A review: Pancreatic enzymes in the treatment of chronic pancreatic insufficiency in companion animals.

The purpose of this review was to analyze the scientific literature on exocrine pancreatic insufficiency (EPI) in dogs and cats and our own research on porcine model to compare animal- and microbial-derived enzymes in the treatment of animals with this disease. Clinical signs of EPI occur when more than 85% of the pancreatic parenchyma is non-functional. EPI can be a consequence of various diseases. The insufficient activity or deficiency of pancreatic enzymes leads to impaired digestion and absorption, and consequently, to malnutrition. The primary treatment for enzyme insufficiency is pancreatic enzyme replacement therapy (PERT). PERT in animals with EPI is a lifetime therapy. Most commercially available products are of animal origin (processed pancreata obtained from a slaughter house) and contain lipases, alpha-amylase, and proteases. Enzymes of microbial and plant origin seem to be a promising alternative to animal-derived enzymes, but to date there are no registered preparations containing all enzymes simultaneously for use in clinical practice to treat EPI. Results from some previous studies have highlighted the "extra-digestive" functions of pancreatic enzymes, as well as the actions of pancreatic-like microbial enzymes. For example, trypsin activates protease-activated receptor and provokes maturation of enterocytes and enterostatin inhibits fat absorption. It has been postulated that intrapancreatic amylase is the main component of the acini-islet-acinar axis-the reflex which down regulates insulin release, while gut and blood amylase exhibit anti-incretin actions "per se." Additionally, high but still physiological blood amylase activity coincide with physiological glucose homeostasis and a lack of obesity.

Diclofenac and dexamethasone modulate the effect of cannabidiol on the rat colon motility ex vivo.

Introduction: Due to the growing interest in the use of cannabinoids in supportive therapies, they are increasingly used together with anti-inflammatory drugs. Cannabinoids inhibit gastrointestinal motility, while steroidal and nonsteroidal anti-inflammatory drugs influence motility in other ways. The aim of the research was to study the interactions between cannabidiol (CBD) and these two classes of anti-inflammatory drugs in the context of gastrointestinal motility. Dexamethasone (DEX) was selected as a steroidal drug and diclofenac (DCF) as a nonsteroidal counterpart. Material and Methods: The experiments were performed on isolated rat colon strips in isometric conditions. The contractile response to acetylcholine (ACh) (1 µM) was measured with no substance applied as a control value and was measured after application of CBD (80 µM), DEX (100 µM), DCF (100 µM), or a combination of these substances. Results: Cannabidiol strongly inhibited intestinal motility mediated by ACh application, DCF inhibited it non-significantly, while DEX intensified it. When CBD was co-administered with DEX, the combination inhibited intestinal motility non-significantly relative to the ACh-only control. Co-administration of CBD with DCF inhibited motility more than when these substances were administered separately. Conclusion: Inhibition of the intestinal response to ACh is likely due to the synergistic effect of CBD and endogenous cannabinoids. Dexamethasone lessened the inhibitory effect of CBD, likely because of diminished availability of the arachidonic acid necessary for endogenous cannabinoid synthesis. However, diclofenac may increase endogenous cannabinoid synthesis, because of the greater availability of arachidonic acid caused by DCF blocking the cyclooxygenation pathway.

Cardiological Reference Intervals in Adult American Staffordshire Terrier Dogs.

The aim of this study was to determine reference intervals (RI) for echocardiography, electrocardiography (ECG), vertebral heart score (VHS) measurement, and arterial systolic blood pressure (SBP) in American Staffordshire Terrier dogs. The study population included 29 clinically healthy AST dogs of different ages, genders, and body weights. SBP measurement, ECG, thoracic radiography, and echocardiography were performed on each dog. Compared to RIs available for the general population of dogs, the duration of the P wave and QRS complex was longer and VHS was higher. Moreover, the left ventricular dimension in diastole and systole, left atrial dimension, and end point to septal separation values were higher, while the interventricular septum in diastole and systole and aortic root diameter were lower compared to general similar average body weight. The AST breed has a different heart shape, which in this breed is more rounded compared to other dog breeds, especially the deep chest. The specific body structure and the shape of the heart had an impact on the results of the cardiological examination.

