Background: Separation-related disorders are prevalent in dogs and can result in problem behaviours such as inappropriate elimination, destructive behaviour, and distress vocalizations which occur when an owner is abs...Background: Separation-related disorders are prevalent in dogs and can result in problem behaviours such as inappropriate elimination, destructive behaviour, and distress vocalizations which occur when an owner is absent or perceived as absent. Numerous natural products are marketed for managing anxiety in dogs;however, few, if any, of these products have documented evidence of efficacy in veterinary behaviour. Purpose: This open, multicentric clinical trial aimed to assess the efficacy of a new supplement containing alpha-casozepine and fish muscle hydrolysate on the separation-related problem in dogs and the impact on the quality of life of dogs and their pet owners. Material and Methods: Fifty-one dogs with clinical signs compatible with separation-related problems were included. The dogs were treated with a complementary feed (Zylkene Plus, Vetoquinol, France) for 30 days. Separation-related behaviour assessment (SRP), emotional disorders assessment (EDED) and the quality of life (QoL) of the pets and their pet owner were assessed at baseline and the end of the study by the veterinarians. In addition, owners were asked to rate their dog’s SRP weekly after enrolment using the same questionnaire that the veterinarian used at the beginning of the study. Results: There was a statistically significant improvement in the SRP, EDED, and QoL scores (P Conclusion: The results of this study support the hypothesis that the combination of alpha-casozepine and fish muscle hydrolysate is a promising and effective option to reduce the frequency and severity of separation-related behaviour in dogs and improve the quality of dogs and their owners.展开更多
Antinuclear antibodies are found in animals suffering from Systemic Lupus Erythematosus (SLE) and some other diseases, their presence in the blood is determined by antinuclear antibody (ANA) test using indirect immuno...Antinuclear antibodies are found in animals suffering from Systemic Lupus Erythematosus (SLE) and some other diseases, their presence in the blood is determined by antinuclear antibody (ANA) test using indirect immunofluorescence (IF) with HEp2 cells as a substrate. In this work, an immunoperoxidase (IP) assay was developed to evaluate the ANAs in canine sera, using canine kidney cell lines (MDCK) and compared with a commercial immunofluorescence test on Hep2 cells for this system, a fluoresceinated anti-canine Ig antibody was standardized. The study was performed on 50 sera from dogs submitted to the laboratory with different clinical diagnoses of autoimmune-associated diseases. The procedures on both cells were unified to perform comparisons of the reactions, direct sera or at different dilutions were added to a monolayer of permeabilized MDCK cells, followed by a peroxidized anti-canine IgG conjugate, and a substrate for the IP reaction. The same sera were tested on the commercial IF assay on Hep2 cell system. In 22/50 cases, the presence of LE cells in peripheral blood was determined. A high correlation was found in the detection of antinuclear antibodies between both cell lines and techniques, however there were differences in the reaction patterns in the nucleus and cytoplasm between cell lines. The diffuse nuclear pattern observed in MDCK cells was more related to the presence of high percentages of LE cells in peripheral blood. The differences found in the results were possibly associated with the presence of homologous antigens between the MDCK cells and the dog. In addition, the methodology and standardization for the use and interpretation of a reference serum was developed to unify the interpretation criteria in the laboratory.展开更多
Canine parvovirus(CPV)is a high morbidity and lethality virus which causes severe enteric disease in dogs.In an attempt to gain canine-origin neutralizing antibodies against CPV,single chain variable fragment(scFv)bac...Canine parvovirus(CPV)is a high morbidity and lethality virus which causes severe enteric disease in dogs.In an attempt to gain canine-origin neutralizing antibodies against CPV,single chain variable fragment(scFv)bacteria display libraries against the protective antigen VP2 were constructed and screened.VP2 specific scFvs were selected following three rounds of screening procedures.Selected scFvs were characterized by FCM and ELISA.Seven scFvs showed high affinity and specific binding to CPV.Moreover,the neutralizing activity of the antibody was preliminarily identified by CPV(100 TCID_(50))in vitro and scFv-2 showed the neutralizing ability with a titer of 32768.This study generated the first canine-origin neutralizing scFv against CPV.The neutralizing scFv would be constructed into full-length antibody in the future.This study laid the foundation for the generation of an effective therapeutic reagent with long half-life and no immunological rejection for the prevention and treatment of CPV infection.展开更多
Objective:To study the various processes involved in transcellular transport(TT) of huperzine A alone or in combination with ginkgolide B in Caco-2 and Madin-Darby canine renal(MDCK)cell monolayer.Methods:The transepi...Objective:To study the various processes involved in transcellular transport(TT) of huperzine A alone or in combination with ginkgolide B in Caco-2 and Madin-Darby canine renal(MDCK)cell monolayer.Methods:The transepithelial passage was assayed in the apical-to-basolateral(AP to BL) direction and opposite direction(BL to AP) in both cell lines.The determination of huperzine A and ginkgolide B were performed by high performance liquid chromatography(HPLC).The passage rates of huperzine A and ginkgolide B were calculated.Bi-directional TT(absorption and secretion) were taken in huperzine A and ginkgolide B in Caco-2 and MDCK cell monolayer.Results:TT absorption and secretion kinetics of huperzine A and ginkgolide B across two cells existed at the same time.The passage rates of huperzine A were increased significantly with adding different concentrations of ginkgolide B.Conclusions:The compound preparations of HA in combination with CB for dementia caused by cerebral ischemic have synergistic effects on the pharmacodynamics,and improve the bioavailability through BBB.展开更多
Background: Appropriate preclinical evaluation of abioartificial liver assist device （BAL） demands alarge animal model, as presented here, that demon-strates many of the clinical features of acute liverfailure and t...Background: Appropriate preclinical evaluation of abioartificial liver assist device （BAL） demands alarge animal model, as presented here, that demon-strates many of the clinical features of acute liverfailure and that is suitable for clinical qualitative andquantitative evaluation of the BAL. A lethal canineliver failure model of acute hepatic failure that re-moves many of the artifacts evidenced in prior caninemodels is presented.Methods: Six male hounds, 24-30 kg, under isoflu-rane anesthesia, were administered 1.5 g/kg D-galactosamine intravenously. Canine supportive carefollowed a well-defined management protocol thatwas guided by electrolyte and invasive monitoringconsisting of arterial pressure, central venous pres-sure, extradural intracranial pressure （ICP）, pul-monary artery pressure, and end-tidal CO<sub>2</sub>. Theanimals were treated until death-equivalent, definedas inability to sustain systolic blood pressure】80mmHg for 20 minutes despite maximal fluids and 20μg·kg<sup>-1</sup>·min<sup>-1</sup> dopamine infusion.Results: The mean survival time was 43.7±4.6hours （mean±SE）. All animals showed evidence ofprogressive liver failure characterized by increasingliver enzymes （aspartate transaminase from 26 to5977 IU/L; alanine transaminase from 32 to 9740IU/L）, bilirubin （0.25 to 1.30 mg/dl）, ammonia（19. 8 to 85. 3 μmol/L）, and coagulopathy （pro-thrombin time from 8.7 to 46 s）. Increased labilityand elevations in intracranial pressures were ob-served. All animals were refractory to maintenanceof cerebral perfusion pressure even with only mode-rately elevated intracranial pressure. Severe neuro-logic obtundation, seen in 2 of 6 animals, was associ-ated with elevations of ICP above 50 mmHg. Post-mortem liver histology showed evidence of massivehepatic necrosis. Postmortem blood and ascites mi-crobial growth was consistent with possible transloca-tion of intestinal microbes.Conclusions: The improved lethal canine liver failuremodel presented here reproduces many of the clinicalfeatures of acute liver failure. The model may proveuseful for qualitative and quantitative evaluation ofBALs.展开更多
