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. 2023 May 23;85(6):2689-2694.
doi: 10.1097/MS9.0000000000000900. eCollection 2023 Jun.

The optimum oxygen level in hypoxic culture conditions of ligament derived stem cells: experimental research

Sholahuddin Rhatomy  1 , Dwikora N Utomo  2 , Cita R S Prakoeswa  3 , Heri Suroto  2 , Damayanti Tinduh  4 , Hari B Notobroto  5 , Nur Arfian  6 , Fedik A Rantam  7   8 , Ferdiansyah Mahyudin  2
Affiliations

The optimum oxygen level in hypoxic culture conditions of ligament derived stem cells: experimental research

Sholahuddin Rhatomy et al. Ann Med Surg (Lond). .

Abstract

The hypoxic condition is a physiological norm for various stem cells. The natural microenvironment contains lower oxygen pressures. Recent studies reported significant increases in the cultured cells' proliferation in the presence of a low oxygen pressure.

Objective: This study aimed to investigate the optimum oxygen level for rabbit cruciate ligament fibroblast cells culture and Ligament Derived Conditioned Medium/LD-CM (Secretome) preparation in vitro.

Materials and methods: Fibroblasts were isolated from the cruciate ligament of the rabbit's knee. Cultured of rabbit cruciate ligament Fibroblast Cells (fifth passage) were assigned to the slight (5% O2), middle (3% O2), and severe hypoxia (1% O2) groups and the normoxia (21% O2) group. Measurement of growth factors: TGF-β1, PDGF, FGF, and VEGF in LD-CM (Secretome) used an enzyme-linked immunosorbent assay.

Results: The highest number of cultured cells were in the 5% O2 group compared to the normoxia, 1 and 3% groups. The hypoxia 5% group also had increased productions of PDGF, FGF, and VEGF proteins in LD-CM (secretome) compared to the 1, 3%, and normoxia groups. TGF-β1 production was slightly higher in the 3 group than the 5% group.

Conclusion: The hypoxic precondition of 5% oxygen was the optimum condition for ligament culture and ligament derived conditioned medium (secretome) preparation in vitro.

Keywords: hypoxia; ligament derived conditioned medium; normoxia; proliferation; secretome.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Morphology of rabbit Cruciate Ligament Fibroblast Cells (rCLFCs); Inverted microscopy showed flat fusiform or dendroid shapes that were arranged in a monolayer with normal cell spacing; original magnification: ×100.
Figure 2
Figure 2
Cell number per group; data presented as mean±SD.
Figure 3
Figure 3
TGF β1 measurement results.
Figure 4
Figure 4
PDGF measurement results.
Figure 5
Figure 5
FGF measurement results.
Figure 6
Figure 6
VEGF measurement results.
See this image and copyright information in PMC

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