Sains Malaysiana 49(2)(2020): 429-437

http://dx.doi.org/10.17576/jsm-2020-4902-22

Nasal Fibroblast Conditioned Medium Promotes Cell Attachment and Migration of Human Respiratory Epithelium

(Medium Terkondisi Fibroblas Hidung Menggalakkan Perlekatan dan Migrasi Sel Epitelium Pernafasan)

ROHAINA CHE MAN¹*, YOGESWARAN LOKANATHAN¹, RABIATUL ADAWIYAH RAZALI², SHIPLU ROY CHOWDURY¹, AMINUDDIN BIN SAIM³ & RUSZYMAH BINTI HJ IDRUS²

¹Tissue Engineering Centre, Faculty of Medicine, Universiti Kebangsaan Malaysia Medical Centre, Jalan Yaacob Latif, Bandar Tun Razak, 56000 Kuala Lumpur, Federal Territory, Malaysia

²Department of Physiology, Faculty of Medicine, Universiti Kebangsaan Malaysia Medical Centre, Jalan Yaacob Latif, Bandar Tun Razak, 56000 Kuala Lumpur, Federal Territory, Malaysia

³Ear, Nose & Throat Consultant Clinic, Ampang Puteri Specialist Hospital, 68000 Ampang, Selangor Darul Ehsan, Malaysia

Received: 17 March 2019/Accepted: 30 October 2019

ABSTRACT

Endoscopic sinus surgery (ESS) is a well-known surgical treatment for chronic rhinosinusitis disease after failed medical and antibiotics treatment. However, improper wound healing might induce synechiae or adhesion. Conditioned medium from cultured cells is known to promote wound healing and potentially able to accelerate wound healing in ESS and other airway epithelial injuries. This study was to investigate the effect of human nasal fibroblast conditioned medium on the attachment, proliferation and migration of respiratory epithelial cells (RECs) in an in vitro model. RECs and fibroblasts were co-cultured in Defined Keratinocytes Medium and F-12 and Dulbecco's Modified Eagle's Medium. Once confluent, the fibroblasts were removed, leaving the colonies of RECs to reach confluency. RECs and fibroblasts were cultured separately and the conditioned medium was acquired by culturing fibroblast either in DKSFM or F12: DMEM, denoted as NFCM_DKSFM and NFCM_FD, respectively. RECs were supplemented with 20% conditioned medium for attachment, proliferation and migration assay. The results showed significantly higher cell attachment in NFCM_DKSFM (3452.77±588.1 cell/cm²) compared to NFCM_FD (2336.1±440.4 cell/cm²) and DKSFM alone (2819.8±509.5 cell/cm²). After 7 days, the specific growth rate was higher in DKSFM (0.019±5.16×10^-4 h^-1) compared to NFCM_DKSFM (0.015±8.94×10^-4 h^-1) and NFCM_FD (0.013±1.03×10^-3 h^-1). The mean of migration rate was significantly higher in NFCM_DKSFM (4341.81±385.7 µm²/hr) compared to NFCM_FD (1803.38±408.1 µm²/hr) and DKSFM (1933.48±271.9 µm²/hr). Hence, NFCM_DKSFM supplementation provides suitable culture conditions for RECs through increased cell attachment and migration, which suggest that the factors secreted in conditioned medium may play a major role in enhancing airway epithelial wound healing.

Keywords: Conditioned culture media; epithelial cells; fibroblast; wound healing

ABSTRAK

Pembedahan sinus endoskopi (ESS) adalah rawatan yang terkenal untuk penyakit rhinosinusitis kronik selepas kegagalan rawatan perubatan dan antibiotik. Walau bagaimanapun, penyembuhan luka yang tidak sempurna boleh menyebabkan synechiae atau lekatan. Medium terkondisi (MT) daripada sel yang dikultur didapati boleh menggalakkan penyembuhan luka dan berpotensi mempercepatkan penyembuhan luka disebabkan oleh ESS dan kecederaan lain pada epitelium saluran pernafasan. Kajian ini adalah untuk mengkaji kesan MT daripada fibroblas hidung manusia untuk perlekatan, percambahan dan migrasi sel epitelium pernafasan (RECs) dalam model in vitro. RECs dan fibroblas telah dibiakkan dalam Defined Keratinocytes Medium (DKSFM) dan F-12 danDulbecco's Modified Eagle's Medium (FD). Selepas konfluen, fibroblas ditanggalkan dan koloni RECs terus dikultur sehingga padat. RECs dan fibroblas dibiakkan secara berasingan dan MT diperoleh dengan mengkultur fibroblas sama ada dalam DKSFM atau FD, dengan dilabelkan sebagai NFCM_DKSFM dan NFCM_FD. RECs telah ditambah dengan 20% medium terkondisi untuk perlekatan, percambahan dan migrasi sel. Keputusan menunjukkan perlekatan sel yang lebih tinggi dalam NFCM_DKSFM (3452.77±588.1 sel/cm²) berbanding NFCM_FD (2336.1±440.4 sel/cm²) dan DKSFM sahaja (2819.8±509.5 sel/cm²). Selepas 7 hari dikultur, kadar pertumbuhan khusus adalah lebih tinggi dalam DKSFM (0.019±5.16×10^-4/jam) berbanding NFCM_DKSFM (0.015±8.94×10^-4/jam) dan NFCM_FD (0.013±1.03×10^-3/jam). Purata kadar migrasi sel jauh lebih tinggi dalam NFCM_DKSFM (4341.81±385.7 μm²/jam) berbanding NFCM_FD (1803.38±408.1 μm²/jam) dan DKSFM (1933.48±271.9 μm²/jam). Oleh itu, penambahan NFCM_DKSFM menyediakan persekitaran yang sesuai untuk RECs melalui peningkatan perlekatan dan migrasi sel, yang menunjukkan bahawa faktor yang dirembeskan dalam MT boleh memainkan peranan utama dalam meningkatkan penyembuhan luka pada epitelium saluran pernafasan.

Kata kunci: Fibroblas; medium kultur terkondisi; penyembuhan luka; sel epitelium

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*Corresponding author; email: ruszyidrus@gmail.com