Calf Lung Research Articles

The fascinating story of surfactant.

The fascinating story of surfactant.

Prevalence of different OmpH-types among Pasteurella multocida isolated from lungs of calves with respiratory problems.

Pasteurella multocida is among the most important respiratory pathogens of cattle. Outer-membrane protein (OmpH) constitutes an essential bacterial antigen and is well studied in avian bacterial strains. Studies on isolates from cattle with signs of respiratory disease caused by Pasteurella multocida serotypes A and D have not yet been covered in the literature. The objective of this study was a comparative analysis of the ompH gene sequences from 83 isolates and four Russian reference strains of P.multocida to assign them to the allelic variants of the gene (OmpH-types). In addition, the above P.multocida strains have been characterized on the basis of capsular serotypes and virulence-associated genes. The isolates were classified into the OmpH -types based on allele specific PCR and gene fragment sequencing. The isolates of capsular serotype A have been subdivided into 6 OmpH -types, of which the most common types identified were A1 and A2. All capsular serotype D isolates belong to the same OmpH-type (D1). On 16 of a total of 23 farms all isolates belong to only one OmpH-type, on 4 farms - to 2, and on 3 farms - to three OmpH-types. The tbpA and pfhA genes were found more often in the isolates of capsular group А as compared to capsular group D (p≤0.05). OmpH-types of serogroup А differ significantly (p≤0.05) among themselves by the prevalence of the pfhA and hgbB genes.

Histological Study of Pneumonic Lungs of Calves in Batna Slaughterhouse

93 lung fragments were taken from 31 calves, aged approximately from 1 to 2 years, with pulmonary lesions in order to define the nature of the lesions and to highlight the prevalence of respiratory diseases in young bulls. Results showed a prevalence of 4.67%. The seasonal impact of the lesions has been highly marked. Whatever the season, the most common lung injury is red hepatization injury (or consolidation) (38.70%) followed by grey hepatization (6.45%), emphysema (19.35%), pneumonia (32.25%) and finally bronchopneumonia (3.22%). Our study showed that lesions were preferentially localized in the right apical lobe. This investigation demonstrated the strong spread of atypical pneumonia in the cattle population (young bulls) at the slaughterhouse of Batna justifying stunting and losses recorded in cattle farms of the region. For this, it was deemed urgently the appreciation of the extent and severity of macroscopic and microscopic tissue changes in order to limit this increased infection.

Impact of Enzymes and Toxins Potentiality of Four Aspergillus Species to Cause Aspergillosis

Aspergillus species are the main causing agents of invasive apergillosis chest disease. Eighty isolates of Aspergillus species, A. flavus (20), A. fumigatus (15), A. niger (30) and A. terreus (15 isolates), previously isolated and identified from aspergillosis suspected patients at our lab in Assiut university hospitals, were assayed for their enzymes and toxins profile. The results revealed that, all of the tested isolates were able to utilize calf lung tissue and produce catalase and peroxidase enzymes. Meanwhile, 82.5-90% of the fungal isolates had the ability to produce protease, lipase, urease and phospholipase, whereas, 70% of isolates exhibited hemolytic activities. Thin layer chromatography (TLC) of the cleaned extracts of the tested isolates exhibited the ability of all A. flavus isolates assayed to produce aflatoxins B1 and G1, 53%, moreover, A. fumigatus isolates produced fumagillin and gliotoxin. On the other side, 43.3% and 23.3% of A. niger isolates produced ochratoxins and gliotoxin respectively. Virulence assay of 10 μl of gliotoxin standard and cleaned extracts of A. fumigatus toxic isolates showed necrotic area on Guinea Pigs lungs compared to the control. Thus, opportunistic fungi isolated from aspergillosis patients possess high enzymatic and toxic profile that might play an important role in their mycopathy.

Tensiometric and Phase Domain Behavior of Lung Surfactant on Mucus-like Viscoelastic Hydrogels.

