The role of melanogenesis in regulation of melanoma behavior: melanogenesis leads to stimulation of HIF-1α expression and HIF-dependent attendant pathways.

To study the effect of melanogenesis on HIF-1α expression and attendant pathways, we used stable human and hamster melanoma cell lines in which the amelanotic vs. melanotic phenotypes are dependent upon the concentration of melanogenesis precursors in the culture media. The induction of melanin pigmentation led to significant up-regulation of HIF-1α, but not HIF-2α, protein in melanized cells for both lines. Similar upregulation of nuclear HIF-1α was observed in excisions of advanced melanotic vs. amelanotic melanomas. In cultured cells, melanogenesis also significantly stimulated expression of classical HIF-1-dependent target genes involved in angiogenesis and cellular metabolism, including glucose metabolism and stimulation of activity of key enzymes in the glycolytic pathway. Several other stress related genes containing putative HRE consensus sites were also upregulated by melanogenesis, concurrently with modulation of expression of HIF-1-independent genes encoding for steroidogenic enzymes, cytokines and growth factors. Immunohistochemical studies using a large panel of pigmented lesions revealed that higher levels of HIF-1α and GLUT-1 were detected in advanced melanomas in comparison to melanocytic nevi or thin melanomas localized to the skin. However, the effects on overall or disease free survival in melanoma patients were modest or absent for GLUT-1 or for HIF-1α, respectively. In conclusion, induction of the melanogenic pathway leads to robust upregulation of HIF-1-dependent and independent pathways in cultured melanoma cells, suggesting a key role for melanogenesis in regulation of cellular metabolism.

A quantitative method for assessing stages of the rat estrous cycle.

The impact of gender and/or hormone variations on a wide variety of neural functions makes the choice between studying males or females (or both) of a given species difficult. Although female rats are widely used experimentally, few studies control for the stage of estrus. More detailed information about how to distinguish the various stages of the estrous cycle is needed. For the present study, vaginal smears were obtained once a day and stained using an adaptation of the Papanicolaou (PAP) procedure. Images are provided of unstained "wet" samples and the corresponding PAP stained smears illustrating the cellular profile for each stage of the cycle as well as post-ovariectomy. The different cell populations across the cycle were quantified and ratios determined to show trends between the predominant and other cell types in each stage of the estrous cycle. Both stained and unstained images and cell quantification data provide valuable guidelines for distinguishing the stages of the estrous cycle.

Comparison of practical treatment methods to eradicate pinworm (Dentostomella translucida) infections from Mongolian gerbils (Meroines unguiculatus).

This study evaluates the efficacy of various treatment methods to eradicate Dentostomella translucida from Mongolian gerbil colonies. The following five treatment methods were instituted in naturally infected groups of 10 gerbils each: topical ivermectin misting, ivermectin-medicated drinking water, piperazine citrate-medicated drinking water, fenbendazole-medicated feed, and a combination of ivermectin-medicated drinking water and fenbendazole-medicated feed. Treatment success was assessed by using weekly fecal flotations, with necropsy examinations performed on fecal-negative gerbils (except those in the misted group) at 5 weeks after the last treatment. Topical ivermectin misting left 40% of gerbils fecal-positive. With piperazine citrate-medicated drinking water, 60% of the gerbils were fecal-positive; the remaining 40% had adult worms in their digestive tract at necropsy. Ivermectin-medicated drinking water caused 80% of the gerbils to be negative on fecal flotation. On necropsy, however, all but one of these gerbils harbored adult pinworms. Treatments with fenbendazole-supplemented feed alone or in combination with ivermectin-treated water resulted in no fecal shedding or evidence of adult pinworms on necropsy examination. Of the five treatments evaluated, only those using fenbendazole-medicated feed (150 ppm) provided a practical and reliable treatment method to eradicate pinworm infections in Mongolian gerbil colonies.

Antipsychotic medications and the elderly: effects on cognition and implications for use.

