Two novel c-abl mRNAs are expressed in rat parotid salivary glands during in vivo beta-adrenergic receptor stimulation.

The c-abl proto-oncogene is transcribed in most cell lines and tissues into two mRNAs of 6.5 and 5.3 kb, which have different 5' ends and encode two 150 kDa proteins that are largely colinear, but have different N-termini. We show here that two unusually short and abundant c-abl-related mRNAs of 1.5 and 1.3 kb appear in rat parotid salivary glands, within 1 day of in vivo administration of the beta-adrenergic receptor agonist isoproterenol. These transcripts are not found in the submandibular salivary gland or in the heart and they are too short to encode the known c-abl proteins. RNA blot, S1 nuclease protection and primer extension analysis suggest that the isoproterenol inducible parotid gland mRNAs do not contain the kinase domain, but represent part of the C-terminal segment of the abl reading frame.

"Inert" vehicles do affect wound healing.

The effect of a single daily application of U.S.P. petrolatum, an oil-in-water vanishing cream or a lotion on the rate of epidermal wound healing was determined in domestic white pigs. The superficial wounds were made with a dermatome and were not infected. In these studies, applications of U.S.P. white petrolatum retarded the rate of epidermal healing by 17% compared to untreated control wounds. Applications of an oil-in-water vanishing cream increased the rate of epidermal healing by 24% and a lotion increased the rate 15% compared to untreated control wounds.

New methods for assessing epidermal wound healing: the effects of triamcinolone acetonide and polyethelene film occlusion.

Epidermal healing of superficial, excised wounds in domestic white pigs was evaluated visually and histologically after separation of the epidermis and dermis. The visual determination of epidermal healing correlated well with the histologic studies of surface re-epithelialization. Wounds healed 40% faster when occluded with polyethelene film. Topical triamcinolone acetonide treatment delayed healing (62% slower than control).

Collagenase in wound healing: effect of wound age and type.

Collagenase is believed to be important for cell migration and collagen remodeling during tissue repair and regeneration. We have investigated collagenase concentrations in different types of surgically inflicted wounds in pigs. Collagenase was extracted from tissue homogenates of wounds by heating to 60 degrees C for 6 min in 0.1 M CaCl2. The molecular weight of latent collagenase was about 52 kDa. Activated collagenase produced the characteristic 3/4 fragment of collagen. Collagenase was assayed by the use of radiolabeled telopeptide-free collagen. To detect maximal collagenase activity, extracts were reduced and alkylated to destroy inhibitors, then activated with aminophenylmercuric acetate. Sutured incisions showed peak collagenase content on postoperative day 1 and thereafter steadily declining concentrations. Granulation tissue from non-sutured large defect full-thickness wounds showed high collagenase content on postoperative day 5 and then a sharp decline to day 7 followed by a slowly declining curve to postoperative day 21. Partial-thickness wounds exhibited a different time course, with collagenase increasing to peak concentrations on postoperative days 3-5; however, a large proportion of the detected collagenase was due to the adherent scab. By day 7 collagenase concentrations approached the low concentrations of normal skin when epithelialization was complete and the scab rejected. In general, collagenase shows an early maximum and then declines with postoperative time, with the sharpest decline occurring when epithelialization is complete.

The healing of superficial skin wounds is stimulated by external electrical current.

We studied the effects of direct electric current supplied by an energized silver-coated electrode on dermal and epidermal wound healing. Keratome-induced wounds (0.3 mm deep) on the skin of young domestic pigs were treated with either an energized (50-300 microA) electrode (DC), an unenergized electrode (placebo), or left untreated. Wounds were excised on days 1-7 after wounding and the epidermis was separated from the dermis. The epidermal sheet was evaluated for reepithelialization and the dermis was assayed for collagen biosynthetic capacity. Dermal collagen production among treatments did not differ markedly on days 1-4 after wounding. However, a highly significant increase (p less than 0.001) in the collagen synthetic capacity was observed on days 5, 6, and 7 in wound treated with DC. There was no significant difference in collagen synthesis among treatments when collagen production was corrected for DNA content. The rate of wound epithelialization was also significantly accelerated (p less than 0.05) in DC-treated wounds. These results suggest that the proliferative and/or migratory capacity of epithelial and connective tissue cells involved in repair and regeneration can be affected by an electrical field.

