Browsing by Author "Huang, G. Steve"

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A nanodevice for rapid modulation of proliferation, apoptosis, invasive ability, and cytoskeletal reorganization in cultured cells
(2010)
Hung, Yao-Ching
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Pan, Hsu-An
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Tai, Shih-Ming
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Huang, G. Steve
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材料科學與工程學系奈米科技碩博班
We have fabricated a nanodevice composed of a matrix of nine nanodot arrays with various dot sizes, ranging from a flat surface to 10 nm, 50 nm, 100 nm, and 200 nm arrays. HELA, C33A, ES2, PA-1, TOV-112D, TOV-21G, MG63, and NIH-3T3 cells were seeded onto the device and cultured for three days. To evaluate the size-dependent effect of nanodot arrays on cell growth, indices corresponding to cell proliferation, apoptosis, cell adhesion, and cytoskeletal organization were defined. VD(50) is defined as the diameter of nanodots on which 50% of the cell population remains viable. AD(50) is defined as the diameter of nanodots on which 50% of the cell population appears to have an apoptosis-like morphology. FD(50) is the diameter of nanodots that promotes the formation of 50% of the focal adhesions compared to cells grown on a flat surface. CD(50) is defined as the diameter of nanodots on which cells have half the amount of microfilament bundles compared to cells grown on a flat surface. We were able to distinguish between the invasive ability of HELA versus later-staged C33A cells. Ovarian cancer cell lines (ES2, PA-1, TOV-112D, and TOV-21G) also exhibited differential growth parameters that are associated with cell type, grade, and stage. Modulation of the growth of MG63 cells was also achieved. More broadly, we have established a platform that can be used to assess basic parameters of cell growth. A simplified fabrication process ensures mass production and lowers cost. According to our results, the device is capable of distinguishing among cancer cell lines at various stages and also provides basic design parameters for artificial implants. Our device will serve as a convenient and fast tool for tissue engineering and cancer treatment.
A Nanodot Array Modulates Cell Adhesion and Induces an Apoptosis-Like Abnormality in NIH-3T3 Cells
(2009-08-01)
Pan, Hsu-An
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Hung, Yao-Ching
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Su, Chia-Wei
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Tai, Shih-Ming
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Chen, Chiun-Hsun
Micro-structures that mimic the extracellular substratum promote cell growth and differentiation, while the cellular reaction to a nanostructure is poorly defined. To evaluate the cellular response to a nanoscaled surface, NIH 3T3 cells were grown on nanodot arrays with dot diameters ranging from 10 to 200 nm. The nanodot arrays were fabricated by AAO processing on TaN-coated wafers. A thin layer of platinum, 5 nm in thickness, was sputtered onto the structure to improve biocompatibility. The cells grew normally on the 10-nm array and on flat surfaces. However, 50-nm, 100-nm, and 200-nm nanodot arrays induced apoptosis-like events. Abnormality was triggered after as few as 24 h of incubation on a 200-nm dot array. For cells grown on the 50-nm array, the abnormality started after 72 h of incubation. The number of filopodia extended from the cell bodies was lower for the abnormal cells. Immunostaining using antibodies against vinculin and actin filament was performed. Both the number of focal adhesions and the amount of cytoskeleton were decreased in cells grown on the 100-nm and 200-nm arrays. Pre-coatings of fibronectin (FN) or type I collagen promoted cellular anchorage and prevented the nanotopography-induced programed cell death. In summary, nanotopography, in the form of nanodot arrays, induced an apoptosis-like abnormality for cultured NIH 3T3 cells. The occurrence of the abnormality was mediated by the formation of focal adhesions.
