Histopathology of insulin-induced laminitis in ponies

Reasons for performing study: Ponies with laminitis associated with insulin resistance and hyperinsulinaemia lack systemic and/or intestinal inflammatory signs, suggesting a different pathogenesis potentially reflected in differing histopathology. Objectives: To describe the histological appearance and quantify morphological changes in primary and secondary epidermal lamellae (PEL and SEL) of laminitis lesions from ponies with insulin‐induced laminitis. Methods: Equine hoof lamellar tissue was obtained from 4 control ponies and 5 ponies with laminitis induced following infusion of insulin (1036 ± 55 µU/ml) while maintaining euglycaemia for 55.4 ± 5.5 h. Sections from all 4 hooves were stained and examined by a veterinary pathologist. Measurements of lamellar length (PEL and SEL) were made in mid‐dorsal sections of the right forefeet by 2 blinded observers. Immunolabelling for calprotectin was performed using a monoclonal antibody. Results: No lesions were detected in normal ponies. Lesions detected in ponies with laminitis were variable in severity between ponies. Within ponies, SEL lesions were more severe along the axial region of PEL. Lesions included swelling, disorganisation and abnormal keratinisation of epidermal cells, increased mitotic activity and apoptosis. Separation of basement membranes was minimal. Immunostaining revealed inflammatory cells within the lamellar dermis. SEL were significantly elongated in laminitic hooves relative to controls, with the greatest elongation in those attached to abaxial and middle regions of PEL. Conclusions: Laminitis induced by prolonged infusion of insulin lacked widespread basement membrane disintegration, and increases in epidermal cellular proliferation at axial aspects were marked for this acute stage of disease. Potential relevance: Defining equine laminitis entirely in terms of separation of the basement membrane may not be appropriate for laminitis associated with hyperinsulinaemia.     

Effect of prolonged water immersion on equine hoof epidermis in vitro

OBJECTIVE: To evaluate the effect of prolonged water exposure on tissue mass and solutes of outer and inner layers of the stratum medium, sole, frog, and the stratum medium (SMZA) zona alba layer of horses' hooves. SPECIMEN POPULATION: 10 hooves from 10 horses without foot abnormalities. PROCEDURES: Hoof wall tissue specimens were obtained and immersed for 10 days in distilled deionized water. Serial changes in mass were recorded during the immersion period. Subsequently, osmolarity and Na+, K, Cl-, and protein concentrations of the immersion solution were quantified. RESULTS: Fully cornified outer hoof wall, sole, and frog epidermal structures increased in mass, whereas the SMZA lost mass when immersed in water. All hoof structures had a variable loss of crystalloids during immersion, but none of the specimens lost proteins. The frog epidermis was distinct in that total solute lost during immersion could not be ascribed to N+, K+, and Cl-. CONCLUSIONS AND CLINICAL RELEVANCE: Data support a 2-compartment model for the fully cornified outer stratum medium, frog, and sole that permits the exchange of crystalloids, but not proteins, across the cell membrane and infers that topical agents containing proteins cannot benefit the hoof. The unique osmotic behavior of the SMZA relative to other hoof structures suggests the hypothesis that it is composed of transitional epithelial cells. The solutes lost from frog epithelium are interpreted to reflect its unique lipid composition.     