Recruitment of mononuclear cells by endothelial cell binding into wounded skin is a selective, time-dependent process with defined molecular interactions.

Wound healing involves a complex series of interactions between cells in the dermis and epidermis, and important relationships exist between keratinocytes and resident dermal cells. Monocytes and lymphocytes secrete cytokines that are capable of stimulating dermal repair and influencing keratinocyte and fibroblast migration and proliferation, although the mechanism by which mononuclear cells are recruited into the wound is unknown. We have tested the hypothesis that in wounded skin specialized endothelial cells are induced to mediate peripheral blood mononuclear cell (PBMC) emigration from the vasculature into the dermis. For this purpose, partial-thickness wounds made with a keratome on the backs of domestic pigs were excised 0 to 9, 12, 15, and 21 d after wounding. The biopsies were then tested for the capacity to adhere selectively to PBMC. The results indicated that PBMC overlaid onto sections of wounds from day 4 to 15 adhered selectively to dermal endothelium, with two distinct peaks of adherence observed on day 7 and day 12. In contrast, PBMC did not adhere to the tissue sections when overlaid onto frozen sections of normal skin or 0-, 1-, 2-, 3-, and 21-d-old wounded skin. Additional studies on the binding properties of PBMC subsets revealed that monocytes adhered maximally at day 7, whereas T cells adhered optimally at day 12 post-wounding. Furthermore, the adhesion process was energy and magnesium dependent but not calcium dependent and involved surface protein and carbohydrate moieties on PBMC surface. Pre-treatment of PBMC with monoclonal antibodies against the LFA-1 adhesive receptors inhibited the binding by greater than 80%, suggesting that LFA-1 adhesive receptors play an important role in the binding process. These studies provide evidence that the recruitment of monocytes and lymphocytes into wounds is an active, dynamic, and regulated process mediated at least in part by specific adhesive interactions between mononuclear leukocytes and dermal endothelial cells.

Characterization of a 54-nucleotide gap region in the 28S rRNA gene of Schistosoma mansoni.

We have analyzed 572 bp in the 28S rDNA of the human blood fluke Schistosoma mansoni which correspond to expansion segment 5 of domain IV as defined by Clark et al. for the Xenopus laevis 28S rRNA. S1 nuclease mapping and primer extension analysis comparing this region with the mature 28S rRNA indicate that there are 54 nucleotides present in the 28S rDNA which are absent from the mature rRNA. This defines a gap that creates two 28S rRNA subunits (28S alpha and 28S beta). Comparison of the S. mansoni sequence with rDNAs of other organisms which contain gaps in their 28S rRNA shows that the overall features are conserved except that the S. mansoni gap is less A + T-rich. The conserved features include: (1) the location of the gap within the 28S rRNA; (2) the predicted secondary structure of the gap, containing a stem-loop with a UAAU sequence within the loop; and (3) a conserved CGAAAGGG on the 3' side of the gap.

Effects of ionizing radiation and beta-adrenergic stimulation on the expression of early response genes in rat parotid glands.