Despite being frequently prescribed in the elderly, antipsychotic medications are commonly associated with adverse effects in this population, including sedative, orthostatic and extrapyramidal adverse effects. Growing evidence suggests that antipsychotics can also cause deleterious cognitive effects in some elderly patients. Preclinical and growing clinical evidence indicates that inhibitory effects on dopaminergic, cholinergic and histaminergic neurochemical systems may account for antipsychotic-associated cognitive impairment in the elderly. A review of published reports of the cognitive effects of antipsychotics in the elderly suggests that newer antipsychotic medications may possess a more favourable cognitive profile than that of traditional agents in this population. The cognitive effect that a specific antipsychotic will have in the elderly, however, is likely better predicted by considering the pharmacodynamic action of an individual agent in combination with the pathophysiology of the condition being treated. Agents with relatively weak dopamine inhibiting effects (e.g. clozapine and quetiapine), for example, would theoretically have a cognitive profile superior to that of agents with higher degrees of dopaminergic inhibition (all traditional agents, risperidone, olanzapine and ziprasidone) when used for conditions associated with diminished dopamine function (e.g. idiopathic Parkinson's disease). Drugs with weak anticholinergic effects (high-potency traditional agents, risperidone, quetiapine and ziprasidone) would theoretically be less likely to cause cognitive impairment than agents with high degrees of cholinergic receptor blocking actions (clozapine and olanzapine) when treating patients with impaired cholinergic function (e.g. Alzheimer's disease). Cholinergic agonist effects of clozapine and olanzapine may, however, mitigate potential adverse cognitive effects associated with the cholinergic blocking actions of these agents. Large, rigorous trials comparing the cognitive effects of antipsychotics with diverse pharmacodynamic actions are lacking in the elderly and are needed.

Evaluation of commercially available Escherichia coli J5 bacterin as protection against experimental challenge with Pasteurella multocida in rabbits.

OBJECTIVE: To evaluate the ability of commercially available Escherichia coli J5 bacterin to protect rabbits from experimental challenge with Pasteurella multocida. ANIMALS: 40 P multocida-free New Zealand White rabbits. PROCEDURES: Rabbits were assigned to 1 of 4 groups of 10 rabbits each. Three of the groups were inoculated SC with J5 bacterin at 8 weeks old. Inoculation was repeated 3 and 6 weeks later. The fourth group was not inoculated and served as controls. Groups 1, 2, and 3 were given 10(9), 10(8), and 10(7) colony forming units (CFU), respectively. Response was monitored by titer assessment, using an E coli J5 antigen capture ELISA. Five weeks after the last inoculation, all rabbits were challenged with P multocida and observed for an additional 5 weeks. Clinical, hematologic, serologic, culture, and necropsy data were collected. RESULTS: Inoculation of rabbits with 10(9) CFU of E coli J5 bacterin-induced titers that were significantly greater than titers of rabbits vaccinated with 10(8) or 10(7) CFU or those in controls. The incidence of acute bacteremia was lower in rabbits with high titers. At necropsy, prevalence of lesions typical of P multocida was not significantly different among groups. Prevalence of histologic lesions was also not significantly different among groups. CONCLUSIONS AND CLINICAL RELEVANCE: Although the bacterin induced considerable antibody response and possibly reduced the rate of bacteremia, antibodies were not protective against long-term colonization or infection of the frontal sinuses or tympanic bullae by the challenge strain of P multocida. This bacterin in its currently available form is unlikely to aid in reducing the prevalence of pasteurellosis in rabbits.

The observation of reactive thrombocytosis in New Zealand white rabbits in response to experimental Pasteurella multocida infection.

Reactive thrombocytosis is an increase in the circulating thrombocyte count secondary to a physiologic process within the body, often an infection. Reactive thrombocytosis is different than primary or essential thrombocytosis which is usually related to myeloproliferative neoplasia. Essential thrombocytosis is most common in adults, whereas reactive thrombocytosis is most frequently observed in children. Reactive thrombocytosis has been occasionally reported in cats, dogs and horses but has not been previously reported in the rabbit. Rabbits were challenged with virulent Pasteurella multocida. Hematologic, clinical, and culture assessments were performed prior to challenge, enabling each animal to serve as its own control. The questions asked were whether reactive thrombocytosis was a consistent phenomena and whether its presence and/or intensity was related to disease severity. All challenged rabbits demonstrated some degree of thrombocytosis in response to the infection, but individual rabbits were varied in their pattern of thrombocytosis. Elevations varied from intense to mild to undulating with durations of 1 to 11 days above 500 x 10(9)/L and 0 to 5 days above 650 x 10(9)/L. Correlation analysis was unable to demonstrate significant association between thrombocytosis, body temperature, leukocyte count, or the granulocyte lymphocyte ratio (all r < 0.2). No significant association between intensity of thrombocytosis and degree or type of pathologic lesions was observed. Thrombocytosis does not appear predictive of disease intensity or outcome. The data indicate that in the rabbit thrombocytosis is a consistent response to infection with P. multocida. Rabbits may serve as a model for the study of reactive thrombocytosis, in humans especially in children infected with Haemophilus sp., which are also a members of the bacterial family Pasteurellaceae.

