Cytokine profile of a Holstein calf with bovine leukocyte adhesion deficiency during the acute-phase inflammatory response

Changes in interleukin (IL)-1beta, IL-6 and IL-8 in serum, and their mRNA expression on neutrophils from a 4.6-month old Holstein young calf with bovine leukocyte adhesion deficiency (BLAD) during the acute phase were evaluated. IL-1beta concentrations in the serum of the calf with BLAD at age 143-162 days ranged from 8.7 to 16.6 ng/ml, whereas the values were less than 2.7 ng/ml in control calves. Serum IL-6 (0.04 ng/ml) was only detected on the 1st day when the animal was diagnosed with the BLAD. IL-1beta and IL-8 mRNA expression on neutrophils from the affected calf appeared to be similar to those of controls. Serum cytokine levels and their mRNA expression on neutrophils from the calf with BLAD appeared to be little affected by the deficient expression of beta(2)-integrin on leukocytes, and are considered to be modulated by the inflammatory stimuli.     

Systemic AL amyloidosis associated with multiple myeloma in a horse

AL amyloidosis is the most common type of systemic amyloidosis in humans, and it is frequently associated with multiple myeloma. But, AL amyloidosis is very rare in domestic animals. A 16-year-old Quarter horse gelding was diagnosed with systemic AL amyloidosis associated with multiple myeloma. Clinical problems were rapid weight loss, muscle atrophy, soft unformed stool, and ventral edema. Grossly, diffuse gastrointestinal hemorrhage, markedly thickened jejunal mucosa, and splenomegaly were present. Microscopically, diffuse severe amyloid deposits were present in the lamina propria of glandular stomach, duodenum, and jejunum. Much of the spleen and sternal bone marrow was replaced by neoplastic round cells, and multiple foci of amyloid were also present in the spleen and bone marrow. Electron microscopy revealed the neoplastic round cells to be of plasma cell origin, and the amyloid showed a strongly positive immunoreactivity with polyclonal anti-human immunoglobin lambda light-chain antisera. To our knowledge, this is the second report describing systemic AL amyloidosis in domestic animals-associated plasma cell neoplasia and the first associated with multiple myeloma, as is common in humans.     

Detection of bovine leukocyte adhesion deficiency (BLAD) in Turkish native and Holstein cattle

The purpose of this work was to study whether the bovine leukocyte adhesion deficiency (BLAD) allele is present in native cattle breeds and the Holstein breed in Turkey. Blood samples were obtained from 120 Holstein, 20 Brown Swiss, 20 Anatolian Black, 20 Turkish Grey, 20 South Anatolian Red and 20 East Anatolian Red cattle. The isolated DNA materials were multiplied in PCR using the primer developed by Kriegesmann et al. (1997). In order to determine the area of mutation in PCR products, the PCR products were digested with TaqI endonuclease enzyme. The resulting fragments were analysed on 2% agarose gel for the absence of a TaqI restriction site. It was found that two of the Holstein cattle (a bull and a cow) were heterozygote BLAD carriers. There was no homozygote BLAD animal. The BLAD allele was not found in the other breeds used in the study. The mutant BLAD allele frequency in the 120 Holstein cattle calculations was 0.0084.     

Screening for bovine leukocyte adhesion deficiency,deficiency of uridine monophosphate synthase,complex vertebral malformation,bovine citrullinaemia,and factor XI deficiency in Holstein cows reared in Turkey

Background

Bovine leukocyte adhesion deficiency (BLAD), deficiency of uridine monophosphate synthase (DUMPS), complex vertebral malformation (CVM), bovine citrullinaemia (BC) and factor XI deficiency (FXID) are autosomal recessive hereditary disorders, which have had significant economic impact on dairy cattle breeding worldwide. In this study, 350 Holstein cows reared in Turkey were screened for BLAD, DUMPS, CVM, BC and FXID genotypes to obtain an indication on the importance of these defects in Turkish Holsteins.