Introduction:Parallel stent-stent grafting is a major endovascular technique used to preserve the supra-aortic branches during thoracic endovascular aortic repair(TEVAR)of aortic pathologies involving the aortic arch....Introduction:Parallel stent-stent grafting is a major endovascular technique used to preserve the supra-aortic branches during thoracic endovascular aortic repair(TEVAR)of aortic pathologies involving the aortic arch.The short-and mid-term results of this technique are satisfactory;however,endoleak remains a major concern.Thus,here we designed a new chimney stent-graft to decrease the endoleak rate.Aim:To testify the feasibility and safety of the new chimney stent-graft system in a canine model.Material and methods:Six Labrador retrievers were used.Pre-operative data were collected and all operations were performed under general anesthesia.The main and chimney stent-grafts were implanted through the abdominal aorta and left subclavian artery approaches,respectively.Completion digital subtraction angiography(DSA)was performed to confirm the immediate outcomes.All dogs were fed separately for 6 months and sacrificed after aortic angiography.The thoracic aorta and the main and chimney stent-grafts were harvested for histopathologic examination.Results:No complications were found in follow-up DSA.All branch arteries were patent.In?ammatory responses were observed around the stent-grafts in 3 experimental animals,and slight hyperplasia was observed in the surrounding tissues compared with the normal vessels.There was no mural thrombus in the stent,endothelial cells were noted on the inner surface of the stent,and thrombus was formed in the outer skirt and gutter area.The histopathologic examinations revealed similar results to those of gross necropsy observations.Conclusions:This study demonstrated the feasibility and safety of the Longuette stent-graft and the first to report a revised stent-graft specific for chimney technique.展开更多
AIM: To test whether intra-articular injection of porcine adipose-derived stem cells(ADSCs) can treat canine osteoarthritis(OA).METHODS: To enroll in this study dogs must have stifle joint OA that had lasted ≥ 3 mo a...AIM: To test whether intra-articular injection of porcine adipose-derived stem cells(ADSCs) can treat canine osteoarthritis(OA).METHODS: To enroll in this study dogs must have stifle joint OA that had lasted ≥ 3 mo and been treated with OA medication without significant improvement. Three dogs fulfilled these criteria and were thus subjects for ADSCs treatment. ADSCs were isolated from abdominal adipose tissue of a 2-mo-old female Yorkshire pig. Their stem cell marker expression was examined by immunofluorescence staining. For treatment, 5 million ADSCs were injected into the diseased joint of each dog. In the next 48 h, the patient was observed for signs of inflammatory and allergic reactions. Thepatient was then discharged to the owner and, at 2, 6, and 12 wk, followed up with orthopedic assessment, owner questionnaire, X-ray imaging, and force-plate gait analysis.RESULTS: Porcine ADSCs expressed mesenchymal stem cell markers CD90 and CD105. Injection of porcine ADSCs into canine stifle joints did not cause any inflammatory or allergic reactions. Orthopedic evaluation found improvements in two dogs, particularly at the longest time point. Owners' evaluation found increased capacity and decreased pain in all three dogs' activities such as walking and running. Radiographic evaluation did not find statistically significant differences before and after treatment. Force-plate analysis found significant improvements in all three dogs after treatment.CONCLUSION: Xenotransplantation of ADSCs for the treatment of OA is feasible. Further studies are needed to validate this novel treatment modality, which can then be implemented for the routine treatment of OA in veterinary medicine.展开更多
The chromosomal number variations & structural aberrations of the MDCK cell line, primary feline or canine kidney cell(FKC or CKC) and Hela cell line were investigated and their karyotypes of conventional chromoso...The chromosomal number variations & structural aberrations of the MDCK cell line, primary feline or canine kidney cell(FKC or CKC) and Hela cell line were investigated and their karyotypes of conventional chromosome bands were analyzed. The carcinogenesis or tumorigenicity testing of these cell lines in about 232 nude mice and for colony formation in soft agarose and for haemagglutination under different concentration of plant lectins of these cells were carried out. Under the prerequisite that the incidence of cancer or tumor in negative-control nude mice inoculated subcutaneously with primary feline or canine kidney cell cultures purified in vitro at passage 3 was 0 (0/22) and 0 (0/10), respectively. The incidence of the progressively-growing malignant tumor(MT) in positive-control nude mice inoculated subcutaneously with Hela cell cultures of KB, X, or NM20/X strain was 10/10, 25/25 and 5/51, respectively. The results showed that the incidence of tumor in nude mice with tetrapioid YA strain of MDCK cell during 20 - 45 passages, with hypodiploid JB strain of MDCK cell on passage 25, with di-and hypoploid JC strain of MDCK cell during 2 - 15passages or with hypoploid M strain of MDCK cell during 9 - 27 passages was 28/58, 1/5, 4/18 and 0/31,respectively. The chromosomal analysis results showed that the ratio of difference in the rate of modal chromosome number between high (mcs + n) and lowest (mcs)passages was not more than 5 % - 15 % and the structure aberrations was generally 0 - 3%. These results proved that the genetic characteristics of chromosomal number of cell lines determines their tumorigenicity, but it is species-specific. MDCK line has tumorigenicity no matter what its chromosome karyotype is, at least it has very low tumorigenicity even when its modal chromosome number is hypoploid. The repeatedly frozen, thawed and split controls of tumorigenicity-positive cell lines(X strain of Hela, M strain of BHK-21, JA strain of Vero, YA strain of MDCK) have much lower tumorigenicity or are even non-carcinogenesis, and the repeatedly frozen, thawed and split controls of very low tumorigenicity cell lines (M or JC strain of MDCK) are certainly non-carcinogenic and never have increased tumorigenicity.It is thus evident that MDCK cell of M, JB or JC strain can be approved as substrate for the preparation of attenuated viral vaccines, but MDCK cell of YA strain can not be approved as substrate for the preparation of comattenuated viral vaccines. In summary, all strains of MDCK cell line have tunorigenicity, at least have low tumorigencity, never have non-cancinogenic MDCK, but very low tumorigenicity MDCK cell strains can certainly be used for the approval production of canine viral vaccines if the DNA content in viral cell cultures was remarkably decreased through conventional means in manufacturing process. Therefore, the master cell stock and working cell bank of MDCK line used for vaccine manufacture were established in China, which are free of infectious agents, and described with respect to cytogenetic characteristics and tumorigenicity. Tests showed that there were correlations among cell line chromosome number variations, anchorage independence in soft agarose, haemagglutination under plant lectins, and tumor-forming ability in nude mice, thus all the in vitro tests are economic, simple and reliable means for monitoring the tumor-forming ability of MDCK line in nude mice.展开更多
