Effect of selective nutrients in medium on human skin fibroblasts growth and metabolism

Effect of selective nutrients in medium on human skin fibroblasts growth and metabolism

A thorough understanding of cell metabolism and physiology is necessary for medium optimization, where cells can improve their yield and increase their efficiency of medium utilization or minimize the formation of toxic by-products. The objectives of the present study are to investigate the effect o...

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Main Author: Ting, Lee Yu
Format: Thesis
Language:English
Published: 2008
Subjects:
TP Chemical technology
Online Access:http://eprints.utm.my/id/eprint/9646/1/TingLeeYuMFKT2008.pdf
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spelling my-utm-ep.96462018-10-14T07:19:58Z Effect of selective nutrients in medium on human skin fibroblasts growth and metabolism 2008-06 Ting, Lee Yu TP Chemical technology A thorough understanding of cell metabolism and physiology is necessary for medium optimization, where cells can improve their yield and increase their efficiency of medium utilization or minimize the formation of toxic by-products. The objectives of the present study are to investigate the effect of culture conditions on the growth of human skin fibroblasts, and to characterize human skin fibroblasts growth and metabolism. Growth profiles of human skin fibroblasts by using various donor skin biopsies, seeding densities (SD), medium volume to cell growth area ratio (VAR), interval between medium changes (IMC), and way medium changes (WMC) were studied. Experiments were also conducted to determine the consumption or production of glucose, glutamine, amino acid, lactate and ammonia by fibroblasts. Human skin fibroblasts were cultured and used after three passages. Cell proliferation was measured using trypan blue exclusion test and 3-(4,5- dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) assay. Glucose, lactate and glutamine were measured using YSI biochemistry analyzer; amino acids were measured by gas chromatography; and ammonia was determined by enzymatic assay. The results show no significant difference on growth of human skin fibroblasts isolated from different donor skin biopsy. Fibroblasts with higher SD (1×104cell/cm2 and 2×104cell/cm2) have shorter lag phase and population doubling time, and higher saturation density than the lower SD (1×103cell/cm2 and 2×103cell/cm2). Results also shown that fibroblasts cells could grow in VAR between 0.1-1.0ml/cm2. Higher cell proliferation was obtained by fully changing the medium at IMC two days. Conditioned medium tested by WCM did not show any proliferative effect on fibroblasts. Percentage of nutrients consumption was 12.6% for glucose and 14.3% for glutamine; and percentage of metabolite production was 305.7% for lactate and 55.8% for ammonia. The overall apparent yield of lactate from glucose, Y’Lac,Glc (mmol mmol-1) and overall apparent yield of ammonia from glutamine, Y’Amm,Gln (mmol mmol-1), was calculated to be 2.3 and 0.96 respectively. 2008-06 Thesis http://eprints.utm.my/id/eprint/9646/ http://eprints.utm.my/id/eprint/9646/1/TingLeeYuMFKT2008.pdf application/pdf en public http://dms.library.utm.my:8080/vital/access/manager/Repository/vital:1077 masters Universiti Teknologi Malaysia, Faculty of Chemical and Natural Resources Engineering Faculty of Chemical and Natural Resources Engineering
institution Universiti Teknologi Malaysia
collection UTM Institutional Repository
language English
topic TP Chemical technology
spellingShingle TP Chemical technology
Ting, Lee Yu
Effect of selective nutrients in medium on human skin fibroblasts growth and metabolism
description A thorough understanding of cell metabolism and physiology is necessary for medium optimization, where cells can improve their yield and increase their efficiency of medium utilization or minimize the formation of toxic by-products. The objectives of the present study are to investigate the effect of culture conditions on the growth of human skin fibroblasts, and to characterize human skin fibroblasts growth and metabolism. Growth profiles of human skin fibroblasts by using various donor skin biopsies, seeding densities (SD), medium volume to cell growth area ratio (VAR), interval between medium changes (IMC), and way medium changes (WMC) were studied. Experiments were also conducted to determine the consumption or production of glucose, glutamine, amino acid, lactate and ammonia by fibroblasts. Human skin fibroblasts were cultured and used after three passages. Cell proliferation was measured using trypan blue exclusion test and 3-(4,5- dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) assay. Glucose, lactate and glutamine were measured using YSI biochemistry analyzer; amino acids were measured by gas chromatography; and ammonia was determined by enzymatic assay. The results show no significant difference on growth of human skin fibroblasts isolated from different donor skin biopsy. Fibroblasts with higher SD (1×104cell/cm2 and 2×104cell/cm2) have shorter lag phase and population doubling time, and higher saturation density than the lower SD (1×103cell/cm2 and 2×103cell/cm2). Results also shown that fibroblasts cells could grow in VAR between 0.1-1.0ml/cm2. Higher cell proliferation was obtained by fully changing the medium at IMC two days. Conditioned medium tested by WCM did not show any proliferative effect on fibroblasts. Percentage of nutrients consumption was 12.6% for glucose and 14.3% for glutamine; and percentage of metabolite production was 305.7% for lactate and 55.8% for ammonia. The overall apparent yield of lactate from glucose, Y’Lac,Glc (mmol mmol-1) and overall apparent yield of ammonia from glutamine, Y’Amm,Gln (mmol mmol-1), was calculated to be 2.3 and 0.96 respectively.
format Thesis
qualification_level Master's degree
author Ting, Lee Yu
author_facet Ting, Lee Yu
author_sort Ting, Lee Yu
title Effect of selective nutrients in medium on human skin fibroblasts growth and metabolism
title_short Effect of selective nutrients in medium on human skin fibroblasts growth and metabolism
title_full Effect of selective nutrients in medium on human skin fibroblasts growth and metabolism
title_fullStr Effect of selective nutrients in medium on human skin fibroblasts growth and metabolism
title_full_unstemmed Effect of selective nutrients in medium on human skin fibroblasts growth and metabolism
title_sort effect of selective nutrients in medium on human skin fibroblasts growth and metabolism
granting_institution Universiti Teknologi Malaysia, Faculty of Chemical and Natural Resources Engineering
granting_department Faculty of Chemical and Natural Resources Engineering
publishDate 2008
url http://eprints.utm.my/id/eprint/9646/1/TingLeeYuMFKT2008.pdf
_version_ 1747814768553492480

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