Comparison of Two Methods for Presurgical Disinfection of the Equine Hoof

OBJECTIVES: To determine for equine hooves the normal resident aerobic bacterial population and the efficacy of 2 methods of disinfection. Study Design-Measurement of total bacterial, gram-positive bacterial, and gram-negative bacterial surface populations from the frog, sole, and hoof wall after each step of 2 different preoperative surgical disinfection techniques. ANIMALS: Six adult horses. METHODS: Hoof wall, sole, and frog samples were collected for quantitative bacteriology before, during, and after 2 multistep antiseptic preparation techniques: Method A-6-minute scrub with povidone-iodine soap, followed by 24-hour submersion in povidone-iodine solution-soaked cotton; and Method B-initial removal of superficial layer of hoof capsule before completing Method A disinfection procedures. RESULTS: Removal of the superficial hoof layer, application of the povidone iodine scrub, and completion of the povidone-iodine soak all significantly (P < .0008) decreased total bacterial numbers. Method B had significantly lower bacterial counts than method A at each consecutive step. Final total bacterial counts remained greater than 10(5) bacteria per gram of tissue regardless of preparation method. CONCLUSIONS: The hoof surface hosts a broad spectrum of aerobic gram-positive and -negative bacteria, many of which are potential pathogens. Bacterial numbers can be significantly reduced by removal of the superficial hoof surface, by application of a povidone-iodine scrub, and by use of a 24-hour povidone-iodine soak. However, bacterial populations >10(5) g per tissue persist after these disinfection procedures. CLINICAL RELEVANCE: Regardless of the preparation methods used in this study, bacterial populations capable of inducing wound infection remain on the hoof capsule.     

Suspected selenium poisoning in a horse

A case of selenium poisoning was suspected following the administration of 325 micromol selenium a day as sodium selenate (25 mg Se) for five consecutive days. The main signs were confined to changes in the integument which were loss of hair from the mane and tail, disintegration of the skin of the anus, lips, prepuce and scrotum, and separation of the hooves from the coronary corium. Periodic analyses of blood, hair and hoof parings from the sole correlated strongly with blood and identified horn and hair as important additional routes of excretion.     

Experimental infection of ponies with Borrelia burgdorferi by exposure to Ixodid ticks

Seven specific-pathogen-free (SPF) ponies, 1-5 years old, were exposed to Borrelia burgdorferi-infected adult ticks while being treated with dexamethasone over 5 consecutive days. One SPF pony (pony No. 178) was first exposed to laboratory-reared nymphs without B. burgdorferi infection and 3 weeks later was exposed to B. burgdorferi-infected adult ticks with concurrent dexamethasone treatment for 5 consecutive days. Four uninfected ponies treated with dexamethasone, exposed to laboratory-reared ticks without B. burgdorferi infection served as uninfected controls. Clinical signs, bacteriologic culture, polymerase chain reaction (PCR) for bacterial DNA, immunologic responses, and gross lesions and histopathologic changes were investigated during the experiment or at necropsy 9 months after tick exposure. In all of the seven challenged ponies, infection with B. burgdorferi was detected from monthly skin biopsies and various tissues at postmortem examination by culture and by PCR. However, pony No. 178 exposed to laboratory-reared nymphs (without B. burgdorferi infection) and challenged with B. burgdorferi-infected adult ticks 2 months later did not develop a B. burgdorferi infection. All of the infected ponies seroconverted. Control ponies and pony No. 178 were negative by culture, PCR, and serology. Except for skin lesions, we failed to induce any significant histopathologic changes in this study. This is the first report of successful tick-induced experimental infection in ponies by exposure to B. burgdorferi-infected ticks. This Lyme disease model will be very useful to evaluate efficacy of vaccines against the Lyme agent and the effect of antibiotic therapy on horses infected with B. burgdorferi.     

