Expression of matrix metalloproteinases in bovine luteal cells induced by prostaglandin F2α, interferon γ and tumor necrosis factor α

We recently demonstrated that luteal cells flow out from the ovary via lymphatic vessels during luteolysis. However, the regulatory mechanisms of the outflow of luteal cells are not known. Matrix metalloproteinases (MMPs) can degrade the extracellular matrix and basal membrane, and tissue inhibitors of matrix metalloproteinases (TIMPs) inhibit the activity of MMPs. To test the hypothesis that MMP expression in luteal cells is regulated by luteolytic factors, we investigated the effects of prostaglandin F2α (PGF), interferon γ (IFNG) and tumor necrosis factor α (TNF) on the mRNA expression of MMPs and TIMPs in cultured luteal cells. Luteal cells obtained from the CL at the mid-luteal stage (days 8–12 after ovulation) were cultured with PGF (0.01, 0.1, 1 μM), IFNG (0.05, 0.5, 5 nM) and TNF (0.05, 0.5, 0.5 nM) alone or in combination for 24 h. PGF and IFNG significantly increased the expression of MMP-1 mRNA. In addition, 1 μM PGF in combination with 5 nM IFNG stimulated MMP-1 and MMP-9 mRNA expression significantly more than either treatment alone. In contrast, IFNG significantly decreased the level of MMP-14 mRNA. The mRNA expression of TIMP-1, which preferentially inhibits MMP-1, was suppressed by 5 nM INFG. One μM PGF and 5 nM IFNG suppressed TIMP-2 mRNA expression. These results suggest a new role of MMPs: luteal MMPs stimulated by PGF and IFNG break down the extracellular matrix surrounding luteal cells, which accelerates detachment from the CL during luteolysis, providing an essential prerequisite for outflow of luteal cells from the CL to lymphatic vessels.     

Expression and Activity of Collagenases in the Digital Laminae of Horses with Carbohydrate Overload‐Induced Acute Laminitis

Background

Matrix metalloproteinases (MMP) are hypothesized to degrade structurally important components of the laminar extracellular matrix (ECM) in horses with laminitis.

Objective

To compare levels of expression of stromelysin‐1 (MMP‐3), collagenases (MMP‐1, ‐13), and membrane type‐MMPs (MMP‐14, ‐15, ‐16), and the distribution of their ECM substrates, in laminae of healthy horses and horses with carbohydrate overload laminitis.

Animals

Twenty‐five adult horses.

Methods

Gene and protein expression were determined in extracts of laminae using real‐time quantitative polymerase chain reaction and Western blotting after sodium dodecylsulfate polyacrylamide gel electrophoresis. Distribution of MMP‐13 and ECM components was determined using indirect immunofluorescent microscopy of nonfixed frozen sections. ECM morphology was assessed by hematoxylin and eosin staining.

Results

Of the genes studied, only those encoding MMP‐1 and ‐13 were upregulated in CHO‐induced laminitis; MMP‐1 at Obel grade (OG)1 lameness and MMP‐13 at OG3 lameness. Laminar MMP‐1 was present as 52 kDa proenzyme only. MMP‐13 was present as pro‐ (61 kDa) and processed (48 kDa) enzyme. MMP‐13 localized to the basal epithelium of the secondary epidermal laminae and its increased expression were accompanied by the appearance in secondary dermal laminae (SDL) of multiple foci that were devoid of collagen I, fibronectin, chondroitin and keratan sulfate glycosaminoglycans, and eosin‐staining material.

Conclusions and Clinical Relevance

MMP‐13 is upregulated in laminae of horses with CHO‐induced OG3 lameness and, by degrading components of the ECM, may contribute to the formation of ECM‐free lesions (gaps or tears) that appear in the SDL with OG3 lameness.     

Influence of persistent canine distemper virus infection on expression of RECK, matrix-metalloproteinases and their inhibitors in a canine macrophage/monocytic tumour cell line (DH82)

A morbillivirus infection of tumour cells is known to exert oncolytic activity, but the mechanism of this inhibitory action has not been well defined. Matrix metalloproteinases (MMPs) are important enzymes degrading the extracellular matrix and are often upregulated in malignant neoplasms. Recent studies have demonstrated that RECK may potently suppress MMP-2 and -9 activity, thus inhibiting angiogenesis and metastasis. In this study, real time quantitative polymerase chain reaction (RT-qPCR) was used to determine the effect of persistent infection with canine distemper virus (CDV) infection on the expression of MMPs and their inhibitors (TIMPS) in a canine macrophage/monocytic tumour cell line (DH82). The activity of proMMP-2 and proMMP-9 was also verified zymographically. Following CDV infection, MMP-2, TIMP-1 and TIMP-2 were down-regulated, while RECK was upregulated. These findings suggest that CDV infection restores RECK expression in tumour cells and may interfere with the intracellular processing of MMPs and TIMPs, thus possibly influencing tumour cell behaviour beneficially for the host. However, this needs to be verified in in vivo studies.     

