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首頁> 外文學(xué)位 >Composite diaphragm inflation: A method for probing the rheological functions of cell-cell anchoring junctions and cytoskeletal networks within a living normal human epidermal keratinocyte sheet.
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Composite diaphragm inflation: A method for probing the rheological functions of cell-cell anchoring junctions and cytoskeletal networks within a living normal human epidermal keratinocyte sheet.

機(jī)譯：復(fù)合隔膜充盈：一種探測正常人表皮角質(zhì)形成細(xì)胞片中細(xì)胞-細(xì)胞錨定連接和細(xì)胞骨架網(wǎng)絡(luò)的流變功能的方法。

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Normal human epidermal keratinocytes (NHEKs), like most all nucleated human cells, possess a filamentous cytoskeleton, composed of actin microfilaments (MFs), microtubules (MTs), and intermediate filaments (IFs). Distinct from connective tissues cells like the dermal fibroblast, however, keratinocytes in vivo are organized as a multicellular epithelium. In the absence of extracellular matrix for mechanical support, keratinocytes express specialized cell-cell anchoring junctions that interconnect MFs and IFs within adjacent cells in the formation of a mesoscopic network cytoarchitecture. Despite advances in our understanding of the proteins that regulate cytoskeletal filament and anchoring junction assembly, the biophysical mechanisms by which these structures provide the epidermis an innate mechanical resilience are, at present, not fully understood.; In this thesis, we validate a new method for the exploration of keratinocyte rheology, referred to as the technique of composite diaphragm inflation (CDI). Sheets of living NHEKs were reconstituted in vitro on tensed but highly compliant, freestanding polydimethylsiloxane (PDMS) elastomer membranes, 5.0 mm in diameter and 10.0 mum thick. NHEK-PDMS composite diaphragm (CD) specimens were then subjected to a series of quasi-static axisymmetric inflation tests to examine the stress response of the epithelial sheet at physiologically severe deformations (∼50% nominal biaxial strains). During these experiments, living NHEK sheets exhibited several unique rheological behaviors, including viscoelasticity, plasticity, and the process of biological adaptation and recovery or a restitutio ad integrum. In addition to a rigorous accounting of the experimental instrumentation and protocol distinct to CDI, a finite elasticity model is proposed for analyzing the mechanics of the associated inflation test. Numerical solution procedures are formulated to predict the quasi-static load-deformation response of a prestretched clamped circular isotropic incompressible hyperelastic membrane inflated into a horizontally semi-infinite incompressible liquid reservoir of finite vertical depth. Assuming a Mooney-Rivlin (MR) constitutive model, we quantitatively demonstrate a new non-traditional regression analysis for estimating values of the MR material parameters and residual membrane tension that best describe a set of experimental inflation response data. Combining improved culture techniques with the more advanced tools of the molecular cell biologist, CDI experiments can potentially transform morphological observations of NHEK cytoarchitecture into well-posed boundary value problems amenable to mechanical experimentation and hypothesis testing.
機(jī)譯：像大多數(shù)所有有核人細(xì)胞一樣，正常人表皮角質(zhì)形成細(xì)胞（NHEK）具有絲狀細(xì)胞骨架，由肌動(dòng)蛋白微絲（MFs），微管（MTs）和中間絲（IFs）組成。與結(jié)締組織細(xì)胞（如真皮成纖維細(xì)胞）不同，但是體內(nèi)的角質(zhì)形成細(xì)胞被組織為多細(xì)胞上皮。在缺乏用于機(jī)械支持的細(xì)胞外基質(zhì)的情況下，角質(zhì)形成細(xì)胞表達(dá)專門的細(xì)胞-細(xì)胞錨定連接，該連接在介觀網(wǎng)絡(luò)細(xì)胞結(jié)構(gòu)的形成中將相鄰細(xì)胞內(nèi)的MF和IF相互連接。盡管我們對調(diào)節(jié)細(xì)胞骨架絲和錨定連接裝配的蛋白質(zhì)的理解有了進(jìn)步，但目前尚不完全了解這些結(jié)構(gòu)為表皮提供先天機(jī)械彈性的生物物理機(jī)制。在本文中，我們驗(yàn)證了一種探索角質(zhì)形成細(xì)胞流變學(xué)的新方法，稱為復(fù)合隔膜擴(kuò)張技術(shù)（CDI）。在張緊但高度順應(yīng)的獨(dú)立式聚二甲基硅氧烷（PDMS）彈性體膜上，將活性NHEK片體外重構(gòu)，該膜直徑為5.0毫米，厚度為10.0微米。然后，對NHEK-PDMS復(fù)合膜片（CD）標(biāo)本進(jìn)行一系列準(zhǔn)靜態(tài)軸對稱膨脹測試，以檢查上皮片在生理性嚴(yán)重變形（約50％名義雙軸應(yīng)變）下的應(yīng)力響應(yīng)。在這些實(shí)驗(yàn)中，活潑的NHEK片材表現(xiàn)出幾種獨(dú)特的流變行為，包括粘彈性，可塑性以及生物適應(yīng)和恢復(fù)或恢復(fù)原狀的過程。除了對CDI以外的實(shí)驗(yàn)儀器和協(xié)議進(jìn)行嚴(yán)格的考慮之外，還提出了有限彈性模型來分析相關(guān)的充氣測試的力學(xué)。制定了數(shù)值求解程序，以預(yù)測預(yù)膨脹的夾緊的圓形各向同性不可壓縮超彈性膜的準(zhǔn)靜態(tài)載荷-變形響應(yīng)，該膜膨脹到水平垂直無限的半無限不可壓縮儲(chǔ)液罐中。假設(shè)Mooney-Rivlin（MR）本構(gòu)模型，我們定量地展示了一種新的非傳統(tǒng)回歸分析，用于估算MR材料參數(shù)和殘余膜張力的值，該分析最能描述一組實(shí)驗(yàn)性充氣反應(yīng)數(shù)據(jù)。將改進(jìn)的培養(yǎng)技術(shù)與分子細(xì)胞生物學(xué)家的更先進(jìn)工具相結(jié)合，CDI實(shí)驗(yàn)可以將NHEK細(xì)胞結(jié)構(gòu)的形態(tài)學(xué)觀察潛在地轉(zhuǎn)化為適合機(jī)械實(shí)驗(yàn)和假設(shè)檢驗(yàn)的恰當(dāng)定位的邊值問題。

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