Geometric symmetry of the solar surface of hooves of thoroughbred racehorses

OBJECTIVE: To define a 3-dimensional (3-D) coordinate system with clear definitions of origins and axes relative to hoof anatomic features and determine whether solar surfaces of Thoroughbred racehorse hooves have geometric asymmetry in the mediolateral and dorsopalmar directions. SAMPLE POPULATION: Left forelimb hooves from 20 Thoroughbred racehorse cadavers. PROCEDURE: A right-handed 3-D coordinate axes system centered on the collateral sulci was defined for the left front hoof. Orthogonal distances of anatomic features from the dorsopalmar axis and the plane coincident with the ground were measured and compared between medial and lateral sides and between dorsal and palmar regions of the hoof. RESULTS: The hoof was wider and had a greater radius laterally than medially. The most distal part of the lateral bar of the frog was further from the dorsopalmar axis than that of the medial bar. Overall, mediolateral asymmetries in depth were not observed. The sole at the perimeter was deeper medially in the dorsal part of the hoof and laterally in the palmar part, with depth overall being greater palmarly than dorsally. Most features had dorsopalmar asymmetry. CONCLUSIONS AND CLINICAL RELEVANCE: When the angle bisected by the collateral sulci is used to determine the dorsopalmar axis of the hoof, most central structures (bars and collateral sulci) have mediolateral symmetry. However, the hoof wall and sole have some mediolateral asymmetries and most structures have dorsopalmar asymmetry. These findings may assist the development of devices for attachment to hooves and studies of the interaction of hooves with bearing surfaces.     

Acute experimentally induced aflatoxicosis in the weanling pony

Nineteen weanling ponies and 1 adult pony were given a single oral dose of aflatoxin B1 (AFB1). Dosages were: 0, 0.5, 1, 2, 4, 5, 6, and 7.4 mg of AFB1/kg of body weight. Vital signs were monitored, and whole blood and serum collected for analysis of serum enzymes, prothrombin time, blood cell counts, and serum urea nitrogen. Ponies that died were examined for gross lesions, and tissues were collected for histopathologic examination and analysis of AFB1 and AFM1 residues. Two of the 4 ponies given the 2 mg/kg dose and all ponies given the larger dosages died within 76 hours. Clinical signs included increased rectal temperature, faster heart and respiratory rates, abdominal straining, bloody feces, and tetanic convulsions. At necropsy, ponies that died of acute aflatoxicosis showed visceral petechiae and hepatic focal lesions. Histopathologic changes included severe hepatic necrosis, vacuolation, and bile duct hyperplasia. Aflatoxins B1 and M1 were recovered from liver, kidney, skeletal muscle, and gastrointestinal contents. One other pony given the 2 mg/kg dose died 32 days after dosing, and 1 control pony died after 70 days. Continuous elevations in prothrombin time and serum aspartate aminotransferase, alanine aminotransferase, and gamma-glutamyl transpeptidase levels were observed in ponies dosed at 4 mg/kg or more. Significant (P less than 0.05) elevations in these values, which peaked 2 to 3 days after dosing, were seen in ponies given the 2 mg/kg dose. This group also had significant increases over controls in PCV and hemoglobin concentration 5 days after dosing.     

Hoof wall wound repair

REASONS FOR PERFORMING STUDY: Surgical stripping of the hoof wall results in a wound that heals remarkabley well. In contrast, lamellae recovering from laminitis are often deformed. Investigating lamellar wound healing may aid understanding of laminitis. OBJECTIVES: To document temporal changes in the lamellar basement membrane (BM), dermis and epidermis after surgery. METHODS: Wall strips were made in the dorsal hoof wall midline of 6 mature horses. Immunohistochemistry was used to document changes in the basement membrane (BM) and detect proliferation of epidermal cells in lamellar tissues harvested at intervals. A conforming metal plate was screwed to the hoof wall to maintain alignment of the wound edges. RESULTS: Wall stripping caused lamellar tips to snap and remain behind in the dermis along with the majority of the lamellar BM and some lamellar basal cells. Three days later the BM was intact and new lamellae had been reconstructed by proliferation of surviving epidermal cells. By 5 days the surface of the stripped zone was covered with yellow epidermis that subsequently thickened and hardened. Eventually the hoof wall deficit was replaced by new wall growing down from the coronet. The conforming metal plate and post operative analgesic ensured minimal lameness. CONCLUSIONS AND POTENTIAL RELEVANCE: In wall stripped lamellae the BM survives virtually intact and is used as a template for proliferating cells, from snapped-off lamellar tips, to migrate and quickly achieve repair to near normality. In laminitis epidermal dysadhesion and lamellar BM destruction occurs and lack of a functional BM template may explain the prolonged and abnormal repair of affected lamellae.     