Characterisation of three novel canine osteosarcoma cell lines producing high levels of matrix metalloproteinases

Three canine osteosarcoma cell lines were established from spontaneous pelvic and radial osteosarcomas. The cell populations cultured exhibited characteristics of malignancy and consisted of adherent, pleomorphic, mostly large spindle-shaped or polyhedral cells, characterised by the presence of numerous cytoplasmic granules and vacuoles. The main ultrastructural features included the presence of abundant rough endoplasmic reticulum and numerous cytoplasmic vesicles, deposit vacuoles and small cytoplasmic protrusions. Zymography showed that the cell lines produce high levels of MMP-2 and MMP-9, enzymes directly involved in crucial aspects of the metastatic process. Consistent with their osteoblastic lineage and malignant phenotype, all cell lines were immunoreactive to vimentin, osteopontin, PCNA, p53, MMP-2 and MMP-9, while they were negative for cytokeratin, desmin, SMA, Factor VIII, NSE, GFAP, Rb and p21 protein. No retroviral particles or RNA were detected ultrastructurally or with RT-PCR, although the possibility of viral involvement in osteosarcoma cannot be excluded. The new cell lines provide excellent in vitro models that may allow further studies on the pathobiology of canine osteosarcoma to be undertaken.     

人工培植狗宝的研究

通过手术方法在21只犬的膀胱内植入核心异物，实验组饲喂钙、磷不平衡饲料并添加小苏打使尿液碱化。对照组饲喂钙磷平衡饲料。定期测定血液、尿液。实验组尿液呈乳白色浑浊，尿沉渣增多，尿蛋白及尿潜血呈阳性反应。尿Ca^2 降低，无机磷升高。血钙升高、血磷降低。经过180d，在实验犬的膀胱内形成了狗宝。狗宝呈乳白色、卵圆形、坚硬。在扫描电镜下呈层状、颗粒状及蜂窝状。狗宝中含有铵盐、磷酸盐以及Ca^2 ,Mg^2 ,K^ 等离子，还含有Fe,Sr,Mn,Ba,Zn等元素及多种氨基酸。狗宝具有极显著的抗炎、解热和促进小肠推进运动的作用。     

Pigmented Plaques Associated with Papillomavirus Infection in Dogs: Is this Epidermodysplasia Verruciformis?

Abstract— Two Pugs and two Miniature Schnauzers with multiple pigmented epidermal nevi were investigated. The four dogs had pigmented cutaneous maculae and plaques. Histopathological evaluation showed papillated or digitated epidermal hyperplasia with hypermelanosis and giant keratohyalin granules in the stratum granulosum. Immunohistochemical staining revealed papillomavirus group-specific antigen in the skin specimens from all four dogs. Electron microscopic study of the specimens from two dogs revealed numerous round viral particles within the nuclei of the keratinocytes in the upper stratum granulosum. It was suspected that papillomavirus was the etiologic agent of the lesions, and that Pugs and Miniature Schnauzers might be predisposed to infection. These findings indicate this canine dermatosis resembles epidermodysplasia verruciformis (EV) of humans, a rare chronic disease caused by human papillomavirus. The potential for transformation of the lesions to squamous cell carcinoma is also suspected and discussed. Résumé— Deux Carlins et deux Schnauzers nains présentant de multiples naevi épidermiques pigmentés sont examinés. Les quatre chiens présentent des macules et des plaques pigmentées. Les lésions histopathologiques montrent une hyperplasie épidermique papillaire ou digitée avec une hypermélanose et la présence de grains de kératohyaline dans le stratum corneum. Les colorations immunohistochimiques révèlent des antigènes spécifiques du groupe des papillomas virus dans les biopsies des quatre chiens. L'étude ultrastructurale à partir des biopsies de deux chiens montrent de nombreuses particules virales rondes dans les noyaux des kératinocytes des couches supérieures du stratum granulosum. II a été suspecté que le papilloma virus était l'agent causal des lésions et que les Carlins et les Schnauzers nains pouvaient être prédisposés à cette infection. Ces éléments font que cette dermatose observée chez le chien ressemble à l'épidermodysplasie verruciforme de l'homme, une dermatose chronique rare causée par un papilloma virus humain. La potentialité de transformation des lésions en épithélioma spinocellulaire est aussi suspectée et discutée. [Nagata, M., Nanko, H., Moriyama, A., Washizu, T., Ishida, T. Pigmented plaques associated with papilloma virus infection in dogs: Is this epidermodysplasia verruciformis? (Plaques hyperpigmentées associées à une infection à papilloma virus chez le chien: est-ce épidermodysplasie verruciforme?). Resumen— Se investigó dos perros de raza Pug y dos de raza Schnauzer Miniatura con múltiples nevos epidérmicos pigmentados. Los cuatro perros presentaban máculas y placas cutáneas pigmentadas. El estudio histológico mostró hiperplasia epitelial con papilas y digitaciones, así como hipermelanosis y gránules de queratohialina gigantes en el estrato granuloso. Las tinciones immunohistoquimicas detectaron antígeno grupo-específico de papilomavirus en las muestras de los cuatro animales. Mediante estudios de microscopía electrónica en muestras de dos de los perros se observaron numerosas particulas virales redondas en el núcleo de los queratinocitos más superficiales del estrato granuloso. Se sospechó que Papilomavirus era el agente etiológico de estas lesiones y que los Pugs y los Schnauzer Miniatura podrian estar predispuestos a la infección. Estos hallazgos indican que esta dermatosis canina es parecida a la Epidermodisplasia Verruciforme (EV) de la especie humana, una enfermedad crónica rara causada por el Papilomavirus Humano. Se sospecha y discute la posible transformación de estas lesiones a carcinoma de células planas. [Nagata, M., Nanko, H., Moriyama, A., Washizu, T., Ishida, T. Pigmented plaques associated with papilloma virus infection in dogs: Is this epidermodysplasia verruciformis? (Placas pigmentadas asociadas con infección por papillomavirus en el perro: se trata de epidermodisplasia verruciforme?)     