There is little known about the regulation of gene expression in rat parotid glands after exposure to ionizing radiation. The present studies investigate the effects of in vivo ionizing radiation, with subsequent stimulation of beta-adrenergic receptors by isoproterenol, on parotid gland function and on the expression of the early response genes, c-fos, c-jun, and jun B. Ionizing radiation diminished parotid gland weight and saliva output. Treatment of irradiated rats with isoproterenol increased the gland weight to levels similar to those in nonirradiated rats. However, such treatment had no effect on saliva output as indicated by measurements of parotid salivary flow rate. Irradiation alone increased the expression of c-fos, c-jun, and jun B. The combination of irradiation and isoproterenol had an additional effect on the levels of c-fos and jun B mRNAs and proteins particularly at earlier experimental times (1 to 8 h). Isoproterenol alone induced high levels of c-fos and jun B mRNA but not of c-jun mRNA. However, c-jun mRNA was induced markedly by radiation and 8 h of isoproterenol treatment, indicating a combined effect on c-jun gene expression. These observations suggest that the expression of the proto-oncogenes c-fos, c-jun, and jun B is probably regulated through differential signal transduction pathways which may be activated by these external stimuli and may be associated with functional changes induced in the rat parotid gland by ionizing radiation and by ionizing radiation and isoproterenol.

The effect of a semiocclusive dressing on the microbial population in superficial wounds.

Superficial wounds in a Yorkshire pig were treated with a semiocclusive polyurethane film dressing (PUD) or left open. The number and types of microflora present in the wounds each day was determined by a scrub technique, selective media, and the spiral plating system. In wounds covered with the PUD, the number of organisms increased, and there were more gram-negative pathogens. We concluded that microorganisms in wound beds multiply and survive better beneath a semiocclusive dressing than with air exposure.

Occlusive dressings. Does dressing type influence the growth of common bacterial pathogens?

We studied the effect of different occlusive dressings and of air exposure on the growth of four pathogenic bacteria in wounds. Partial-thickness wounds on domestic pigs were inoculated with Staphylococcus aureus, Clostridium perfringens, Bacteroides fragilis, or Pseudomonas aeruginosa. Each wound was covered with three dressings (DuoDERM, Opsite, or Vigilon), or left exposed to air. Groups of wounds were sampled at 24, 48, and 72 hours. Staphylococcus aureus reached high levels beneath all of the dressings and in the air-exposed wounds. The numbers of C perfringens and B fragilis were greatly reduced in the air-exposed wounds and slightly reduced in the Opsite-covered wounds. The numbers of P aeruginosa were greatest in the Opsite- and Vigilon-covered wounds. The results indicate that occlusive dressings are not indicated in wounds that clinically appear to be grossly contaminated or that may contain anaerobic organisms.

Povidone-iodine in polyethylene oxide hydrogel dressing. Effect on multiplication of Staphylococcus aureus in partial-thickness wounds.

We studied the ability of a polyethylene oxide hydrogel dressing (Vigilon) containing povidone-iodine to prevent Staphylococcus aureus proliferation in partial-thickness wounds. We previously reported that a single application of povidone-iodine on wounds challenged with 2 X 10(6) S aureus was not effective in reducing the number of S aureus after 24 hours. It was therefore hypothesized that povidone-iodine might be effective if it was available continuously and applied to wounds containing a smaller number of bacteria. To test this hypothesis, we made multiple partial-thickness wounds (5 X 7 X 0.3 mm) on six domestic pigs. We then inoculated the wounds by scrubbing them with either a low concentration (log 3.5) or a high concentration (log 7) of S aureus suspension. The wounds were either treated with Vigilon or Vigilon containing povidone-iodine or left air exposed. Wounds from each of these treatment groups were cultured by the scrub technique for S aureus with a 0.5% sodium thiosulfate-polysorbate (Tween) 80 solution 5 minutes, 30 minutes, 24 hours, or 48 hours later. A significant reduction in the number of S aureus recovered from wounds treated with Vigilon containing povidone-iodine was seen with the group inoculated with a low bacterial concentration after 24 hours; but no reductions were observed when wounds were inoculated with the higher bacterial concentration of log 7. We found Vigilon containing povidone-iodine to be an effective inhibitor of S aureus in wounds over a 24-hour period when the organism was present in low numbers.

Studies on the repopulation of Langerhans cells in partial-thickness wounds. Air exposed and occlusively dressed.