A Photometric Method for Recording Hoof Diseases in Cattle,with Special Reference to Haemorrhages of the Sole

Bergsten, C: A photometric method for recording hoof diseases in cattle, with special reference to haemorrhages of the sole. Acta vet. scand. 1993,34,281-286.– After cleaning and trimming, the sole surface and interdigital space of the hooves of cows are photographed in a standardized way. The colour slides are projected on to a screen and the haemorrhages of the sole (reflecting Pododermatitis aseptica diffusa) and the lesions of Dermatitis interdigitalis and Erosio ungulae are scored according to their severity and extent by 2 people who do not know the identity of the cows. In a repeatability test the correlation coefficients between 2 assessments of the haemorrhages in the sole ranged between 0.78 and 0.88 for assessments of the front hooves, the hind hooves and all the hooves together.     

Light and electron microscopy of keratinization in the laminar epidermis of the equine hoof with reference to laminitis

The laminar epidermis (epidermis parietis) of hooves from 14 clinically normal horses, 6 months to 15 years old, was examined by light and electron microscopy and immunofluorescence to measure the contributions of this region to the formation of the hoof wall. By their progressive keratinization to form primary epidermal laminae, the secondary epidermal laminae ultimately contributed about 20% of the thickness of the hoof wall (as revealed in the white line [zona alba]). The keratinized, primary epidermal laminae were developed to a height of 4 mm during their proximodistal-course, much of this obscured because of their basal portion being embedded in the cap horn epidermis. From evaluation of structural changes accompanying keratinogenesis in the cell and determination of the contribution of the laminar epidermis to the formation of laminar horn, cap horn, connecting horn, terminal horn, and the white line, we concluded that the sterile bed concept of a nongerminative role for the secondary epidermal laminae is no longer tenable.     

A Cross-Sectional Survey of Forelimb Hoof Conformation and the Prevalence of Flat Feet in a Cohort of Thoroughbred Racehorses in New Zealand

Racehorses in New Zealand predominantly train counter clockwise. This training pattern has been associated with between forelimb differences in bone mineral density profile and asymmetrical limb loading after training. At present, there is limited data on the hoof conformation of these racehorses. Distal forelimb and digital hoof conformation data were collected from 75 Thoroughbred racehorses (2–5 years old) from two training yards. Digital conformation was subjectively graded, and multiple hoof measurements were made with a modified tire gauge (sole and sulci depth) and from digital photographs. All the horses were shod by two registered master farriers within a median of 15 (interquartile range [IQR], 1–25) days before measurement. There were few distal limb conformation abnormalities scored. Most (62/75) horses presented with some deviation from normal hoof parameters, with 2 (IQR, 1–3) abnormalities reported per horse. The most common hoof abnormality was uneven sulci, which was identified in 43 horses and 59 affected hooves, followed by higher medial hoof wall height in 38 horses and 53 affected hooves. Many of the linear and hoof angle measurements and their ratios were within the bounds reported within the literature and indicative of a balanced foot. The length and width measurements increased with horse age. The dorsal hoof wall (DHW) length:heel length ratios were consistently less than 3:1, and the absolute difference between toe and heel angle was generally greater than 5°. Between limb hoof variation was identified for a number of the morphologic measurements including frog length and sole length and the ratio of sole width:sole length. Flat feet (lack of concave solar surface) were identified in 21/75 (28%) horses and in 28/150 (19%) forelimb hoofs. More horses had a flat left foot (10/75) than right foot (4/75), but seven horses had both feet classified as being flat. Flat feet had 2.4 (1.1–5.6, P = .036) greater odds of presenting with uneven sulci. These data indicate that uneven sulci depth and flatter hooves with may be a typical presentation of Thoroughbred feet. Asymmetry in measurements between limb may reflect the greater loading of the left forelimb when race training counter clockwise.     

