Dog Serum Proteins Research Articles

Proteomics identification of overexpressed serum proteins in dogs with Babesiacanis infection.

Canine babesiosis, caused by the protozoan parasite Babesia canis, is characterized by clinical manifestations, including hemolytic anemia, thrombocytopenia, multiple organ failure, and may result in death. This disease is detected using conventional blood smears, which are time-consuming and have low sensitivity. This study aimed to investigate a more rapid and sensitive method for detecting B. canis infection in dogs by examining the expressed serum protein profiles using proteomics. We collected six sera samples from three healthy and three B. canis-infected dogs diagnosed using blood smear and polymerase chain reaction. We analyzed the proteins using two-dimensional gel electrophoresis. The candidate spots from the gel were subjected to protein identification using a nano-liquid chromatography system coupled to an ion-trap mass spectrometer equipped with an electrospray ionization nano-sprayer. We found that 10 protein spots were overexpressed in the serum samples from infected dogs compared with healthy dogs, which corresponded to three proteins: serotransferrin, serotransferrin isoforms X1, and hemopexin. Furthermore, analysis of the protein-protein interaction network confirmed that they strongly interacted with each other. This study suggests that high levels of serotransferrin and hemopexin are related to B. canis infection, making these proteins potential candidates for the development of diagnostic molecules or vaccines.

Identification of serum proteins in dogs naturally infected with Anaplasma phagocytophilum and Borrelia burgdorferi: a pilot study

Serum proteomic analysis would aid in better understanding the pathophysiology of several diseases. The aim of this study was to identify the serum proteomes of dogs with anaplasmosis and Lyme disease using a proteomic approach. Diseases were diagnosed by a commercial rapid in-clinic ELISA. Borrelia antibodies were evaluated by IFAT. Four groups were designated: symptomatic dogs with anaplasmosis (n=5), dogs with Lyme disease (n=5), dual-positive dogs (n=5), and healthy control dogs (n=5). Serum samples were collected before treatment. Two-dimensional electrophoresis of pooled samples in each group were run in triplicate. Ten out of 57 differentially expressed spots between groups were evaluated for identification by mass spectrometry. Compared to those of controls, levels of vitamin D-binding protein (VDBP), glycoprotein-9 (GP9) and kininogen-1 (KGN-1) decreased, while haptoglobin (Hp) and immunoglobulin (Ig) heavy chain levels increased in dual infection group. Serum apolipoprotein-A1 (Apo-A1) levels decreased in dogs with anaplasmosis, Lyme disease and dual infections compared to those in control dogs. Serum clusterin levels decreased in dogs with anaplasmosis but were not differentially expressed in dogs with Lyme disease or dogs with dual infections compared to those in control dogs. Calpain-3 decreased in dogs with anaplasmosis and Lyme disease. This study showed that many protein levels might be changed in dogs with naturally acquired anaplasmosis and Lyme disease. Understanding the role of these proteins in different biological processes can provide information of interest for diagnostic and therapeutic approaches for these clinical conditions.

The electrophoretic pattern of serum proteins in dogs with babesiosis

This study was aimed at the evaluation of the electrophoretic pattern of serum proteins in dogs naturally infected withBabesia canis. Blood samples were collected from 37 dogs infected withB. canisand showing clinical signs consistent with the disease. The sick animals were classified as dogs with physiologic and decreased red blood cell (RBC) values. Twenty-five healthy dogs formed the control group. The concentrations of total proteins and protein fractions were measured in blood serum. The values of total proteins, albumin and albumin/globulin (A/G) ratio in dogs with babesiosis were significantly lower than in healthy ones (P< 0.001). In the globulin fractions, significantly higher relative concentrations of α1-, β1- and β2-globulins (P< 0.01), and non-significantly higher values of α2- and γ-globulins were found in dogs with babesiosis with a double α2-zone in six out of 37 animals. Marked differences were observed also between the two groups of sick animals, with significantly lower values of albumin and A/G ratio (P< 0.05), and significantly higher values of α1- and β1-globulins in dogs with decreased RBC (P< 0.05 andP< 0.01, respectively). Presented results indicate marked alterations in the electrophoretic pattern of serum proteins in dogs with babesiosis suggesting its usefulness for the evaluation of pathophysiological changes caused by the disease and for diagnostic of disease severity.