Assessment of the In Vivo Toxicity of Gold Nanoparticles
(2009-08-01)
Chen, Yu-Shiun
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Hung, Yao-Ching
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Liau, Ian
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Huang, G. Steve
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材料科學與工程學系
The environmental impact of nanoparticles is evident; however, their toxicity due to their nanosize is rarely discussed. Gold nanoparticles (GNPs) may serve as a promising model to address the size-dependent biological response to nanoparticles because they show good biocompatibility and their size can be controlled with great precision during their chemical synthesis. Naked GNPs ranging from 3 to 100 nm were injected intraperitoneally into BALB/C mice at a dose of 8 mg/kg/week. GNPs of 3, 5, 50, and 100 nm did not show harmful effects; however, GNPs ranging from 8 to 37 nm induced severe sickness in mice. Mice injected with GNPs in this range showed fatigue, loss of appetite, change of fur color, and weight loss. Starting from day 14, mice in this group exhibited a camel-like back and crooked spine. The majority of mice in these groups died within 21 days. Injection of 5 and 3 nm GNPs, however, did not induce sickness or lethality in mice. Pathological examination of the major organs of the mice in the diseased groups indicated an increase of Kupffer cells in the liver, loss of structural integrity in the lungs, and diffusion of white pulp in the spleen. The pathological abnormality was associated with the presence of gold particles at the diseased sites, which were verified by ex vivo Coherent anti-Stoke Raman scattering microscopy. Modifying the surface of the GNPs by incorporating immunogenic peptides ameliorated their toxicity. This reduction in the toxicity is associated with an increase in the ability to induce antibody response. The toxicity of GNPs may be a fundamental determinant of the environmental toxicity of nanoparticles.
Control of In Vivo Transport and Toxicity of Nanoparticles by Tea Melanin
(2012)
Chen, Yu-Shiun
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Hung, Yao-Ching
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Hong, Meng-Yeng
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Onischuk, Andrei Aleksandrovich
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Chiou, Jin Chern
"Nanoparticles are unfamiliar to researchers in toxicology. Toxicity may be generated simply due to the reduction in size. Compounds that prevent or cure toxic materials may not work on nanoparticles. Furthermore, as there are more and more applications of nanoparticles in drug delivery and in vivo imaging, controlling the transport and toxicity will be primary concerns for medical application of nanoparticles. Gold nanoparticles (GNPs) if injected intraperitoneally into mice can enter hippocampus and induce cognitive impairment. GNPs caused a global imbalance of monoamine levels, specifically affecting the dopaminergic and serotonergic neurons. Pretreatment of tea melanin significantly prevented the deposition of GNPs in mouse brains, especially in the hippocampus. Pretreatment of melanin completely alleviated GNP-induced impairment of cognition. Pre-administration of melanin stably maintained monoamines at normal profiles. Melanin completely prevented the invasion of GNPs into the Cornu Ammonis region of the hippocampus shown by coherent anti-Stoke Raman scattering microscopy. Here we show that the administration of tea melanin prevented the accumulation of Au in brain, the imbalance of monoamines, and the impairment of cognition in mice. The current study provides a therapeutic approach to toxicity of nanoparticles and a novel strategy to control the transport of GNP in mouse brain."
Gold nanoparticles caused learning impairment in mice
(2009)
Chen, Yu-Shiun
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Hung, Yao-Ching
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Liau, Ian
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Lin, Li-Wei
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Hong, Meng-Yeng
Aims : To investigate the toxic effect of gold nanoparticles (GNPs) to the learning and memory of mice. Main methods: Naked GNPs of 17 nm and 37 nm were synthesized, purified, and injected intraperitoneally into BALB/C mice at a dose of 8 mg/kg/week. Passive avoidance test was performed to detect the possible leaning impairment. Biochemical analysis to examine levels of monoamine and acetylcholine in mouse brain was also performed. ICP-MS was performed to examine the presence of GNP in brain. Localization of GNP at hippocampus was surveyed by ex vivo Coherent anti-Stoke Raman scattering (CARS) microscopy. Transmission electron microscopy (TEM) was also performed to verify the cellular location of GNPs at hippocampus. Key findings: Both 17 nm and 37 nm GNPs induced severe sickness in rnice. However, only 17 rim GNP impaired the learning and memory of mice. GNP treatment elevated levels of dopamine from 114.5 ng/mouse brain to 143.6 ng/mouse brain for 17 nm (p<0.01) and to 138.2 for 37 nm GNP (p<0.05). Serotinin was significantly reduced by 17 nm GNP treatment from 57.2 ng/mouse brain to 44.3 ng/mouse brain (p<0.05). ICP-MS indicated the presence of GNPs in every part of the brain. CARS microscopy showed that 17 nm GNP was located at the Cornu Amonis regions of hippocampus where neuronal cells clustered, while 37 nm GNP was excluded from the cell clustered region. TEM and EDAX verified the presence of 17 nm GNP in the cytoplasm and in the dendrite of pyramidal cell, while 37 rim GNP was found in the extracellular region of neuronal cells. TEM image also indicated that both endocytosis and free diffusion coexisted for the invasion of 17 nm GNP. Significance: The current study provided evidence that nanoparticles were capable of entering into brain and affecting normal brain function. The size-dependent invading ability of GNPs provides an extra dimension for drug delivery.
