In living organisms, tissue injury and repair processes evolved to protect organisms from various tissue damage and insults that arise from both extrinsic and intrinsic factors. Unlike many non-vertebrates and some vertebrates such as salamanders, tissue repair capabilities in mammals are relatively limited. In humans, understanding how appropriate tissue regeneration processes occur without organ function impairment and systemic adverse effects has important clinical implications in the prognosis and treatment of disease. Tissue regeneration processes in vertebrates require stem cells which are maintained and regulated through complex interactions between the stem cell and cellular and humoral components of the surrounding milieu. Here, I investigated the role of paracrine fibroblast growth factors (FGFs) in tissue injury and regeneration, specifically in the intestine and muscle.
Ionizing radiation has been used in various model organisms as an experimental paradigm for acute tissue injury and repair processes. In my studies, I assessed the gain of function effects of FGF2 in gastrointestinal tract regeneration after radiation injury. In another setting, aging is known to act as a modifier of regenerative processes in skeletal muscle tissue. Clinically, age-dependent decline in muscle regenerative function has been suggested to contribute to sarcopenia, a decrease in muscle mass and function, in older people. To explore a potential role for FGFs in muscle regeneration, I investigated the loss-of-function effects of FGF1, which has recently been shown to be a regulator of adipose tissue homeostasis, in skeletal muscle regeneration after chemical injury.
I assessed the mitigation effect of FGF2 in gastrointestinal tract injury after whole body irradiation. In IEC-6 cell based screening, FGF2 showed possible mitigative effects by reducing apoptosis among FGF1, FGF2, FGF19. For in vivo experiments, 12 Gy of whole body irradiation to maximize acute gastrointestinal syndrome in mice. After 12 Gy whole body irradiation, adjuvant FGF2 with bone marrow transplantation (BMT) showed significantly increased 2-weeks survival rates, over other experimental groups. Treating mice with FGF2 after BMT augmented proliferation, decreased apoptosis and improved crypt regeneration, while there was no significant improvement with FGF2 alone or BMT alone. Furthermore, intestinal hyperpermeability, a key measure of radiation-induced gastroinstestinal damage, was absent by combined treatment of FGF2 and BMT, accompanied by improved cytokine profiles. In addition, several genes associated with intestinal barrier integrity and stem cell markers were significantly up-regulated with adjuvant FGF2 combined with BMT. These results suggested that FGF2 following BMT could mitigate intestinal irradiation damage through stem cell regeneration, maintaining barrier integrity maintenance, and cytokine modulation.
In contrast to the gain-of-function role of extrinsically administered FGF2 in gastrointestinal tract as a model to study tissue repair, I investigated the intrinsic role and loss-of-function phenotypes of FGF1, another paracrine FGF, in skeletal muscle regeneration. In FGF1 muscle specific knock-out (mKO) mice, muscle regeneration was impaired after chemical injury, and cross-section area of regenerating muscle fiber was smaller in FGF1 mKO compared to the wild type (WT) mice. Disorganized regenerating fiber and adipocyte infiltration in the regenerating area was also observed in FGF1 mKO mice. To probe the functional mechanism of FGF1 in myogenesis, I examined the effect of FGF1 signal blockade in a C2C12 myoblast differentiation model. Among FGF receptors (FGFRs), FGFR4 specific small molecule inhibitor potently inhibited myotube formation. Similarly, treatment of FGFR4 specific inhibitor led to formation of disorganized muscle fibers and fatty infiltration in regenerating muscle after chemical injury. On the other hand, injured muscle regenerated normally with FGFR1 specific blockade. In C2C12 myoblast differentiation model, I identified FGFR4-Wnt pathway as a possible downstream mediator and PPAR$\delta$ as a possible upstream regulator of FGF1 during myogenesis.
Taken together, I show that FGF2 has beneficial effects on irradiation induced gastrointestinal injury, and that FGF1 is required for appropriate skeletal muscle regeneration, supporting the importance of paracrine FGFs in tissue repair process after injury.
다양한 임상적 상황에서 조직 손상이 유발되며, 내, 외과적 치료의 예후에 있어서 조직의 기능을 저하시키지 않으며 전신의 합병증을 초래하지 않는 조직의 재생은 중요한 역할을 한다. 조직의 재생에서는 해당 조직에 존재하는 줄기 세포와 주변 환경간의 상호작용에 의하여 조직의 형성 과정을 거치게 되며, 이러한 과정에 있어서 여러가지 곁분비 물질과 마찬가지로, 섬유아세포성장인자 (FGF)의 역할이 존재할 것으로 생각되지만 아직까지 성체의 조직 재생에서 FGF 1과 2의 구체적인 역할은 잘 연구되어 있지 않은 실정이다.
본 연구에서는 줄기세포의 계대 배양에 널리 사용되는 FGF2 의 방사선 유발 장 손상에서의 역할과, 과거 지방 조직 리모델링 과정에서의 역할이 알려진 바 있는 FGF1의 골격근 손상 후 재생에서의 역할을 규명하고자 하였다.
전신 방사선 조사 후 FGF2 에 의한 장 손상의 완화효과를 확인하였다. 세포 실험을 통하여 FGF1, 2, 19 중 FGF2의 효과가 확인되었으며, 이에 동물 실험에서 FGF2를 이용하였다. 동물 실험에서, 12 Gy 의 방사선 조사 후 골수이식에 FGF2를 추가로 투여한 경우, 골수이식 단독군에 비하여 2주간의 생존률이 우수하였다. FGF2와 골수이식의 병용이 선와의 증식을 촉진하고 세포 사멸을 억제하며 선와 재생을 촉진하였으며, 장 투과성과 혈중 염증성 사이토카인이 FGF2 와 골수이식 병용 그룹에서 감소되었다. 또한, 장의 완결성과 관련된 유전자와 장 줄기세포 관련 유전자의 발현이 FGF2 병용에 의하여 증가되었다.
근육 조직에서는 FGF1 기능 소실 모델의 표현형을 분석하였다. FGF1 근육특이 KO (mKO) 생쥐에서 화학적 손상 후 근육 재생이 정상적으로 이루어지지 않았으며, 재생 근섬유의 단면적이 감소되어 있었으며 근섬유의 형태가 비정상적이었다. 또한 지방 세포 침윤이 FGF1 mKO 마우스에서 관찰되었다. FGF1 의 하위 경로 중 FGFR4 를 특이적으로 억제하였을 때에 근육아세포 분화가 지연되었으며, 생체에서 FGFR4 를 억제하였을 때에 FGF1 mKO 의 표현형이 재연되었다. 또한, 근육아세포를 통한 기전 연구를 통하여 PPAR$\delta$ 가 FGF1 의 근육 생성 과정의 상위 인자임이 확인되었다.
이와 같은 연구는 곁분비 성장인자인 FGF2 가 방사선 조사 장 손상에 유익한 효과를 보이며, FGF1 이 존재하지 않을 경우 골격근 재생이 정상적으로 이루어지지 않음을 시사한다