Seroprevalence of antibodies against gram-negative core antigens in rabbits, using an Escherichia coli J5 antigen-capture enzyme-linked immunosorbent assay.

OBJECTIVE: To determine the seroprevalence of antibodies to gram-negative core antigens (GNCA) in specific-pathogen-free (SPF) rabbits (ie, free of Pasteurella multocida) and rabbits of undefined bacterial status (conventional). SAMPLE POPULATION: Serum samples were obtained from 7 groups of rabbits. The SPF rabbits comprised 2 adult groups and 1 immature group, whereas the 4 groups of conventional rabbits were all adults. PROCEDURE: A seroprevalence survey was conducted on rabbit sera for antibodies against GNCA, using an Escherichia coli J5 antigen-capture ELISA. RESULTS: Collective geometric mean titer (GMT) of adult rabbits was 1:6,463. The GMT of each of the 6 groups of adult rabbits was 1:956, 1:1,133, 1:4,525, 1:5,338, 1:7,669, and 1:25,600. Titers of populations differed significantly. CONCLUSION: Data analysis revealed there were anti-GNCA antibodies in rabbits. Similar to other species, the prevalence of IgM and IgG anti-GNCA antibodies increased with age. The IgG response was more marked than the IgM response. The SPF rabbits had lower IgG anti-GNCA titers than conventional rabbits, indicating possible cross-reactive epitopes between P multocida and Enterobacteriaceae. Rabbits with the highest anti-GNCA titers were those used in polyclonal antibody production, possibly stemming from endotoxin contamination of antigen or adjuvant. CLINICAL RELEVANCE: The possible cross-reactive antibodies directed at homologous wall components of Pasteurellaceae and Enterobacteriaceae could prove to be a possible heterotypic vaccination strategy for the protection of rabbits against pasteurellosis. Investigators should determine whether antigen impurity (endotoxin contamination) influences epitope focus during polyclonal antibody production and whether it affects sera variability among rabbits.

Serologic survey of free-living nestling prairie falcons (Falco mexicanus) for selected pathogens.

Serum samples from 34 free-living nestling prairie falcons (Falco mexicanus) in southwestern Idaho were negative for antibodies to avian influenza virus, Newcastle disease virus, and three Aspergillus species. Serum from a single bird had hemagglutinating inhibition activity in response to Mycoplasma synoviae, and another bird's serum had slight activity in response to M. gallisepticum.

A pathological and immunohistochemical study of goat kids undergoing septicaemic disease caused by Mycoplasma capricolum subsp. capricolum, Mycoplasma mycoides subsp. capri and Mycoplasma mycoides subsp. mycoides (large colony type).

This paper studies the pathological and immunohistochemical findings in 12 kids experimentally infected with Mycoplasma capricolum subsp. capricolum (Mcc), M. mycoides subsp. capri (Mmc) and M. mycoides subsp. mycoides (large colony type), (MmmLC). For the demonstration of Mcc, Mmc and MmmLC antigens an immunoperoxidase technique based on the labelled streptavidin biotin method was used in the 12 kids inoculated with mycoplasmas and the three control kids inoculated with mycoplasma medium. All 12 kids, inoculated by different routes, developed subcutaneous swelling at the point of inoculation and terminated in fatal septicaemia from 1 to 5 days post inoculation. The histopathological findings consisted of cellulitis at the point of inoculation, acute diffuse interstitial pneumonia, arthritis and multifocal necrotic purulent splenitis in some kids. The Mcc, Mmc and MmmLC antigens were detected immunohistochemically in all kids with specific punctiform labelling inside the cytoplasm of the leucocytes or extracellularly at the inoculation point, respiratory airways, spleen, liver, joints, tonsils and lymph nodes. The results obtained in this study showed that the inoculation of these mycoplasmas by parenteral routes caused mycoplasmaemia. Moreover, the immunohistochemical results appear fully to confirm that the mycoplasmas were the cause of the death of the kids because of a septicaemic state.

Incidence and risk factors for bumblefoot (pododermatitis) in rehabilitated raptors.