Cardiovascular, acid-base, electrolyte, and plasma volume changes in ponies developing alimentary laminitis.

Twelve Shetland ponies were fed a high-starch ration. Seven ponies which had a transitory metabolic acidosis developed laminitis 56 hours (+/- 3.5, SEM) after overfeeding. These ponies also developed persistent hypokalemia, hyperthermia, and increased heart rate 24 hours before the onset of lameness. Serum sodium, serum chloride, hematocrit, plasma volume, and blood volume were unchanged. At the onset of clinical signs of laminitis, cardiac output and blood pressure increased, but total peripheral resistance was unchanged. None of the measured or calculated values predicted the onset of laminitis. Hypertension appeared to be a response to, rather than a cause of, lameness. Three of the remaining ponies apparently died of shock 29.3 +/- 2.7 hours after overfeeding. All 3 had severe metabolic acidosis; decreased cardiac output, systemic arterial pressure, and plasma volume; and increased hematocrit, total peripheral resistance, and pulmonary vascular resistance. The 11th pony was unaffected and the 12th pony was euthanatized.     

Prevalence and interrelationships of hoof lesions and lameness in Swedish dairy cows

The prevalence of hoof lesions and lameness in 4899 heifers and cows was determined at claw trimming one time in a cross-sectional study on 101 Swedish dairy farms, 1996-1998. The percentage of affected animals was 41% for heel-horn erosion, 30% for sole haemorrhages, 27% for erosive dermatitis, 21% for abnormal claw shape, 14% for white-line haemorrhages, 8.8% for white-line fissures, 8.6% for sole ulcers, 3.3% for double soles, 2.3% for verrucose dermatitis, and 1.8% for interdigital hyperplasia (IH). Seventy-two percent of all animals had at least one hoof lesion. The prevalence of lameness was 5.1%; most hoof lesions did not cause lameness. Differences between herds were substantial; the herd-specific, animal-level prevalence of lesions ranged from 25 to 98% and of lameness from 0 to 33%. Sole haemorrhages were found in all herds. The proportion of variance at the between-herd level was particularly high for heel-horn erosion (62%) and abnormal claw shape (54%). Strong correlations between lesions were found within hooves (and animals), e.g. for heel-horn erosion and dermatitis (Spearman's rank correlation, r(s)=0.36 and 0.37, respectively) and for sole and white-line haemorrhages (r(s)=0.25 and 0.28). Most hoof lesions affected hind and front hooves bilaterally, whereas the correlation between hind and front hooves generally was lower. Herds that ranked high for prevalence of sole ulcer also ranked high for sole haemorrhages and for abnormal claw shape and herds that ranked high for dermatitis also ranked high for heel-horn erosion, verrucose dermatitis and IH. Abnormal claw shape was strongly associated with sole ulcer (r(s)=0.41 at cow level)-suggesting the importance of maintaining a correct claw shape for the prevention of hoof-horn lesions.     

Density and morphologic features of primary epidermal laminae in the feet of three-year-old racing Quarter Horses

OBJECTIVE: To quantify the density of primary epidermal laminae (PELs) around the solar circumference and evaluate the relationship between regional PEL density and hoof capsule morphology in horses. SAMPLE POPULATION: Forefeet from nine 3-year-old Quarter Horse cadavers. PROCEDURES: Data pertaining to gross features of hoof morphology and PEL variables, including number, density, and distribution patterns around the perimeter of the hoof wall and number of bar PELs, were collected. Tissues of the laminar junction were examined histologically. RESULTS: No significant differences were found between left and right forefeet with respect to gross hoof morphologic measurements. Mean +/- SD number of PELs, including those at the bars of the hoof, was 551 +/- 30. Primary epidermal laminar density in the toe was significantly higher, compared with that in the quarter and heel regions, and was higher on the lateral aspect of the foot, compared with the medial aspect. Feet were significantly wider on the lateral aspect of the foot, compared with the medial aspect, as measured across the widest point of the solar surface. Histologic examination revealed atypical laminar morphology at the toe. CONCLUSIONS AND CLINICAL RELEVANCE: Variations were detected in PEL density and morphology around the solar circumference of hooves from 3-year-old racing Quarter Horses. A better understanding of relationships between laminar density, laminar morphology, and gross morphology of the hoof capsule in different populations of horses may aid practitioners in diagnosis and treatment of disease involving the hoof wall in horses.     