Size-dependent impairment of cognition in mice caused by the injection of gold nanoparticles
(2010-12-03)
Chen, Yu-Shiun
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Hung, Yao-Ching
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Lin, Li-Wei
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Liau, Ian
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Hong, Meng-Yeng
We explored the size-dependent impairment of cognition in mice caused by the injection of gold nanoparticles (GNPs). GNPs of 17 and 37 nm in diameter were injected intraperitoneally into BALB/c mice at doses ranging from 0.5 to 14.6 mg kg(-1). ICP-MS was performed on brain tissue collected 1, 14 and 21 days after the injection. A passive-avoidance test was performed on day 21. Monoamine levels were determined on day 21. The microscopic distribution of GNPs in the hippocampus was examined using coherent anti-Stokes Raman scattering (CARS) microscopy and transmission electron microscopy (TEM). The results indicated that 17 nm GNPs passed through the blood-brain barrier more rapidly than 37 nm GNPs. Treatment with 17 nm GNPs decreased the latency time, which was comparable to the effect of scopolamine treatment, while 37 nm GNPs showed no significant effect. Dopamine levels and serotonin levels in the brain were significantly altered by the injection of 17 and 37 nm GNPs. GNPs affected dopaminergic and serotonergic neurons. CARS microscopy indicated that 17 nm GNPs entered the Cornu Ammonis (CA) region of the hippocampus, while 37 nm GNPs were excluded from the CA region. TEM verified the presence of 17 nm GNPs in the cytoplasm of pyramidal cells. In this study, we showed that the ability of GNPs to damage cognition in mice was size-dependent and associated with the ability of the particles to invade the hippocampus. The dosage and duration of the treatment should be taken into account if GNPs are used in the future as vehicles to carry therapeutic agents into the brain.
Topographic control of the growth and function of cardiomyoblast H9c2 cells using nanodot arrays
(2012-01-01)
Pan, Hsu-An
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Hung, Yao-Ching
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Sui, Yu-Ping
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Huang, G. Steve
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材料科學與工程學系
Cardiovascular stents require optimised control for the enhancement or inhibition epithelial and smooth muscle cell growth in close contact with the implant. Here we propose that the surface topology in contact with the living cells could be designed to control and optimise the growth and function of such cells. The cardiomyoblast H9c2 was cultured on nanodot arrays with dot diameters ranging between 10 and 200 nm. On the 50-nm nanodot arrays H9c2 showed maximum attachment and proliferation with largest cell area and extended lamellipodia. In contrast, 53.7% and 72.6% reductions of growth were observed on the 100- and 200-nm nanodot arrays after 3 days. Immunostaining indicated that nanodots smaller than 50-nm induced cell adhesion and cytoskeleton organization. Expression of genes associated with fibrosis and hypertrophy was up-regulated in cells grown on 100-nm nanodots. Western blot data showed high levels of expression for vinculin and plasminogen activator inhibitor-1 for cells cultured on 50-nm nanodots. Nanotopography controls cell adhesion, morphology and proliferation. By adjusting the diameter of the nanodots, we could modulate the growth and expression of function-related genes and proteins of H9c2 cardiomyoblasts. The current study provides insights for improved design of artificial implants and parameters that affect biocompatibility. (C) 2011 Elsevier Ltd. All rights reserved.