Methods

Genomic DNA was obtained from blood and the amplicons of BLAD, DUMPS, CVM, BC and FXID were obtained by using PCR. PCR products were digested with TaqI, AvaI and AvaII restriction enzymes for BLAD, DUMPS, and BC, respectively. These digested products and PCR product of FXID were analyzed by agarose gel electrophoresis stained with ethidium bromide. CVM genotypes were detected by DNA sequencing. Additionally, all genotypes were confirmed by DNA sequencing to determine whether there was a mutant allele or not.

Results

Fourteen BLAD, twelve CVM and four FXID carriers were found among the 350 Holstein cows examined, while carriers of DUMPS and BC were not detected. The mutant allele frequencies were calculated as 0.02, 0.017, and 0.006 for BLAD, CVM and FXID, respectively with corresponding carrier prevalence of 4.0% (BLAD), 3.4% (CVM) and 1.2% (FXID).

Conclusion

This study demonstrates that carriers of BLAD, CVM and FXID are present in the Turkish Holstein population, although at a low frequency. The actual number of clinical cases is unknown, but sporadic cases may appear. As artificial insemination is widely used in dairy cattle breeding, carriers of BLAD, CVM and FXID are likely present within the population of breeding sires. It is recommended to screen breeding sires for these defective genes in order to avoid an unwanted spread within the population.     

牛白细胞粘附缺陷病携带牛CD18的部分基因序列分析

牛白细胞粘附缺陷病（bovine leukocyte adhesion deficiency ,BLAD）是一种常染色体单基因隐性遗传疾病。目前虽已确定BLAD病因与人的白细胞粘附缺陷病（leukocyte adhesion deficiency, LAD），为白细胞表面的一种称为整合素CD18亚单位表达缺陷所致，但至今关于CD18基因突变情况研究较少。为了深入研究和探讨BLAD发病机制，本研究将在我国黑龙江省东部地区调查发现的6头BLAD携带牛CD18部分基因进行了克隆和序列比较分析。     

The clinical and pathological findings in two Friesian calves with leukocyte adhesion deficiency

Leukocyte adhesion deficiency is an inherited defect of neutrophil function reported in man, dogs and cattle. In cattle it was first described in 1983 and to date is found only in Holstein Friesians where it is inherited as an autosomal recessive trait. The neu-trophils of affected calves are less able to migrate from blood vessels to sites of inflammation, thus greatly compromising their ability to fight infection.     

An improved DNA test for bovine leucocyte adhesion deficiency

A modified DNA test, based on the polymerase chain reaction, was developed for the monogenic recessive disease bovine leucocyte adhesion deficiency ( ). The test was improved by the selection of new primers which facilitated the interpretation of the results. An easily scorable banding pattern makes the test useful in cattle breeding schemes and for clinical diagnosis. A total of 2381 samples was analysed over a period of three years. The carrier rate among young bulls at artificial insemination ( ) stations decreased from 11·6 per cent in 1993 to 9·9 per cent in the first five months of 1995. Continuous screening of young bulls before entering is still recommended unless both parents are proven to be genetically free of . The carrier rate among clinically suspect animals was not increased, and carriers are therefore not expected to be immunodeficient. Despite all efforts to eradicate the disease, calves with were still observed in 1995.     

Canine leukocyte adhesion deficiency colony for investigation of novel hematopoietic therapies

The genetic immunodeficiency disease canine leukocyte adhesion deficiency (CLAD) was originally described in juvenile Irish Setters with severe, recurrent bacterial infections. CLAD was subsequently shown to result from a mutation in the leukocyte integrin CD18 subunit which prevents leukocyte surface expression of the CD11/CD18 complex. We describe the development of a mixed-breed CLAD colony with clinical features that closely parallel those described in Irish Setters. We demonstrate that the early identification of CLAD heterozygotes and CLAD-affected dogs by a combination of flow cytometry and DNA sequencing allows the CLAD-affected animals to receive life-saving antibiotic therapy. The distinct clinical phenotype in CLAD, the ability to detect CD18 on the leukocyte surface by flow cytometry, and the history of the canine model in marrow transplantation, enable CLAD to serve as an attractive large-animal model for the investigation of novel hematopoietic stem cell and gene therapy strategies.     