Comparison of a Vegetable-Based Dental Chew to 2 Other Chews for Oral Health Prevention.

Giving dental chews to dogs is part of the passive homecare that helps prevent the formation of plaque and tartar. The objectives of these studies were to assess the effectiveness of a vegetable-based dental chew (VF) to maintain oral health, and to compare it to 2 different reference chews (RC) with a proven effectiveness. The first study was conducted on 45 small dogs (<10 kg) and the second on 60 larger dogs (15-30 kg) who were randomly assigned to 3 different groups. During 30 days, one group received no chew (control) while the second and third group received either one RC (RC1 or RC2) or one VF per day. All dogs had their teeth scaled on Day 0. On Day 30, scores were given for plaque and calculus. Gingival parameters were also assessed. Statistical analysis (analysis of variance and Tukey tests ± Bonferroni's adjustment) were performed to compare groups with α set at .05 for significance.The 3 types of chews were found to be efficacious to reduce plaque and calculus formation and the gingival bleeding compared to control (P < .05). There was no significant difference between RCs and VF in both trials except for the gingival bleeding parameters which showed a greater improvement with VF. Therefore, daily administration of the VF is effective to reduce plaque and calculus formation and gingival bleeding and has a better efficacy on gingival bleeding than the other reference products tested. It can therefore be used with confidence at home for preventative dental care.

Influence of Dietary Supplementation With a Powder Containing A.N. ProDen™ (Ascophyllum Nodosum) Algae on Dog Saliva Metabolome.

The objective of this placebo-controlled, double-blind, randomized study (designed according to evidence-based medicine standards) was to determine the effect of 30-day administration of powdered brown algae, Ascophyllum nodosum (ProDen PlaqueOff, SwedenCare AB, Sweden), on saliva metabolomes in dogs. Sixty client-owned dogs underwent professional dental cleaning and were randomly subdivided into two groups receiving daily powdered brown algae A. nodosum, or a placebo (microcrystalline cellulose in powder), adjusted to their bodyweight. After a comprehensive oral health assessment and professional dental cleaning, which were both performed under general anesthesia, clinical assessments for gingivitis, plaque, and calculus were conducted. Saliva samples were collected at Day 0 and Day 30 of supplementation. Whole saliva is a mixed fluid that is derived predominantly from the major salivary glands but it also contains numerous other constituents. Additionally, its composition varies on whether salivary secretion is basal or stimulated. Authors put efforts to avoid contamination of saliva by other constituents and character of saliva was basal. Quadrupole time-of-flight (QTOF) mass spectrometer was used to conduct analysis of the saliva samples. Metabolomic analyses identified clear changes after 30 days of supplementation, and the direction of these changes was completely different than in dogs that received a placebo treatment during the same period. The positive clinical effect of 30 days of A. nodosum supplementation on oral health status in dogs described in previous publication combined with the absence of some metabolites in the saliva of dogs on day 30 of supplementation suggest that brown algae inhibit or turn off some pathways that could enhance plaque or calculus development. The exact mechanism of A. nodosum is still unclear and warrants further study.

Impact of selected individual dog traits on echocardiographic parameters obtained in 1-dimensional (M-mode) and 2-dimensional (2D) imaging.

The popularity and availability of echocardiography in veterinary practice for companion animals have substantially increased in recent years. The results obtained during the procedure are compared to reference values established for the general dog population or to standards developed for a specific dog breed. The aim of this study was to determine whether individual dog traits, such as body weight, chest structure, and level of physical activity and performance, affect the reference values for echocardiographic parameters. Published reference values for echocardiographic examination parameters for 32 dog breeds were analyzed and the relationship between individual echocardiographic parameters and body weight, chest structure, and level of physical activity and performance was then statistically analyzed. It was found that echocardiographic parameters are affected by the dog's weight and physical activity. There was no significant relationship between heart size and chest structure. The great variety of dog breeds means that echocardiographic findings should be individually interpreted rather than establishing reference ranges for each breed in population studies. This will allow for a more accurate interpretation of the results obtained in the echocardiographic examination and consequently lead to earlier diagnosis of changes in myocardial morphology.

