本文選題：非特異性筋膜結(jié)締組織 + 機(jī)械牽拉刺激　； 參考：《南方醫(yī)科大學(xué)》2008年碩士論文

【摘要】： 背景: 細(xì)胞是人體的最基本功能單位;功能類似的細(xì)胞構(gòu)成組織;不同的組織構(gòu)成具有一定形態(tài)和功能的器官;多個功能類似的器官組成特定功能的系統(tǒng):消化、呼吸、泌尿、生殖、循環(huán)、運(yùn)動、感官、神經(jīng)和內(nèi)分泌。根據(jù)胚胎發(fā)育的三胚層原理、發(fā)育生物學(xué)理論和現(xiàn)代醫(yī)學(xué)生物學(xué)的研究結(jié)果,結(jié)合我們在數(shù)字人研究過程中的發(fā)現(xiàn),中胚層間充質(zhì)分化成多個器官系統(tǒng)后,所遺留的部分形成遍布全身的非特異筋膜結(jié)締組織支架,非特異筋膜結(jié)締組織以干細(xì)胞為中心,為其他功能細(xì)胞的再生提供細(xì)胞儲備和支持——稱為支持儲備系統(tǒng);該支架在神經(jīng)系統(tǒng)和免疫系統(tǒng)的作用下,參與機(jī)體內(nèi)環(huán)境變化的檢測與功能狀態(tài)的調(diào)控——稱為自體監(jiān)控系統(tǒng)。因此,我們提出了人體內(nèi)存在第十個功能系統(tǒng)——支持儲備與自體監(jiān)控系統(tǒng)的新學(xué)說。 經(jīng)絡(luò)是整個中醫(yī)理論的基礎(chǔ)與核心,從現(xiàn)代生物學(xué)的角度揭示經(jīng)絡(luò)實質(zhì)是中醫(yī)理論現(xiàn)代化的關(guān)鍵。我們在數(shù)字人研究的過程中,對近年來經(jīng)絡(luò)研究中受關(guān)注較多的人體筋膜結(jié)構(gòu)進(jìn)行了分割、標(biāo)記和三維重建。發(fā)現(xiàn)了與古代經(jīng)絡(luò)記載走行相似的粗細(xì)不等的立體結(jié)構(gòu),認(rèn)為其中較粗的部位相當(dāng)于經(jīng)絡(luò)上的“穴位”,較細(xì)的部位相當(dāng)于經(jīng)絡(luò)的“經(jīng)脈”。通過對整個人體的筋膜進(jìn)行重建,顯示出筋膜在人體內(nèi)部所形成的支架,從淺入深有5種結(jié)構(gòu)構(gòu)成:1、真皮致密結(jié)締組織;2、皮下疏松結(jié)締組織;3、肌肉表面疏松結(jié)締組織;4、肌間隔和肌間隙結(jié)締組織;5、內(nèi)臟器官門、被膜和內(nèi)部間隔結(jié)締組織。通過研究國標(biāo)人體14條經(jīng)絡(luò)361個穴位的進(jìn)針部位和手法,發(fā)現(xiàn)人體穴位的針刺部位均位于筋膜的不同層次,其中位于肌間隔和肌間隙結(jié)締組織者最多,其次是真皮致密結(jié)締組織層和皮下疏松結(jié)締組織層,少數(shù)位于肌肉表面疏松結(jié)締組織(深筋膜)和內(nèi)臟器官門、被膜和內(nèi)部間隔結(jié)締組織。從筋膜學(xué)的角度,我們認(rèn)為中醫(yī)療法的基本點是通過包括針灸、刮痧、梅花針在內(nèi)的各種物理方式,刺激筋膜結(jié)締組織以調(diào)整人體的機(jī)能狀況和生命代謝;通過中藥湯劑的內(nèi)服等生物活性成分的作用,改變?nèi)梭w結(jié)締組織中功能細(xì)胞的活性,以調(diào)整機(jī)能狀況和生命代謝。