Comparison of the effects between animal-derived trypsin and recombinant trypsin on human skin cells proliferation, gene and protein expression

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• 作者：Maarof Manira (1)
  Khairoji Khairul Anuar (1)
  Wan Tai Seet (1)
  Abd Wahab Ahmad Irfan (1)
  Min Hwei Ng (1)
  Kien Hui Chua (1) (2)
  Mohd. Yunus Mohd Heikal (1) (2)
  Bin Saim Aminuddin (1) (3)
  Bt Hj Idrus Ruszymah (1) (2)
  
• 关键词：Keratinocytes ; Fibroblasts ; Animal ; derived trypsin ; Recombinant trypsin ; Tissue engineering
• 刊名：Cell and Tissue Banking
• 出版年：2014
• 出版时间：March 2014
• 年：2014
• 卷：15
• 期：1
• 页码：41-49
• 全文大小：2,411 KB
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• 作者单位：Maarof Manira (1)
  Khairoji Khairul Anuar (1)
  Wan Tai Seet (1)
  Abd Wahab Ahmad Irfan (1)
  Min Hwei Ng (1)
  Kien Hui Chua (1) (2)
  Mohd. Yunus Mohd Heikal (1) (2)
  Bin Saim Aminuddin (1) (3)
  Bt Hj Idrus Ruszymah (1) (2)
  
  1. Tissue Engineering Centre, Universiti Kebangsaan Malaysia Medical Centre, Jalan Yaacob Latiff, Bandar Tun Razak, 56000, Cheras, Kuala Lumpur, Malaysia
  2. Department of Physiology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Bangi, Malaysia
  3. Ear Nose and Throat Consultant Clinic, Ampang Puteri Specialist Hospital, Selangor, Malaysia
  
• ISSN：1573-6814

文摘

Animal-derivative free reagents are preferred in skin cell culture for clinical applications. The aim of this study was to compare the performance and effects between animal-derived trypsin and recombinant trypsin for skin cells culture and expansion. Full thickness human skin was digested in 0.6?% collagenase for 6?h to liberate the fibroblasts, followed by treatment with either animal-derived trypsin; Trypsin EDTA (TE) or recombinant trypsin; TrypLE Select (TS) to liberate the keratinocytes. Both keratinocytes and fibroblasts were then culture-expanded until passage 2. Trypsinization for both cell types during culture-expansion was performed using either TE or TS. Total cells yield was determined using a haemocytometer. Expression of collagen type I, collagen type III (Col-III), cytokeratin 10, and cytokeratin 14 genes were quantified via RT-PCR and further confirmed with immunocytochemical staining. The results of our study showed that the total cell yield for both keratinocytes and fibroblasts treated with TE or TS were comparable. RT-PCR showed that expression of skin-specific genes except Col-III was higher in the TS treated group compared to that in the TE group. Expression of proteins specific to the two cell types were confirmed by immunocytochemical staining in both TE and TS groups. In conclusion, the performance of the recombinant trypsin is comparable with the well-established animal-derived trypsin for human skin cell culture expansion in terms of cell yield and expression of specific cellular markers.