Lung surfactant has been observed at all surfaces of the airway lining fluids and is an important contributor to normal lung function. In the conducting airways, the surfactant film lies atop a viscoelastic mucus gel. In this work, we report on the characterization of the tensiometric and phase domain behavior of lung surfactant at the air-liquid interface of mucus-like viscoelastic gels. Poly(acrylic acid) hydrogels were formulated to serve as a model mucus with bulk rheological properties that matched those of tracheobronchial mucus secretions. Infasurf (Calfactant), a commercially available pulmonary surfactant derived from calf lung extract, was spread onto the hydrogel surface. The surface tension lowering ability and relaxation of Infasurf films on the hydrogels was quantified and compared to Infasurf behavior on an aqueous subphase. Infasurf phase domains during surface compression were characterized by fluorescence microscopy and phase shifting interferometry. We observed that increasing the bulk viscoelastic properties of the model mucus hydrogels reduced the ability of Infasurf films to lower surface tension and inhibited film relaxation. A shift in the formation of Infasurf condensed phase domains from smaller, more spherical domains to large, agglomerated, multilayer structures was observed with increasing viscoelastic properties of the subphase. These studies demonstrate that the surface behavior of lung surfactant on viscoelastic surfaces, such as those found in the conducting airways, differs significantly from aqueous, surfactant-laden systems.

The Adult Calfactant in Acute Respiratory Distress Syndrome Trial

Surfactant has been shown to be dysfunctional in ARDS, and exogenous surfactant has proven effective in many forms of neonatal and pediatric acute lung injury (ALI). In view of the positive results of our studies in children along with evidence that surfactant-associated protein B containing pharmaceutical surfactants might be more effective, we designed a multiinstitutional, randomized, controlled, and masked trial of calfactant, a calf lung surfactant, in adults and children with ALI/ARDS due to direct lung injury. Adult subjects within 48 h of initiation of mechanical ventilation for direct ARDS were randomized to receive up to three interventions with instilled calfactant vs air placebo. The primary outcome was 90-day all-cause mortality. Three hundred seventeen subjects were enrolled, 308 of whom could be evaluated. There were no significant baseline differences between groups. Calfactant administration was not associated with improved survival, lengths of stay, or oxygenation. Calfactant instillation was frequently associated with transient hypoxia and hypotension. The study was stopped at the first interim analysis at the sponsor's request. Administration of calfactant was not associated with improved oxygenation or longer-term benefits relative to placebo in this randomized, controlled, and masked trial. At present, exogenous surfactant cannot be recommended for routine clinical use in ARDS. ClinicalTrials.gov; No.: NCT00682500; URL: www.clinicaltrials.gov.

Clinico-Pathological Studies in Some Respiratory Diseases in Newly Born Buffalo Calves

A Total of 40 buffalo calves aged from 2 weeks to 3 months were used in this study. The animals were divided into two main groups the first group (group I) represented as a control group. The second group (group II) showed the clinical signs of pneumonia (natural diseased non- treated). Group (II) was subdivided into two subgroups: the first subgroup (group A) naturally diseased and treated with florofenicol (Nuflor), and the second subgroup (group B) naturally diseased and treated with tulathromycin (Draxxin). Concerning the hematological findings of the diseased non treated group (II) revealed anemia in addition to neutrophilic leukocytosis with increase in both TNF- α, IL-6 and decrease in both IgG and IgM levels was observed compared to control healthy group. While subgroups (A & B) revealed increase in RBCs count, Hb concentration, PCV, MCHC, IgG and IgM and decrease in MCV, MCH, TNF- α and IL-6 compared with diseased non treated group (II). Diseased treated subgroups (A&B) revealed decrease in T.L.C, neutrophil and monocytes with increase of lymphocytes, basophils, eosinophils and platelets count compared with diseased non treated group (II). The diseased treated subgroup (B) with Draxxin exhibited improvement in hematological and immunological parameters analysis compared with the diseased treated subgroup (A) with Nuflor. The lungs of dead calves with respiratory diseases were examined microscopically. The obtained results were lung edema, hemorrhage, collapse and different types of pneumonia such as catarrhal, suppurative, fibrinous and interstitial pneumonia. Associated bronchial lymph nodes showing congestion of blood vessels and hyperplasia of lymphocyte cells.