A novel method for rapid and reliable detection of complex vertebral malformation and bovine leukocyte adhesion deficiency in Holstein cattle

Background: Complex vertebral malformation (CVM) and bovine leukocyte adhesion deficiency (BLAD) are two autosomal recessive lethal genetic defects frequently occurring in Holstein cattle, identifiable by single nucleotide polymorphisms. The objective of this study is to develop a rapid and reliable genotyping assay to screen the active Holstein sires and determine the carrier frequency of CVM and BLAD in Chinese dairy cattle population. Results: We developed real-time PCR-based assays for discrimination of wild-type and defective alleles, so that carriers can be detected. Only one step was required after the DNA extraction from the sample and time consumption was about 2 hours. A total of 587 Chinese Holstein bulls were assayed, and fifty-six CVM-carriers and eight BLAD-carriers were identified, corresponding to heterozygote carrier frequencies of 9.54% and 1.36%, respectively. The pedigree analysis showed that most of the carriers could be traced back to the common ancestry, Osborndale Ivanhoe for BLAD and Pennstate Ivanhoe Star for CVM. Conclusions: These results demonstrate that real-time PCR is a simple, rapid and reliable assay for BLAD and CVM defective allele detection. The high frequency of the CVM allele suggests that implementing a routine testing system is necessary to gradually eradicate the deleterious gene from the Chinese Holstein population.     

Bovine leukocyte adhesion deficiency: in vitro assessment of neutrophil function and leukocyte integrin expression.

Bovine leukocyte adhesion deficiency (BLAD) was identified in a two-month-old Holstein heifer calf using DNA-polymerase chain reaction analysis of the affected calf and other clinical parameters. Neutrophil integrin expression (CD18, CD11a, CD11c), aggregation, and transendothelial migration were studied in vitro. Neutrophils were isolated from the affected calf and from normal, healthy, age-matched control Holstein calves. Neutrophils isolated from the affected BLAD calf had decreased expression of leukocyte integrins on their cell surface, decreased ability to aggregate in response to chemotactic stimuli, and decreased ability to migrate across bovine endothelial cell monolayers in vitro. Transendothelial migration of neutrophils from normal calves was reduced to levels comparable to the BLAD neutrophils by treatment with an anti-CD18 monoclonal antibody (MAb 60.3). Peripheral-blood lymphocytes from the BLAD calf also expressed negligible levels of leukocyte integrins, similar to their neutrophil counterparts. Our experimental findings in vitro correlate well with the clinical observations of decreased leukocyte trafficking and diminished host defense in leukocyte adhesion-deficient animals. The syndrome of BLAD may be a suitable model for one of the human leukocyte adhesion deficiency disorders.     

Analysis of surface antigen expression and host defense function in leukocytes from calves heterozygous or homozygous for bovine leukocyte adhesion deficiency

OBJECTIVE: To analyze surface antigen expression and functional responses of leukocytes from calves heterozygous and homozygous for bovine leukocyte adhesion deficiency (BLAD). ANIMALS: 8 clinically normal calves, 4 calves heterozygous for BLAD, and 4 calves homozygous for BLAD. PROCEDURE: Surface antigen expression was examined by flow cytometric analysis of leukocytes stained with monoclonal antibodies. Neutrophil function analyses included phagocytosis and killing of Candida albicans and measurement of respiratory burst activity using cytochrome c and dihydrorhodamine 123 assays. Differential leukocyte counts also were performed. RESULTS: Leukocytes from heterozygous calves were similar to those of clinically normal calves with respect to surface antigen expression, C albicans phagocytosis and killing, and respiratory burst activity. In contrast, neutrophils from calves homozygous for BLAD had significantly reduced phagocytic and yeast-killing capacity but had higher respiratory burst activity than cells from clinically normal or heterozygous calves. Homozygous calves also had extreme neutrophilia and significantly more immature neutrophils. CONCLUSIONS: The heterozygous BLAD genotype does not cause detectable functional differences in leukocytes, compared with those of clinically normal calves. In contrast, leukocytes from homozygous calves seem to upregulate alternative host defense capabilities (eg, respiratory burst activity) to partially compensate for the lack of typical adherence-dependent host defense functions.     