La popularité et la disponibilité de l'échocardiographie dans la pratique vétérinaire pour les animaux de compagnie ont considérablement augmenté ces dernières années. Les résultats obtenus au cours de la procédure sont comparés à des valeurs de référence établies pour la population canine générale ou à des normes développées pour une race de chien spécifique. Le but de cette étude était de déterminer si les caractéristiques individuelles du chien, tels que le poids corporel, la structure de la poitrine et le niveau d'activité physique et de performance, affectent les valeurs de référence des paramètres échocardiographiques. Les valeurs de référence publiées pour les paramètres d'examen échocardiographique pour 32 races de chiens ont été analysées et la relation entre les paramètres échocardiographiques individuels et le poids corporel, la structure thoracique et le niveau d'activité physique et de performance a ensuite été analysée statistiquement. Il a été constaté que les paramètres échocardiographiques sont affectés par le poids et l'activité physique du chien. Il n'y avait pas de relation significative entre la taille du coeur et la structure thoracique. La grande variété des races de chiens signifie que les résultats échocardiographiques devraient être interprétés individuellement plutôt que d'établir des plages de référence pour chaque race dans des études de population. Cela permettra une interprétation plus précise des résultats obtenus à l'examen échocardiographique et conduira par conséquent à un diagnostic plus précoce des modifications de la morphologie myocardique.(Traduit par Docteur Serge Messier).

Small intestinal development in suckling rats after enteral obestatin administration.

This study investigated the effect of enteral administration of obestatin on the development of small intestine, as well as oxidative stress markers and trancriptomic profile of gastrointestinal genes. Suckling rats were assigned to 3 groups treated with: C-saline solution; OL-obestatin (125 nmol/kg BW); OH-obestatin (250 nmol/kg BW) administered twice daily, from the 14th to the 21st day of life. Enteral administration of obestatin in both studied doses had no effect neither on the body weight of animals nor the BMI calculated in the day of euthanasia. Compared to the control group (C), treatment with obestatin resulted in significant changes in the histometry of the small intestinal wall as well as intestinal epithelial cell remodeling. The observed changes and their possible implications for intestinal development were dependent on the dosage of peptide. The enteral administration of high dose (OH) of obestatin significantly decreased its expression in the stomach and increased markers of oxidative stress. The gene profile revealed MAPK3 (mitogen-activated protein kinase-3) as the key regulator gene for obestatin action in the gastrointestinal track. In conclusion, we have showed that enteral administration of obestatin influences the gut mucosa remodeling. It is also suggested that the administration of high dose (OH) has inhibitory effect on the intestinal maturation of suckling rats.

Effects of Edible Treats Containing Ascophyllum nodosum on the Oral Health of Dogs: A Double-Blind, Randomized, Placebo-Controlled Single-Center Study.

The objective of this placebo-controlled, double-blind, randomized study (designed according to evidence based medicine standards) was to determine the effect of 90-day administration of edible treats containing the brown algae, Ascophyllum nodosum, on plaque and dental calculus accumulation on the teeth of dogs, as well as on other parameters characterizing canine oral health status, including: plaque index (PI), calculus index (CI), oral health index (OHI), gingival bleeding index (GBI), and volatile sulfur compound (VSC) concentration. Sixty client-owned dogs, including Japanese chin, miniature Schnauzer, Chihuahua, Pomeranian, and West Highland White Terrier (WHWT) breeds, underwent professional dental cleaning and were randomly subdivided into two groups receiving daily edible treats containing the brown algae A. nodosum, or placebo, adjusted to their bodyweight. After a comprehensive oral health assessment, including a professional dental cleaning, which were both performed under general anesthesia, clinical assessments of PI, CI, OHI, GBI, and VSC concentration were performed under sedation after 30, 60, and 90 days of treatment. Oral administration of edible treats containing A. nodosum significantly improved PI, CI, and VSC concentration, compared with the placebo-treated group. The consumption of edible treats containing A. nodosum efficiently decreased plaque and calculus accumulation in the investigated dogs. Dogs treated with A. nodosum also exhibited significantly lower concentrations of VSC and better oral health status (e.g., OHI and GBI) than those in the placebo-control group.