Immunohistochemical Labelling of Cyclooxygenase-2 in Lung Lesions of Calves Infected with Mycoplasma bovis

The pathogenesis and persistence of Mycoplasma bovis (Mb) infection of the respiratory tract is incompletely understood. Cyclooxygenase (COX)-2 is overexpressed during inflammatory responses by different cell types in the lung. This study evaluated COX-2 expression immunohistochemically in the inflammatory lesions of calves with naturally occurring and experimentally induced Mb pneumonia. Experimentally infected lungs showed catarrhal bronchointerstitial pneumonia and varying degrees of peribronchiolar mononuclear cell cuffing. Lesions in calves with spontaneously arising disease included exudative bronchopneumonia and extensive foci of coagulative necrosis surrounded by inflammatory cells. Mb antigen was located in epithelial and inflammatory cells in the airway lumina and surrounding areas of necrosis. COX-2 protein was detected in the lung of all infected calves and was localized to goblet cells, bronchial, bronchiolar and alveolar epithelial cells and macrophages. COX-2 protein was overexpressed during Mb infection and was always associated with areas of pneumonia and with the presence of Mb antigen.

Cytokine expression in lungs of calves spontaneously infected with Mycoplasma bovis.

Cytokine expression in the lung can play an important role during Mycoplasma bovis infection through leukocyte recruitment and activation, and the induction of a broad array of inflammatory mediators. To gain further insight into the pathogenesis of M. bovis-associated pneumonia, cytokine expression was examined, by immunohistochemical methods in formalin-fixed, paraffin wax-embedded tissues, in the lung of 20 calves spontaneously infected. Immunolabelling for tumor necrosis factor alpha (TNF)-α, interleukin-4 (IL-4), IL-10 and interferon-gamma (IFN-γ), was usually associated with pneumonia, particularly in macrophages and lymphocytes, and with the presence of M. bovis antigen. The expression was minimal in lungs from negative controls. The results demonstrated consistent upregulation of TNF-α, IL-4, IL-10 and IFN-γ expression during M. bovis-associated pneumonic lesions. These cytokines can participate in the immune and inflammatory responses during the pulmonary defense mechanisms against M. bovis infection.

Effects of Preinfection With Bovine Viral Diarrhea Virus on Immune Cells From the Lungs of Calves Inoculated With Bovine Herpesvirus 1.1.

The aim of this work was to study the interstitial aggregates of immune cells observed in pulmonary parenchyma of calves preinfected with bovine viral diarrhea virus and challenged later with bovine herpesvirus 1. In addition, the intent of this research was to clarify the role of bovine viral diarrhea virus in local cell-mediated immunity and potentially in predisposing animals to bovine respiratory disease complex. Twelve Friesian calves, aged 8 to 9 months, were inoculated with noncytopathic bovine viral diarrhea virus genotype 1. Ten were subsequently challenged with bovine herpesvirus 1 and euthanized at 1, 2, 4, 7, or 14 days postinoculation. The other 2 calves were euthanized prior to the second inoculation. Another cohort of 10 calves was inoculated only with bovine herpesvirus 1 and then were euthanized at the same time points. Two calves were not inoculated with any agent and were used as negative controls. Pulmonary lesions were evaluated in all animals, while quantitative and biosynthetic changes in immune cells were concurrently examined immunohistochemically to compare coinfected calves and calves challenged only with bovine herpesvirus 1. Calves preinfected with bovine viral diarrhea virus demonstrated moderate respiratory clinical signs and histopathologic evidence of interstitial pneumonia with aggregates of mononuclear cells, which predominated at 4 days postinoculation. Furthermore, this group of animals was noted to have a suppression of interleukin-10 and associated alterations in the Th1-driven cytokine response in the lungs, as well as inhibition of the response of CD8+ and CD4+ T lymphocytes against bovine herpesvirus 1. These findings suggest that bovine viral diarrhea virus preinfection could affect the regulation of the immune response as modulated by regulatory T cells, as well as impair local cell-mediated immunity to secondary respiratory pathogens.