A retrospective cohort study was conducted on 821 raptors of 12 representative species, admitted to the California Raptor Center (CRC), during 1980-1990. The incidence rate for bumblefoot was 52 cases per 100 bird-years at risk. Eagles and hawks (buteos) were more likely to develop bumblefoot, and did so earlier during their captivity than other species. Also, raptors admitted with a limb fracture had the greatest risk (OR = 4.2) of developing bumblefoot than any other condition on entry. Median time from admission to development of bumblefoot was 52 days, and median duration of bumblefoot was 23 days.

Polymerase chain reaction and restriction endonuclease digestion for selected members of the "Mycoplasma mycoides cluster" and Mycoplasma putrefaciens.

A specific diagnostic method using the polymerase chain reaction, together with restriction endonuclease digestion patterns, was developed for members of the "Mycoplasma mycoides cluster" that normally occur in the United States (i.e., Mycoplasma mycoides subsp. mycoides Large Colony and Mycoplasma capricolum subsp. capricolum in addition to "cluster" mycoplasma, bovine serogroup 7, and Mycoplasma putrefaciens. The digestion of "cluster" polymerase chain reaction DNA (1,225 bp) amplification products with restriction enzymes AseI and SspI gave mycoplasma species-specific patterns for all strains of M. mycoides subsp. mycoides Large Colony, M. capricolum subsp. capricolum, and bovine group 7 tested. Moreover, we found a nonspecific amplification product for M. putrefaciens that occurred with the oligonucleotide primers used for the "M. mycoides cluster" reaction. However, the restriction endonuclease digestion patterns observed with the restriction enzymes AluI, AseI, and SspI for M. putrefaciens were different than the digestion patterns obtained for the other "cluster" mycoplasmas. This report confirms the usefulness of polymerase chain reaction DNA amplification allied with restriction enzyme digestion profile analysis for the rapid and specific identification of mycoplasmas belonging to the "M. mycoides cluster" and M. putrefaciens.

Lesions associated with Pasteurella multocida infection in raptors.

Several case reports attest to the pathogenicity of Pasteurella multocida in raptors; however, the pathologic syndromes have not been fully described. We describe here the lesions encountered in 22 avian cholera cases in raptors. Besides septicemia-related lesions, a unique syndrome of esophageal abscesses was noted in 8 of the 11 (73%) Buteo hawks that succumbed to avian cholera. Esophageal abscesses were not noted in birds belonging to the order Strigiformes (owls) or family Falconidae (falcons and their relatives). Thus, the presence of white plaques in both the oropharynx and esophagus of Buteo hawks may indicate a possible P. multocida infection and should be considered in the differential diagnosis. This study also documents the first cases of avian cholera in a rough-legged hawk (Buteo lagopus) and a flammulated owl (Otus flammeolus).

Pasteurella multocida in psittacines: prevalence, pathology, and characterization of isolates.

Although the pathogenicity of Pasteurella multocida for psittacines (parrots and their relatives) has been documented in several case reports, the associated pathologic syndromes have not been well defined nor have the isolates been characterized. In addition, the prevalence of P. multocida in psittacines has not been determined. Three hundred twenty-eight psittacines (253 clinically healthy and 75 clinically ill) were cultured for P. multocida. Pasteurella multocida was not isolated from the pharynx, choana, or cloaca of psittacines. However, in five dead psittacines submitted for necropsy, P. multocida was isolated. These isolates were characterized, and all belonged to either somatic serotype 3 or 4,7. Pasteurella multocida somatic serotype 3 was isolated from psittacines with septicemia, whereas P. multocida somatic serotype 4,7 was isolated from psittacines with cutaneous lesions. The majority (four out of five) of the P. multocida isolates belonged to the subspecies multocida, and all isolates were susceptible to penicillin G, sulfisoxazole, gentamicin, erythromycin, tetracycline, and trimethoprim-sulfamethoxazole but resistant to streptomycin. DNA fingerprints demonstrated that isolates belonging to the same somatic serotype were genetically related. The isolate from a cockatiel that had been caught by a cat belonged to somatic serotype 3 and was not genetically related to the other two isolates belonging to this somatic serotype.

Pasteurella multocida in raptors: prevalence and characterization.

