An optical detection technique is developed to detect and measure the resonant vibration of the cellular membrane. Biological membranes are active components of living cells and play a complex and dynamic role in life processes. They are believed to have oscillation modes of frequencies in the range of 1 to 1000 GHz. To measure such a high-frequency vibration, a linear laser cavity is designed to produce a train of femtosecond pulses of adjustable repetition rate. The method is then directly applied to liposomes, "artificial membrane", stained with a liphophilic potential sensitive dye. The spectral behavior of a selection of potential sensitive dyes in the membrane is also studied.

Thesis (B.S.)--Massachusetts Institute of Technology, Dept. of Electrical Engineering and Computer Science, 1982. / MICROFICHE COPY AVAILABLE IN ARCHIVES AND ENGINEERING / Includes bibliographical references. / by Stephen F. Bart. / B.S.

Cell wounding, the loss of plasma membrane integrity, is a common event in the life of many cell types. Most cells are subjected to physiological events during normal functions that can lead to disruption of their plasma membranes, especially cells in the load bearing organs such as muscle, skin and bone. The capacity of the cell to repair day-to-day wear-and-tear injuries, as well as traumatic ones, is fundamental for maintaining tissue integrity. / In this thesis, we were trying to uncover single cell wound responses by applying the femtosecond laser (fs laser) technology. A well-characterized tunable fs laser was coupled with a laser scanning confocal microscopic system. Combining real-time observations of the fs laser-induced wound and 3D reconstruction of the cells, post-damage cell and nucleus morphological transformation and wound dynamics were reported. The major findings of this study include: (1) Fs laser could induce a small hole on the plasma membrane of the targeted cell. With the same laser irradiation time, the initial hole size were positively correlated with the laser power. (2) Four typical hole evolution scenarios were reported. Hole resealing was a fast process mostly within 100 seconds in normal condition. Whether a cell could reseal the hole is dependent on the initial hole size. Cells had difficulty to cope with the bigger holes. Three ranges of hole size were given in the thesis to predict the hole resealing result. (3) After fs laser damage, the whole cell underwent a contraction. The post-damage nucleus area, footprint, and each section layer of the cell all shrank, only the thickness remained the same. The nucleus retreated a bit from the damage site after damage. (4) Oxidative stress altered some of the cellular responses to the laser damage. The fs laser- induced holes in oxidative groups were bigger than the normal condition. The cells underwent an overall swelling after fs laser damage instead of the contraction in the normal group. Section layer areas and the thickness of the cell increased after damage. But similar to the normal condition, nucleus shrinkage and retreat from the damage site were also found in the oxidative stress groups. (5) Although both acute and chronic oxidative stresses compromised the integrity of the plasma membrane, chronic oxidative stress compromised more severely with several critical post-damage cell transformations and low resealing ratio. Acute oxidative stress on the other hand may somehow promote the resealing ability of the cells. (6) The section layers closer to the bottom of the cells transformed less than the layers further away from the bottom. This probably suggested that the cell basal attachment provided a constraint force to the plasma membrane for morphological changes. / 細胞創傷，即細胞膜的完整性受損，是多種細胞生命週期中一種常見的現象。細胞在執行正常功能時可能遭遇不同程度的生理性損傷，其中大部分會導致細胞膜的破壞。這一現象對存在於承受壓力器官中的細胞更為頻繁，例如肌肉，皮膚和骨骼。細胞對於日常磨損性傷害以及意外創傷的修復能力，是維持組織完整性的基石。 / 在本論文中，我們通過使用飛秒鐳射技術模擬單細胞創傷，觀察並試圖揭示單細胞對於創傷的反應過程。在實驗中，參數可調的飛秒鐳射器與共軛聚焦顯微鏡整合為一個系統，用於在單細胞膜上進行定點損傷。我們結合了對損傷的即時觀測，細胞的三維結構重建技術，完整記錄了損傷前後的損傷部位，細胞整體以及細胞核的形態變化。以下是本研究的主要發現：（1）飛秒鐳射能夠在目標細胞的細胞膜上進行局部穿孔。在鐳射照射時間相同的情況下，鐳射穿孔的大小與鐳射的平均功率呈正相關。（2）我們發現了鐳射穿孔後穿孔部位有四種不同的變化情況。穿孔後細胞封孔是一相當快的過程，在細胞成功封孔的情況下，大部分細胞將在100秒以內將穿孔部位重新填滿。細胞是否可以將穿孔封住取決於鐳射照射後初始穿孔的大小。細胞很難修復較大的孔。我們將細胞初始穿孔大小分為三個範圍，根據這三個範圍可以利用初始孔的尺寸大致預測穿孔後細胞的封孔情況。（3）飛秒鐳射損傷細胞後，細胞將會收縮，並且細胞核的平面面積，細胞的平面面積（或稱細胞足跡），以及細胞各分層面積都有不同程度的縮小。僅細胞厚度未發生顯著變化。同時，細胞核的位置相對於損傷部位有所後退。（4）細胞在氧化應激過後，對於飛秒鐳射造成的損傷反應有所變化。具體表現為：鐳射穿孔的尺寸比正常情況下更大；穿孔後細胞將會整體腫脹而非收縮。各分層面積和細胞厚度都有不同程度的增大。但是細胞核的反應與正常情況類似，即細胞核將會收縮，並且後退以遠離鐳射損傷部位。（5）儘管急性氧化應激和慢性氧化應激都一定程度上損傷了細胞膜的完整性，但是從細胞對於鐳射創傷的反應觀察，長期慢性氧化應激對於細胞膜的損害更為嚴重，具體表現為鐳射損傷後細胞的嚴重形變和細胞膜修復比例的降低。而另一方面，急性氧化應激在某種程度上可以增強細胞對於鐳射穿孔的修復能力。（6）細胞膜穿孔後，細胞各層的面積變化不一，位置越靠近底層的分層面積變化越小。這可能表明細胞貼壁行為形成了一個對細胞形變的約束力。 / Duan, Xinxing. / Thesis M.Phil. Chinese University of Hong Kong 2014. / Includes bibliographical references (leaves 73-85). / Abstracts also in Chinese. / Title from PDF title page (viewed on 24, October, 2016). / Detailed summary in vernacular field only. / Detailed summary in vernacular field only.