Mitral Regurgitation in Dachshund Dogs without Heart Murmurs.

INTRODUCTION: Older small breed dogs are considered at risk for heart failure secondary to chronic mitral valve disease. However, few data are available on the onset of this disease in such dogs. This study was performed to determine if auscultation alone can be used to eliminate clinically relevant mitral valve regurgitation seen in echocardiography in Dachshund dogs. MATERIAL AND METHODS: Clinical and echocardiographic data were obtained from 107 dogs without heart murmurs. RESULTS: The study revealed that 63.6% of the dogs had mitral regurgitation. Numbers increased with age and a larger percentage of male Dachshunds were affected than female Dachshunds. Mitral valve prolapse and thickening were mild, and the regurgitant area inextensive in most dogs. CONCLUSIONS: The study shows that mitral valve regurgitation is prevalent (63.6%) in Dachshunds without heart murmurs. Typical lesions often become apparent during echocardiographic examinations in dogs under 5 years of age.

Lymphocytic, cytokine and transcriptomic profiles in peripheral blood of dogs with atopic dermatitis.

BACKGROUND: Canine atopic dermatitis (cAD) is a common chronic and pruritic skin disease in dogs. The development of cAD involves complex interactions between environmental antigens, genetic predisposition and a number of disparate cell types. The aim of the present study was to perform comprehensive analyses of peripheral blood of AD dogs in relation to healthy subjects in order to determine the changes which would be characteristic for cAD. RESULTS: The number of cells in specific subpopulations of lymphocytes was analyzed by flow cytometry, concentration of chosen pro- and anti-inflammatory cytokines (IL-4, IL-10, IL-13, TNF-α, TGF-ß1) was determined by ELISA; and microarray analysis was performed on RNA samples isolated from peripheral blood nuclear cells of AD and healthy dogs. The number of Th cells (CD3(+)CD4(+)) in AD and healthy dogs was similar, whereas the percentage of Tc (CD3(+)CD8(+)) and Treg (CD4(+)CD25(+) Foxp3(+)) cells increased significantly in AD dogs. Increased concentrations of IL-13 and TNF-α, and decreased levels of IL-10 and TGF-ß1 was observed in AD dogs. The level of IL-4 was similar in both groups of animals. Results of the microarray experiment revealed differentially expressed genes involved in transcriptional regulation (e.g., transcription factors: SMAD2, RORA) or signal transduction pathways (e.g., VEGF, SHB21, PROC) taking part in T lymphocytes lineages differentiation and cytokines synthesis. CONCLUSIONS: Results obtained indicate that CD8(+) T cells, beside CD4(+) T lymphocytes, contribute to the development of the allergic response. Increased IL-13 concentration in AD dogs suggests that this cytokine may play more important role than IL-4 in mediating changes induced by allergic inflammation. Furthermore, observed increase in Treg cells in parallel with high concentrations of TNF-α and low levels of IL-10 and TGF-ß1 in the peripheral blood of AD dogs point at the functional insufficiency of Treg cells in patients with AD.

Plasma miRNAs as potential biomarkers of chronic degenerative valvular disease in Dachshunds.

BACKGROUND: Endocardiosis is the most common heart disease in Dachshunds and is therefore an important cause of cardiac morbidity and death. In recent years we have observed an increasing interest in the development of new genetic and genomic markers of heart disease. The discovery of miRNAs circulating in biofluids such as plasma or serum aroused researchers' interest in using them as potential biomarkers. In the present study we analysed the expression of 9 miRNAs described in literature as being involved in cardiovascular pathology in the plasma of dogs suffering from endocardiosis. RESULTS: Expression analysis using the Real-time PCR method revealed that two out of nine miRNAs were significantly downregulated: the expression of miR-30b differed between ACVIM stage B and stage A (control) dogs; the expression of mi-133b differed ACVIM stage C and stage A dogs. 5 miRNAs (miR-125, miR-126, miR-21, miR-29b and miR-30b) showed a trend of downregulation in the ACVIM C group. Levels of miR-423 were the same in healthy and diseased dogs. Expression of miR-208a and 208b was not detected. CONCLUSIONS: miR-30b could be a potential biomarker of ACVIM stage B heart failure in Dachshunds with endocardiosis and miR-133b could be a potential biomarker of ACVIM stage C. The lack of expression or lack of significant changes in expression in 7 miRNAs which are potential biomarkers of heart diseases in humans proves that findings from human medicine are not always directly reflected in veterinary medicine.

