSC transplantation, and facilitated NSCs differentiation in vivo. The combination of NSCs with exosomes ameliorated the injury of brain tissue including cerebral infarction, neuronal death, and glial scarring, and promoted the recovery of motor function. To explore the underlying mechanisms, we analyzed the miRNA profiles of NSC-derived exosomes and the potential downstream genes. Our study provided the rationale for the clinical application of NSC-derived exosomes as a supportive adjuvant for NSC transplantation after stroke.
Article activity feed
-
-
Author Response
Reviewer #2 (Public Review):
Major points:
1). This study does not provide any evidence about the cell death of the transplanted cells. The immunostaining of the Caspase-3 or TUNEL staining should be used to address this issue.
We have conducted immunostaining of Caspase-3 at 7 days after transplantation using the human-specific STEM121 antibody to demonstrate the transplanted cells. We have added the results to Figure 3A and modified the text accordingly (Page 8, Line 156-165).
2). The authors showed that the neurological functions (evaluated by balance beam, ladder lung, rotarod test and Modified Neurological Severity Score (mNSS) up to 8 weeks after treatment (Figure 1C)) were significantly improved in the NES+Exo group compared to their control groups. However, these cells (transplanted cells) are progenitors …
Author Response
Reviewer #2 (Public Review):
Major points:
1). This study does not provide any evidence about the cell death of the transplanted cells. The immunostaining of the Caspase-3 or TUNEL staining should be used to address this issue.
We have conducted immunostaining of Caspase-3 at 7 days after transplantation using the human-specific STEM121 antibody to demonstrate the transplanted cells. We have added the results to Figure 3A and modified the text accordingly (Page 8, Line 156-165).
2). The authors showed that the neurological functions (evaluated by balance beam, ladder lung, rotarod test and Modified Neurological Severity Score (mNSS) up to 8 weeks after treatment (Figure 1C)) were significantly improved in the NES+Exo group compared to their control groups. However, these cells (transplanted cells) are progenitors (Nestin+) or undifferentiated cells (Tuj1+) at this stage (Figure 3). Thus, I was curious about that how can the immature neurons play neurological functions? This point should be explained.
We agree with the reviewer’s insightful comments. We have performed immunostaining using antibodies against the post-mitotic mature neuron marker RBFOX3/NeuN, post-synaptic marker PSD-95 and human-specific STEM121 at 4 weeks after transplantation. The results confirmed that NeuN+/STEM121+ and PSD-95+/STEM121+ mature neurons appeared in NSC group and increased in NSC+Exo group (Figure 3B and Figure 3 - supplement 1D). Furthermore, our additional data showed that the expression of presynaptic marker SYN1 was increased in both NSC and NSC+Exo groups at 8 weeks after treatment. Therefore, we believe that there are mature neurons and newly formed synapses involved in neurological functions.
3). The authors used the Golgi staining to show the NES+Exo can improve dendritic density and length. How do you know these neurons are transplanted cells?
Our data show that mature neurons and synapses are generated by the transplanted cells (please also see response to reviewer #2-major ponts #2). We believe that the newly generated neurons partly contribute to the improved dendritic density and length. However, we agree that the neurons with increased dendritic density and length may be both survived local neurons and those generated by the transplanted cells.
4). The cell morphology of tdTomato+ cells is fuzzy and it is difficult to distinguish the cell body. It looks like that these cells out of whack.
We have immunostaining using the human-specific STEM121 antibody to demonstrate the transplanted cells and more neuronal markers such as RBFOX3/NeuN to identify NSC differentiation (Figure 3A and 3B; Figure 3 - supplement 1C and 1D).
-
eLife assessment
The current study employed NSCs derived from human induced pluripotent stem cells (iPSCs) together with NSC-derived exosomes extracted from NSCs to treat cerebral ischemia, and they made an important observation. Remarkably, NSC-derived exosomes could promote NSCs differentiation, reduce the oxidative stress and inflammation, and alleviate the formation of glial scars after ischemia and reperfusion, and as a result, could enhance the therapeutic effects of NSC transplantation, which is compelling. The solid experimental evidence strongly supports their major claims.
-
Reviewer #1 (Public Review):
The therapeutic effects of NSC transplantation is limited by the low survival and differentiation rates of NSCs due to the harsh environment in the brain after ischemic stroke. To solve this technical challenge, the authors employed NSCs derived from human induced pluripotent stem cells (iPSCs) together with NSC-derived exosomes extracted from NSCs to treat cerebral ischemia induced by middle cerebral artery occlusion/reperfusion (MCAO/R) in mice. In the current study, the authors attempt to demonstrate that NSC-derived exosomes could act as a supportive adjuvant for NSC transplantation after stroke. They showed that NSC-derived exosomes significantly reduced the inflammatory response, alleviated oxidative stress after NSC transplantation, and facilitated NSCs differentiation in vivo. The combination of NSCs …
Reviewer #1 (Public Review):
The therapeutic effects of NSC transplantation is limited by the low survival and differentiation rates of NSCs due to the harsh environment in the brain after ischemic stroke. To solve this technical challenge, the authors employed NSCs derived from human induced pluripotent stem cells (iPSCs) together with NSC-derived exosomes extracted from NSCs to treat cerebral ischemia induced by middle cerebral artery occlusion/reperfusion (MCAO/R) in mice. In the current study, the authors attempt to demonstrate that NSC-derived exosomes could act as a supportive adjuvant for NSC transplantation after stroke. They showed that NSC-derived exosomes significantly reduced the inflammatory response, alleviated oxidative stress after NSC transplantation, and facilitated NSCs differentiation in vivo. The combination of NSCs with exosomes ameliorated the injury of brain tissue including cerebral infarct, neuronal death and glial scarring, and promoted the motor function recovery. To explore the underlying mechanisms, they analyzed the miRNA profiles of NSC-derived exosomes and the potential downstream genes. Overall, the study provided solid rationale supporting the application of exosomes during stem cell-based therapy. The data quality is good and convincingly supports the major claims. The impact is high for the NSC-transplantation for treating various neurological diseases.