Traumatic brain injury (TBI) is a major health and socioeconomic concern in the United States and across the globe. Experimental models of TBI are used to study the mechanisms underlying cell dysfunction and death that result from injury, the functional deficits that result from injury, and the potential of various therapies to treat injury. This thesis explores the fundamental mechanical damage associated with brain trauma, investigating the effects of mechanical deformation on neurons at the molecular, cellular, tissue, and animal levels. First, a novel hydrogel system was developed to support 3-D neuronal cultures, and the cultures were studied in an in vitro model of neuronal injury. The dependence of cell viability on hydrogel stiffness and extracellular matrix ligand concentration revealed a role for molecular interactions in the cellular response to injury. Subsequently, in a rat model of TBI neuronal plasma membrane damage was observed coincidentally with cell death within the hippocampus; however not all permeable cells died, suggesting a complex role for plasma membrane damage in neuronal degeneration. The spatial profile of permeable cells in the hippocampus reveals further heterogeneity of neuronal plasma membrane damage, with populations of cells in certain hippocampal subregions exhibiting an increased vulnerability to plasma membrane damage. These observations support recent finite element model predictions of strains in the brain during injury. Finally a system for measuring locomotor disturbances is used for the first time following brain injury. Continued investigation of how neurons deform and fail mechanically will contribute to the understanding of the pathophysiology of brain injury and may help identify potential therapeutic targets.

Although dynamic interactions between plant cells and their environment require adhesion between the cell wall (CW) and the plasma membrane (PM), few plant adhesion molecules have been identified. Therefore, the seed coat mucilage secretory cells (MSCs) of Arabidopsis thaliana (which undergo developmentally regulated changes in adhesion) were developed into a novel model system to study PM-CW adhesion. Twenty-seven candidate genes were identified using data from publicly available and seed-specific microarrays. Mutant plants for these genes were screened for defects in adhesion via plasmolysis, and for changes in MSC morphology that may result from defective adhesion (Chapter 1). Two fasciclin-like arabinogalactan proteins were isolated in this screen. One of these, SOS5, was characterized in detail (Chapter 2). sos5 mutants are sensitive to hyperosmotic conditions and show defects in PM-CW adhesion and MSC mucilage structure. Interestingly, these phenotypes may be attributed to defects in adhesion or to defects in cell wall deposition.

LPA1 lysophosphatidic acid receptors (LPA1Rs) are normally present on the surface of the cell. Our initial findings were that HMG-CoA reductase inhibitors (atorvastatin and mevastatin) induce the sequestration of the G protein-coupled LPA1R in recycling endosomes, most likely by inhibiting the recycling of tonically internalized receptors. Whereas, co-addition of geranylgeranylpyrophosphate (GGPP) or geranylgeraniol (GGOH) prevented atorvastatin-induced sequestration of LPA1Rs, the geranylgeranyltransferase-I inhibitor, GGTI-298, mimicked atorvastatin and induced LPA1R sequestration. This suggested that statin-induced endosomal sequestration was caused by defective protein prenylation. The likely targets of atorvastatin and GGTI-298 are the Rho family GTPases, RhoC and RhoA, since both inhibitors greatly reduced the abundance of these GTPases and since knockdown of endogenous RhoC or RhoA with small interfering RNAs (siRNAs) led to endosomal sequestration of LPA1R. Knockdown of RhoC was much more potent at inducing endosomal sequestration than knockdown of either RhoA or RhoB. In contrast, atorvastatin, GGTI-298, siRNA against RhoA, B, or C did not alter the internalization or recycling of transferrin receptors, indicating that recycling of transferrin receptors is distinct from LPA1Rs. Thus, these results, for the first time, implicate RhoA and RhoC in endocytic recycling of LPA1Rs and identify atorvastatin and GGTI-298 as novel inhibitors of this process. / Per the request of the author and advisor, and with the approval of the Graduate Education office, the following changes were made to this thesis: Replaced original page 1 with Errata Page 2. Replaced original pages 3-28 with Errata Pages 3 – 16. Replaced original pages 69-71 with Errata pages 17 – 19.