Systemic AA amyloidosis in the common marmoset

The common marmoset (Callithrix jacchus) is a small New World primate native to Brazil that has been used extensively in biomedical research. A retrospective analysis of archived hematoxylin and eosin-stained tissue sections and clinical records was conducted at the New England Primate Research Center on 86 marmosets more than 1 year of age that were euthanized during the past decade because of morbidity and failure to thrive. Approximately 17% (15 of 86) were found to have amyloid deposits in one or more organs, including the liver, adrenal glands, kidneys, and intestine. This material was shown by amino acid sequence analysis to be composed of serum amyloid A (SAA)-related protein. This type of amyloidosis, designated AA or "secondary," is associated typically with an inflammatory process that induces elevated levels of the SAA amyloidogenic precursor molecule. Notably, there were no significant pathologic differences or other distinguishing features in animals with amyloid versus those without; furthermore, on the basis of the limited number of serum specimens available for analysis, the SAA concentrations in the two groups were comparable, thus suggesting the possible inheritable nature of the disorder. In this respect, the common marmoset provides a unique experimental model for study of the pathogenesis and treatment of AA and other forms of systemic amyloidosis.     

Immunoglobulin M associated with secretory component and immunoglobulin A deficiency in bovine colostrum.

Immunoglobulin (Ig) M purified from bovine colostrum was examined by an immunodiffusion analysis with antisecretory IgA serum and was found to be associated with a secretory component. Some of the combined proteins were dissociated if treated with 5 M guanidine-HCl and others were not. Another immunodiffusion analysis of 23 specimens of colostrum led to the finding that certain colostrums were deficient in IgA, even though they contained IgG and IgM.     

Systemic cutaneous and subcutaneous amyloidosis in the horse]

A 9-year-old horse had numerous firm, painless nodules of the skin and subcutis. Moderately vascular granulation tissue with numerous uni- or multinuclear reticuloendothelial cells was in the nodules and the regional lymph nodes but not in the viscera. By using special stains and electron microscopy, widespread amyloid deposits, mainly in the cytoplasm of reticuloendothelial cells, were identified. Amyloid was probably produced within the reticuloendothelial cells, then expelled from the dying cell and deposited in the intercellular space.     

Diagnosis of secondary amyloid in bovine renal amyloidosis

In paraffin sections, using histochemical methods and the indirect immunofluorescence technique for protein AA, the amyloid of different types of bovine renal amyloidosis was characterized and compared with amyloid of other species, including man. The bovine amyloid was found to be of the secondary type in all cases.Resume Sur des coupes incluses dans de la paraffine, en utilisant des méthodes histochimiques et l'immunofluorescence indirecte pour la protéine AA, la substance amyloïde de différents types d'amyloïdose rénale bovine a été caractérisée et comparée avec celle d'autres espèces y compris l'homme Dans tous les cas la substance amyloïde a été classée comme une sustance amyloïde secondaire.

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Kurzfassung Bei Anwendung histochemischer Methoden und der indirekten Immunofluoreszenztechnik zum Nachweis des Proteins AA wurde in Paraffinschnitten das Amyloid bei verschiedenen Formen der bovinen Nierenamyloidose charakterisiert und mit Amyloid anderer Spezies einschliesslich des Menschen verglichen. Das bovine Amyloid gehörte in allen Fällen zum sekundären Typ.

