In the present work, we observed that with an EPA/DHA dose of 1.6 and 1.2 g/day respectively, there were significant increases in plasma fatty acids levels for both EPA and DHA from baseline to four weeks for individuals in the active group, which were basically consistent with several previous studies.16
In addition, there were significantly diminished quantities of AA and ratio of total ω-6–ω-3 fatty acids in individuals consuming the EPA/DHA supplement for 4 weeks, which were outcomes also aligned with some earlier research findings.14,29
Collectively, the plasma fatty acid results from the present study validated the use of the supplements by participants in the active group during the study interval and that the supplied dose was adequate to raise plasma fatty acid levels from baseline. There were no significant changes in plasma fatty acid levels for participants in the control group who consumed the placebo, mineral oil, for 4 weeks.
Interestingly, a recent study revealed that when EPA and DHA were provided as supplements to the diet in a total dose as low as 1 g/day in the form of ethyl esters, which are taken up much more slowly than the more rapidly absorbed triacyglycerols from fish, EPA, and DHA plasma levels rose from 0.6% to 1.4% and 2.9% to 4.3% respectively within only 10 days.30 Additionally, both EPA and DHA values returned to near baseline measures within 10 days of discontinuing the supplements. In the current study, the decision to use the chosen EPA/ DHA dose was based on work by Caughey et al.,14
which reported plasma proinflammatory cytokine inhibition after 4 weeks with the same dose. It may be beneficial in future wound studies to include smaller EPA/DHA dosages, in the form of both ethyl esters and triacyglycerols, for shorter time periods in a stratified design. Correlating data from this type of design may clarify the minimal EPA/DHA doses that affect the target mediators, such as proinflammatory cytokines, that control inflammatory responses during wound healing.
Not surprisingly, we found that IL-β, IL-6, and TNF-α proinflammatory cytokine levels in blister fluid were significantly increased across time from 5 to 24 hours after blister wound initiation in both groups in response to tissue trauma, which is consistent with earlier studies using suction blister models.11,24,25
Although the findings of significantly elevated IL-1 levels at 24 hours postblistering in the active group over the placebo group were not consistent with our original hypothesis, the results were in concert with a study that observed higher IL-1 expression in EPA treated nonirradiated and UVB-irradiated keratinocytes.31
Interleukin-1 assists in regulating fibroblast proliferation and collagen synthesis2
therefore, it can be posited that its initial upregulation at the wound site, as a result of EPA/DHA dietary supplementation, could be a pathway to regulate collagen formation. Improved production of healthy collagen is advantageous for efficient healing of skin with minimal scarring and for providing strength for connective tissues such as ligaments.17
In addition, initial IL-1 expression increases keratinocyte growth, important for reepithelialization,32
induces the production of IL-6 and has a stimulatory effect on angiogeneses.
Unexpectedly, the other primary predictor variable that contributed to the 47.4% of the variance in IL-1 values at 24 hours postblistering was gender (males), which could be related to the possible modulatory effects of local and systemically produced steroid hormones. Though the understanding of the role of estrogen in cytokine production is limited, it has been postulated that estrogen has an inhibitory effect on proinflammatory cytokine activity that could vary with tissue type.33
This is one explanation for the naturally occurring sexual dimorphisms of various inflammatory-related neurological disorders. It may be beneficial for future wound healing studies that explore the effects of EPA/DHA consumption on proinflammatory cytokine production to include measures of estrogen and testosterone, so that potential variations in data correlated to gender can be analyzed more thoroughly.
Although not statistically significant, higher quantities of IL-6 and TNF-α were also found in the wound fluid in the active group at 24 hours postblistering when compared with those in the placebo group. These identified trends may have translated to statistical significance if a larger sample size had been utilized. Again, these findings were not aligned with our original hypotheses, but were similar to a few previous studies that investigated the effects of EPA and DHA on tissue healing in animal models and cell cultures.20,31
For example, a study by Hankenson et al.20
found that EPA significantly elevated IL-6 release in medial collateral ligament cells in a bovine model. The researchers also found a significant linear correlation between IL-6 levels and collagen production. This suggests that EPA dietary supplementation may facilitate healing in tissues that particularly benefit from increased collagen production, such as ligaments. An interesting result in the present study was that increased levels of TNF-α in blister fluid at 24 hours postblistering were associated with slightly slower healing in the active group, which differs from an earlier study that demonstrated diminished local levels of TNF-α in healing-impaired glucocorticoid-treated mice.11
In the present study, we established that the time to complete wound healing (100% closure) for all eight blisters did not differ significantly between the two groups, however, those in the active group who consumed the EPA/DHA supplement for 4 weeks took approximately 1 day longer to heal than those in the placebo group. Considering the small size of the blister wounds, this finding could be of clinical significance. The higher levels of proinflammatory cytokines in blister fluid, found at 24 hours postblistering in the active group, suggest a more vigorous early inflammatory reaction. Therefore, it may have taken longer for the exudative stage of the inflammatory response to resolve, which may have resulted in a slightly longer time to complete wound closure. A previous study that measured wound healing in dog models also observed a longer time for reepithelialization in surgical wounds after ω-3 PUFA supplementation when compared with other dietary alterations.34
Furthermore, another research paper reported that topical administration of ω-3 PUFA to surgical wounds of mice was associated with slower wound closure in the first 10 days after surgery.35
Conversely, Ruthig et al.36
found improved reconstitution of epithelial integrity with both ω-3 and ω-6 PUFA-treated intestinal cells in rats following mucosal injury. Once again, these variable findings support the need for additional research studies to elucidate the effects PUFA have on wound healing and compare outcomes among similarly designed endeavors.
In summary, this study is the first to examine the effects of ω-3 (EPA/DHA) fatty acid dietary supplements on proinflammatory cytokine production in blister wound fluid and subsequently on cutaneous healing in a healthy human population. The results presented in this paper linked the EPA/DHA dietary supplements, taken by the participants in the active group, to a decreased plasma AA:EPA ratio from baseline and a significantly higher production of the proinflammatory cytokine IL-1β at blister wound sites at 24 hours postblistering, when compared with the placebo group, and nonsignificantly slower wound healing. Although not consistent with the original hypotheses, the study findings support the proposition that dietary ω-3 PUFA demonstrate ability to affect the local production of inflammatory mediators, such as proinflammatory cytokines, that regulate the wound healing process. Future research will hopefully clarify whether they are beneficial or detrimental to various wound types and populations of patients. This knowledge is desirable because the use of fish oil supplements, as well as other forms of complementary/ alternative medical (CAM) or integrative therapies, is growing briskly in the US
In 2002, 36% of adults in the US used some form of CAM therapy, such as dietary supplements, for health reasons.37
Approximately 12% of the adult population adds fish oil/ω-3 fatty acids to their diet and that number is on the rise.38
Essential fatty acid sales increased 50.3% in 2003 over 2002 and represented 11.5% of supplements’ total dollar sales.39
Thus, many individuals are consuming varying doses of EPA and DHA through supplements, which may alter cellular and molecular activities that influence wound healing. As our understanding of their bioactivity expands, it may be found increasingly important for clinicians to evaluate patients’ ω-3 fatty acid use when a wound is anticipated or already present, particularly if negative effects are identified.