Changes in the plasma membrane phospholipid composition may alter the structure and/or fluidity and lead to a variety of changes in membrane functions. Dietary fats are known to influence the composition of lipids in the plasma membrane. The purpose of this investigation was to compare effects of dietary linoleic and stearic acid on the composition of the phospholipids in the plasma membranes of mammary tumors in A/St mice.Plasma membranes were isolated and lipids were extracted. Phospholipids were separated by thin-layer chromatography and identified by detection with molybdenum blue reagent. The Rf values and integration of optical densities were used to compare phospholipid composition in membranes of tumors from mice fed experimental diets. It appears that the amount of dietary fat, but not the type, affects the phospholipid distributions. The phosphatidylinositol was the phospholipid most affected, representing the smallest amount in membranes from tumors in mice fed the low fat diets. / Department of Biology

Access to abstract permanently restricted. / Aldol condensation to synthesize substituted chalcones -- Synthesis and testing of substituted 4,5-dihydropyrazoles -- Biological testing of synthesized 4,5-dihydropyrazoles. / Access to thesis permanently restricted. / Department of Chemistry

It has been suggested that plasma membranes play a role in tumor production. Changes in plasma membrane lipid composition may change membrane fluidity and disrupt cellular communication. These changes in membrane lipid composition appear to be related to the fatty acid content of the animal's diet. The purpose of this investigation was to compare the effect of dietary linoleic and stearic acids and tumor size on cholesterol/phospholipid ratios and fatty acids in membranes of mammary adenocarcinomas.Plasma membranes of mammary tumors from Strain A/St mice were isolated by centrifugation and the lipids extracted. Phospholipid content was assayed by the method of Bartlett (102); cholesterol and fatty acids by gas liquid chromatography.Plasma membranes of tumors from mice fed the high linoleic diet (SAFF) were found to have the highest cholesterol/phospholipid ratios (Mean=0.396); mice fed high stearic acid diets produced tumors with the lowest ratios (0.280). Membranes of tumors from mice fed SA-4 and Stock diets had intermediate ratios (0.0.341 and 0.0.346, respectively). / Department of Biology

The possibility that changes in membrane transport systems may contribute to the pathophysiology of the uraeraic syndrome has not been extensively studied. This thesis presents a study of eight erythrocyte membrane transport systems, namely the Na/K pump, the amino acid systems y⁺, ASC, gly, L and T, the nucleoside and choline transporters. The results indicate that, compared to normal controls, K⁺ flux through the Na/K pump was reduced in chronic renal failure patients (CRF), on haemodialysis (HD), and on continuous ambulatory peritoneal dialysis (CAPD), but was normal in functional transplant (FT) patients' erythrocytes. The number of Na/K pumps per erythrocyte was decreased in CRF and CAPD but showed no differences between HD, FT and Normal controls. The mean turnover rate per pump site was reduced in patients on HD, whereas other groups were not significantly different from controls. Cross-incubation experiments suggest that the lowered pump flux seen in the HD group was due to plasma factors since reversibility of the defect was achieved when those cells were incubated in normal plasma. The defect was completely reversed with a successful transplant. Erythrocytes from haemodialysis patients exhibited an increased uptake of L-lysine through the y⁺ system. The uptake of L-serine was decreased and the affinity of the ASC system for L-serine was increased in these patients compared with controls. The glycine transporter showed a significant increase in affinity for glycine. The flux of L-leucine and L-tryptophan showed no differences from control cells. Erythrocyte membrane transport of uridine was similar in normal control cells and in those obtained from uraemic patients. Choline influx rates were significantly increased and affinity of the transporter for choline reduced in dialysis patients' erythrocytes. Renal transplant and CRF patients showed variable influx rates which gave a significant negative correlation with creatinine clearance. These results show that there are selective abnormalities in some membrane transport system of the erythrocyte in patients with renal failure. The mechanism and possible significance of these changes are discussed.