Objective To investigate the desensitization of acetylcholine (ACh) on the inhibition effects of blood pressure (BP) in anesthe tized canine and build a model for studying desensitization in vivo. Methods Through chan...Objective To investigate the desensitization of acetylcholine (ACh) on the inhibition effects of blood pressure (BP) in anesthe tized canine and build a model for studying desensitization in vivo. Methods Through changing the intervals (120, 100, 80, 60, 40, 20 seconds) of twice ACh administration (each was 15μg·kg -1,i.v.), the desensitization on the effect of systemic blood pressure of the first ACh in jection towards the subsequent ACh administration was observed. Results When ACh administration intervals were 40, 60, 80 , 100 seconds, the percentages of desensitization of ACh on systemic blood press ure were significantly increased (P<0.05). However, as the intervals were 20 and 120 seconds, the effects of twice ACh administration had no significant dif ference (P>0.05). Conclusion The results indicated that ACh contents in blo od might influence the action of next ACh administration. To some extent, the hi gher the concentration of ACh in blood, the bigger the ratio of desensitization of exogenous ACh is. In addition, this method of twice drug administration could be used as a model of studying desensitization in vivo.展开更多
Objective:To study the canine echinococcosis by coproantigen ELISA method.Methods:During the present investigation experimental infection was established using evaginated worms of Echinococcus granulosus(E.granulosus)...Objective:To study the canine echinococcosis by coproantigen ELISA method.Methods:During the present investigation experimental infection was established using evaginated worms of Echinococcus granulosus(E.granulosus).To check cross reactivity two pups were infected with Taenia hydatigena(T.hydatigena).In order to detect the presence of antigen,hyperimmune sera were raised against excretory-secretory products of adult worms E.chinococcus granulosus. Faecal sample collected either from experimentally infected pups or from other sources were heated at 70℃to detect heat stable soluble antigen.Results:Pups harbouring less than 104 worms showed negative results.Samples collected from 14 days onwards from experimentally infected animals harbouring more than 104 worms showed positive value.The maximum positive samples were detected in samples collected from in and around slaughter house and the least number of samples were detected positive maintained by dog squad.Conclusions:The affinity purified IgG exhibited promising results for detection of canine echinococcosis by indirect ELISA.展开更多
Pyometra is one of the most important and common disorders in canine. The disease results in life threatening condition associated with septicemia and toxemia. The condition commonly occurs during luteal phase of estr...Pyometra is one of the most important and common disorders in canine. The disease results in life threatening condition associated with septicemia and toxemia. The condition commonly occurs during luteal phase of estrous cycle, generally 4 weeks to 4 months of estrous cycle. The age, parity, breeds, administration of hormones are some predisposing factors of the condition. However, interaction between potentially pathogenic bacteria and hormonally primed uterus is believed to result in pyometra. The disease is initiated in the form of cystic endometrial hyperplasia, which later progresses into purulent uterine content following bacterial infection resulting in pyometra. The disturbance in antioxidant and pro-oxidant balance succeeding bacterial infection results in oxidative stress. The resultant stress further induces endometrial degeneration, immunosuppression and additionally aggravates the condition. An important clinical sign is malodorous, sanguineous or mucopurulent vaginal discharge with general symptoms such as vomiting, polyuria, polydipsia, depression, anorexia and occasionally fever. Till date, ultrasonography is the best diagnostic method while radiology and serological test are also helpful. Recently, the estimation of inflammatory mediators (acute phase proteins and prostaglandin metabolites) has appeared as marker for diagnosis of the pyometra. Determination of levels of inflammatory mediators has emerged as an important diagnostic method because it can be helpful in prognosis of the condition. The pyometra can be best treated by surgical manipulation (ovariohysterectomy);however, hormonal treatment can be given in less severe cases which aimed at reducing the effect of progesterone on the reproductive tract. Together, provision of antioxidants could be helpful in disease recovery along with the determined treatment. In this review, incidence, predisposing factors, etio-pathogenesis, oxidative stress associated with condition, diagnostic methods and treatment are discussed.展开更多
Objective:To investigate the effects of Azadirachta(A.)indica methanol seed extract on canine male fertility in vitro.Methods:The effect of the A.indica methanol seed extract(2.5,5.0,10.0 and 20.0 mg/mL,respectively)o...Objective:To investigate the effects of Azadirachta(A.)indica methanol seed extract on canine male fertility in vitro.Methods:The effect of the A.indica methanol seed extract(2.5,5.0,10.0 and 20.0 mg/mL,respectively)on canine spermatozoa was evaluated in vitro at 1.0,2.5,5.0 and 10.0 min post-treatment.The two controls were comprised of sperm sample in phosphate-buffered saline,and sperm sample in phosphate-buffered saline and 1%dimethyl sulfoxide,respectively.The experiment was repeated(n=3)in duplicates with semen collected from 3 adult Basenjis.The effect of A.indica methanol seed extract treatments on canine spermatozoal total and progressive motility(phase-contrast microscopy),sperm vitality(eosin-nigrosin vital staining method)and total abnormalities(phase contrast microscopy,eosin-nigrosin staining and Papanicolaou staining methods)were determined.Results:A.indica methanol seed extract treatment caused a concentration and time-dependent decrease in sperm percentage total motility,progressive motility and vitality.The absence of sperm revival post-treatment provided evidence of a permanent or irreversible incapacitation of sperm by A.indica methanol seed extract.Conclusions:The inhibition of sperm motility and significant spermicidal effect observed post-treatment suggest that A.indica methanol seed extract may produce a potent antifertility or contraceptive effect in dogs if applied as an intra-vaginal cream pre-coitus.展开更多
Aurora kinases play an important role in the cell cycle. These enzymes help establish mitotic spindles by directing centrosome duplication and separation and by regulating the spindle assembly checkpoint thereby helpi...Aurora kinases play an important role in the cell cycle. These enzymes help establish mitotic spindles by directing centrosome duplication and separation and by regulating the spindle assembly checkpoint thereby helping control cytokinesis. An over-expression of aurora kinases has been reported in a variety of human tumors. In this study, we identified the expression of aurora-A and aurora-B kinases in canine malignant lymphoid cells. We also evaluated the effects of the aurora kinase inhibitor (ZM447439), and found that this inhibitor decreases cell viability, increases DNA content change, and leads to apoptosis in canine B- and T-cell lymphoid cell lines. The lymphotoxicity induced by ZM447439 in these canine lymphoid cell lines suggests that further in vivo evaluation of aurora kinase inhibitors as a potential treatment for canine malignant lymphoid tumors is warranted.展开更多
