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Sains Malaysiana 54(8)(2025): 1985-1994
http://doi.org/10.17576/jsm-2025-5408-09
Evaluating the Efficacy of Cryopreservation Media for
the Preservation and Short-Term Storage of Human Dermal Fibroblast
(Menilai Keberkesanan Media Pengkrioawetan untuk Pemeliharaan dan Penyimpanan Jangka Pendek Fibroblas Dermal Manusia)
TITHTEEYA RATTANACHOT1,
NUR RASYIDAH HAZIMAH MOHD ROSDI1, NUSAIBAH SALLEHUDDIN1,
FAUZI MH BUSRA1,2 & MANIRA MAAROF1,2,3,*
1Department of Tissue Engineering and
Regenerative Medicine, Faculty of Medicine, Universiti Kebangsaan Malaysia, 56000 Cheras, Kuala Lumpur,
Malaysia
2Advance Bioactive Materials-Cells UKM
Research Group, Universiti Kebangsaan Malaysia, 43600 UKM Bangi, Selangor, Malaysia
3Ageing and Degenerative Disease UKM
Research Group, Universiti Kebangsaan Malaysia, 43600 UKM Bangi, Selangor, Malaysia
Received: 20
November 2024/Accepted: 17 June 2025
Abstract
Cryopreservation is a vital process
for long-term preservation of cells without compromising their viability and functionality.
Continuous cell culturing can lead to reduction in cell viability, higher risk
of contamination, and increased reagent consumption. Optimizing short-term
storage is essential to minimize cell damage and enhance cell adaptability for
applications requiring brief storage duration. This study evaluates the effects
of short-term storage on human dermal fibroblasts cryopreserved in different cryopreservation
media. Redundant skin samples were obtained from surgeries with patient
consent, processed, and sub-cultured to passage three (P3). Confluent cells
were trypsinised and cryopreserved in three cryopreservation media: Foetal bovine serum with 10% dimethyl sulfoxide
(FBS+10%DMSO), CryoStor10 (CS10), and cryo freezing
serum-free media (CF-SFM). Cells were stored at -80 °C for 7 days, 14 days, and
1 month. Fibroblasts maintained their spindle-shaped, elongated morphology
across all groups post-storage. The total number of live cells slightly
decreased after 1 month, but no significant differences were found between the
groups. Cell viability in CS10 after 1 month was significantly lower compared
to the other storage durations, while no significant differences were observed
in the other two media groups. Immunocytochemistry showed positive collagen
type I (Col-1) and Ki67 expression at all storage durations. These findings
suggest that fibroblasts retain their characteristics after short-term storage
at -80 °C in different cryopreservation media. However, further studies are
needed to examine the impact of long-term storage on other cell types.
Keywords: Cell characteristics; cryopreservation;
fibroblasts; short-term storage
Abstrak
Pengkrioawetan merupakan proses penting untuk pemeliharaan jangka panjang sel tanpa menjejaskan daya hidup dan fungsinya. Kultur sel berterusan boleh menyebabkan pengurangan daya hidup sel, peningkatan risiko pencemaran dan penggunaan reagen yang tinggi. Pengoptimuman penyimpanan jangka pendek adalah penting untuk mengurangkan kerosakan dan meningkatkan kebolehsuaian sel bagi aplikasi yang memerlukan tempoh penyimpanan yang singkat. Penyelidikan ini menilai kesan penyimpanan jangka pendek terhadap fibroblas dermal manusia yang dikrioawet dalam media krioawetan berbeza. Sampel kulit berlebihan diperoleh daripada pembedahan dengan kebenaran pesakit, diproses dan dikultur sehingga subkultur 3 (P3). Sel yang mencapai konfluensi akan ditripsinkan dan dibekukan dalam tiga jenis media berbeza: serum janin lembu dengan 10% dimetil sulfoksida (FBS+10%DMSO),
CryoStor10 (CS10) dan media bebas serum pembekuan kriogenik (CF-SFM). Sel ini disimpan pada suhu -80 °C selama 7 hari, 14 hari dan 1 bulan. Fibroblas mengekalkan morfologi berbentuk gelendong yang memanjang selepas penyimpanan. Jumlah keseluruhan sel hidup menurun sedikit selepas 1 bulan, namun tiada perbezaan yang ketara ditemui antara kumpulan. Keviabelan sel dalam CS10 selepas 1 bulan adalah lebih rendah berbanding tempoh penyimpanan yang lain, manakala tiada perbezaan diperhatikan dalam dua kumpulan media yang
lain. Analisis imunositokimia menunjukkan ekspresi positif kolagen jenis I (Col-1) dan Ki67 pada semua tempoh penyimpanan. Penemuan ini menunjukkan bahawa fibroblas dapat mengekalkan cirinya selepas penyimpanan jangka pendek pada suhu -80 °C dalam media krioawetan berbeza. Namun, kajian lanjut diperlukan untuk menilai kesan penyimpanan jangka panjang pada jenis sel lain.
Kata kunci: Ciri sel; fibroblas; pengkrioawetan; penyimpanan jangka pendek
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