-
Reviewer #2 (Public Review):
In the submitted manuscript under the title "NSC-derived exosomes enhance therapeutic effects of NSC transplantation on cerebral ischemia in mice", Zhang et al. applied human induced pluripotent stem cells (iPSCs) together with exosomes extracted from NSCs to treat cerebral ischemia induced by middle cerebral artery occlusion/reperfusion (MCAO/R) in mice. They reported that NSC-derived exosomes can ease the inflammatory response, alleviated oxidative stress after NSC transplantation, and facilitated NSCs differentiation in mouse brain. Using the NSC together with their exosomes can ameliorate the injury of brain tissue including cerebral infarct, neuronal death and glial scarring, and promoted the motor function recovery. Finally, they speculated that the miRNA(s) in the exosomes is the key factor to improve …
Reviewer #2 (Public Review):
In the submitted manuscript under the title "NSC-derived exosomes enhance therapeutic effects of NSC transplantation on cerebral ischemia in mice", Zhang et al. applied human induced pluripotent stem cells (iPSCs) together with exosomes extracted from NSCs to treat cerebral ischemia induced by middle cerebral artery occlusion/reperfusion (MCAO/R) in mice. They reported that NSC-derived exosomes can ease the inflammatory response, alleviated oxidative stress after NSC transplantation, and facilitated NSCs differentiation in mouse brain. Using the NSC together with their exosomes can ameliorate the injury of brain tissue including cerebral infarct, neuronal death and glial scarring, and promoted the motor function recovery. Finally, they speculated that the miRNA(s) in the exosomes is the key factor to improve the treatment of the NSC transplantation for the stroke. This is an interesting study that contributes important findings which will be of interest to the researcher in the field of the NSC transplantation. However, there are some key points should be further explained.
Major points:
1). This study does not provide any evidence about the cell death of the transplanted cells. The immunostaining of the Caspase-3 or TUNEL staining should be used to address this issue.2). The authors showed that the neurological functions (evaluated by balance beam, ladder lung, rotarod test and Modified Neurological Severity Score (mNSS) up to 8 weeks after treatment (Figure 1C)) were significantly improved in the NES+Exo group compared to their control groups. However, these cells (transplanted cells) are progenitors (Nestin+) or undifferentiated cells (Tuj1+) at this stage (Figure 3). Thus, I was curious about that how can the immature neurons play neurological functions? This point should be explained.
3). The authors used the Golgi staining to show the NES+Exo can improve dendritic density and length. How do you know these neurons are transplanted cells?
4). The cell morphology of tdTomato+ cells is fuzzy and it is difficult to distinguish the cell body. It looks like that these cells out of whack.
-
Reviewer #3 (Public Review):
This study investigates the efficacy of exosomes of neuronal stem cells (NSC) derived from human iPSCs) in improving NSC therapy for neuroprotection in mouse stroke model. The results show that at one-week post-stroke, administration of NSCs through lateral ventricle injections in combination with exosomes significantly improved post-stroke survival, neurological function recovery and brain lesion attenuation in mice at 8-week post treatment. The strengths of this study include: 1) the positive outcomes from this combinatory treatment delivered at the subacute phase; 2) multiple assessments of neurological function impairments; 3) non-invasive, unbiased assessment of brain lesion with MRI. However, the evaluation of the possible underlying mechanisms is weak, which included reduction of reactive astrocytes, …
Reviewer #3 (Public Review):
This study investigates the efficacy of exosomes of neuronal stem cells (NSC) derived from human iPSCs) in improving NSC therapy for neuroprotection in mouse stroke model. The results show that at one-week post-stroke, administration of NSCs through lateral ventricle injections in combination with exosomes significantly improved post-stroke survival, neurological function recovery and brain lesion attenuation in mice at 8-week post treatment. The strengths of this study include: 1) the positive outcomes from this combinatory treatment delivered at the subacute phase; 2) multiple assessments of neurological function impairments; 3) non-invasive, unbiased assessment of brain lesion with MRI. However, the evaluation of the possible underlying mechanisms is weak, which included reduction of reactive astrocytes, increased NeuN+ cells, and possible roles of anti-inflammatory miRNA profiles of exosomes from NSCs in the study. Further strengthening of the relationship in the above phenomena will be beneficial for developing cell therapy for ischemic stroke.
-