This study was undertaken to examine which factors contributed to the correction of crowding in two patients who underwent nonextraction orthodontic treatment. A study model analysis was conducted to determine the eff...This study was undertaken to examine which factors contributed to the correction of crowding in two patients who underwent nonextraction orthodontic treatment. A study model analysis was conducted to determine the effects of the orthodontic treatment for crowding with high canines on crown angulation and dental arch width in two patients. The results showed that the crown angulation was significantly increased, indicating distal tipping in the maxillary dental arch. This tendency was most commonly observed in the premolars among the lateral teeth. With respect to the dental arch width, the largest change was evident in the first molar and first premolar regions in cases 1 and 2, respectively. On the basis of these results, up-righting of mesially tipped lateral teeth and expansion of narrow dental arches could prove to be the keys to the success of space regaining or correction of high canines and mild crowding.展开更多
Measurement of fecal cytokines has been used as a marker of intestinal inflammation in people and correlates with endoscopic findings. The aim of this study was to evaluate the use of canine-specific enzyme-linked imm...Measurement of fecal cytokines has been used as a marker of intestinal inflammation in people and correlates with endoscopic findings. The aim of this study was to evaluate the use of canine-specific enzyme-linked immunosorbant assays (ELISAs) for quantification of cytokines in canine fecal samples as a non-invasive biomarker. Interleukin (IL)-6, -8, -10, -23/12p40 and TNF-a were assessed by using spiked fecal samples from 3 healthy dogs. Standard curve validation was performed, and the impact of time to freeze, duration of storage and number of freeze-thaw cycles on cytokine concentration were also examined. All the cytokines assayed could be detected, with varying accuracy. The mean coefficient of variation (CV) for all standard curves ranged from 2.95%-9.8%. The mean intra-assay CV ranged from 3.1%-11.14%, and inter-assay CV from 4.36%-18.83%. Recovery of IL-23 was poor (7.23%-17.12%), precluding further interpretation of stability studies. Mean recovery did not appear to be affected by time to freeze and repeat freeze-thaw cycles in all cytokines investigated. Recovery for all cytokines after short-term storage of 30 days at -80℃ showed a recovery of 70% or >130%. In conclusion, although fecal IL-6, -8, -10, and TNF-a could be used as biomarkers of intestinal inflammation in the dog, the quality of laboratory performance and poor recovery at lower concentrations limit their application. Bench-top and freeze-thaw stability was acceptable, and samples should ideally be analyzed within a week. Investigation involving dogs with acute and chronic inflammatory intestinal disease is required to determine the role of this methodology in a clinical setting.展开更多
Canine adipose derived stem cells (ASCs) hold a great promise for the therapy of osteoarthritis in veterinary medicine. Current therapy is an autologous, stromal vascular fraction. Allogeneic ASCs provide many advanta...Canine adipose derived stem cells (ASCs) hold a great promise for the therapy of osteoarthritis in veterinary medicine. Current therapy is an autologous, stromal vascular fraction. Allogeneic ASCs provide many advantages, including efficient, cost-effective treatments while eliminating a surgical procedure in a diseased animal. Cultured ASCs can be expanded and characterized, allowing selection of desirable qualities. Use of allogeneic ASCs requires selection of a culture medium that provides consistent, desirable cellular products. The supplements within a medium can greatly influence cellular phenotypes. We hypothesized that medium type influenced cellular phenotype, allowing selection of a specified cellular product for clinical applications. We evaluated ASCs derived from adipose tissue of six dogs, assessing mRNA expression of proinflammatory: interleukin-1b, cyclooxygenase-2, and anti-inflammatory mediators: tissue inhibitor metalloproteinase-2 and interleukin -1 receptor antagonist, via quantitative RT-PCR prior to, and following culture in five cell culture media: basic cell growth medium (BGM), Keratinocyte N acetyl-L-cysteine supplemented (KNAC) medium, Multipotent Adult Progenitor Cell (MAPC) medium, serum free medium (SFM) and xeno-free medium. Major histocompatability complex I (MHCI), major histocompatability complex II (MHCII), CD44 and CD90 immunophenotypes were assessed via flow cytometry analysis. Tri-lineage differentiation (bone, adipose and cartilage tissue) was utilized to verify multipotency. SFM and xeno-free culture conditions did not produce cell expansion sufficient to assess phenotype. ASCs prior to culture had wide variability in all mediator levels, while culturing in the remaining conditions resulted in more predictable expression levels of inflammatory mediators, with a decrease in all levels. Cultured ASCs retained expression of cell surface markers MHCI, CD44 and CD90, while decreasing MHCII expression levels. KNAC and MAPC medium conditions consistently produced tri-lineage differentation;BGM, SFM and xeno-free medium did not. Culture condition will influence phenotype of ASCs, and should be selected according to the intended therapeutic effect.展开更多
Canine coronavirus(CCoV),a member of the genus Alphocoronavirus,is an enveloped,single-stranded positive-sense RNA virus that responsible for gastroenteritis in dogs.In this study,two CCoV isolates were successfully p...Canine coronavirus(CCoV),a member of the genus Alphocoronavirus,is an enveloped,single-stranded positive-sense RNA virus that responsible for gastroenteritis in dogs.In this study,two CCoV isolates were successfully propagated from 53 CCoV-positive clinical specimens by serial passaging in A-72 cells.These two strains,CCoV JS1706 and CCoV JS1712,caused cytopathic effects in A-72 cells.The sizes of virus plaque formed by them differed in early passages.Electron microscopy revealed a large quantity of typical coronavirus particles with 80-120 nm in diameter in cell culture media and cytoplasm of infected cells,in which they appeared as inclusion bodies.RT-PCR analysis of 5 gene indicated that these two isolates were belonged to CCoV lla subtype.Homology of RdRp,S,M and N proteins between the two strains were 100,99.6,99.2 and 100.0%,respectively,whereas they were 99.4-100%,83.1-95.2%,88.5-99.2% and 91.9-99.7%identity compared to CCoV II reference strains.Phylogenetic analysis of RdRp,S,M and N protein showed that they were closely related to CCoV II strains.These two subtype lla isolates will be useful for evaluating the pathogenesis and evolution of CCoV and for developing diagnostic reagents and vaccines.展开更多
Objective: To update the recent knowledge of the microbiological causes of canine otitis externa in Campania Region(Italy) and the antibiotic susceptibility patterns of the isolated strains.Methods: A total of 122 dog...Objective: To update the recent knowledge of the microbiological causes of canine otitis externa in Campania Region(Italy) and the antibiotic susceptibility patterns of the isolated strains.Methods: A total of 122 dogs were examined by otoscopy, and auricular swab samples were collected from both ears in 74 dogs presenting clinical bilateral otitis and from single ears in 48 dogs displaying clinical unilateral otitis. Cytological examination,bacteriological analysis and antimicrobial susceptibility tests were performed.Results: Thirty-one out of 122 dogs were positive for yeast species(25.4%, 95% confidence interval(CI): 18.2%–34.2%) with a higher prevalence of Malassezia pachydermatis(21/31 isolates, 67.7%, CI: 48.5%–82.7%), and a total of 91 out of 122 dogs were positive for bacterial species(74.6%; CI: 65.8%–81.8%) with a higher prevalence of Staphylococcus pseudintermedius(45/143 isolates, 31.5%, CI: 24.1%–39.8%). These results are the first description of Streptococcus agalactiae-associated otitis. The yeasts isolated showed high levels of susceptibility to all antifungal agents tested; on the contrary all the isolated bacterial strains were highly resistant to at least four out of ten antimicrobial classes. Both Gram-positive and Gram-negative bacteria showed high resistance to amoxicillin/clavulanate and kanamycin hence they are not recommended as initial empirical therapy for the otitis treatment.Conclusions: This update illustrates an increase in antibiotic resistances providing an insight into the current knowledge of the therapeutic procedures followed on canine otitis externa in Italy. It also emphasizes the importance of considering the results of the microbiological and sensitivity tests to decide on an appropriate antibiotic therapy.展开更多