Effect of biotin supplementation on hoof health and ceramide composition in dairy cattle

The effect of biotin supplementation on various foot lesions and hoof ceramide composition of toe (wall) and sole portions of hooves was studied in crossbred dairy cattle. Biotin supplementation was done for five months in 14 cattle at a farm and the other 14 animals kept as control. A significant decline was observed in heel erosions and sole avulsions along with total disappearance of white line fissures and double soles in the biotin supplemented cattle resulting in decrease in the overall disease score. Thin layer chromatographs of the hoof lipids revealed 11 types of ceramides in sole lipids and 6 types of ceramides in toe (wall) lipids. The ceramides were typed and identified according to their Rf values. A qualitative increase in the density of thin layer chromatographs of sole lipids was observed in biotin supplemented cattle whereas a non-significant difference in density of thin layer chromatographs of toe lipids was observed after supplementation of biotin.     

Sequential mesenteric arteriography in pony foals during repeated inoculations of Strongylus vulgaris and treatments with ivermectin

Semiselective mesenteric arteriography was performed at regular intervals (inoculation weeks [IW] 0, 11, 18, and 24) in 9 of 10 pony foals raised to be free of parasites. Fifty infective larvae (L3) of Strongylus vulgaris were administered weekly for 4 weeks, then every 2 weeks through the 20th week. Three ponies were given ivermectin (oral paste, 0.2 mg/kg of body weight) treatment at IW 8, 16 and 24. Four ponies were inoculated, but did not receive ivermectin, and a third group of 2 ponies acted as uninoculated controls. Control ponies did not have gross or arteriographic lesions, whereas the inoculated untreated ponies had gross and progressive arteriographic lesions typical of verminous arteritis. Arteriographic lesions in the ivermectin-treated inoculated ponies were not as severe those in the untreated inoculated group, and there was either a partial resolution or a lack of progression of arteriographic lesions in all treated ponies. One untreated inoculated pony did not have progressive arterial lesions as did the 3 others in the group, and may develop resistance to the parasite.     

Acute hemolytic anemia after oral administration of L-tryptophan in ponies

The hematologic and pathologic effects of orally administered L-tryptophan and indoleactic acid and of L-tryptophan administered IV were studied in ponies. Sixteen adult Shetland ponies were allotted into 4 experimental groups. Group 1 consisted of 5 ponies (1-5) given 0.6 g of tryptophan/kg of body weight in a water slurry via stomach tube. Group 2 included 4 ponies (6-9) given 0.35 g of tryptophan/kg orally. Group-3 ponies (10-13) were given 0.35 g of indoleacetic acid/kg orally. Group 4 consisted of 3 ponies (14-16) given a single 4-hour IV infusion of 0.1 g of tryptophan/kg. Restlessness, increased respiratory rate, hemolysis, and hemoglobinuria were detected in 4 of the 5 group-1 ponies. Only pony 7 in group 2 developed hemolysis, hemoglobinuria, and a significant increase in respiratory rate. Renal pathologic lesions, consistent with hemoglobinuric nephrosis, were seen in ponies 2, 4, 5, and 7. Bronchiolar degeneration was evident in 4 of 9 ponies given tryptophan orally. The importance of these respiratory lesions was unknown. Clinical or pathologic abnormalities were not noticed in the ponies of groups 3 and 4. Mean plasma tryptophan values increased significantly in groups 1 and 2 at 6 hours after dosing. A second peak of tryptophan was detected in both groups at 12 hours. Values returned to predose values by 48 hours. Plasma indole and 3-methylindole concentrations were detectable in only 2 ponies (4 and 7). In vitro incubations of cecal fluid from ponies 6, 8, and 9 yielded a percentage conversion of tryptophan to indole of 16.75%, 5.84%, and 7.96%, respectively. 3-Methylindole was not produced. These results suggested that indole was the major metabolite of orally administered tryptophan in these ponies.     