Changes in serum proteins in dogs with Ehrlichia canis infection

The aim of this study was the identification of proteins differentially represented in the serum proteome of seropositive dogs with (Group 1) and without (Group 2) clinical-pathologic signs consistent with ehrlichiosis compared to healthy control dogs. Serum samples were collected from 20 dogs of various breeds with naturally occurring ehrlichiosis (10 dogs belonged to Group 1 and 10 to Group 2) and 10 healthy dogs. Two-dimensional electrophoresis (2DE) of pooled serum for each of the group of dogs were run in triplicate. 2D image analysis showed 39 spots differently expressed between Group 1 and Group 2 compared with healthy ones. Mass spectrometry analysis allowed identification of 6 proteins: albumin, haptoglobin (Hp), alpha-1-antitrypsin (AAT), Retinol Binding Protein 4 (RBP-4), alpha-1-acid glycoprotein (AGP) and vitamin D-binding protein (VDBP). When a confirmatory study was performed for albumin, Hp, AAT and RBP-4 by using different assays, significant differences (P < 0.05) between diseased and healthy groups were observed. It can be concluded that there are significant changes in the serum proteome of dogs with ehrlichiosis with modifications in proteins related with the acute phase response such as Hp, albumin and AGP, with vitamin A transport such as RBP-4, with inhibitors of serine proteases and anti-inflammatory proteins such as AAT, and vitamin D metabolism and actin scavengers such as VDBP.

Concentration of Lipocalin Region of Collagen XXVII Alpha 1 in the Serum of Dogs with Hemangiosarcoma

Hemangiosarcoma (HSA) is a common malignancy of dogs with characteristic early, aggressive metastasis. Diagnosis of HSA is challenging because of lack of sensitive and specific diagnostic tests. Specific proteins that are increased in serum of dogs with HSA might represent useful biomarkers of the disease. Thirty-four dogs with HSA and 42 healthy dogs from the Ontario Veterinary College Teaching Hospital. This case-control study compared serum proteins in dogs with HSA and healthy dogs. Proteins were separated by 2-dimensional difference gel electrophoresis and identified by liquid chromatography and tandem mass spectrometry. Western blot analysis showed that serum collagen XXVII peptide concentration in serum of dogs with large metastatic HSA burdens (1,488, 231-3,754 DU; median, minimum-maximum); was, on average, 9.5-fold higher than in healthy dogs (156; 46-2,101 DU). While concentrations for dogs with osteosarcomas (678; 124-3,251 DU), lymphomas (423; 92-2,777 DU), carcinomas (1,022; 177-3,448 DU), and inflammatory disease were also increased, values were consistently lower than those for HSA. Receiver operating characteristic curves revealed an estimated area under the curve of 83% for HSA cases whereas areas for other neoplastic and nonneoplastic diseases were nondiscriminatory. Serum collagen XXVII peptide concentration before splenectomy (1,350; 1,156-1,929 DU) was reduced after tumor removal (529; 452-562 DU) and chemotherapy but increased in 2 dogs with tumor recurrence (511-945 DU; 493-650 DU). Collagen XXVII peptide might be useful for diagnosis and monitoring of advanced HSA.

Pharmacokinetics of a new quinolone, AM-833, in mice, rats, rabbits, dogs, and monkeys.

