陳國慶

(武漢體育學(xué)院健康科學(xué)學(xué)院,武漢430079)

骨骼肌衛(wèi)星細(xì)胞是一種未分化的成肌前體細(xì)胞,于1961年被Mauro首先從蛙類骨骼肌中分離出來[1],在骨骼肌生長(zhǎng)發(fā)育、損傷修復(fù)以及骨骼肌重塑等生理病理過程中具有重要的作用。肌衛(wèi)星細(xì)胞自被發(fā)現(xiàn)以來,已經(jīng)在組織工程、基因治療、運(yùn)動(dòng)醫(yī)學(xué)等領(lǐng)域展示了良好的應(yīng)用前景,為臨床治療各種骨骼肌和心肌疾病奠定堅(jiān)實(shí)的基礎(chǔ)。適宜的運(yùn)動(dòng)訓(xùn)練是提高骨骼肌功能的有效手段,其作用機(jī)制目前尚未完全明確。本文就骨骼肌衛(wèi)星細(xì)胞及運(yùn)動(dòng)訓(xùn)練對(duì)骨骼肌衛(wèi)星細(xì)胞的調(diào)控機(jī)制作一綜述。

1 骨骼肌衛(wèi)星細(xì)胞的來源、形態(tài)特征及特異性標(biāo)記

1.1 骨骼肌衛(wèi)星細(xì)胞的來源

骨骼肌衛(wèi)星細(xì)胞的來源,目前存在體節(jié)來源和非體節(jié)來源兩種假說[2]。體節(jié)來源假說最先來自禽類模型的嵌合體或種間移植實(shí)驗(yàn),即將鵪鶉的胚胎中胚層生肌節(jié)移植到雞的胚胎中,且被移植的鵪鶉細(xì)胞有明顯的形態(tài)特征,可以觀察到這些細(xì)胞從移植的中胚層生肌節(jié)遷徙到胚胎發(fā)育的肢體,并組成出生后雞骨骼肌中肌衛(wèi)星細(xì)胞群[3～4]。此后的各種研究也證明了肌衛(wèi)星細(xì)胞來自體壁中胚層生肌節(jié)間充質(zhì)細(xì)胞的這一假說。而Angelis等[5]從胚胎背側(cè)主動(dòng)脈分離到具有與肌衛(wèi)星細(xì)胞相似的形態(tài)和基因表達(dá)特征的細(xì)胞,將這種主動(dòng)脈來源的細(xì)胞移植到新生小鼠后,發(fā)現(xiàn)這種細(xì)胞參與出生后肌肉的生長(zhǎng)和再生,并可與中胚層生肌節(jié)來源的肌纖維融合,因此認(rèn)為肌衛(wèi)星細(xì)胞可能起源于胚胎血管祖細(xì)胞。此外,還有研究發(fā)現(xiàn)將骨髓干細(xì)胞衍生的成肌細(xì)胞注射入靜脈,發(fā)現(xiàn)其也能夠參與骨骼肌的再生,表明骨髓干細(xì)胞衍生的成肌細(xì)胞也具有骨骼肌衛(wèi)星細(xì)胞相似的功能特征[6～7]。

1.2 骨骼肌衛(wèi)星細(xì)胞的形態(tài)學(xué)特征

成體的骨骼肌衛(wèi)星細(xì)胞位于肌纖維的基底膜處,通常處于靜息的非增殖狀態(tài),主要有以下特征: 1)明確定位于肌膜和基底膜之間;2)細(xì)胞的核漿比高,細(xì)胞器很少;3)細(xì)胞核體積較小;4)與肌細(xì)胞核相比,衛(wèi)星細(xì)胞核的異染色質(zhì)含量較多[8]。上述形態(tài)學(xué)特征與衛(wèi)星細(xì)胞相對(duì)靜止和缺乏轉(zhuǎn)錄活性相符合。經(jīng)過劇烈運(yùn)動(dòng)或是受到外界刺激之后,成體骨骼肌衛(wèi)星細(xì)胞會(huì)被激活,激活狀態(tài)下的衛(wèi)星細(xì)胞則另有其特征:1)細(xì)胞突起數(shù)目增多;2)胞漿內(nèi)的細(xì)胞器數(shù)量增多;3)細(xì)胞核異染色質(zhì)減少[8]。在光學(xué)顯微鏡下,傳統(tǒng)的組織學(xué)方法不易區(qū)別肌衛(wèi)星細(xì)胞和肌細(xì)胞核。但是通過甲苯氨藍(lán)染色和免疫組織化學(xué)的方法則可以有效鑒定肌衛(wèi)星細(xì)胞。