BACKGROUND AND DESIGN: The use of occlusive dressings on partial-thickness wounds has been shown to promote early epithelization and connective tissue regeneration. Because Langerhans cells (LC) have been implicated in epidermal homeostasis we studied the rate of repopulation of LC in air-exposed vs occlusively dressed wounds. Partial-thickness wounds on the backs of pigs were treated with occlusive dressings (Tegaderm) for 3 days or left air exposed. On days 3, 5, 7, and 11 after keratome wounding, epidermal sheets from the regenerating wounds were isolated and stained for LC using indirect immunofluorescence. The LC populations were quantified in the interfollicular regions and expressed as average number of cells per square millimeter of epidermis. RESULTS: Normal skin control had 1024 +/- 93 LC/mm2 distributed uniformly. On day 3 after wounding occlusive-dressing-treated wounds had an LC repopulation of 46% of the original value. Langerhans cells in air-exposed skin could not be evaluated until epithelization occurred at day 5. Langerhans cells in both air-exposed and occlusive-dressing-treated wounds were 46% to 51%, 65% to 71%, and 91% of normal value, respectively, on days 5, 7, and 11. CONCLUSIONS: We conclude that at least in regenerating epidermis, the degree of repair of the new epidermis apparently plays a limited role in the migration of LC, as does the earlier growth of blood vessels.

Benzoyl peroxide and epidermal wound healing.

The effectiveness of 10%, 20%, and 50% benzoyl peroxide in a lotion, 20% benzoyl peroxide in a gel, and the effect of the vehicles alone on wound reepithelialization were evaluated in young domestic pigs. Twenty percent benzoyl peroxide suspension in a lotion base substantially increased the rate of reepithelialization by 33% over a seven-day evaluation period. Twenty percent benzoyl peroxide suspension in a gel base and 10% benzoyl peroxide suspension in a lotion base slightly enhanced epidermal resurfacing, while 50% benzoyl peroxide suspension in a lotion base and the vehicle gel retarded healing. Variations in the rate of reepithelialization were observed when different lots of 20% benzoyl peroxide lotions were compared. Chemical analysis of each of the 20% benzoyl peroxide preparations tested disclosed great differences in zinc, magnesium, and sodium content.

Ultraviolet irradiation-induced inflammation: effects of steroid and nonsteroid anti-inflammatory agents.

The effect of combined topical applications of a steroid and nonsteroid anti-inflammatory agent on erythema induced by ultraviolet-B irradiation (UV-B) was evaluated. In human volunteers, the combination more effectively suppressed UV-B--induced erythema than either agent alone. When applied singly, the nonsteroid agent was far more effective than the steroid. The combination, or either agent alone, was most effective when applied immediately after irradiation. This study demonstrates that for the treatment of UV-B--induced erythema, the anti-inflammation effects of these two classes of anti-inflammatory agents are greater when used in combination than when either agent is used alone. However, the effect of the combination in this study is not sufficiently long-lasting to be therapeutically useful.

Wound healing. The effects of topical antimicrobial agents.

The effect of four commonly used topical antimicrobial agents on the rate of reepithelialization of clean wounds was evaluated in white domestic pigs. Neosporin Ointment was found to significantly increase the rate of reepithelialization by 25%, while Furacin significantly retarded the healing rate by 24%. Pharmadine, a preparation containing povidone-iodine, did not affect the rate of healing. Both Silvadene and its vehicle significantly increased the rate of reepithelialization by 28% and 21%, respectively. The effects of these agents cannot be explained on the basis of their antimicrobial activity.

Optimal use of an occlusive dressing to enhance healing. Effect of delayed application and early removal on wound healing.