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Riassunto Usando metodi istochimici e la tecnica dell'immunofluorescenza indiretta per la proteina AA, la sostanza amiloide nei diversi tipi di amiloidosi renale del bovino, in sezioni di paraffina, è stata caratterizzata e confrontata con la sostanza amiloide di altre specie, uomo compreso. La sostanza amiloide del bovino è risultata essere di tipo secondario in tutti i casi.

     

Systemic xanthomatosis associated with hyperchylomicronaemia in a cat

We report a case of systemic xanthomatosis in a 4-month-old domestic cat. The kitten presented with multiple cutaneous lesions and 'cream tomato soup' coloured blood. Necropsy revealed multiple, whitish, nodular lesions, compatible with xanthomas, on most of the abdominal organs (liver, spleen, kidney, adrenal glands, mesentery and colon). The diagnosis was confirmed by histopathological examination. This is the first report of granulomatous colitis associated with feline xanthomatosis.     

Cell adhesion molecules, leukocyte trafficking, and strategies to reduce leukocyte infiltration

Leukocyte-endothelial cell interactions are mediated by various cell adhesion molecules. These interactions are important for leukocyte extravasation and trafficking in all domestic animal species. An initial slowing of leukocytes on the vascular endothelium is mediated by selectins. This event is followed by (1) activation of beta2 integrins after leukocyte exposure to cytokines and pro-inflammatory mediators, (2) adherence of leukocyte beta2 integrins to vascular endothelial ligands (eg, intercellular adhesion molecule-1 [ICAM-1]), (3) extravasation of leukocytes into tissues through tight junctions of endothelial cells mediated by platelet and endothelial cell adhesion molecule-1 (PECAM-1), and (4) perivascular migration through the extracellular matrix via beta1 integrins. Inhibiting excessive leukocyte egress and subsequent free radical-mediated damage caused by leukocyte components may attenuate or eliminate tissue damage. Several methods have been used to modify leukocyte infiltration in various animal models. These methods include nonspecific inhibition of pro-inflammatory mediators and adhesion molecules by nonsteroidal anti-inflammatory drugs (NSAIDs) and glucocorticoids, inhibition of cytokines and cytokine receptors, and inhibition of specific types of cell adhesion molecules, with inhibitors such as peptides and antibodies to beta2 integrins, and inhibitors of selectins, ICAMs, and vascular cell adhesion molecule-1 (VCAM-1). By understanding the cellular and molecular events in leukocyte-endothelial cell interactions, therapeutic strategies are being developed in several animal models and diseases in domestic animal species. Such therapies may have clinical benefit in the future to overcome tissue damage induced by excessive leukocyte infiltration.     

The frequency of the canine leukocyte adhesion deficiency (CLAD) allele within the Irish Setter population of Australia

OBJECTIVE: To determine the frequency of the 107G-->C canine leukocyte adhesion deficiency (CLAD) mutation in Irish Setters from the Australian breeding population. METHOD: Genomic DNA was isolated from 87 Irish Setter blood samples and a region of the beta-2 integrin gene (ITGB2), encompassing the mutation, was amplified using real-time Polymerase Chain Reaction (PCR). Two fluorescently labelled probes were hybridised to the fragment, and fluorescence resonance energy transfer (FRET) was used to detect the 107G-->C mutation responsible for CLAD. RESULTS: Three new heterozygotes were identified among 87 healthy Irish Setters from Australia. All originated from a litter sired by a known heterozygote. A total of seven heterozygotes have now been identified in 92 dogs (7.6%), representing over 90% of all major breeding stock in five Australian states. Two of the heterozygotes were recently imported adult dogs and the others were their offspring. CONCLUSIONS: The frequency of the 107C allele in the Australian population of Irish Setters is lower than that in Europe. Selective breeding programs should be adopted to eliminate the mutant allele presently in two breeding lines.