1.3 骨骼肌衛(wèi)星細(xì)胞的特異性標(biāo)記

隨著研究的深入,骨骼肌衛(wèi)星細(xì)胞特異表達(dá)的標(biāo)志蛋白相繼被發(fā)現(xiàn),通過對(duì)骨骼肌衛(wèi)星細(xì)胞表達(dá)的特異性蛋白的研究,可以明確骨骼肌衛(wèi)星細(xì)胞的激活、分化狀態(tài)以及衛(wèi)星細(xì)胞在增殖和分化過程中的分子調(diào)控。盡管目前對(duì)靜息狀態(tài)和激活狀態(tài)的骨骼肌衛(wèi)星細(xì)胞的基因表達(dá)譜尚未完全明確,但已有多種衛(wèi)星細(xì)胞特異性表達(dá)的標(biāo)記蛋白被確定,通過這些標(biāo)記蛋白可以明確區(qū)分各種靜息狀態(tài)、激活狀態(tài)或增殖分化狀態(tài)的肌衛(wèi)星細(xì)胞。這些標(biāo)記蛋白主要包括:1)細(xì)胞膜蛋白:M-Cadherin、Syndecan-3、Syndecan-4、c-Met、VCAM-1、NCAM、CD34;2)細(xì)胞骨架蛋白:Desmin;3)細(xì)胞轉(zhuǎn)錄因子:Pax-3、Pax-7、M yf5、M yoD、MNF、M yostain、IRF-2、M sx1[9]。

2 運(yùn)動(dòng)訓(xùn)練對(duì)骨骼肌衛(wèi)星細(xì)胞的影響

2.1 長(zhǎng)期訓(xùn)練對(duì)骨骼肌衛(wèi)星細(xì)胞的影響

長(zhǎng)期運(yùn)動(dòng)訓(xùn)練可有效增加骨骼肌衛(wèi)星細(xì)胞的數(shù)目。研究表明,經(jīng)過多年訓(xùn)練的優(yōu)秀舉重運(yùn)動(dòng)員斜方肌衛(wèi)星細(xì)胞較對(duì)照人群高出70%,而服用合成類固醇并不能有效增加骨骼肌衛(wèi)星細(xì)胞[10～11]。

2.2 短期訓(xùn)練對(duì)骨骼肌衛(wèi)星細(xì)胞的影響

依據(jù)骨骼肌細(xì)胞核域理論,一個(gè)肌細(xì)胞核只能控制一部分肌漿內(nèi)的m RNA轉(zhuǎn)錄和蛋白質(zhì)的合成,因此,隨著運(yùn)動(dòng)訓(xùn)練導(dǎo)致的肌纖維增粗(肥大),必然伴隨著骨骼肌細(xì)胞核的增多,而這些增多的細(xì)胞核則可能來源于骨骼肌衛(wèi)星細(xì)胞[12]。Roth等[13]對(duì)不同年齡的男子和婦女進(jìn)行了9周的單腿屈膝訓(xùn)練后發(fā)現(xiàn),骨骼肌衛(wèi)星細(xì)胞顯著增多,在訓(xùn)練的早期達(dá)到峰值。在另一項(xiàng)進(jìn)行16周的力量訓(xùn)練研究中發(fā)現(xiàn),伴隨著骨骼肌纖維肥大,衛(wèi)星細(xì)胞數(shù)目在第4周、第8周、第16周分別增加22%、40%、27%,肌肉活檢發(fā)現(xiàn)衛(wèi)星細(xì)胞CyclinD1和P21m RNA表達(dá)上調(diào),表明衛(wèi)星細(xì)胞增殖增多[14]。諸多的研究都證實(shí)在進(jìn)行數(shù)周至數(shù)十周的力量訓(xùn)練后,骨骼肌衛(wèi)星細(xì)胞數(shù)目均不同程度地增多。除了力量訓(xùn)練可以激活肌衛(wèi)星細(xì)胞外,研究還發(fā)現(xiàn)耐力訓(xùn)練同樣可以使衛(wèi)星細(xì)胞數(shù)目增多,在對(duì)11名男子進(jìn)行14周的有氧訓(xùn)練后,最大攝氧量提高25%,骨骼肌衛(wèi)星細(xì)胞數(shù)目增加29%[15];在對(duì)11名70～80歲老年男子的研究中發(fā)現(xiàn),在以最大攝氧量65%的強(qiáng)度訓(xùn)練14周后,最大攝氧量、檸檬酸合成酶活性、Ⅱ(a)肌纖維含量及股外側(cè)肌衛(wèi)星細(xì)胞數(shù)目顯著上升[15]。有氧耐力訓(xùn)練導(dǎo)致骨骼肌衛(wèi)星細(xì)胞數(shù)目增加的幅度較之力量訓(xùn)練要小。在機(jī)體衰老過程中,由于骨骼肌衛(wèi)星增殖能力的下降,衛(wèi)星細(xì)胞數(shù)目隨年齡的增加而減少,而進(jìn)行有氧訓(xùn)練則可以減緩由于年齡增加而導(dǎo)致的骨骼肌衛(wèi)星細(xì)胞數(shù)目減少以及骨骼肌機(jī)能水平下降的趨勢(shì)[16]。