We examined the effect of delayed application and early removal of a polyurethane dressing on excisional wounds in swine. Backs of pigs were wounded with an electrokeratome, and wounds were divided into the following treatment groups: (1) air exposed; (2) dressings applied immediately after wounding and kept on until wounds were evaluated; (3) dressings applied immediately after wounding and removed at 6, 24, or 48 hours; and (4) dressings applied 2, 6, and 24 hours after wounding. Wounds were excised on days 3 through 7 and incubated in sodium bromide to allow separation of the epidermis and dermis. Specimens were considered healed if no defect was present. To promote optimal resurfacing in superficial wounds, polyurethane dressings need to be applied within two hours after wounding and should be kept in place for at least a 24-hour period.

Topical mupirocin treatment of impetigo is equal to oral erythromycin therapy.

Topical antimicrobial therapy has not been effective in the past against cutaneous bacterial infections. In this study, a new topical antibiotic ointment, mupirocin, was compared with oral erythromycin ethylsuccinate in the treatment of impetigo. Seventy-five patients clinically diagnosed as having impetigo and with positive cultures of Staphylococcus aureus, Streptococcus pyogenes, or both were examined in an investigator-blinded study. Patients used topical mupirocin applied three times daily or the usual oral dose of erythromycin ethylsuccinate (30 to 50 mg/kg per day). Patients' lesions were examined clinically and cultured bacteriologically on days 0, 3, and 8, and 1 week after treatment. Susceptibility testing was performed on pathogens isolated to determine antibiotic resistance. Mupirocin treatment produced similar clinical results to oral erythromycin and was superior in the eradication of S aureus, including antibiotic-resistant S aureus. These results show topical mupirocin to be a safe and effective alternative to oral antibiotic therapy in the treatment of impetigo.

Topical antibiotic treatment of impetigo with mupirocin.

Because the effectiveness of topical antimicrobials in the treatment of ecthyma, impetigo, and pyoderma is not well established, the US Food and Drug Administration has recently proposed guidelines for tests of topical antimicrobial efficacy in primary skin infections. The guidelines require both comparison with the agent's base and microbiologic documentation of efficacy. These guidelines were followed in this double-blind, eight-day evaluation of impetigo/ecthyma treated with mupirocin, a new agent that is only active topically. All cultures, before and after therapy, were taken using swabs dipped in neutralizing broth plus 10% fetal bovine serum to minimize antimicrobial "carry over" to the culture plate. Staphylococcus aureus, which was isolated from 94% of the patients before therapy, was eliminated in 88% of the mupirocin-treated patients and 47% of the vehicle-treated patients. Group A beta-hemolytic streptococci were eliminated in 100% of the mupirocin-treated and 0% of the vehicle-treated patients. To our knowledge, this is the first topical antibacterial treatment for primary skin infections proved superior to its vehicle using the proposed US Food and Drug Administration guidelines.

A new in vivo model for the evaluation of topical antiseptics on superficial wounds. The effect of 70% alcohol and povidone-iodine solution.

A preliminary evaluation of the antiseptic activity of 70% alcohol and 10% povidone-iodine solution was carried out in an animal model. After the inoculation of partial-thickness wounds with Staphylococcus aureus, 0.1 mL of the antiseptic agent (70% alcohol or povidone-iodine solution) was rubbed into the wound for 30 s. The agent was left on the wound for one minute, three minutes, and 24 hours, and then the wounds were cultured for bacteria. After one minute, the treatments did not reduce the number of pathogens. After three minutes, both 70% alcohol and povidone-iodine solution produced a slight reduction, and after 24 hours, povidone-iodine solution slightly reduced the number of pathogenic bacteria that could be cultured. However, after 24 hours, neither agent reduced the number of pathogens below 10(5) colony-forming units per milliliter. These preliminary data suggest that single applications of 70% alcohol or povidone-iodine may have very limited efficacy as antiseptic agents for the treatment of superficial wounds.

Systemic antibiotic therapy of secondary infected dermatitis.

Systemic cloxacillin therapy of secondarily infected dermatitis cloxacillin therapy become apparent produces a significant increase in healing when compared to a placebo. The effects of systemic after five days of treatment. A single pretreatment culture was not helpful in directing therapy.