中醫(yī)針灸刺激的解剖學(xué)基礎(chǔ)為人體非特異性筋膜結(jié)締組織(如:肌間隔、肌間隙等在旋轉(zhuǎn)針體時能牽動較大范圍筋膜結(jié)構(gòu),可達(dá)骨膜,產(chǎn)生較強(qiáng)生物信息的部位),刺激點(穴位)與非刺激點(非穴位)其實只是信息“量”的區(qū)別而沒有“質(zhì)”的不同。中藥通過改善筋膜的微循環(huán)狀況(活血化淤)和上皮基膜的通透性(補(bǔ)氣)調(diào)節(jié)功能細(xì)胞的再生和活性。 從生物進(jìn)化的過程中可以了解到結(jié)締組織是從間充質(zhì)演化而來的,間充質(zhì)是從次級間充質(zhì)演化而來,次級間充質(zhì)是從初級間充質(zhì)演化而來。概括來說多細(xì)胞生物——細(xì)胞外液——維持生物內(nèi)環(huán)境穩(wěn)定和外形,修復(fù)損傷細(xì)胞、調(diào)控生物自身代謝;三胚層動物——間充質(zhì)——分化為各種維持內(nèi)環(huán)境穩(wěn)定的器官系統(tǒng),修復(fù)損傷細(xì)胞、調(diào)控自身代謝;高等動物——筋膜——支持、固定、分隔、修復(fù)損傷細(xì)胞、調(diào)控生物自身代謝。 間充質(zhì)干細(xì)胞(mesenchymal stem cells,MSCs)具有自我更新、多向分化的能力。最初是由Friendenstein等發(fā)現(xiàn),并證明其在體外可以分化為成骨細(xì)胞及脂肪細(xì)胞,而后其它的研究小組發(fā)現(xiàn)骨髓來源的MSCs不僅可以分化為中胚層來源的組織,而且可以分化為內(nèi)胚層和外胚層來源的組織�，F(xiàn)在已有利用間充質(zhì)干細(xì)胞移植治療心肌梗死、先天性骨發(fā)育不全、神經(jīng)系統(tǒng)疾病、糖尿病的報道。目前研究比較多的是骨髓來源的MSCs,但是MSCs廣泛分布于其它組織中,包括臍血、外周血、肌肉、皮膚、肝臟、胰腺和脂肪等。間充質(zhì)干細(xì)胞的多組織來源性與筋膜結(jié)締組織支架的全身分布性是一致的,其縱向、橫向分化的生物學(xué)特性與筋膜學(xué)提出的筋膜結(jié)締組織對機(jī)體的支持儲備作用也是一致的。2007年,UCLA(University of California Los Angeles)研究人員與哈佛大學(xué)研究人員合作,通過向小鼠成纖維細(xì)胞添加能夠與DNA特定位點結(jié)合的4種轉(zhuǎn)錄因子,包括Nanog和OCT-4,可以將成纖維細(xì)胞再程序化為多能細(xì)胞,多項檢測結(jié)果證實這些再程序化后的細(xì)胞與胚胎干細(xì)胞從結(jié)構(gòu)到功能均非常相似。 總之,為證實筋膜結(jié)締組織以干細(xì)胞為中心對機(jī)體產(chǎn)生的支持儲備作用,可以從以下幾個方面進(jìn)行初步研究:1、筋膜結(jié)締組織中是否存在間充質(zhì)干細(xì)胞;2、物理刺激對筋膜結(jié)締組織中功能細(xì)胞產(chǎn)生怎樣的效應(yīng);3、物理刺激作用產(chǎn)生的細(xì)胞效應(yīng)如何對機(jī)體發(fā)揮支持儲備作用。 目的 研究機(jī)械牽拉刺激對非特異性筋膜結(jié)締組織細(xì)胞生物活性的影響,初步探討非特異性筋膜結(jié)締組織對機(jī)體產(chǎn)生支持儲備作用的細(xì)胞生物學(xué)基礎(chǔ): 1、判斷非特異性筋膜結(jié)締組織中是否存在間充質(zhì)干細(xì)胞; 2、研究機(jī)械牽拉刺激能否對筋膜結(jié)締組織細(xì)胞增殖活性產(chǎn)生影響; 3、研究機(jī)械牽拉刺激能否改變筋膜結(jié)締組織中相關(guān)細(xì)胞因子基因表達(dá)。 