The pharmacokinetics of AM-833 [6,8-difluoro-1-(2-fluoroethyl)-1, 4-dihydro-7-(4-methyl-1-piperazinyl)-4-oxo-3-quinolinecarboxylic acid] were studied in mice, rats, rabbits, dogs, and monkeys by reversed-phase high-performance liquid chromatography. AM-833 was rapidly and completely absorbed from the digestive tracts of mice, rats, and dogs. About half of AM-833 bound to rat and dog serum proteins. Drug levels in lung, spleen, liver, and kidney tissues of rats and dogs were greater than the respective levels in serum but lower in brain tissue. Drug levels in tissues declined with the decrease in levels in serum. AM-833 penetrated rapidly and well into inflammatory exudate of rats. Elimination half-lives in serum were species dependent, ranging from 1.57 h in rabbits to 9.42 h in dogs. Profiles of drug levels in serum were dose related over a single dose range from 2 to 40 mg/kg and not modified significantly during multiple dosing in dogs. Unchanged AM-833 was excreted in urine and bile in both rats and dogs. The metabolism of AM-833 was suggested by evidence that 24-h total recovery of unchanged AM-833 in urine and bile accounted for about half of the intravenous dose in rats.

Breed-specific dog hypersensitivity in humans

The contribution of allergenic components of dander and serum from individual dog breeds in human hypersensitivity and the different responses to the various breeds in vivo and in vitro were examined. Differences in IgE antibodies directed against 13 individual dog breed danders and five different breed-specific sera were measured in groups sensitive and nonsensitive to dogs by the radioallergosorbent test (RAST). Cross-reactivity between dander extracts and sera was examined by RAST inhibition. Mean RAST binding to dog danders and sera was 15.9% and 14%, respectively, for the dog-sensitive group versus 0.6% for both danders and sera in the nonsensitive group. The mean difference in binding between groups was statistically different for all dander extracts and sera. The variances in binding were statistically greater for danders than sera in six of eight dog-reactive subjects. RAST binding and intradermal skin tests at a 1:10,000 w v dilution correlated well (p < 0.05) for seven of nine breed-specific extracts. RAST inhibition studies showed that dog serum proteins were relatively poor inhibitors of dander extracts, and vice versa. Variability of skin test and RAST responses to different dog breed dander extracts support the existence of breed-specific allergens.

Characterization and serum inhibition of neutral collagenase from cultured dog gingival tissue

1. 1. Explants of dog gingiva, maintained in culture for 9 days in the absence of serum, released a collagenase (EC 3.4.24.3) into the medium. The yield of active enzyme reached a maximum after 5–8 days with concomitant release of collagen degradation products from the explants. 2. 2. The enzyme attacked undenatured collagen in solution at 25°C resulting in a 58% loss of specific viscosity and producing the two characteristic products TC A ( 3 4 ) and TC B ( 1 4 ) . Electron microscopy of segment-long-spacing crystallites of these reaction products showed the cleavage locus of the collagen molecule at interband 40. 3. 3. Optimal enzyme activity was observed over the pH range 7.5–8.5 and a molecular weight of approximately 35 000 was derived from gel filtration studies. EDTA, 1,10-phenanthroline, cysteine and dithiothreitol all inhibited collagenase activity. Proteoglycan derived from porcine and human cartilage did not inhibit the enzyme. 4. 4. The enzyme was inhibited by the dog serum proteins α 2-macroglobulin and a smaller component of molecular weight approximately 40 000. This smaller component was purified by column chromatography utilising Sephadex G-200, DEAE A-50, and G-100 (superfine grade). Agarose electrophoresis of the purified component showed it to represent a β-serum protein. α 1-Antitrypsin did not inhibit the enzyme. 5. 5. The physiological importance of the natural serum inhibitors and gingival collagenase are discussed in relation to latent enzyme and periodontal disease.

Immunochemical studies of the protein fraction of pulmonary surface active material.

SUMMARy _ Rabbits were immunized to pulmonary surface active material that was isolated from lungs of dogs. The resulting antiserum was directed against 3 antigenic components, 2 of which were im­ munologically identical to dog serum proteins. The third antigen was not contained in dog serum or plasma. Immunofluorescent labeling indicated that this antigen originated in the alveoli of dog lungs.

