The tripeptide-copper complex glycyl-L-histidyl-L-lysineCu2+ (GHK-Cu) was first described as a growth factor for differentiated cells. Recent in vitro data showed that it possesses several properties of a potential activator of wound repair. We investigated the effects of GHK-Cu in vivo, using the wound chamber model described previously (Schilling, J. A., W. Joel, and M. T. Shurley, 1959 Stainless steel wire mesh cylinders were implanted subcutaneously on the back of rats. The animals were divided into groups that received sequential injections into the wound chamber of either saline (control group) or various concentrations of GHK-Cu. At the end of the experiments, rats were killed, wound chambers were collected, and their content was analyzed for dry weight, total proteins, collagen, DNA, elastin, glycosaminoglycans, and specific mRNAs for collagens and TGFfI. In the GHK-Cu-injected wound chambers, a concentration-dependent increase of dry weight, DNA, total protein, collagen, and glycosaminoglycan contents was found. The stimulation of collagen synthesis was twice that of noncollagen proteins. Type I and type III collagen mRNAs were increased but not TGFft mRNAs. An increase ofthe relative amount ofdermatan sulfate was also found. A control tripeptide, L-glutamyl-L-histidyl-Lproline, had no significant effect. These results demonstrate that GHK-Cu is able to increase extracellular matrix accumulation in wounds in vivo. (J. Clin. Invest. 1993.92:2368-2376

We elected to examine the efficacy of the topically applied immunosuppressive agent FK506 (Prograf) in the treatment of alopecia areata (AA) using the Dundee experimental bald rat (DEBR) model. Thirty lesional DEBR rats were allocated to five groups of six. Group 1 rats received 0.1 mL of a 0.25% solution of FK506 within a 2 x 2 cm marked area on one bald flank twice a week (125 micrograms FK506/cm2 per week) for 8 weeks, while the contralateral flank was left untreated. In group II, 0.05 mL of a 0.1% solution of FK 506 was applied 5 days per week on one flank (62.5 micrograms FK506/ cm2 per week) and control vehicle to the opposite flank for 8 weeks. Group III rats were treated as in group II except that drug and vehicle were applied twice a week (25 micrograms FK506/cm2 per week) for 4 weeks. A positive control group received orally administered cyclosporin A (CsA) (10 mg/kg daily) for 8 weeks and a further group was left untreated. Rats were regularly examined and photographed with skin biopsies taken from groups II and III. All FK 506-treated rats regrew hair at the site of drug application within 14-21 days. Growth continued for 3 weeks beyond termination of treatment after which gradual hair loss was observed. No hair growth was seen as a result of vehicle application and hair loss continued on untreated areas and in the untreated control group. Immunohistology revealed a drastic reduction in the follicular inflammatory infiltrate at the site of the FK506 application. The oral CsA group responded by simultaneous regrowth of hair over the whole body. Our findings suggest that FK506 may have considerable potential as a topical treatment for AA.

The laryngeal chemoreflex (LCR) was investigated in 21 piglets (ages 6 to 80 days old) with the use of physiologic and histologic techniques. The central projection of the superior laryngeal nerve (SLN) was determined in 14 animals by use of horseradish peroxidase-wheat germ agglutinin. Ipsilateral labeling of the solitary tract nucleus was seen. The caudal extent of the labeling varied with age. Sensory labeling of the nucleus ambiguus was present bilaterally in three younger animals and unilaterally in older piglets. Bilateral labeling of the nucleus dorsomedialis was seen in all ages. Apneic and cardiovascular response to water stimulation of laryngeal mucosa and the laryngeal adductor reflex (LAR) were examined in 16 piglets. Blunting of the apneic and cardiovascular response was seen after sacrifice of a single SLN. Hypoxia did not significantly affect the LAR or apnea duration in animals with only one intact SLN. A contralateral LAR was found in younger animals. Relevance to the LCR is also discussed.

In recent years there has been increased interest in the possibility of reinnervating paralyzed muscles, particularly the muscles of the face and of the larynx. There has been dispute over the relative effectiveness of different approaches. This study was undertaken to compare direct nerve implantation with the transfer of a neuromuscular pedicle using denervated strap muscles in a rabbit model. Reinnervation was carried out both immediately after denervation and after a delay period. Evidence of return of function was obtained by a method of long-term electromyographic monitoring and was confirmed electrically by evoked electromyography and muscle tension measurement and histologically by brain stem uptake of horseradish peroxidase. Both methods of nerve transfer produced consistently functional neuromuscular units, with physiological activity and muscle strength comparable with those in normally innervated controls. Return of function was apparent within 6 weeks of both nerve implantation and neuromuscular transfer; with this model, neither method showed a clear advantage over the other.

The decision to use an end-to-end versus end-to-side anastomosis remains a controversial issue in microvascular surgery, although it is generally accepted that certain clinical situations require skills in both techniques (eg, vessel size discrepancy or the need to preserve donor vessel distal flow). Using rats, this investigation presents a unique free flap model comparing not only arterial end-to-end and end-to-side anastomoses, but venous anastomoses as well. There were 15 animals in each anastomosis group with a greater than 80% flap survival rate in each group. This supports findings of earlier studies that suggested no difference in survival when these two anastomosis types were compared. Equally important was the assessment of dermofluorometry as an immediate postoperative perfusion monitoring device. In this model, fluorometry was not found to be useful for predicting flap viability in an early period after completion of the anastomoses.