Biologically active substances-enriched diet regulates gonadotrope cell activation pathway in liver of adult and old rats.

According to the Hippocrates' theorem "Let food be your medicine and medicine be your food", dietary interventions may induce changes in the metabolic and inflammatory state by modulating the expression of important genes involved in the chronic disorders. The aim of the present study was to evaluate the influence of long-term (14 months) use of biologically active substances-enriched diet (BASE-diet) on transcriptomic profile of rats' liver. The experiment was conducted on 36 Sprague-Dawley rats divided into two experimental groups (fed with control or BASE-diet, both n = 18). Control diet was a semi-synthetic diet formulated according to the nutritional requirements for laboratory animals. The BASE-diet was enriched with a mixture of polyphenolic compounds, ß-carotene, probiotics, and n-3 and n-6 polyunsaturated fatty acids. In total, n = 3,017 differentially expressed (DE) genes were identified, including n = 218 DE genes between control and BASE groups after 3 months of feeding and n = 1,262 after 14 months. BASE-diet influenced the expression of genes involved particularly in the gonadotrope cell activation pathway and guanylate cyclase pathway, as well as in mast cell activation, gap junction regulation, melanogenesis and apoptosis. Especially genes involved in regulation of GnRH were strongly affected by BASE-diet. This effect was stronger with the age of animals and the length of diet use. It may suggest a link between the diet, reproductive system function and aging.

The transcriptomic profile of peripheral blood nuclear cells in dogs with heart failure.

BACKGROUND: In recent years advances have been made in the investigative methods of molecular background of canine heart disease. Studies have been conducted to identify specific genes which, when pathologically expressed, could lead to the dysfunction of the canine heart or are correlated with heart failure. For this purpose genome wide microarray experiments on tissues from failing hearts have been performed. In the presented study a whole genome microarray analysis was used for the first time to describe the transcription profile of peripheral blood nuclear cells in dogs with heart failure. Dogs with recognized heart disease were classified according the ISACHC (International Small Animal Cardiac Health Council) classification scheme as class 1 (asymptomatic)--13 dogs, class 2 (mild to moderate heart failure)--13 dogs and class 3 (severe heart failure)--12 dogs. The control group consisted of 14 healthy dogs. The clinical picture of the animals included: animal history, clinical examination, echocardiographic examination and where applicable electrocardiographic and radiographic examinations. RESULTS: In the present study we identified four sets of differentially expressed genes, namely heart-failure-specific genes and ISACHC1-specific genes, ISACHC2-sepcific genes and ISACHC-3 specific genes. The most important set consisted of genes differentially expressed in all dogs with heart failure, despite the ISACHC stage. We identified 71 heart-failure-specific genes which were involved in two statistically significant receptor signalling pathways, namely angiotensinR - > CREB/ELK-SRF/TP53 signalling and ephrinR - > actin signalling. The number of ISACHC1-specific genes was 83; ISACHC2-specific genes--1247 and ISACHC3-specific--200. CONCLUSIONS: The transcriptomic profile of peripheral blood nuclear cells in dogs with heart failure seems to reflect the presence of clinical signs of the disease in patients based on the observation that the largest number of differentially expressed genes was identified in ISACHC 2 group of patients. This group consists of dogs just starting to show clinical signs of heart failure. A set of genes was also found to have changed expression in all dogs with heart failure, despite the stage of the disease.

Relationship between clinical data and gene expression in the HER2/ErbB2-dependent signaling pathway in patients with acute heart failure.