方法 1、取成年大鼠皮下筋膜結(jié)締組織,酶消化法分離、培養(yǎng)筋膜結(jié)締組織源間充質(zhì)干細(xì)胞。通過形態(tài)學(xué)、功能學(xué)、流式細(xì)胞術(shù)鑒定非特異性筋膜結(jié)締組織源細(xì)胞是否符合間充質(zhì)干細(xì)胞的特性,以判斷非特異性筋膜結(jié)締組織中是否含有間充質(zhì)干細(xì)胞。 2、機(jī)械牽拉刺激筋膜結(jié)締組織,BrdU摻入法標(biāo)記增殖細(xì)胞,免疫熒光法觀察細(xì)胞增殖活性,并進(jìn)行統(tǒng)計學(xué)分析;并觀察牽拉刺激對筋膜結(jié)締組織結(jié)構(gòu)和形態(tài)的影響。 3、機(jī)械牽拉刺激筋膜結(jié)締組織,通過RT-PCR擴(kuò)增基因片段,觀察細(xì)胞增殖因子(a-fgf);血管生成因子(VEGF);胚胎干細(xì)胞特異基因:Nanog,OCT-4基因表達(dá)是否增高。 結(jié)果 1、非特異筋膜結(jié)締組織源細(xì)胞具有長梭形、多角形的形態(tài)學(xué)特征,可以形成細(xì)胞集落,體外誘導(dǎo)可以分化成成骨細(xì)胞、脂肪細(xì)胞,絕大多數(shù)細(xì)胞(＞96%)處于細(xì)胞周期的G0/G1期。 2、機(jī)械拉伸刺激筋膜結(jié)締組織后,實驗組大鼠腹股溝脂肪墊失去正常結(jié)構(gòu),切口有較多滲血,細(xì)胞增殖活性顯著提高,有統(tǒng)計學(xué)意義。 3、機(jī)械拉伸刺激筋膜結(jié)締組織后,細(xì)胞增殖因子a-fgf、血管生成因子VEGF基因表達(dá)量顯著增高,未能檢測到Nanog、OCT-4的基因表達(dá)。 結(jié)論 1、非特異性筋膜結(jié)締組織源細(xì)胞在形態(tài)學(xué)上與其他組織來源的間充質(zhì)干細(xì)胞無明顯差別;可以誘導(dǎo)成骨、成脂肪,與間充質(zhì)干細(xì)胞多向分化功能一致;細(xì)胞增殖周期檢測中,絕大多數(shù)處于靜止?fàn)顟B(tài),符合間充質(zhì)干細(xì)胞特性�？梢耘袛嘣诔墒齑笫蟮慕钅そY(jié)締組織中存在間充質(zhì)干細(xì)胞。 2、機(jī)械牽拉刺激可以顯著提高非特異性筋膜結(jié)締組織細(xì)胞的增殖活性,初步證實筋膜學(xué)觀點:物理刺激改變筋膜結(jié)締組織功能細(xì)胞增殖活性,通過功能細(xì)胞數(shù)量和性質(zhì)的改變,對機(jī)體產(chǎn)生支持儲備及調(diào)控作用。 3、機(jī)械牽拉刺激可以顯著提高筋膜結(jié)締組織中細(xì)胞增殖相關(guān)因子FGF、血管生成因子VEGF基因表達(dá)量,可以促進(jìn)細(xì)胞增殖,及血管生成。證實物理刺激改變細(xì)胞基因表達(dá),為其對機(jī)體產(chǎn)生的支持、儲備作用提供了可能。但是本實驗未能檢測到干細(xì)胞相關(guān)因子Nanog、OCT-4基因表達(dá),因此尚無法從干細(xì)胞相關(guān)因子方面去證實筋膜結(jié)締組織的以干細(xì)胞為中心的支持儲備功能,需要繼續(xù)研究下去。
[Abstract]:Background :