Objective:To observe other hemoprotozoan diseases with canine ehrlichiosis und to evaluate the clinical and hematological aspects of dogs naturally infected with ehrlichiosis with other hemoprotozoun diseases.Methods:...Objective:To observe other hemoprotozoan diseases with canine ehrlichiosis und to evaluate the clinical and hematological aspects of dogs naturally infected with ehrlichiosis with other hemoprotozoun diseases.Methods:Blood was collected for hematological value and Giemsa stained blood smear was made for diagnosis of Ehrlichia sp.and other hemoprotozoan parasites from naturally infected dogs.Case history was taken from the owner and clinical signs and symptoms were noted.Results:A total of 47 cases of ehrlichiosis in dogs were reported with babesiosis(8.51%)and hepatozoonosis(6.38%)hemoprotozoan diseases.Ehrlichia cams,Ehrlichia ewingii,Brucella canis,Babesia gibsoni and Hepatozoon canis were observed under oil immersion lense of microscope in Giemsa stained peripheral blood smears.Marked anaemia and neutrophilic leukocytosis were observed.Conclusions:The results of this study stated that clinical and haeinatological changes occurred in canine ehrlichiosis with babesiosis and hepatozoonosis due to parasitemia.In mixed infection,the disease more severe,and also it depended on immunity of animals.Babesia gibsoni and Hepatozoon canis with Ehrlichia sp.were first reported from West Bengal state of India by this study.展开更多
文摘Background: Separation-related disorders are prevalent in dogs and can result in problem behaviours such as inappropriate elimination, destructive behaviour, and distress vocalizations which occur when an owner is absent or perceived as absent. Numerous natural products are marketed for managing anxiety in dogs;however, few, if any, of these products have documented evidence of efficacy in veterinary behaviour. Purpose: This open, multicentric clinical trial aimed to assess the efficacy of a new supplement containing alpha-casozepine and fish muscle hydrolysate on the separation-related problem in dogs and the impact on the quality of life of dogs and their pet owners. Material and Methods: Fifty-one dogs with clinical signs compatible with separation-related problems were included. The dogs were treated with a complementary feed (Zylkene Plus, Vetoquinol, France) for 30 days. Separation-related behaviour assessment (SRP), emotional disorders assessment (EDED) and the quality of life (QoL) of the pets and their pet owner were assessed at baseline and the end of the study by the veterinarians. In addition, owners were asked to rate their dog’s SRP weekly after enrolment using the same questionnaire that the veterinarian used at the beginning of the study. Results: There was a statistically significant improvement in the SRP, EDED, and QoL scores (P Conclusion: The results of this study support the hypothesis that the combination of alpha-casozepine and fish muscle hydrolysate is a promising and effective option to reduce the frequency and severity of separation-related behaviour in dogs and improve the quality of dogs and their owners.
文摘Antinuclear antibodies are found in animals suffering from Systemic Lupus Erythematosus (SLE) and some other diseases, their presence in the blood is determined by antinuclear antibody (ANA) test using indirect immunofluorescence (IF) with HEp2 cells as a substrate. In this work, an immunoperoxidase (IP) assay was developed to evaluate the ANAs in canine sera, using canine kidney cell lines (MDCK) and compared with a commercial immunofluorescence test on Hep2 cells for this system, a fluoresceinated anti-canine Ig antibody was standardized. The study was performed on 50 sera from dogs submitted to the laboratory with different clinical diagnoses of autoimmune-associated diseases. The procedures on both cells were unified to perform comparisons of the reactions, direct sera or at different dilutions were added to a monolayer of permeabilized MDCK cells, followed by a peroxidized anti-canine IgG conjugate, and a substrate for the IP reaction. The same sera were tested on the commercial IF assay on Hep2 cell system. In 22/50 cases, the presence of LE cells in peripheral blood was determined. A high correlation was found in the detection of antinuclear antibodies between both cell lines and techniques, however there were differences in the reaction patterns in the nucleus and cytoplasm between cell lines. The diffuse nuclear pattern observed in MDCK cells was more related to the presence of high percentages of LE cells in peripheral blood. The differences found in the results were possibly associated with the presence of homologous antigens between the MDCK cells and the dog. In addition, the methodology and standardization for the use and interpretation of a reference serum was developed to unify the interpretation criteria in the laboratory.
基金Supported by the National Key R&D Program of China(2017YFD0501102,2016YFD0501003)Science and Technology Department of Heilongjiang Province(GX18B018)Education Department of Heilongjiang Province(TSTAU-R2018017)。
文摘Canine parvovirus(CPV)is a high morbidity and lethality virus which causes severe enteric disease in dogs.In an attempt to gain canine-origin neutralizing antibodies against CPV,single chain variable fragment(scFv)bacteria display libraries against the protective antigen VP2 were constructed and screened.VP2 specific scFvs were selected following three rounds of screening procedures.Selected scFvs were characterized by FCM and ELISA.Seven scFvs showed high affinity and specific binding to CPV.Moreover,the neutralizing activity of the antibody was preliminarily identified by CPV(100 TCID_(50))in vitro and scFv-2 showed the neutralizing ability with a titer of 32768.This study generated the first canine-origin neutralizing scFv against CPV.The neutralizing scFv would be constructed into full-length antibody in the future.This study laid the foundation for the generation of an effective therapeutic reagent with long half-life and no immunological rejection for the prevention and treatment of CPV infection.
基金supported by Clinical Special Funds of China University Medical Journals(11321611)
文摘Objective:To study the various processes involved in transcellular transport(TT) of huperzine A alone or in combination with ginkgolide B in Caco-2 and Madin-Darby canine renal(MDCK)cell monolayer.Methods:The transepithelial passage was assayed in the apical-to-basolateral(AP to BL) direction and opposite direction(BL to AP) in both cell lines.The determination of huperzine A and ginkgolide B were performed by high performance liquid chromatography(HPLC).The passage rates of huperzine A and ginkgolide B were calculated.Bi-directional TT(absorption and secretion) were taken in huperzine A and ginkgolide B in Caco-2 and MDCK cell monolayer.Results:TT absorption and secretion kinetics of huperzine A and ginkgolide B across two cells existed at the same time.The passage rates of huperzine A were increased significantly with adding different concentrations of ginkgolide B.Conclusions:The compound preparations of HA in combination with CB for dementia caused by cerebral ischemic have synergistic effects on the pharmacodynamics,and improve the bioavailability through BBB.