2.3 單次急性運(yùn)動(dòng)對(duì)骨骼肌衛(wèi)星細(xì)胞的影響

單次急性運(yùn)動(dòng)也可以使骨骼肌衛(wèi)星細(xì)胞數(shù)目增多,8名男子在進(jìn)行單次高強(qiáng)度單腿跳躍訓(xùn)練后4至8 d,骨骼肌衛(wèi)星細(xì)胞仍顯著高于對(duì)側(cè)肌肉,但是肌肉活檢卻未發(fā)現(xiàn)有新的肌漿球蛋白合成以及成肌細(xì)胞增殖分化的標(biāo)志蛋白M yogenin和Desmin的表達(dá),表明單次的急性運(yùn)動(dòng)可以促進(jìn)衛(wèi)星細(xì)胞數(shù)目增多,但卻不能誘導(dǎo)衛(wèi)星細(xì)胞向成肌細(xì)胞分化[17]。

2.4 停訓(xùn)對(duì)骨骼肌衛(wèi)星細(xì)胞的影響

在進(jìn)行系統(tǒng)的運(yùn)動(dòng)訓(xùn)練后,隨著停訓(xùn)時(shí)間的延長(zhǎng),骨骼肌衛(wèi)星細(xì)胞數(shù)目開始逐步減少,通過對(duì)衛(wèi)星細(xì)胞CyclinD1和P21 m RNA表達(dá)的分析表明停訓(xùn)后衛(wèi)星細(xì)胞活化終止[14]。運(yùn)動(dòng)訓(xùn)練不僅可以使衛(wèi)星細(xì)胞池得以維持,還可使衛(wèi)星細(xì)胞數(shù)目增多,促進(jìn)骨骼肌衛(wèi)星細(xì)胞的自我更新。

3 運(yùn)動(dòng)訓(xùn)練影響骨骼肌衛(wèi)星細(xì)胞的調(diào)控機(jī)制

運(yùn)動(dòng)訓(xùn)練能夠激活骨骼肌衛(wèi)星細(xì)胞,并使之增殖甚至向成肌細(xì)胞分化已經(jīng)在諸多的人體實(shí)驗(yàn)和動(dòng)物模型中得到證實(shí),然而目前對(duì)于運(yùn)動(dòng)訓(xùn)練引起的骨骼肌衛(wèi)星細(xì)胞激活的確切機(jī)制尚未完全清楚。一般認(rèn)為運(yùn)動(dòng)訓(xùn)練激活骨骼肌衛(wèi)星細(xì)胞主要通過三種途徑:1)運(yùn)動(dòng)導(dǎo)致局部骨骼肌的微細(xì)損傷;2)運(yùn)動(dòng)刺激導(dǎo)致炎性因子釋放;3)運(yùn)動(dòng)導(dǎo)致局部生長(zhǎng)因子和細(xì)胞因子的釋放增多。運(yùn)動(dòng)負(fù)荷的刺激導(dǎo)致骨骼肌微細(xì)損傷并引起免疫應(yīng)答反應(yīng),巨噬細(xì)胞進(jìn)入損傷部位,一方面清除可能壞死的肌纖維,另一方面分泌各種炎性因子和生長(zhǎng)因子激活衛(wèi)星細(xì)胞,促進(jìn)其活化增殖、遷移、分化及融合到鄰近的肌纖維,調(diào)節(jié)骨骼肌生長(zhǎng)[18]。

3.1 細(xì)胞生長(zhǎng)因子

目前已經(jīng)確認(rèn)的對(duì)骨骼肌衛(wèi)星細(xì)胞具有調(diào)控作用的生長(zhǎng)因子和細(xì)胞因子主要有胰島素樣生長(zhǎng)因子(IGF)、肝細(xì)胞生長(zhǎng)因子(HGF)、成纖維細(xì)胞生長(zhǎng)因子(FGF)、轉(zhuǎn)化生長(zhǎng)因子β(TGF-β)、機(jī)械生長(zhǎng)因子(M GF)等。

骨骼肌主要表達(dá)IGF-Ⅰ和IGF-Ⅱ,體外的細(xì)胞培養(yǎng)研究中發(fā)現(xiàn)IGF-Ⅰ和IGF-Ⅱ可以促進(jìn)衛(wèi)星細(xì)胞增殖。在損傷肌肉的肌膜下注射外源性IGF-Ⅰ,可以促使肌衛(wèi)星細(xì)胞增殖,骨骼肌纖維肥大[19～20]。IGF-Ⅰ通過多種信號(hào)通道調(diào)節(jié)骨骼肌衛(wèi)星細(xì)胞群,主要包括Calcineurin/NFA T、MAPK激酶及PI3K信號(hào)通路[21-23]。