Molecular and pathological insights into Chlamydia pecorum-associated sporadic bovine encephalomyelitis (SBE) in Western Australia

BackgroundDespite its global recognition as a ruminant pathogen, cases of Chlamydia pecorum infection in Australian livestock are poorly documented. In this report, a C. pecorum specific Multi Locus Sequence Analysis scheme was used to characterise the C. pecorum strains implicated in two cases of sporadic bovine encephalomyelitis confirmed by necropsy, histopathology and immunohistochemistry. This report provides the first molecular evidence for the presence of mixed infections of C. pecorum strains in Australian cattle.Case presentationAffected animals were two markedly depressed, dehydrated and blind calves, 12 and 16 weeks old. The calves were euthanized and necropsied. In one calf, a severe fibrinous polyserositis was noted with excess joint fluid in all joints whereas in the other, no significant lesions were seen. No gross abnormalities were noted in the brain of either calf. Histopathological lesions seen in both calves included: multifocal, severe, subacute meningoencephalitis with vasculitis, fibrinocellular thrombosis and malacia; diffuse, mild, acute interstitial pneumonia; and diffuse, subacute epicarditis, severe in the calf with gross serositis. Immunohistochemical labelling of chlamydial antigen in brain, spleen and lung from the two affected calves and brain from two archived cases, localised the antigen to the cytoplasm of endothelium, mesothelium and macrophages. C. pecorum specific qPCR, showed dissemination of the pathogen to multiple organs. Phylogenetic comparisons with other C. pecorum bovine strains from Australia, Europe and the USA revealed the presence of two genetically distinct sequence types (ST). The predominant ST detected in the brain, heart, lung and liver of both calves was identical to the C. pecorum ST previously described in cases of SBE. A second ST detected in an ileal tissue sample from one of the calves, clustered with previously typed faecal bovine isolates.ConclusionThis report provides the first data to suggest that identical C. pecorum STs may be associated with SBE in geographically separated countries and that these may be distinct from those found in the gastrointestinal tract. This report provides a platform for further investigations into SBE and for understanding the genetic relationships that exist between C. pecorum strains detected in association with other infectious diseases in livestock.

New Measurements of Lung Surfactant Interfacial Tension with Micropipette Manipulation Technique

Approximately 50,000-190,000 adults and 24,000 newborn per year in the U.S. develop Acute- and Neonatal-Respiratory Distress Syndrome (ARDS&NRDS), respectively. For more than a couple of decades, many researchers have been tackling the problem of how to investigate lung surfactant behavior, especially of the relationships between the lipids and the natural lung surfactant proteins, by using techniques such as the Langmuir-Trough or Pulsating Bubble Surfactometer (PBS). Currently, we are developing a new method of the micropipette manipulation technique for lung surfactant surface tension measurement. The technique builds on previous studies that used a simple tapered micropipet to measure liquid-gas and liquid-liquid interfacial tension [1-2]. Working with micropipettes that contain the aqueous phase but viewed in air allows us to measure interfacial tensions at controlled pressures and interfacial radii when lung surfactant material is introduced, as in delivery of liquids to the lung alveoli. The new measurements of calf lung surfactant, Infasurf, at the aqueous-air interface showed how the interfacial tension γ at 37 degree Celsius was rapidly reduced from 70.3 to 21.1 ± 0.1 mN/m. One of the advantages of this technique is that by using a second delivery pipette it can be used to observe kinetic processes like adsorption, condensation and desorption of interfacial materials. In order to provide a reversed cone shape for the delivery measurements, we modified the pipette shape to be trumpet-shaped. We will present more details at the meeting regarding lung surfactants and synthetic systems that have been studied.References[1–2] Lee, S., D. H. Kim, and D. Needham, Langmuir, 2001. 17: 5537 & 5544.