The Opsonins of Normal and Immune Sera

Summary and Conclusions No relationship exists between the hemolytic complementary and opsonic powers of the guinea-pig, human, monkey (rhesus), dog, cat and sheep sera toward staphylococci. Normal sheep and bovine sera show the highest opsonic values. The normal opsonins of guinea-pig, human, monkey, sheep, dog, cat and rabbit sera are for the most part thermolabile. The CO2-soluble fraction of the guinea-pig serum which contains the endpiece and its associated fourth component of complement carries also the antistaphylococcic opsonic agent of this serum. In the human serum the CO2-soluble fraction also contains the greater portion of the normal human serum antistaphylococcic opsonin. In the serum of the rhesus monkey the antistaphylococcic opsonins appear to be distributed between the CO2-soluble and CO2-insoluble fractions, the major portion being in the CO2-soluble fraction. Sheep-serum antistaphylococcic opsonins are readily destroyed by heat as well as by the relatively mild agents employed in the fractionation of the guinea-pig serum. One-third of the sheep-sera examined showed exceedingly high opsonic values. A separation of the dog-serum proteins by CO2 demonstrates that the anti-staphylococcic opsonic factor or factors are not carried independently by either the CO2-precipitate or by the CO2-soluble fractions. A combination of these two fractions yields a full opsonic value. The antistaphylococcic opsonins of cat-serum appear to be distributed between the CO2-soluble and the CO2-insoluble fractions, the major portion occurring in the CO2-soluble fraction. The treatment of rabbit-serum with zymin greatly enhances its antistaphylococcic opsonic value. Ammonia does not destroy this opsonic activity. CO2, however, markedly destroys the antistaphylococcic opsonin of this serum. The treatment of staphylococci with concentrated human serum (1:1) and subsequently with an antistaphylococcic serum also in a 1:1 dilution leads to a reduction in the number of cocci ingested by human leucocytes. A similar result is obtained when the cocci are first heated with the immune serum. In the latter case, however, and at the optimal concentration of the immune serum, the addition of the inactivated human serum definitely increases the number of cocci ingested. In general, rhesus monkey-serum gives the same results. At optimal dilutions of the monkey-serum and the antiserum an actual enhancement of phagocytosis is observed. Phagocytosis of staphylococci treated with concentrated sheep-serum and later with a highly diluted antistaphylococcic serum is markedly enhanced. This is, however, not the case when the cocci are first treated with the immune serum prior to the addition of the diluted normal serum. Dilution of the sheep-serum or inactivation of this serum leads to a great reduction in the opsonic effects of the combined sera. The treatment of staphylococci with a 1:40 normal dog-serum and 1:64 anti-staphylococcic serum results in an enhancement of phagocytosis. Inactivated dog-serum does not show this synergistic effect. Cat-serum shows a distinct enhancement of phagocytosis in the proper concentrations of cat-serum and the immune serum. Even inactivated cat-serum in the presence of optimal amounts of the immune serum, shows this enhancing action. Rabbit-serum also shows a less marked enhancing effect when the cocci are first treated with the optimal concentration of the antiserum followed by a very dilute normal rabbit-serum. Inactivated rabbit-serum does not enhance phagocytosis in the presence of the antistaphylococcic serum. The differences encountered in the human, monkey- and cat-sera as compared with the sera of the other species investigated are probably due to the fact that these sera contain along with the thermolabile opsonin also a thermostable opsonin. It is therefore difficult to differentiate the effects of these two opsonins.

A PRECIPITIN REACTION OF THE DIGESTIVE MUCOSA OF THE DOG

By simply extracting the mucous membrane of the dog stomach or intestine in physiologic sodium chlorid solution, antigens are obtained that differ in precipitin reaction from the proteins of dog serum. After two days without food, the dog is etherized, bled to death from the aorta, and thoroughly perfused with salt solution after the cava and portal vein have been opened. The mucous membrane is scraped off, washed in water, and extracted for twenty-four hours in a salt solution to which a little thymol has been added. The extracts are neutral in reaction, and with rare exceptions contain measurable quantities of dissolved protein substances, which can be separated into globulin and albumin fractions by means of ammonium sulphate. Prepared as described, the extracts of dog stomach or intestine may not give any reactions at all with precipitin serums for dog serum or for the isolated dog serum proteins (euglobulin, pseudoglobulin,