Roles of astrocyte on ventral midbrain- derived neural stem cell differentiation, dopamine neuron maturation, and survival

Abstract

Parkinson's disease (PD), characterized by specific loss of dopamine (DA) neurons in the midbrain substantia nigra, is the most prime target disorder for stem cell-based cell therapeutic approaches. Neural stem cells (NSCs) derived from ventral midbrain (VM)(VM-NSCs) are an appropriate cell source for PD cell therapy and studies have been performed to test therapeutic efficacy by VM-NSC transplantation in PD animal models. However, therapeutic efficacy by NSC transplantation does not reach the level to be clinically applied for PD patients, mainly because of donor poor cell survival, DA neuron differentiation, and functions in the grafted host brain environments. Astrocytes are the major glial cell type in the CNS, exerting physiologic functions to support neuronal cell survival and functions. In order to know if the neurotropic functions of astrocytes can be utilized to improve NSC transplantation outcomes, I examined a series of VM-NSC behaviors such as mDA neuron differentiation, neuronal maturation, midbrain-specific marker expression, survival and resistance against toxic stimuli, which are critical for successful NSC transplantation, in the VM-NSC cultures mixed with astrocytes derived from VM, cortical tissues or NSCs derived from embryonic cortices (Ctx-NSCs, control). Here I show that astrocytes promote VM-NSC differentiation towards DA neurons with greater neuronal maturity, midbrain-specific marker expressions, and resistance against toxic stimulus in a paracrine manner. VM-astrocytes were superior to Ctx-astrocytes in those neurotropic actions. I further show that forced expression of the midbrain-specific transcription factors Nurr1 and Foxa2 in VM-astrocytes striking reduced proinflammatory cytokine synthesis from the astrocytes and thus enhanced the neuroprotective actions. Collectively, this study suggest VM-astrocytes, especially those overexpressing Nurr1 and Foxa2 are the best cell type, co-transplantation of which can improve the therapeutic efficacy in VM-NSC-based transplantation approach for PD. |파킨슨병은 뇌의 중뇌의 흑색질 (substantia nigra)에 분포하는 도파민 신경세포의 소실로 인하여 발병하는 질환으로써 이의 치료를 위해 줄기세포를 이용하는 세포치료가 시행되고 있다. 이러한 세포치료는 파킨슨병 동물에 중뇌에서 유래된 neural stem cells (NSCs)의 세포 이식치료 연구가 활발히 진행되고 있다. 하지만 이러한 세포이식치료의 효율이 파킨슨병 환자에 적용할 수준까지 이르고 있지 못하고 있는 한계가 존재하고 있는데 이에 대한 원인을 살펴본다면 host brain으로 이식 되어지는 NSCs의 생존능력이 낮고 이식 후에 도파민 신경세포로의 분화와 기능이 제대로 이루어지지 않는 문제가 존재한다. 따라서 이에 대한 문제의 해결을 위하여 본 연구에서는 신경세포의 생존능력과 기능을 증가시키는데 도움을 주는 astrocyte의 neurotropic기능에 초점을 두어 세포이식치료의 효율의 향상을 목표로 하였다. 이러한 astrocyte의 효과를 확인하기 위하여 cortex(Ctx), VM astrocyte와 대조군인 Ctx NSCs를 도파민 신경세포와의 공배양을 통하여 도파민 신경세포의 분화와 성숙, 생존 정도를 조사하였고 astrocytes가 paracrine하게 도파민 신경세포의 분화와 성숙, 생존능력을 향상시켜주는 확인할 수 있었다. 또한 이러한 neurotropic효과가 VM-astrocytes에서 가장 뛰어남을 알 수 있었다. 더욱이 VM-astrocytes에서 중뇌 특이 전사인자인 Nurr1과 Foxa2의 과발현이proinflammatory cytokine의 합성을 감소시켜 도파민 신경세포의 보호효과를 향상시키는 것을 확인할 수 있었다.; Parkinson's disease (PD), characterized by specific loss of dopamine (DA) neurons in the midbrain substantia nigra, is the most prime target disorder for stem cell-based cell therapeutic approaches. Neural stem cells (NSCs) derived from ventral midbrain (VM)(VM-NSCs) are an appropriate cell source for PD cell therapy and studies have been performed to test therapeutic efficacy by VM-NSC transplantation in PD animal models. However, therapeutic efficacy by NSC transplantation does not reach the level to be clinically applied for PD patients, mainly because of donor poor cell survival, DA neuron differentiation, and functions in the grafted host brain environments. Astrocytes are the major glial cell type in the CNS, exerting physiologic functions to support neuronal cell survival and functions. In order to know if the neurotropic functions of astrocytes can be utilized to improve NSC transplantation outcomes, I examined a series of VM-NSC behaviors such as mDA neuron differentiation, neuronal maturation, midbrain-specific marker expression, survival and resistance against toxic stimuli, which are critical for successful NSC transplantation, in the VM-NSC cultures mixed with astrocytes derived from VM, cortical tissues or NSCs derived from embryonic cortices (Ctx-NSCs, control). Here I show that astrocytes promote VM-NSC differentiation towards DA neurons with greater neuronal maturity, midbrain-specific marker expressions, and resistance against toxic stimulus in a paracrine manner. VM-astrocytes were superior to Ctx-astrocytes in those neurotropic actions. I further show that forced expression of the midbrain-specific transcription factors Nurr1 and Foxa2 in VM-astrocytes striking reduced proinflammatory cytokine synthesis from the astrocytes and thus enhanced the neuroprotective actions. Collectively, this study suggest VM-astrocytes, especially those overexpressing Nurr1 and Foxa2 are the best cell type, co-transplantation of which can improve the therapeutic efficacy in VM-NSC-based transplantation approach for PD.