Clinico-pathological Responses of Calves Associated with Infection of <em>Pasteurella Multocida</em> Type B and the Bacterial Lipopolysaccharide and Outer Membrane Protein Immunogens

The current study aims to investigate the Clinico-pathological responses of calves associated with the infections of <em>Pasteurella Multocida</em> type B and the bacterial lipopolysaccharide and outer membrane protein immunogens. Alterations in the behavior of animals and pathological lesions observed following innate or experimental infections usually divulge extensive and detrimental changes in the clinical signs, organs and tissues of the animals afflicted with the disease. These alterations are imperative for Veterinary evaluation of herd health. Eight clinically healthy, non-pregnant and non-lactating Brangus cross heifers weighing 150±50 kg were used in the study. The heifers (n = 8) were divided into 4 groups of 2 calves per group. The control calves in group 1 were inoculated intramuscularly with 10 mL of sterile Phosphate Buffered Saline (PBS). Calves in group 2 were inoculated intramuscularly with 10 mL of 10¹² colony forming unit (cfu) of wild-type <em>P. multocida</em> and calves in group 3 were inoculated intravenously with 10 mL of LPS broth extract. Calves in group 4 were inoculated intramuscularly with 10 mL of OMP broth extract. All animals were observed for 48 h for clinical signs, changes in behavior and mortality pattern, including the time of death. The results divulged significant differences in the Clinico-pathological alterations. Calves inoculated with whole cell <em>P. multocida</em> type B: 2 showed a significant (p<0.05) increased in rectal temperature. The affected calves showed significant severe dullness (p<0.000) and significant rumen hypomotility (p<0.000) was also exhibited. The calves showed signs of hypersalivation at 14 h. There is no significant difference (p = 0.240) in pulmonary oedema in the Calves of group 2 compared to control group 1. Calves of group 4 also showed no significant difference in pulmonary oedema (p = 0.612) compared to control group 1. Calves of group 3 showed significantly moderate pulmonary oedema (p<0.000). All the three treatment groups showed significant (p<0.05) differences in the presence of inflammatory cells in the lung. All the three treatment groups showed significant (p<0.05) in the presence of degeneration and necrosis of cells in the lung. Calves of group 2 showed significantly severe haemorrhage (p<0.000) in the lung including groups 3 and 4 (p<0.000) respectively. Calves in group 2 showed significantly (p<0.000) mild thrombus formation. There is no significant thrombus formation in the lung of calves in groups 3 (p = 0.352) and 4 (p = 0.184) respectively. In conclusion, the pathophysiological changes in cattle will assist in the improvement of the vaccines and the vaccination methods that are currently employed in controlling this important disease in Malaysia.

Hydrogen Peroxide Production and Free Radical-mediated Cell Stress in Mycoplasma bovis Pneumonia

Mycoplasma bovis causes chronic pneumonia and polyarthritis in feedlot cattle. M. bovis infects the lungs of most feedlot cattle, but the majority of calves never develop disease. Competing explanations are that some strains of M. bovis are more virulent than othersor, alternatively, that calves require some other abnormality to be present in order for M. bovis to cause disease. We hypothesize that H2O2 production is an important virulence factor of M. bovis, causing oxidative injury to lung tissue. A second hypothesis is that isolates associated with caseonecrotic bronchopneumonia have an increased capacity for H2O2 production. Immunohistochemical markers of oxidative stress (4-hydroxynonenal, HN) and nitrative stress (3-nitrotyrosine, NT) were compared in lungs of calves with caseonecrotic bronchopneumonia characteristic of M. bovis infection, with other forms of bronchopneumoniaor with non-inflamed lungs. HN and NT were identified in M. bovis pneumonia, mainly in foci of caseous necrosis. HN was not observed in inflamed non-necrotic tissue in lesions typical of pneumonic pasteurellosis. H2O2 production by M. bovis was identified, but the levels did not differ in isolates from calves with caseonecrotic bronchopneumonia compared with those with non-inflamed lungs or other forms of pneumonia. These findings provide evidence that oxidative and nitrative injury contribute to the formation of the caseonecrotic lesions that are characteristic of M. bovis pneumoniaand that production of H2O2 by M. bovis may contribute to this oxidative injury.

