A mutation in the CLN8 gene in English Setter dogs with neuronal ceroid-lipofuscinosis

A heritable neurodegenerative disease of English Setters has long been studied as a model of human neuronal ceroid-lipofuscinosis (NCL). Megablast searches of the first build of the canine genome for potential causative genes located the CLN8 gene near the q telomere of canine chromosome 37, close to a marker previously linked to English Setter NCL. Sequence analysis of the coding region from affected dogs revealed a T-to-C transition in the CLN8 gene that predicts a p.L164P missense mutation. Leucine 164 is conserved in four other mammalian species. The C allele co-segregated with the disease phenotype in a two-generation English Setter family in a pattern consistent with autosomal recessive inheritance. All four NCL-affected family members were C/C homozygotes and all four obligate carriers were C/T heterozygotes; whereas, 103 unrelated dogs were all T/T homozygotes. These findings indicate that the CLN8 T-to-C transition is the likely cause of English Setter NCL.     

Expression of versican in relation to chondrogenesis-related extracellular matrix components in canine mammary tumors

Versican plays a role in tumor cell proliferation and adhesion and may also regulate cell phenotype. Furthermore, it is one of the pivotal proteoglycans in mesenchymal condensation during prechondrogenesis. We have previously demonstrated accumulation of versican protein in myoepithelial-like spindle cell proliferations and myxoid tissues of complex and mixed mammary tumors of dogs. The objective of this study was to investigate whether the high expression of versican relates to prechondrogenesis in these tissues. Therefore, we aimed to identify cartilage markers, such as collagen type II and aggrecan both at mRNA and protein level in relation to versican. The neopitope of chondoitin-6-sulphate (3B3) known to be generated in developing cartilage has been investigated by immunohistochemisty and a panel of antibodies were used to characterize the phenotype of cells that are involved in cartilage formation. In addition, co-localization of versican with hyaluronan and link protein was studied. RT-PCR revealed upregulation of genes of versican, collagen type II and aggrecan in neoplastic tissues, especially in complex and mixed tumors. Immunohistochemistry showed the expression of cartilage biomarkers not only in the cartilagenous tissues of mixed tumors, but also in myoepitheliomas and in the myoepithelial-like cell proliferations and myxoid areas of complex and mixed tumors. The results show the cartilagenous differentiation of complex tumors and myoepitheliomas and indicate that the myxoid tissues and myoepithelial-like cell proliferations are the precursor tissues of the ectopic cartilage in mixed tumors. Furthermore, we suggest that cartilage formation in canine mammary tumors is a result of (myo)epithelial to mesenchymal transition.     

Defining functional gene‐circuit interfaces in the mouse nervous system

Complexity in the nervous system is established by developmental genetic programs, maintained by differential genetic profiles and sculpted by experiential and environmental influence over gene expression. Determining how specific genes define neuronal phenotypes, shape circuit connectivity and regulate circuit function is essential for understanding how the brain processes information, directs behavior and adapts to changing environments. Mouse genetics has contributed greatly to current percepts of gene‐circuit interfaces in behavior, but considerable work remains. Large‐scale initiatives to map gene expression and connectivity in the brain, together with advanced techniques in molecular genetics, now allow detailed exploration of the genetic basis of nervous system function at the level of specific circuit connections. In this review, we highlight several key advances for defining the function of specific genes within a neural network .     

Comparative Genomics of X-linked Muscular Dystrophies: The Golden Retriever Model

Duchenne muscular dystrophy (DMD) is a devastating disease that dramatically decreases the lifespan and abilities of affected young people. The primary molecular cause of the disease is the absence of functional dystrophin protein, which is critical to proper muscle function. Those with DMD vary in disease presentation and dystrophin mutation; the same causal mutation may be associated with drastically different levels of disease severity. Also contributing to this variation are the influences of additional modifying genes and/or changes in functional elements governing such modifiers. This genetic heterogeneity complicates the efficacy of treatment methods and to date medical interventions are limited to treating symptoms. Animal models of DMD have been instrumental in teasing out the intricacies of DMD disease and hold great promise for advancing knowledge of its variable presentation and treatment. This review addresses the utility of comparative genomics in elucidating the complex background behind phenotypic variation in a canine model of DMD, Golden Retriever muscular dystrophy (GRMD). This knowledge can be exploited in the development of improved, more personalized treatments for DMD patients, such as therapies that can be tailor-matched to the disease course and genomic background of individual patients.     

Heterologous Expression and Functional Characterization of Matrix Metalloproteinase-11 From Canine Mammary Tumor

Matrix metalloproteinases (MMPs) are reported to be involved in tumor growth, apoptosis, angiogenesis, invasion, and development of metastases. These are zinc containing metalloproteases, known for their role in extracellular matrix degradation. MMP-11 (stromelysin3) is reported to be highly expressed in breast cancer, therefore it may act as marker enzyme for breast cancer progression. The present work was carried out to produce recombinant canine (Canis lupus familiaris) MMP-11 lacking the signal and propeptide in E. coli by optimizing its expression and purification in biologically active form and to functionally characterize it. A bacterial protein expression vector pPROEX HTc was used. The MMP-11 mature peptide encoding gene was successfully cloned and expressed in E. coli and the purified recombinant enzyme was found to be functionally active. The recombinant enzyme exhibited caseinolytic activity and could be activated by Trypsin and 4-Amino phenyl mercuric acetate (APMA). However Ethylene diamine tertra acetate (EDTA) inhibited the enzyme's caseinolytic activity. The recombinant enzyme degraded extracellular matrix constituents and facilitated migration of MDCK (Madin-Darby canine kidney) cells through BD Biocoat Matrigel invasion chambers. These results suggest that in vivo MMP-11 could play a significant role in the turnover of extracellular matrix constituents.     