Failure of Calcium Channel Blockade to Prevent Intra-abdominal Adhesions in Ponies

Intra-abdominal adhesions were created by localized serosal trauma in 11 adult ponies at three locations on the small intestine. Six ponies received verapamil hydrochloride (0.2 mg/ kg) subcutaneously every eight hours for three days, and five ponies received an equal volume of saline solution at the same intervals. The investigators were not informed which treatments the ponies received. Systolic, diastolic, and mean carotid arterial pressures and heart rates were measured six hours before surgery, and then 0.5, 1, 1.5, and 8 hours after the first treatment on each day for three days. One pony was euthanatized on day 13 because of colic, and the other 10 ponies were euthanatized 14 days after surgery. Scoring methods were used to assess the severity of adhesion formation and to grade the histologic appearance of the abraded sites. No significant differences were found for rectal temperature, packed cell volume, total plasma proteins, heart rate, and systolic, diastolic, or mean arterial pressures between control and verapamil-treated ponies. No significant differences were detected between the treatment groups for adhesion scores per abraded site, total adhesion scores per pony, the total number of adhesions per pony, or in the histologic scores.     

Cutaneous DNA vaccination against Ebola virus by particle bombardment: histopathology and alteration of CD3-positive dendritic epidermal cells

We analyzed the localization of gold particles, expression of immunogenic protein, and histopathologic changes after vaccinating guinea pigs and mice with a DNA vaccine to the Ebola virus glycoprotein administered by cutaneous particle bombardment. Gold particles were deposited in all layers of the epidermis and in the dermis. Those in the epidermis were lost as the damaged layers sloughed, while those in the dermis were phagocytized by macrophages. Glycoprotein was demonstrated by immunohistochemistry primarily in keratinocytes in the epidermis and hair follicle epithelium and less frequently in dermal macrophages, fibroblasts, sebocytes, and cells that appeared to be Langerhans cells. The number of cells that expressed glycoprotein increased between 4 and 8 hours postvaccination, then decreased to near zero by 48 hours. The vaccine sites were histologically divisible into three zones. The central portion, zone 1, contained the most gold particles in the dermis and epidermis and had extensive tissue damage, including full-thickness epidermal necrosis. Zone 2 contained fewer gold particles in the epidermis and dermis and had less extensive necrosis. The majority of cells in which glycoprotein was expressed were in zone 2. Zone 3 contained gold particles only in the epidermis and had necrosis of only a few scattered cells. Regeneration of the epidermis in damaged areas was evident at 24 hours postvaccination and was essentially complete by day 5 in the mice and day 10 in the guinea pigs. Inflammatory changes were characterized by hemorrhage, edema, and infiltrates of neutrophils initially and by infiltrates of lymphocytes and macrophages at later times. In zone 1, inflammation affected both the epidermis and dermis. Peripherally, inflammation was relatively limited to the epidermis. CD3-positive dendritic epidermal cells were demonstrated in the epidermis and superficial hair follicles of unvaccinated immunocompetent mice and beige mice but not of SCID mice. These cells disappeared from all but the most peripheral portions of the vaccine sites of vaccinated mice within 24 hours. They reappeared slowly, failing to reach numbers comparable with unvaccinated mice by 35 days postvaccination. The epidermis of control guinea pigs also had CD3-positive cells, but they did not have dendrites. These findings should contribute to a better understanding of the mechanisms operating in response to DNA vaccination by particle bombardment.