Pathogenic mechanisms implicated in the intravascular coagulation in the lungs of BVDV-infected calves challenged with BHV-1

Resistance to respiratory disease in cattle requires host defense mechanisms that protect against pathogens which have evolved sophisticated strategies to evade them, including an altered function of pulmonary macrophages (MΦs) or the induction of inflammatory responses that cause lung injury and sepsis. The aim of this study was to clarify the mechanisms responsible for vascular changes occurring in the lungs of calves infected with bovine viral diarrhea virus (BVDV) and challenged later with bovine herpesvirus type 1 (BHV-1), evaluating the role of MΦs in the development of pathological lesions in this organ. For this purpose, pulmonary lesions were compared between co-infected calves and healthy animals inoculated only with BHV-1 through immunohistochemical (MAC387, TNFα, IL-1α, iNOS, COX-2 and Factor-VIII) and ultrastructural studies. Both groups of calves presented important vascular alterations produced by fibrin microthrombi and platelet aggregations within the blood vessels. These findings were earlier and more severe in the co-infected group, indicating that the concomitance of BVDV and BHV-1 in the lungs disrupts the pulmonary homeostasis by facilitating the establishment of an inflammatory and procoagulant environment modulated by inflammatory mediators released by pulmonary MΦs. In this regard, the co-infected calves, in spite of presenting a greater number of IMΦs than single-infected group, show a significant decrease in iNOS expression coinciding with the presence of more coagulation lesions. Moreover, animals pre-inoculated with BVDV displayed an alteration in the response of pro-inflammatory cytokines (TNFα and IL-1), which play a key role in activating the immune response, as well as in the local cell-mediated response.

Neonatal Calf Infection with Respiratory Syncytial Virus: Drawing Parallels to the Disease in Human Infants

Respiratory syncytial virus (RSV) is the most common viral cause of childhood acute lower respiratory tract infections. It is estimated that RSV infections result in more than 100,000 deaths annually worldwide. Bovine RSV is a cause of enzootic pneumonia in young dairy calves and summer pneumonia in nursing beef calves. Furthermore, bovine RSV plays a significant role in bovine respiratory disease complex, the most prevalent cause of morbidity and mortality among feedlot cattle. Infection of calves with bovine RSV shares features in common with RSV infection in children, such as an age-dependent susceptibility. In addition, comparable microscopic lesions consisting of bronchiolar neutrophilic infiltrates, epithelial cell necrosis, and syncytial cell formation are observed. Further, our studies have shown an upregulation of pro-inflammatory mediators in RSV-infected calves, including IL-12p40 and CXCL8 (IL-8). This finding is consistent with increased levels of IL-8 observed in children with RSV bronchiolitis. Since rodents lack IL-8, neonatal calves can be useful for studies of IL-8 regulation in response to RSV infection. We have recently found that vitamin D in milk replacer diets can be manipulated to produce calves differing in circulating 25-hydroxyvitamin D3. The results to date indicate that although the vitamin D intracrine pathway is activated during RSV infection, pro-inflammatory mediators frequently inhibited by the vitamin D intacrine pathway in vitro are, in fact, upregulated or unaffected in lungs of infected calves. This review will summarize available data that provide parallels between bovine RSV infection in neonatal calves and human RSV in infants.

Bronchopneumonia in two dairy calves associated with Mannheimia species cluster V infection

The pathological, bacteriological, and molecular findings of two 3-week-old Holstein calves with bronchopneumonia are presented. Heavy pure growth of a Mannheimia species most closely aligned with the unnamed cluster V strains on the basis of 16S ribosomal RNA sequencing was detected in the lungs of both calves in association with Bovine respiratory syncytial virus infection. While Mannheimia species closely related to cluster V strains have occasionally been reported in association with pneumonia, meningitis, and abortion in cattle, the current report provides a description of the gross and histopathological lesions produced by a cluster V strain of Mannheimia species. Lesions in the lung were found to be typical of those described for Mannheimia haemolytica with the absence of areas of coagulation necrosis rimmed by leukocytes and more pronounced intra-alveolar hemorrhage. Lesions were linked to the presence of leukotoxin A based on phenotypic hemolysis and molecular demonstration of the leukotoxin A gene.