Anticancer treatment with the human epidermal growth factor receptor (HER) 2 inhibitors can lead to significant myocardial dysfunction. The primary aim of the study was to estimate the possible association between gene expression in the ErbB signaling pathway and selected clinical event data in patients with acute heart failure. Twenty-four patients (19 males), aged 68.6 ± 12.3 years, were diagnosed and treated due to acute heart failure. The globaltest method was used for the correlation between blood nuclear cells' gene expression in the ErbB pathway (KEGG pathway id 04012) and important clinical data. Decreased expression of ErbB2/HER2 was found to be associated with the release of troponin and the need for inotropic support, whereas decreased neuregulin 1 (NRG1) expression was found to be associated with a decrease of ejection fraction below 40 % (globaltest p-value < 0.05). In summary, the ErbB signaling pathway and, especially, HER2/ErbB2 receptor expression are significantly associated with some of the recognized, clinically significant parameters of patients with acute heart failure. Evaluation of the molecular function of the HER2 receptor may be essential for the prognosis and targeted therapy of heart diseases.

A retrospective study of clinical signs and epidemiology of chronic valve disease in a group of 207 Dachshunds in Poland.

BACKGROUND: Chronic mitral valve disease is frequently seen in the Dachshund. Dachshunds (n=207) made up 11.73% of the dogs admitted to the Cardiology Service at the Small Animal Clinic, Warsaw University of Life Sciences, Poland (first visits only). RESULTS: Of these, 35 dogs had no clinically detectable heart disease while 172 had chronic valve disease with the mitral valve affected most often (130 dogs), both mitral and tricuspid valves infrequently (39 dogs) and rarely the tricuspid valve (3 dogs). Males were affected more frequently than females and the average age of dogs with chronic valve disease was 11.9 years for females and 11.3 years for males. A majority of the diseased Dachshunds were classified as ISACHC 2 (79), followed by ISACHC 1 (60). Most frequent clinical signs noted by owners included coughing, exercise intolerance, dyspnea and tachypnea. Heart murmurs were generally louder with increased disease severity; however there were 20 dogs in the ISACHC 1 group with no audible heart murmurs. The most frequent electrocardiographic abnormalities included an increased P wave and QRS complex duration, increased R wave amplitude and tachycardia. With increased disease severity, echocardiography revealed an increase in heart size. A higher ISACHC class was related to increased heart size (based on echocardiography) and increased percentage of patients exhibiting enlargement of both left atrium and left ventricle (based on radiography). CONCLUSIONS: The Dachshund is often affected by chronic mitral valvular disease with a late onset of associated clinical signs and few cardiac complications.

White blood cell transcriptome correlates with renal function in acute heart failure.

It is notoriously difficult to classify patients with acute heart failure (AHF) because of variations in clinical presentation, different etiologies, the impact of comorbidities, and variable prognoses. In this study, we used DNA whole-genome microarrays to classify 24 patients with AHF based on the transcriptome of their peripheral blood nuclear cells. The main purpose was to verify whether any transcriptomic sub-clusters had clinical correlations. We identified two distinct groups of transcriptomic profiles that correlated with normal (1.125 mg/dL) and increased (1.783 mg/dL) mean blood creatinine concentrations. These two subgroups of patients (n = 12) differed in the expression of more than 6000 genes and 108 signaling pathways. The most significant regulated signaling pathway was the aldosterone-regulated sodium reabsorption pathway and the most significant regulated genes included the angiotensin-converting enzyme gene. This suggests that kidney impairment in patients with AHF is related to dysregulation of the renin-angiotensin-aldosterone system. The interesting findings of our study were the significant differences in expression of genes belonging to the aldosterone-regulated signaling pathway: Na+/K+ transporting ATPase and NEDD4L (neuronal precursor cell expressed developmentally down-regulated 4-like) between patients with and without renal dysfunction. Future studies of blood-cell transcriptomic profiles in patients with AHF will provide further insights into the molecular pathogenesis of this cardiorenal disorder.

The gene expression profiles of canine mammary cancer cells grown with carcinoma-associated fibroblasts (CAFs) as a co-culture in vitro.

