Abstract
Dermal fibroblasts (DF) constitute one of key cells involved in wound healing. However, the functions they perform in wound conditions remain poorly understood. This study involved exposing DF to low nutrition and to low nutrition + LPS for 5 d as conditions representing the wound. Although DF exhibited increasing metabolic activity in time under all conditions including control, the proliferation did not change in both low nutrition and low nutrition + LPS. Only the low nutrition + LPS was found to potentiate the migration and pro-inflammatory phenotype (IL6 release) of DF. The potential of DF to contract collagen hydrogel declined only under low nutrition as a consequence of low cell number. The expression of α-SMA was reduced under both conditions independently of the cell number. The remodeling capability of DF was affected under both conditions as documented by the enhanced MMP2 activity. Finally, the production of collagen type I was not affected by either condition. The study shows that low nutrition as the single factor is able to delay the healing process. Moreover, the addition of the mild pro-inflammatory stimulus represented by LPS may amplify the cell response in case of decreased α-SMA expression or excite DF to produce IL6 impairing the healing process.
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Acknowledgements
The authors acknowledge the invaluable assistance provided by Iveta Paurova via her support in terms of the provision of laboratory services.
Funding
The study was supported by project no. CZ.02.1.01/0.0/0.0/16_019/0000787 “Fighting Infectious Diseases,” awarded by the Ministry of Education, Youth and Sports of the Czech Republic and financed from the European Regional Development Fund; by project no. NU20-02–00368 awarded by Czech Health Research Council and by the Ministry of Health of the Czech Republic, by project GA UK No. 128417 awarded by the Charles University; and financed from the European Regional Development Fund and by the Cooperation Program, research area MED/DIAG.
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Anna Zavadakova and Lucie Vistejnova designed the experiments and prepared the figures. Anna Zavadakova and Pavla Tonarova performed the experiments and collected the data. Data analysis was performed by all authors. The first draft of the manuscript was written by Anna Zavadakova and all authors commented on previous versions of the manuscript. All authors have read and approved the final manuscript.
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The authors declare no competing interests. The funders had no role in the design of the study; in the collection, analysis or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.
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11626_2022_713_MOESM1_ESM.jpg
Supplementary file1 Fig. S1 Representative images of collagen hydrogels seeded by dermal fibroblasts and monitored for contraction under control, low nutrition and low nutrition + LPS conditions for 13 days.Suppl. (JPG 544 KB)
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Supplementary file2 Fig. S2 Images of whole wells of 96-well test plates with dermal fibroblasts cultured under control, low nutrition and low nutrition + LPS conditions for 5 days. With respect to the quantification of the α-SMA, nine sequential images covering the whole well area were captured by means of an Olympus UPlanFL N 4x/0.13 objective followed by automatic whole well area picture decomposition using VisiView software. The number of nuclei and the area of the α-SMA were determined from the same well by means of ImageJ Fiji (National Institute of Health, Bethseda, USA). The results were expressed as the mean of the α-SMA area at day 5 normalized to the cell count at day 5 ± standard deviation from 8 independent NHDF donors. (JPG 2652 KB)
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Zavadakova, A., Vistejnova, L. & Tonarova, P. Functional responses of dermal fibroblasts to low nutrition and pro-inflammatory stimuli mimicking a wound environment in vitro. In Vitro Cell.Dev.Biol.-Animal 58, 643–657 (2022). https://doi.org/10.1007/s11626-022-00713-7
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DOI: https://doi.org/10.1007/s11626-022-00713-7