基金This study was partially supported by a grant from Excorp Medical, Inc, Oakdale, MN., Steritek J7000 Intracranial Pressure Monitor provided by Ladd Research Industries, Williston, VT., and Datex Capnomac Ultima Anesthesia Monitor provided by Datex, Helsi
文摘Background: Appropriate preclinical evaluation of abioartificial liver assist device （BAL） demands alarge animal model, as presented here, that demon-strates many of the clinical features of acute liverfailure and that is suitable for clinical qualitative andquantitative evaluation of the BAL. A lethal canineliver failure model of acute hepatic failure that re-moves many of the artifacts evidenced in prior caninemodels is presented.Methods: Six male hounds, 24-30 kg, under isoflu-rane anesthesia, were administered 1.5 g/kg D-galactosamine intravenously. Canine supportive carefollowed a well-defined management protocol thatwas guided by electrolyte and invasive monitoringconsisting of arterial pressure, central venous pres-sure, extradural intracranial pressure （ICP）, pul-monary artery pressure, and end-tidal CO<sub>2</sub>. Theanimals were treated until death-equivalent, definedas inability to sustain systolic blood pressure】80mmHg for 20 minutes despite maximal fluids and 20μg·kg<sup>-1</sup>·min<sup>-1</sup> dopamine infusion.Results: The mean survival time was 43.7±4.6hours （mean±SE）. All animals showed evidence ofprogressive liver failure characterized by increasingliver enzymes （aspartate transaminase from 26 to5977 IU/L; alanine transaminase from 32 to 9740IU/L）, bilirubin （0.25 to 1.30 mg/dl）, ammonia（19. 8 to 85. 3 μmol/L）, and coagulopathy （pro-thrombin time from 8.7 to 46 s）. Increased labilityand elevations in intracranial pressures were ob-served. All animals were refractory to maintenanceof cerebral perfusion pressure even with only mode-rately elevated intracranial pressure. Severe neuro-logic obtundation, seen in 2 of 6 animals, was associ-ated with elevations of ICP above 50 mmHg. Post-mortem liver histology showed evidence of massivehepatic necrosis. Postmortem blood and ascites mi-crobial growth was consistent with possible transloca-tion of intestinal microbes.Conclusions: The improved lethal canine liver failuremodel presented here reproduces many of the clinicalfeatures of acute liver failure. The model may proveuseful for qualitative and quantitative evaluation ofBALs.
文摘Introduction:Parallel stent-stent grafting is a major endovascular technique used to preserve the supra-aortic branches during thoracic endovascular aortic repair(TEVAR)of aortic pathologies involving the aortic arch.The short-and mid-term results of this technique are satisfactory;however,endoleak remains a major concern.Thus,here we designed a new chimney stent-graft to decrease the endoleak rate.Aim:To testify the feasibility and safety of the new chimney stent-graft system in a canine model.Material and methods:Six Labrador retrievers were used.Pre-operative data were collected and all operations were performed under general anesthesia.The main and chimney stent-grafts were implanted through the abdominal aorta and left subclavian artery approaches,respectively.Completion digital subtraction angiography(DSA)was performed to confirm the immediate outcomes.All dogs were fed separately for 6 months and sacrificed after aortic angiography.The thoracic aorta and the main and chimney stent-grafts were harvested for histopathologic examination.Results:No complications were found in follow-up DSA.All branch arteries were patent.In?ammatory responses were observed around the stent-grafts in 3 experimental animals,and slight hyperplasia was observed in the surrounding tissues compared with the normal vessels.There was no mural thrombus in the stent,endothelial cells were noted on the inner surface of the stent,and thrombus was formed in the outer skirt and gutter area.The histopathologic examinations revealed similar results to those of gross necropsy observations.Conclusions:This study demonstrated the feasibility and safety of the Longuette stent-graft and the first to report a revised stent-graft specific for chimney technique.
文摘AIM: To test whether intra-articular injection of porcine adipose-derived stem cells(ADSCs) can treat canine osteoarthritis(OA).METHODS: To enroll in this study dogs must have stifle joint OA that had lasted ≥ 3 mo and been treated with OA medication without significant improvement. Three dogs fulfilled these criteria and were thus subjects for ADSCs treatment. ADSCs were isolated from abdominal adipose tissue of a 2-mo-old female Yorkshire pig. Their stem cell marker expression was examined by immunofluorescence staining. For treatment, 5 million ADSCs were injected into the diseased joint of each dog. In the next 48 h, the patient was observed for signs of inflammatory and allergic reactions. Thepatient was then discharged to the owner and, at 2, 6, and 12 wk, followed up with orthopedic assessment, owner questionnaire, X-ray imaging, and force-plate gait analysis.RESULTS: Porcine ADSCs expressed mesenchymal stem cell markers CD90 and CD105. Injection of porcine ADSCs into canine stifle joints did not cause any inflammatory or allergic reactions. Orthopedic evaluation found improvements in two dogs, particularly at the longest time point. Owners' evaluation found increased capacity and decreased pain in all three dogs' activities such as walking and running. Radiographic evaluation did not find statistically significant differences before and after treatment. Force-plate analysis found significant improvements in all three dogs after treatment.CONCLUSION: Xenotransplantation of ADSCs for the treatment of OA is feasible. Further studies are needed to validate this novel treatment modality, which can then be implemented for the routine treatment of OA in veterinary medicine.
文摘The chromosomal number variations & structural aberrations of the MDCK cell line, primary feline or canine kidney cell(FKC or CKC) and Hela cell line were investigated and their karyotypes of conventional chromosome bands were analyzed. The carcinogenesis or tumorigenicity testing of these cell lines in about 232 nude mice and for colony formation in soft agarose and for haemagglutination under different concentration of plant lectins of these cells were carried out. Under the prerequisite that the incidence of cancer or tumor in negative-control nude mice inoculated subcutaneously with primary feline or canine kidney cell cultures purified in vitro at passage 3 was 0 (0/22) and 0 (0/10), respectively. The incidence of the progressively-growing malignant tumor(MT) in positive-control nude mice inoculated subcutaneously with Hela cell cultures of KB, X, or NM20/X strain was 10/10, 25/25 and 5/51, respectively. The results showed that the incidence of tumor in nude mice with tetrapioid YA strain of MDCK cell during 20 - 45 passages, with hypodiploid JB strain of MDCK cell on passage 25, with di-and hypoploid JC strain of MDCK cell during 2 - 15passages or with hypoploid M strain of MDCK cell during 9 - 27 passages was 28/58, 1/5, 4/18 and 0/31,respectively. The chromosomal analysis results showed that the ratio of difference in the rate of modal chromosome number between high (mcs + n) and lowest (mcs)passages was not more than 5 % - 15 % and the structure aberrations was generally 0 - 3%. These results proved that the genetic characteristics of chromosomal number of cell lines determines their tumorigenicity, but it is species-specific. MDCK line has tumorigenicity no matter what its chromosome karyotype is, at least it has very low tumorigenicity even when its modal chromosome number is hypoploid. The repeatedly frozen, thawed and split controls of tumorigenicity-positive cell lines(X strain of Hela, M strain of BHK-21, JA strain of Vero, YA strain of MDCK) have much lower tumorigenicity or are even non-carcinogenesis, and the repeatedly frozen, thawed and split controls of very low tumorigenicity cell lines (M or JC strain of MDCK) are certainly non-carcinogenic and never have increased tumorigenicity.It is thus evident that MDCK cell of M, JB or JC strain can be approved as substrate for the preparation of attenuated viral vaccines, but MDCK cell of YA strain can not be approved as substrate for the preparation of comattenuated viral vaccines. In summary, all strains of MDCK cell line have tunorigenicity, at least have low tumorigencity, never have non-cancinogenic MDCK, but very low tumorigenicity MDCK cell strains can certainly be used for the approval production of canine viral vaccines if the DNA content in viral cell cultures was remarkably decreased through conventional means in manufacturing process. Therefore, the master cell stock and working cell bank of MDCK line used for vaccine manufacture were established in China, which are free of infectious agents, and described with respect to cytogenetic characteristics and tumorigenicity. Tests showed that there were correlations among cell line chromosome number variations, anchorage independence in soft agarose, haemagglutination under plant lectins, and tumor-forming ability in nude mice, thus all the in vitro tests are economic, simple and reliable means for monitoring the tumor-forming ability of MDCK line in nude mice.