The cells are the most basic units of human body ; cells of similar function form tissues ; different tissues form organs with certain morphology and function ; different organs form organs with certain functions and functions : digestive , respiratory , urinary , reproductive , circulatory , sports , sensory , neurological and endocrine .


Based on the theory of traditional Chinese medicine and its core , it is the key to the modernization of traditional Chinese medicine theory from the angle of modern biology .


In the process of biological evolution , it can be understood that connective tissue is evolved from mesenchymal evolution , and the mesenchymal stem from the secondary mesenchymal evolution . The secondary mesenchyme is derived from the evolution of the mesenchymal stem . In general , the multi - cell biological _ extracellular fluid _ maintains the stable and external appearance of the organism , repairs the injured cells , regulates the metabolism of the organism itself ; the three - dimensional animal _ mesenchymal _ differentiation is a variety of internal and stable organ systems , repair the injured cells , regulate the metabolism of the organism ; and repair the injured cells and regulate the biological metabolism .


Bone marrow - derived mesenchymal stem cells ( MSCs ) can be differentiated into osteoblasts and fat cells in vitro .


In conclusion , in order to confirm the support reserve action of the connective tissue of fascia with stem cell as the center , the preliminary research can be carried out from the following aspects : 1 . Whether mesenchymal stem cells exist in the connective tissue of the fascia ; 2 . The effect of physical stimulation on the functional cells in the connective tissue of the fascia ; 3 . How the cell effect generated by the physical stimulation can play a supportive role in the organism .


Purpose


To investigate the effect of mechanical traction stimulation on the biological activity of non - specific fascia connective tissue cells , and to investigate the cellular biological basis of non - specific fascia connective tissue on the organism ' s support reserve .


1 , judging whether mesenchymal stem cells exist in the non - specific fascia connective tissue ;


2 . The effect of mechanical traction stimulation on the proliferation of connective tissue cells was studied .


3 . To study whether the mechanical traction stimulation can change the expression of related cytokine genes in the connective tissue of fascia .


method


1 . The connective tissue of fascia connective tissue was isolated and cultured in adult rats . The mesenchymal stem cells were cultured by morphological , functional and flow cytometry to determine whether the non - specific fascia connective tissue source cells were consistent with the characteristics of mesenchymal stem cells to determine whether the non - specific fascia connective tissue contained mesenchymal stem cells .


2 . mechanically pulling and stimulating the connective tissue of fascia , labeling the proliferating cells by the method of labeled proliferation , observing the proliferation activity of the cells by immunofluorescence , and carrying out statistical analysis ; and observing the influence of the traction stimulation on the structure and the morphology of the fascia connective tissue .


3 . mechanically pulling the connective tissue of fascia , amplifying the gene fragment by RT - PCR , observing the cell proliferation factor ( a - fgf ) , the angiogenesis factor ( VEGF ) , the specific gene of the embryonic stem cell : Nanog , whether the expression of OCT - 4 gene is increased or not .


Results


1 . The non - specific fascia connective tissue source cells have long fusiform and polygonal morphological characteristics , which can form the colony of cells . In vitro induction can differentiate into osteoblasts , fat cells , and most of the cells ( > 96 % ) are in G0 / G1 phase of the cell cycle .


2 . After mechanically stretching and stimulating the connective tissue of fascia , the fat pad of inguinal fat in the experimental group lost its normal structure .


3 . After mechanically stretching and stimulating the connective tissue of fascia , the expression of the vascular endothelial growth factor a - fgf and the VEGF gene was significantly increased , and the gene expression of Nanog and OCT - 4 was not detected .


Conclusion


1 . The non - specific fascia connective tissue source cells have no significant difference in morphology from other tissue - derived mesenchymal stem cells ; it can be induced into bone and fat , and is consistent with the multi - directional differentiation function of mesenchymal stem cells ; in the cell proliferation cycle detection , the majority of the mesenchymal stem cells are in a quiescent state , and the mesenchymal stem cell characteristics are met .


2 . Mechanical traction stimulation can significantly improve the proliferation activity of non - specific fascia connective tissue cells . It is preliminarily confirmed that the physical stimulation changes the proliferation activity of the functional cells of the fascia connective tissue , and supports and controls the organism by the change of the quantity and nature of the functional cells .


3 . The mechanical traction stimulation can significantly improve the expression of FGF and VEGF in the connective tissue of fascia . It can promote the cell proliferation and angiogenesis . It is proved that the physical stimulation changes the expression of the cell gene and provides the possibility for its support and reserve . However , it is not possible to detect the stem cell related factor Nanog , OCT - 4 gene expression . Therefore , it is not possible to confirm the function of the stem cell related factor Nanog , OCT - 4 gene expression . Therefore , it is necessary to continue to study the expression of the stem cell related factor Nanog , OCT - 4 gene .

【學(xué)位授予單位】：南方醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2008
【分類號】：R329

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1 楊林林;機(jī)械牽拉刺激對非特異性筋膜結(jié)締組織細(xì)胞活性影響的研究[D];南方醫(yī)科大學(xué);2008年

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