Abstract
摘要:身體部位受到損壞或是喪失後的再生能力在不同動物之間有相當大的差異。許多無脊椎動物可以很容易再生出失去的部分,如水螅和扁形蟲。至於脊椎動物的再生能力則相對較差,但是其中蠑螈則具有可觀的再生能力,例如肢體、肺、脊髓和一部分受損傷的腦組織。當蠑螈的肢體在肩膀和手之間任何部位截肢,截肢後的前端會形成一群稱為芽基(blastema)的前驅細胞區,由這些細胞群可以再生長出完整的手臂。蠑螈肢體再生的過程會經歷一連串的細胞逆分化,這個過程會使得軟骨、皮膚和肌肉細胞逆分化成外觀似乎是同質的芽基細胞群,這細胞群裡的細胞常常被看作是單一的細胞類型。目前這群細胞的由來主要有兩種說法:一、由截肢附近的結締組織的成熟細胞去分化(dedifferentiation)而來;二、由已經存在於結締組織中的未分化細胞所聚集產生。目前較被接受的概念是前者,由成熟的結締組織細胞來產生芽基細胞,或至少恢復到較未分化的狀態。由於在再生的肢體中具有多個不同類型的組織,科學家長期以來一直認為芽基細胞是多能性(pluripotent)幹細胞,然而Kragl等人的研究證明由不同組織所產生的芽基細胞會保留其原有的記憶,除少數例外,這些芽基細胞只會貢獻到其源自的組織類型。這項發現導致了重要的結論,即芽基細胞是一群被限制只能分化成其源自組織的異源前驅細胞群。儘管先前的研究已經表明長出的肌肉是由肌肉再生而不是由軟骨或表皮轉分化而來,但是我們仍然不了解再生的肌肉細胞是由肌肉前驅細胞或是已分化的肌肉,甚或是兩者共同而來的。在本計劃,我們提出建立一個血統追蹤系統來研究再生肢體的肌肉細胞之起源,這項技術是利用cre誘導DNA重組原理,第一年我們將鑑定並且選殖肌肉前驅細胞和已分化的肌肉細胞之特異基因啟動子以利血統追蹤實驗之標定。第二年,我們將在蠑螈中建立Cre-loxP血統追蹤系統。第三年,我們將探討蠑螈再生肢體之肌肉細胞的起源。如果Cre-loxP系統可以在蠑螈身上被建立,這個系統將會開啟一條新的路幫助我們探討哺乳動物,包括人類,在演化過程中為何喪失了他們的再生能力,並且可能有助於人類再生醫學領域的發展。
Abstract: The ability to regenerate lost or damaged body parts varies between animals. Many invertebrates can regenerate their lost parts, such as hydra and flatworms. As for vertebrates, salamander, unlike mammals, can reconstitute a fully functional limb, lungs, sliced spinal cord and bits of lopped-off brain after damage. When a salamander’s limb is amputated anywhere between the shoulder and the hand, it triggers the formation of a progenitor cell zone called blastema that reforms the missing limb.Salamander’s limb regenerates by undergoing reprogramming events that allow cartilage, skin and muscle into blastema, which appears to be a homogeneous group of cells and has been commonly viewed as a single cell type. Two general proposals for the origin of these blastema cells are: (1) blasteama cells arise by dedifferentiation of the connective tissue cells adjacent to the site of amputation; (2) they arise from pre-existing undifferentiated cells in the connective tissues adjacent to the site of amputation. The prevalent concept is the former. Since there are multiple tissue types in the regenerated limb, scientists had long believed blastema cells were pluripotent. However, Kragl et al. showed that blastema cells originating from the different tissues retain the memory of those tissues when they regenerate, contributing with few exceptions only to the same type of tissue where they came from. This leads to the important conclusion that the blastema is a pool of heterogenously restricted progenitor cells from its outset. Despite previous studies have demonstrated that muscle makes muscle but not cartilage or epidermis, we still wander whether the regenerated muscle is derived from muscle progenitor cells or from dedifferentiation of mature myocytes, or both, because in Kragl’s experiments they used GFP-transgenic salamanders whose muscle progenitor cells and differentiated myocytes were all labeled.In this project, we propose to investigate the origin of regenerated limb’s muscle cells in axolotls by developing a lineage tracing system by using cre-mediate DNA recombination. We will first identify and clone specific promoters of either muscle progenitor cells or differentiated muscle cells for lineage tracing experiment (1st year), and then establish Cre-loxP lineage tracing system for axolotl (2nd year). At last using this model, we will investigate the origin of regenerated limb muscle cells in axolotls (3rd year). If the Cre-loxP system can be established in axolotl, it will open a mysterious box explaining why mammals have lost their re-growth abilities during evolution, and the knowledge may help the field of regenerative medicine in mammals including humans.
Keyword(s)
蠑螈
肢體再生
去分化
芽基
血統追蹤
肌肉祖細胞
再生醫學
salamander
limb regeneration
dedifferentiation
blastema
lineage tracing
muscle progenitor cell
regenerative medicine