文摘Objective To investigate the desensitization of acetylcholine (ACh) on the inhibition effects of blood pressure (BP) in anesthe tized canine and build a model for studying desensitization in vivo. Methods Through changing the intervals (120, 100, 80, 60, 40, 20 seconds) of twice ACh administration (each was 15μg·kg -1,i.v.), the desensitization on the effect of systemic blood pressure of the first ACh in jection towards the subsequent ACh administration was observed. Results When ACh administration intervals were 40, 60, 80 , 100 seconds, the percentages of desensitization of ACh on systemic blood press ure were significantly increased (P<0.05). However, as the intervals were 20 and 120 seconds, the effects of twice ACh administration had no significant dif ference (P>0.05). Conclusion The results indicated that ACh contents in blo od might influence the action of next ACh administration. To some extent, the hi gher the concentration of ACh in blood, the bigger the ratio of desensitization of exogenous ACh is. In addition, this method of twice drug administration could be used as a model of studying desensitization in vivo.
文摘Objective:To study the canine echinococcosis by coproantigen ELISA method.Methods:During the present investigation experimental infection was established using evaginated worms of Echinococcus granulosus(E.granulosus).To check cross reactivity two pups were infected with Taenia hydatigena(T.hydatigena).In order to detect the presence of antigen,hyperimmune sera were raised against excretory-secretory products of adult worms E.chinococcus granulosus. Faecal sample collected either from experimentally infected pups or from other sources were heated at 70℃to detect heat stable soluble antigen.Results:Pups harbouring less than 104 worms showed negative results.Samples collected from 14 days onwards from experimentally infected animals harbouring more than 104 worms showed positive value.The maximum positive samples were detected in samples collected from in and around slaughter house and the least number of samples were detected positive maintained by dog squad.Conclusions:The affinity purified IgG exhibited promising results for detection of canine echinococcosis by indirect ELISA.
文摘Pyometra is one of the most important and common disorders in canine. The disease results in life threatening condition associated with septicemia and toxemia. The condition commonly occurs during luteal phase of estrous cycle, generally 4 weeks to 4 months of estrous cycle. The age, parity, breeds, administration of hormones are some predisposing factors of the condition. However, interaction between potentially pathogenic bacteria and hormonally primed uterus is believed to result in pyometra. The disease is initiated in the form of cystic endometrial hyperplasia, which later progresses into purulent uterine content following bacterial infection resulting in pyometra. The disturbance in antioxidant and pro-oxidant balance succeeding bacterial infection results in oxidative stress. The resultant stress further induces endometrial degeneration, immunosuppression and additionally aggravates the condition. An important clinical sign is malodorous, sanguineous or mucopurulent vaginal discharge with general symptoms such as vomiting, polyuria, polydipsia, depression, anorexia and occasionally fever. Till date, ultrasonography is the best diagnostic method while radiology and serological test are also helpful. Recently, the estimation of inflammatory mediators (acute phase proteins and prostaglandin metabolites) has appeared as marker for diagnosis of the pyometra. Determination of levels of inflammatory mediators has emerged as an important diagnostic method because it can be helpful in prognosis of the condition. The pyometra can be best treated by surgical manipulation (ovariohysterectomy);however, hormonal treatment can be given in less severe cases which aimed at reducing the effect of progesterone on the reproductive tract. Together, provision of antioxidants could be helpful in disease recovery along with the determined treatment. In this review, incidence, predisposing factors, etio-pathogenesis, oxidative stress associated with condition, diagnostic methods and treatment are discussed.
文摘Objective:To investigate the effects of Azadirachta(A.)indica methanol seed extract on canine male fertility in vitro.Methods:The effect of the A.indica methanol seed extract(2.5,5.0,10.0 and 20.0 mg/mL,respectively)on canine spermatozoa was evaluated in vitro at 1.0,2.5,5.0 and 10.0 min post-treatment.The two controls were comprised of sperm sample in phosphate-buffered saline,and sperm sample in phosphate-buffered saline and 1%dimethyl sulfoxide,respectively.The experiment was repeated(n=3)in duplicates with semen collected from 3 adult Basenjis.The effect of A.indica methanol seed extract treatments on canine spermatozoal total and progressive motility(phase-contrast microscopy),sperm vitality(eosin-nigrosin vital staining method)and total abnormalities(phase contrast microscopy,eosin-nigrosin staining and Papanicolaou staining methods)were determined.Results:A.indica methanol seed extract treatment caused a concentration and time-dependent decrease in sperm percentage total motility,progressive motility and vitality.The absence of sperm revival post-treatment provided evidence of a permanent or irreversible incapacitation of sperm by A.indica methanol seed extract.Conclusions:The inhibition of sperm motility and significant spermicidal effect observed post-treatment suggest that A.indica methanol seed extract may produce a potent antifertility or contraceptive effect in dogs if applied as an intra-vaginal cream pre-coitus.
文摘Aurora kinases play an important role in the cell cycle. These enzymes help establish mitotic spindles by directing centrosome duplication and separation and by regulating the spindle assembly checkpoint thereby helping control cytokinesis. An over-expression of aurora kinases has been reported in a variety of human tumors. In this study, we identified the expression of aurora-A and aurora-B kinases in canine malignant lymphoid cells. We also evaluated the effects of the aurora kinase inhibitor (ZM447439), and found that this inhibitor decreases cell viability, increases DNA content change, and leads to apoptosis in canine B- and T-cell lymphoid cell lines. The lymphotoxicity induced by ZM447439 in these canine lymphoid cell lines suggests that further in vivo evaluation of aurora kinase inhibitors as a potential treatment for canine malignant lymphoid tumors is warranted.
文摘This study was undertaken to examine which factors contributed to the correction of crowding in two patients who underwent nonextraction orthodontic treatment. A study model analysis was conducted to determine the effects of the orthodontic treatment for crowding with high canines on crown angulation and dental arch width in two patients. The results showed that the crown angulation was significantly increased, indicating distal tipping in the maxillary dental arch. This tendency was most commonly observed in the premolars among the lateral teeth. With respect to the dental arch width, the largest change was evident in the first molar and first premolar regions in cases 1 and 2, respectively. On the basis of these results, up-righting of mesially tipped lateral teeth and expansion of narrow dental arches could prove to be the keys to the success of space regaining or correction of high canines and mild crowding.
文摘Measurement of fecal cytokines has been used as a marker of intestinal inflammation in people and correlates with endoscopic findings. The aim of this study was to evaluate the use of canine-specific enzyme-linked immunosorbant assays (ELISAs) for quantification of cytokines in canine fecal samples as a non-invasive biomarker. Interleukin (IL)-6, -8, -10, -23/12p40 and TNF-a were assessed by using spiked fecal samples from 3 healthy dogs. Standard curve validation was performed, and the impact of time to freeze, duration of storage and number of freeze-thaw cycles on cytokine concentration were also examined. All the cytokines assayed could be detected, with varying accuracy. The mean coefficient of variation (CV) for all standard curves ranged from 2.95%-9.8%. The mean intra-assay CV ranged from 3.1%-11.14%, and inter-assay CV from 4.36%-18.83%. Recovery of IL-23 was poor (7.23%-17.12%), precluding further interpretation of stability studies. Mean recovery did not appear to be affected by time to freeze and repeat freeze-thaw cycles in all cytokines investigated. Recovery for all cytokines after short-term storage of 30 days at -80℃ showed a recovery of 70% or >130%. In conclusion, although fecal IL-6, -8, -10, and TNF-a could be used as biomarkers of intestinal inflammation in the dog, the quality of laboratory performance and poor recovery at lower concentrations limit their application. Bench-top and freeze-thaw stability was acceptable, and samples should ideally be analyzed within a week. Investigation involving dogs with acute and chronic inflammatory intestinal disease is required to determine the role of this methodology in a clinical setting.