Maturation of mononuclear phagocytes in the lungs of young calves—In vitro study

The influences of age in calves’ immune system are described in their first phase of life. We hypothesized that variations that occur in the main mechanisms of lung innate response can help to identify periods of greater susceptibility to the respiratory diseases that affect calves in the first stage of their life. This study aimed to evaluate the innate immune system. Nine healthy calves were monitored for 3mo and 8 immunologic evaluations were performed. Bronchoalveolar lavage samples were recovered by bronchoscopy. The alveolar macrophages in samples were identified by protein expression of cluster of differentiation 14 (CD14) and underwent functional evaluation of phagocytosis (Staphylococcus aureus stained with propidium iodide and Escherichia coli). Data was assessed by one-way ANOVA (unstacked and parametric) and the Mann-Whitney test (nonparametric). Functional alterations in CD14-positive phagocytes were observed, with punctual higher intensity of phagocytosis in the third week and its decrease starting at 45 d of life. A gradual increase in phagocytosis rate was observed starting at this date. It is concluded that from 45 d of life on, alveolar macrophages have less phagocytic capacity but more cells perform this function. We suggest that this occurs because lung macrophages of calves start to maintain their immune response without passive immunity influence. Until 90 d of life, calves did not achieve the stability to conclude the maturation of local innate immune response.

Aberrant expression of imprinted genes and their regulatory network in cloned cattle

Domesticated animals cloned by somatic cell nuclear transfer (SCNT) generally have poor developmental competency, with many developmental abnormalities attributed to incomplete reprogramming of the nuclear genome and abnormal expression of genes important for regulation of growth and development. To investigate the molecular mechanism leading to the abnormalities of cloned animals, pathologic and histologic analyses were conducted on seven cloned cattle that were oversized at birth and had cardiac and pulmonary abnormalities. Quantitative reverse transcription (RT)-polymerase chain reaction (PCR) analysis of four imprinted genes IGF2, IGF2R, H19, and GRB10, as well as genes from related regulatory networks, were performed in liver, lung, kidney, and muscle to investigate disruption of expression. Expression of IGFBP2 was not detected in morphologically normal cloned cattle, but was detected in the liver, lung, kidney, and thymus of abnormal calves. Expression levels of IGF1 and imprinted genes IGF2 and H19 were substantially higher in these organs of abnormal cattle. In contrast, expression levels of GRB10, CTSD, and TRPV2 were substantially lower in abnormal cattle. Transcript abundance of IGFBP6 was higher in kidney, but lower in liver and lung. In conclusion, we inferred that altered expression of imprinted and related genes may be closely related to increased birth weight and pathologic changes in abnormal cloned cattle.

Expression of pentraxin 3 in the lungs of human, horse, cattle and pig

Bacterial lung diseases cause significant mortality and morbidity, causing enormous economic damage to the animal industry and cost billions of dollars in human health care. The innate immune system plays a critical role in the initiation of immune response in the lung. Pentraxin 3 (PTX3) is an acute phase protein that is produced at sites of inflammation by many cell types upon stimulation by proinflammatory cytokines, and agonists such as endotoxins. PTX3 recognizes and binds to many pathogens, activates the complement cascade, and has a role in the clearance of apoptotic and necrotic cells. Because there are very few data on the expression of PTX3 in the lungs, we examined PTX3 expression in normal and inflamed lungs of the horse, pig, human, and cattle using light and electron microscopic immunochemistry. We show PTX3 staining in bronchial epithelial cells and vascular endothelium in normal and inflamed lungs. Alveolar macrophages and inflammatory cells recruited into the lungs of bacteria‐infected animals stain intensely for PTX3. Immuno‐gold electron microscopy on horse, and calf lungs showed PTX3 in the nuclei, cytoplasm, and vesicular organelles of alveolar macrophages, endothelial cells and pulmonary intravascular macrophages. These data show PTX3 is expressed in normal lungs and its expression is increased in inflamed lungs.Grant Funding Source: Alberta Livestock and Meat Agency