犬繁殖调控技术研究进展

由于犬具有一些独特的生殖生理特点,使犬的繁殖调控技术的发展一直面临着一些困难。目前犬繁殖调控技术的研究远远落后于其他哺乳动物。本文综述了犬诱导发情、精液保存、人工受精、胚胎移植及克隆等犬的繁殖调控技术国内外研究进展。     

犬SLAM受体mRNA荧光定量PCR检测方法的建立和初步应用

目的建立荧光定量PCR方法,检测犬不同组织中SLAM受体mRNA的表达水平。方法以犬GAPDH为内参基因采用△△ct法,分析SLAM受体mRNA在犬体内不同组织中的表达。结果此方法有较高的重复性,变异系数在0．89％-2．35％。以SLAM受体在心脏的表达为1倍值,结果显示受体mRNA在脾脏中表达最高,为38．49倍;肺门淋巴结、肠系膜淋巴结、腹股沟淋巴结中表达次之,分别为9．13、8．58、6．24倍;膀胱中表达最低。结论成功建立了检测SLAM受体mRNA在不同组织中表达水平的荧光定量PCR检测方法。     

Coronary vasoconstrictor effect of ghrelin is not mediated by growth hormone secretagogue receptor 1a type in dogs

Ghrelin (GHR) is a recently discovered endocrine regulatory peptide of gastrointestinal origin with multiple functions including cardiovascular effects. However, contradictory data are available on the vascular actions of GHR in different organs and species. The aim of this study was to characterize the direct effect of the peptide on the canine coronary bed and to evaluate the role of the growth hormone secretagogue receptor (GHS-R) in the effect of GHR on coronary arterioles. The presence of GHS-R1a and 1b subtypes in canine coronary arterioles was investigated using Western blotting and immunohistochemistry. Responses of coronary arterioles with spontaneous and elevated vascular tone (the latter evoked by the thromboxane mimetic agent U46619, 10⁻⁷-10⁻⁶ mol/l) to GHR (10⁻⁹-3 × 10⁻⁷ nmol/l) were recorded by video-microscopy as changes of vessel diameter. Positive immunostaining for both GHS-R subtypes was found in the wall of intramural arterioles. The microarteriographic study results showed that GHR alone could not elicit any significant effect on vessel diameter of arterioles with spontaneous tone. However, when vascular smooth muscle was preconstricted by the thromboxane mimetic agent U46619, administration of GHR induced further constriction (+31 ± 9% increase in contraction p < 0.01). This was not abolished by the specific blockade of GHS-R1a by d-Lys³-GHRP-6 (5 × 10⁻⁶ mol/l). The results suggest that GHR induces tone-dependent constriction of canine coronary arterioles which is mediated by a receptor other than GHS-R1a.     

Evolution of the mandibular third premolar crown in early Australopithecus

The Pliocene hominins Australopithecus anamensis and Australopithecus afarensis likely represent ancestor-descendent taxa—possibly an anagenetic lineage—and capture significant change in the morphology of the canine and mandibular third premolar (P₃) crowns, dental elements that form the canine honing complex in nonhuman catarrhines. This study focuses on the P₃ crown, highlighting plesiomorphic features in A. anamensis. The A. afarensis P₃ crown, in contrast, is variable in its expression of apomorphic features that are characteristic of geologically younger hominins. Temporal variation characterizes each taxon as well. The A. anamensis P₃ from Allia Bay, Kenya expresses apomorphic character states, shared with A. afarensis, which are not seen in the older sample of A. anamensis P₃s from Kanapoi, Kenya, while spatiotemporal differences in shape exist within the A. afarensis hypodigm. The accumulation of derived features in A. afarensis results in an increased level of P₃ molarisation. P₃ molarisation did not evolve concurrent with postcanine megadontia and neither did the appearance of derived aspects of P₃ occlusal form coincide with the loss of canine honing in hominins, which is apparent prior to the origin of the genus Australopithecus. A. afarensis P₃ variation reveals the independence of shape, size, and occlusal form. The evolution of the P₃ crown in early Australopithecus bridges the wide morphological gap that exists between geologically younger hominins on the one hand and extant apes and Ardipithecus on the other.     

犬瘟热病毒TN株血凝基因的克隆与序列分析

用RT PCR方法对分离的TN野毒株H基因进行了扩增 ,并将其克隆到pGEM T载体上 ,进行核苷酸序列测定。结果TN野毒株H基因ORF全长 1 ,81 5bp ,编码 60 4个氨基酸。与GenBank中己报道的 7个CDV毒株相比 ,H基因核苷酸序列的同源性在 92 %～ 99%之间 ,推导的H蛋白氨基酸序列的同源性在 91 %～ 99%之间。TN株H蛋白潜在的N 联糖基化位点为 8个 ,而Onderstepoort弱毒株H蛋白为 4个 ;其中N端第 1 9～ 2 1、 30 9～ 31 1、 5 8