Infection by Eucoleus aerophilus in dogs and cats: Is another extra-intestinal parasitic nematode of pets emerging in Italy?

The occurrence of the infection by the lungworm Eucoleus aerophilus (syn. Capillaria aerophila) in dogs and cats from Italy has been evaluated with conventional diagnostic procedures. Individual faecal samples from 569 dogs and 200 cats were undertaken to faecal flotation with sugar and zinc sulphate solution. Sixteen dogs (2.8%) and 11 cats (5.5%) scored positive for eggs of E. aerophilus when samples were processed with either of the two flotation solutions. Overall 14 of 16 dogs and eight of 11 cats infected by E. aerophilus showed respiratory symptoms and the most common were general respiratory distress, dry cough, wheezing and sneezing. These results indicate that E. aerophilus is not uncommon and that canine and feline capillariosis is of clinical importance. Given the impact that E. aerophilus infections may have upon animal health and its zoonotic potential, it is strongly advisable to routinely include this disease in the differential diagnosis of (cardio)-respiratory diseases of dogs and cats.     

Castration,other management practices and socio-economic implications for dog keepers in Nsukka area,Enugu state,Nigeria

Structured interviews were used to obtain information from 258 respondents from among 625 people who were selected by stratified random sampling from villages in five of seven local government areas of Nsukka area. Information included gender and literacy status of the respondents, whether the respondents had (between 1990 and 1995) presented any of their dogs for castration, the comparative market value of the dogs, and dog use and owner preference for castrates in performance of such duties. Information on bathing, vaccinations, confinements, use of veterinarians and cultural and religious uses of dogs also were sought. Also, 208 (80%) of the respondents had their dogs castrated.

Of the respondents, 23% were literate, 37% semiliterate and 40% illiterate. Of the 367 non-respondents, 63% were not available during the time of contact and 37% resented dog keeping and therefore refused to talk. About 958 dogs were owned by respondents, of these dogs, 56%, 27% and 17% were females, intact males and castrates, respectively.

There was no association between the respondent’s literacy status and dog-sex preference in performance of such uses as security, hunting, and “economic reasons”. The three most-important reasons for dog keeping were security, pet and hunting. However, the use of castrates for security was favoured by most keepers irrespective of literacy status. Most of the respondents agreed that dogs are used as gifts and 72% of the respondents agree that dog meat is a protein source. Castration should be encouraged because of its market value. Usage was low of veterinary services, confinement, and bathing of dogs.     

舒安对犬的麻醉效果观察

以新近研制的中小动物全身麻醉剂－舒安为实验药品，对１０只实验犬进行麻醉效果观察，并试用于９７例犬的手术麻醉，实验结果表明，舒安对犬具有良好的麻醉作用，毒副作用小，有较强的镇静、镇痛和肌松作用，对体温、脉搏、呼吸等生理功能影响小，恢复快。麻醉诱导平衡而迅速，苏醒期无明显兴奋现象。腺体分泌减少，呕吐机率低。     

母犬阴道和子宫的微生物区系

发情周期不同阶段的母犬阴过来样８２只和子日采样７８只中确定需氧与厌氧微生物．平均总细菌数：８２只１００ｍｇ阴道内容物ｌｏｇ５．０±ｌｏｇ１．５（ｌｏｇ２．４－ｌｏｓ８．８）。发情期的细菌数（ｌｏｇ７．８±ｌｏｇ０．７）放非发情期（ｌｏｇ４．４±ｌｏｇ１．０〕、怀孕期（ｌｏｇ５．９±ｌｏｇ１．３）和产后期（ｌｏｇ５．１±ｌｏｇ１．５）的母犬明显为高（Ｐ＞０．０５）。从阴道分离出的普通微生物为类细菌科、待球菌属、巴氏菌属和支原体属。从７８只子宫样品中４８只（占６８％）也分离出微生物。每１００ｍｇ子宫内容物中总细菌数范围为ｌｏｇ１．６－ｌｏｇ８．３,并分离出葡萄球菌属和支原体屁。尽管子宫内微生物与阴道相似,然而子宫内容物培养可进行单一分离出微生物,在大多数子宫内未见到病理变化。显然,阴道中微生物常常会进入子宫,然而它们很少引起子宫的感染。