肝細(xì)胞生長(zhǎng)因子是一種多功能細(xì)胞因子,最近研究發(fā)現(xiàn)HGF及其受體c-Met僅在骨骼肌衛(wèi)星細(xì)胞及其鄰近的肌纖維表達(dá),而在成纖維細(xì)胞上未見有表達(dá),其表達(dá)水平與骨骼肌損傷程度呈高度相關(guān)[24～25]。運(yùn)動(dòng)訓(xùn)練所致的骨骼肌的微細(xì)損傷使得NO和HGF大量釋放,HGF在NO的活化作用下與衛(wèi)星細(xì)胞膜上的HGF受體c-Met結(jié)合,從而激活衛(wèi)星細(xì)胞增殖的信號(hào)傳導(dǎo)通路,并改變衛(wèi)星細(xì)胞與周圍組織的黏附狀態(tài)。此外,HGF還可以通過抑制成肌調(diào)節(jié)因子如M yoD和M yogenin等的轉(zhuǎn)錄來抑制衛(wèi)星細(xì)胞的分化[26]。

Sheehan和A llen通過細(xì)胞培養(yǎng)深入研究了FGF家族在衛(wèi)星細(xì)胞增殖過程中的作用,結(jié)果發(fā)現(xiàn)FGF-1、2、4、6和FGF-9能夠刺激衛(wèi)星細(xì)胞增殖,而FGF-5、7、8則無促進(jìn)衛(wèi)星細(xì)胞增殖的作用[27]。與HGF一樣,FGF家族同樣也能抑制衛(wèi)星細(xì)胞向成肌細(xì)胞的分化[28～29]。Floss等[30]將小鼠的 FGF-6基因剔除,結(jié)果發(fā)現(xiàn)小鼠骨骼肌損傷后出現(xiàn)再生障礙,然而有趣的是FGF-6基因過度表達(dá)的轉(zhuǎn)基因小鼠同樣也出現(xiàn)骨骼肌損傷后再生障礙,目前對(duì)FGF-6對(duì)骨骼肌的調(diào)控機(jī)制尚有待于進(jìn)一步研究。

TGF-β家族通過 SMAD蛋白家族傳導(dǎo)信號(hào)[31]。一般認(rèn)為 TGF-β家族成員可通過抑制M yoD家族成員的轉(zhuǎn)錄激活而抑制骨骼肌衛(wèi)星細(xì)胞增殖與分化[32～33]。IGF-Ⅰ或 FGF與 TGF-β聯(lián)合并不能改變TGF-β導(dǎo)致衛(wèi)星細(xì)胞分化抑制的作用,然而TGF-β卻不能抑制由IGF-Ⅰ或FGF導(dǎo)致的衛(wèi)星細(xì)胞增殖增加的作用[34]。

3.2 Notch信號(hào)通路

運(yùn)動(dòng)刺激或骨骼肌損傷后,Notch配體Δ表達(dá)隨骨骼肌衛(wèi)星細(xì)胞的活化而上調(diào)。Notch配體與靜息的衛(wèi)星細(xì)胞表面Notch受體1結(jié)合,激活衛(wèi)星細(xì)胞成肌轉(zhuǎn)錄因子,從而促進(jìn)衛(wèi)星細(xì)胞增殖分化。應(yīng)用No tch拮抗劑Numb則可顯著抑制衛(wèi)星細(xì)胞的活化[35～37]。

4 小結(jié)

目前,諸多研究已經(jīng)揭示骨骼肌衛(wèi)星細(xì)胞在骨骼肌生長(zhǎng)發(fā)育、損傷修復(fù)和重塑中的重要作用。然而,在運(yùn)動(dòng)訓(xùn)練過程中,骨骼肌衛(wèi)星細(xì)胞從靜息到激活、增殖、終末分化等過程中基因表達(dá)特征及其分子調(diào)控機(jī)理仍未完全清楚;不同強(qiáng)度不同運(yùn)動(dòng)方式對(duì)骨骼肌衛(wèi)星細(xì)胞活化、增殖和分化的影響均未完全明確。隨著研究的深入,衛(wèi)星細(xì)胞活化、增殖及分化過程中新的調(diào)控機(jī)制不斷被發(fā)現(xiàn),如 microRNA、DNA甲基化等調(diào)控衛(wèi)星細(xì)胞分化[38～41]。對(duì)骨骼肌衛(wèi)星細(xì)胞及運(yùn)動(dòng)訓(xùn)練對(duì)其調(diào)控的深入研究,為臨床上治療和預(yù)防骨骼肌衰減、萎縮等退行性疾病提供理論基礎(chǔ)和實(shí)驗(yàn)依據(jù)。

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