文摘Canine adipose derived stem cells (ASCs) hold a great promise for the therapy of osteoarthritis in veterinary medicine. Current therapy is an autologous, stromal vascular fraction. Allogeneic ASCs provide many advantages, including efficient, cost-effective treatments while eliminating a surgical procedure in a diseased animal. Cultured ASCs can be expanded and characterized, allowing selection of desirable qualities. Use of allogeneic ASCs requires selection of a culture medium that provides consistent, desirable cellular products. The supplements within a medium can greatly influence cellular phenotypes. We hypothesized that medium type influenced cellular phenotype, allowing selection of a specified cellular product for clinical applications. We evaluated ASCs derived from adipose tissue of six dogs, assessing mRNA expression of proinflammatory: interleukin-1b, cyclooxygenase-2, and anti-inflammatory mediators: tissue inhibitor metalloproteinase-2 and interleukin -1 receptor antagonist, via quantitative RT-PCR prior to, and following culture in five cell culture media: basic cell growth medium (BGM), Keratinocyte N acetyl-L-cysteine supplemented (KNAC) medium, Multipotent Adult Progenitor Cell (MAPC) medium, serum free medium (SFM) and xeno-free medium. Major histocompatability complex I (MHCI), major histocompatability complex II (MHCII), CD44 and CD90 immunophenotypes were assessed via flow cytometry analysis. Tri-lineage differentiation (bone, adipose and cartilage tissue) was utilized to verify multipotency. SFM and xeno-free culture conditions did not produce cell expansion sufficient to assess phenotype. ASCs prior to culture had wide variability in all mediator levels, while culturing in the remaining conditions resulted in more predictable expression levels of inflammatory mediators, with a decrease in all levels. Cultured ASCs retained expression of cell surface markers MHCI, CD44 and CD90, while decreasing MHCII expression levels. KNAC and MAPC medium conditions consistently produced tri-lineage differentation;BGM, SFM and xeno-free medium did not. Culture condition will influence phenotype of ASCs, and should be selected according to the intended therapeutic effect.
基金This work was supported by the Open Project Program of Jiangsu Key Laboratory of Zoonosis(No.1604,No.1804)the Priority Academic Program Development of Jiangsu Higher Education Institutions(PAPD).
文摘Canine coronavirus(CCoV),a member of the genus Alphocoronavirus,is an enveloped,single-stranded positive-sense RNA virus that responsible for gastroenteritis in dogs.In this study,two CCoV isolates were successfully propagated from 53 CCoV-positive clinical specimens by serial passaging in A-72 cells.These two strains,CCoV JS1706 and CCoV JS1712,caused cytopathic effects in A-72 cells.The sizes of virus plaque formed by them differed in early passages.Electron microscopy revealed a large quantity of typical coronavirus particles with 80-120 nm in diameter in cell culture media and cytoplasm of infected cells,in which they appeared as inclusion bodies.RT-PCR analysis of 5 gene indicated that these two isolates were belonged to CCoV lla subtype.Homology of RdRp,S,M and N proteins between the two strains were 100,99.6,99.2 and 100.0%,respectively,whereas they were 99.4-100%,83.1-95.2%,88.5-99.2% and 91.9-99.7%identity compared to CCoV II reference strains.Phylogenetic analysis of RdRp,S,M and N protein showed that they were closely related to CCoV II strains.These two subtype lla isolates will be useful for evaluating the pathogenesis and evolution of CCoV and for developing diagnostic reagents and vaccines.
基金Supported in part by the Second University of Naples and University of Naples"Federico II",Fondi di Ateneo,by a fellowship from the University of Naples‘Federico II’(2012-4/STV-Project FORGIARE),and co-funded by"Compagnia San Paolo"of Turin,Italy
文摘Objective: To update the recent knowledge of the microbiological causes of canine otitis externa in Campania Region(Italy) and the antibiotic susceptibility patterns of the isolated strains.Methods: A total of 122 dogs were examined by otoscopy, and auricular swab samples were collected from both ears in 74 dogs presenting clinical bilateral otitis and from single ears in 48 dogs displaying clinical unilateral otitis. Cytological examination,bacteriological analysis and antimicrobial susceptibility tests were performed.Results: Thirty-one out of 122 dogs were positive for yeast species(25.4%, 95% confidence interval(CI): 18.2%–34.2%) with a higher prevalence of Malassezia pachydermatis(21/31 isolates, 67.7%, CI: 48.5%–82.7%), and a total of 91 out of 122 dogs were positive for bacterial species(74.6%; CI: 65.8%–81.8%) with a higher prevalence of Staphylococcus pseudintermedius(45/143 isolates, 31.5%, CI: 24.1%–39.8%). These results are the first description of Streptococcus agalactiae-associated otitis. The yeasts isolated showed high levels of susceptibility to all antifungal agents tested; on the contrary all the isolated bacterial strains were highly resistant to at least four out of ten antimicrobial classes. Both Gram-positive and Gram-negative bacteria showed high resistance to amoxicillin/clavulanate and kanamycin hence they are not recommended as initial empirical therapy for the otitis treatment.Conclusions: This update illustrates an increase in antibiotic resistances providing an insight into the current knowledge of the therapeutic procedures followed on canine otitis externa in Italy. It also emphasizes the importance of considering the results of the microbiological and sensitivity tests to decide on an appropriate antibiotic therapy.
基金Supported by Department of Veterinary Microbiology.West Bengal University of Animal and Fishery Sciences.37.K.B.Sarani.Kolkata-700037.West Bengal.India(Grant No.F.O.-VAS/Micro-11-12/165)
文摘Objective:To observe other hemoprotozoan diseases with canine ehrlichiosis und to evaluate the clinical and hematological aspects of dogs naturally infected with ehrlichiosis with other hemoprotozoun diseases.Methods:Blood was collected for hematological value and Giemsa stained blood smear was made for diagnosis of Ehrlichia sp.and other hemoprotozoan parasites from naturally infected dogs.Case history was taken from the owner and clinical signs and symptoms were noted.Results:A total of 47 cases of ehrlichiosis in dogs were reported with babesiosis(8.51%)and hepatozoonosis(6.38%)hemoprotozoan diseases.Ehrlichia cams,Ehrlichia ewingii,Brucella canis,Babesia gibsoni and Hepatozoon canis were observed under oil immersion lense of microscope in Giemsa stained peripheral blood smears.Marked anaemia and neutrophilic leukocytosis were observed.Conclusions:The results of this study stated that clinical and haeinatological changes occurred in canine ehrlichiosis with babesiosis and hepatozoonosis due to parasitemia.In mixed infection,the disease more severe,and also it depended on immunity of animals.Babesia gibsoni and Hepatozoon canis with Ehrlichia sp.were first reported from West Bengal state of India by this study.
