Arginine is considered an essential amino acid under periods of severe stress. In 1976 Seifter, Barbul, Rettura, and Levenson first studied the role of arginine in injured animals. This was based on the rationale that a growing animal requires more arginine than a mature animal. In the growing animal, much of the arginine is used for the synthesis of connective tissue proteins. In the injured animal, an increase in arginine would be expected to synthesize reparative connective tissue. When subjected to minor trauma, such as dorsal skin incision and closure, rats have shown increased postoperative weight loss and increased mortality compared to rats fed a similar defined diet containing arginine. Furthermore, the arginine deficiency results in decreased wound breaking strength and wound collagen accumulation. Subsequent experiments showed that rats fed a chow diet, not deficient in arginine, and additionally supplemented with 1% arginine also have enhanced wound breaking strength and collagen synthesis when compared to chow-fed controls (Figure 6.2). Similar findings were noted in parenterally fed rats given an amino acid mixture containing high doses (7.5 g/l) of arginine in rats. This effect was observed in elderly rats fed a diet supplemented with a combination of both arginine and glycine. It was noted that these rats had an enhanced rate of wound collagen deposition when compared to controls.5
Two studies have been done so far on the effect of arginine supplementation on collagen accumulation in healthy human subjects (Figure 6.3). A micromodel was
P factor < .001 unless otherwise noted as < .005.
6000 5000 4000 3000 2000 1000 0
4000 3000 2000 1000 0
4000 3000 2000 -1000 -0
FIGURE 6.2 Effect of arginine deficiency (a) and arginine supplementation (b,c) on wound fresh breaking strength (FBS), g; formalin-fixed breaking strength (FxBS), g; and hydroxy-proline accumulation (OHP), |lg/100 mg sponge. Note that the P factor < 0.001 unless otherwise noted as < 0.005.
I I Placebo
I I Placebo
FIGURE 6.3 Effect of arginine on wound healing parameters in healthy elderly human volunteers.
described in which the collagen accumulation occurs in a subcutaneously placed 5 cm segment of polytetrafluoroethylene (PTFE) tubing to study the human fibroblastic response. In the first study, young healthy human volunteers (25 to 35 yr) were found to have a significant increase in wound collagen deposition following oral supplementation with either 30 g of arginine aspartate (17 g of free arginine) or 30 g of arginine HCL (24.8 g of free arginine) daily for 14 d. In a subsequent study of healthy elderly humans (67 to 82 yr), daily supplements of 30 g of arginine aspartate for 14 d resulted in significantly enhanced collagen and total protein deposition at the wound site when compared to placebo controls. This study evaluated the wound response using the PTFE catheters and examined epithelialization on a split thickness wound created on the upper thigh of each subject. The catheters in this study were analyzed for -amino nitrogen content (assessment of total protein content), DNA accumulation (index of cellular infiltration), and hydroxyproline content. There was no enhanced DNA present in the wounds of the arginine-supplemented subjects, suggesting that the effect of arginine is not mediated by an inflammatory mode of action. There was no observed effect in the rate of epithelialization of a superficial skin defect, suggesting that the predominant effect of arginine on wound healing is to enhance wound collagen deposition.6-8
There are several theories on the mechanism of the beneficial effect of arginine on wound healing. Arginine comprises less than 5% of the collagen molecule, and it is possible that supplemental arginine is a necessary substrate for collagen synthesis at the wound site. This may be possible through the direct use of arginine as substrate for the pathway arginine- > ornithine- > glutamic semialdehyde- > proline. Studies have shown that increased deposition of collagen early in the wound healing process correlates with an increase in wound strength. Arginine levels are undetect-able within the wound during the late phases when fibroplasia predominates. However, studies done by Albina showed that although ornithine levels are higher in wound plasma, the rate of conversion of ornithine to proline is quite low. Therefore, this would make the arginine-to-proline mechanism very unlikely.9
It was shown that NO plays a more dominant role in wound healing (Figure 6.4). Arginine is catabolized to NO specifically through iNOS with ultimate production of citrulline. Previous studies have shown that iNOS is upregulated following tissue injury, and that systemic administration of NOS inhibitors, such as S-methyl isothou-ronium, impairs wound tensile strength and collagen deposition in a dose-dependent manner. The iNOS is synthesized in the early phase of wound healing by inflammatory cells, mainly macrophages.10 NO inhibition has been shown to significantly impair wound healing of cutaneous incisional wounds and colonic anastamosis in rodents.11 In vitro studies have noted increased collagen synthesis in association with exogenous NO administration in cultured dermal fibroblasts. Supranormal collagen deposition has been noted after transfection of iNOS DNA into wounds. Conversely, mice lacking the iNOS gene (iNOS knockout mice) have delayed closure of exci-sional wounds, an impairment that is corrected by adenoviral transfer of the iNOS gene to the wound bed. The loss of a functional iNOS gene negates the beneficial effect of arginine in wound healing, while supplemental dietary arginine enhances wound healing in normal mice.12 NO synthesis is also decreased in the wound milieu
of diabetics, and arginine supplementation restored healing by normalizing the NO pathway without affecting the arginase activity.13 This suggests that the metabolism of arginine via the NO pathway is one mechanism responsible for arginine enhancing wound healing (Figure 6.5).
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