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Abstract
Purpose
To investigate the biologic effects of topical calf serum on corneal epithelial cells in vitro.
Methods
The effects of calf serum on the corneal epithelial cells were evaluated using the MTT (3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, and the concentration of IL-1α, TGF-β1 and MMP-9 in the cells was measured. Cell damage was determined using lactate dehydrogenase (LDH), and cellular morphologies were examined by transmission electromicroscopy.
Results
Metabolic activity of the corneal epithelial cells decreased at higher concentrations and longer exposure durations. IL-1α, TGF-β1 and MMP-9 titers were lower in calf serum-treated cells than in the control. LDH and cellular damage to the corneal epithelial cells, such as chromatin margination and cytoplasmic organelle swelling, were prominent in cells treated with 30% calf serum.
Conclusions
Cellular metabolic activity was higher and cellular toxicity was lower in cells treated with 10% calf serum compared to those treated with the 20% and 30% concentrations. Furthermore, inflammatory cytokines were sufficiently inhibited in cells treated with the 10% solution. These results indicate that 10% calf serum could be used clinically.
References
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Figure 1.
The absorption rate of the water-insoluble formazan dye in corneal epithelial cell exposed to calf serum by a scanning spectrometer (ELISA reader). Metabolic activity of corneal epithelial cells decreased, at the higher concentration and longer exposure duration.
Figure 2.
LDH titers of cultured corneal epithelial cells exposed to calf serum. The LDH titers have showed that by dose-and time-dependent response relationship. LDH = lactate dehydrogenase.
Figure 3.
IL-1α concentration of cultured corneal epithelial cells after exposed to calf serum. The concentration of IL-1α decreased significantly compared to that in the control after exposed to all the concentrations of calf serum for up to 4 hours.
Figure 4.
TGF-β1 concentration of cultured corneal epithelial cells after exposed to calf serum. The concentration of TGF-β1 was declined significantly after exposed to all the concentrations of calf serum for up to 4 hours. Especially, TGF-β1 was not detected after exposed to 20, 30 % concentration of calf serum.
Figure 5.
MMP-9 concentration of cultured corneal epithelial cells after exposed to calf serum. The concentration of MMP-9 was lower than in the control after exposed to all the concentrations of calf serum for up to 4 hours.
Figure 6.
Transmission electron micrographs of corneal epithelial cells appeared after 4-hour exposure to (A) control, (B) 10% calf serum, (C) 20% calf serum, and (D) 30% calf serum. (bar length 2 um, original magnification, ×2000–4000). In general, the plasma membrane with microvilli (black arrow head), nuclear membrane, and nuclei of corneal epithelial cells were visible. 30% calf serum had more severe and damaged cellular structures, such as the plasma membranes with microvilli being disrupted (white arrow head), well-developed vacuole formation (black arrow), and chromatin margination of the nucleus (white arrow), rather than 10, 20% calf serum.
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