Several cases dealing with Pasteurella multocida infection have been documented in raptors. However, the isolates have not been fully characterized nor has the prevalence of P. multocida in raptors been determined. Three hundred ninety-eight raptors were cultured for P. multocida. Results indicated that P. multocida was not normally carried in the pharyngeal, choanal, or cloacal regions. However, P. multocida was isolated from raptors with avian cholera. Isolates from eight cases were characterized by biotype, somatic serotype, and antibiogram. Most (six of eight) of the P. multocida isolates belonged to somatic serotype 1. The remaining two P. multocida isolates belonged to somatic serotypes 3 and 3,4. The majority of the isolates belonged to the subspecies multocida. All isolates were susceptible to penicillin G, sulfisoxazole, tetracycline, and trimethoprim-sulfamethoxazole. Various restriction site heterogeneities of P. multocida chromosomal DNA were found among the raptor isolates. Results indicated that isolates of P. multocida somatic serotype 1 from diurnal raptors were genetically related.

Immunohistochemical and ultrastructural characteristics of Mycoplasma capricolum subsp. capricolum in caprine abortion: a case report.

Described in this study are the immunohistochemical and ultrastructural findings in a case of caprine abortion due to the experimental infection of the dam with strain GM13 of Mycoplasma capricolum subsp. capricolum. Mycoplasma antigens were seen mainly in choriallantoic trophoblasts and in the lumen of blood vessels in the allantoic membrane. Examination with an electron microscope showed that the chorioallantoic trophoblasts were filled with typical mycoplasma organisms. No other bacteria were observed in any of the samples. Our results confirm by immunohistochemical and electron microscopic techniques that Mycoplasma capricolum subsp. capricolum can cause caprine abortion and that the process can occur without premonitory signs.

Ixodes pacificus (Acari: Ixodidae) as a vector of Ehrlichia equi (Rickettsiales: Ehrlichieae).

Ehrlichia equi, a rickettsia described from horses in California 30 yr ago, causes equine granulocytic ehrlichiosis throughout the Americas and possibly Europe. Here, we report experimental transmission of E. equi from infected to susceptible horses through bites of western blacklegged ticks, Ixodes pacificus (Cooley & Kohls). In preliminary field studies, only I. pacificus consistently infested horses and vegetation at 3 locations with contemporary cases of equine ehrlichosis, and in particular, I. pacificus was the only species found attached to all of the infected horses. Exposure to bites of ticks in the genus Ixodes poses previously unrecognized and serious health risks to humans and animals.

Experimental oral inoculations in birds to evaluate potential definitive hosts of Neospora caninum.

Experimental oral inoculations to evaluate potential definitive hosts of Neospora caninum were conducted by feeding infected rodent tissues to 9 carnivorous birds of 4 species. Birds included 2 red-tailed hawks (Buteo jamaicensis), 2 turkey vultures (Cathartes aura), 2 barn owls (Tyto alba), and 3 American crows (Corvus brachyrhynchus). The rodents (mice or rats) had been inoculated with 100,000 culture-derived tachyzoites of N. caninum 1-6 mo before feeding to the birds. Fecal samples were collected from each bird daily for 1 mo after feeding rodents and examined for oocysts by fecal flotation. In addition, processed aliquots from all avian fecal samples were fed to BALB/c mice. Five weeks after feeding, mice were bled and sera were tested for antibodies against N. caninum. One to two months later, mice were killed and brain tissue was examined microscopically for protozoal cysts. While occasional oocysts were found in avian fecal samples, these were likely not N. caninum because they were not infective to BALB/c mice. It was concluded that the bird species tested are not likely to be definitive hosts of N. caninum.

Pharmacokinetic properties of methadone hydrochloride after single intramuscular administration in adult dairy goats.

Pharmacokinetic parameters of methadone were studied in adult dairy goats. Five goats were each given methadone hydrochloride as a single 0.2 mg/kg of body weight dosage by intramuscular (IM) administration. Plasma methadone concentrations were determined for 96 h after dosing. Plasma methadone concentrations after IM administration were best described by an open one-compartment model. Overall elimination half-life (t1/2) was 1.38 h. Peak plasma concentrations were reached 0.25 h after dosing, and the actual plasma concentration averaged 37.8 ng/ml (SD = 12.76) at that time. The data obtained from this study suggest that plasma concentrations, similar to those that are analgesic in humans, can be achieved after IM administration of methadone at a dose rate of 0.2 mg/kg of body weight. In addition, these plasma concentrations can be maintained for up to 3 h after a single injection and, therefore, may provide satisfactory analgesia for such period.