BACKGROUND: It is supposed that fibroblasts present in tumour microenvironment increase cancer invasiveness and its ability to metastasize but the mechanisms have not been clearly defined yet. Thus, the current study was designed to assess changes in gene expression in five various cancer cell lines grown as a co-culture with the carcinoma-associated fibroblasts (CAFs) in vitro. RESULTS: A carcinoma-associated fibroblast cell line was isolated from a canine mammary cancer. Then, a co-culture of cancer cells with the CAFs was established and maintained for 72 hrs. Having sorted the cells, a global gene expression in cancer cells using DNA microarrays was examined. The analysis revealed an up-regulation of 100 genes and a down-regulation of 106 genes in the cancer cells grown as a co-culture with the CAFs in comparison to control conditions. The PANTHER binomial statistics tool was applied to determine statistically over-manifested pathways (p < 0.05). Bulk of the up-regulated genes are involved in the adhesion, the angiogenesis, the epithelial-mesenchymal transition (EMT) and generally take part in the developmental processes. These results were further confirmed using real-time qPCR. Moreover, a wound-healing assay and growth characteristics on Matrigel matrix showed that CAFs increase cancer cell migration and matrix invasion. CONCLUSION: The results of the current study showed that the co-culturing of cancer cells and the CAFs caused significant changes to the cancer gene expression. The presence of the CAFs in a microenvironment of cancer cells promotes adhesion, angiogenesis and EMT.

Genomic and genetic aspects of heart failure in dogs - a review.

The most common causes of heart failure in dogs are valvular disease, predominantly endocardiosis, and myocardial disease, predominantly dilated cardiomyopathy. They are related to changes in the expression of several genes in the heart muscle and in peripheral blood nuclear cells which could be considered as prognostic or diagnostic markers of heart disease in dogs. Since many human genetic markers of heart failure have turned out to be useless in dogs, the screening for genomic markers of canine heart failure could give more insight into the molecular pathology of these diseases and aid the development of new treatment strategies.

Gene expression profiling in skeletal muscle of Holstein-Friesian bulls with single-nucleotide polymorphism in the myostatin gene 5'-flanking region.

Myostatin (GDF-8) is a key protein responsible for skeletal muscle growth and development, thus mutations in the mstn gene can have major economic and breeding consequences. The aim of the present study was to investigate myostatin gene expression and transcriptional profile in skeletal muscle of Holstein-Friesian (Black-and-White) bulls carrying a polymorphism in the 5'-flanking region of the mstn gene (G/C transversion at position -7828). Real-time qRT-PCR and cDNA microarray revealed significantly lower mstn expression in muscle of bulls with the CC genotype, as compared to GG and GC genotypes. The direct comparison of skeletal muscle transcriptional profiles between the CC genotype and GG and GC genotypes resulted in identification of genes, of which at least some can be putative targets for myostatin. Using cDNA microarray, we identified 43 common genes (including mstn) with significantly different expression in skeletal muscle of bulls with the CC genotype, as compared to GG and GC genotypes, 15 of which were upregulated and 28 were downregulated in the CC genotype. Classification of molecular function of differentially expressed genes revealed the highest number of genes involved in the expression of cytoskeleton proteins (9), extracellular matrix proteins (4), nucleic acid-binding proteins (4), calcium-binding proteins (4), and transcription factors (4). The biological functions of the largest number of genes involved: protein metabolism and modification (10), signal transduction (10), cell structure (8), and developmental processes (8). The main identified signaling pathways were: Wnt (4), chemokines and cytokines (4), integrin (4), nicotine receptor for acetylocholine (3), TGF-beta (2), and cytoskeleton regulation by Rho GTPase (2). We identified previously unrecognized putatively myostatin-dependent genes, encoding transcription factors (EGR1, Nf1b, ILF1), components of the proteasomal complex (PSMB7, PSMD13) and proteins with some other molecular function in skeletal muscle (ITGB1BP3, Pla2g1b, ISYNA1, TNFAIP6, MST1, TNNT1, CALB3, CACYBP, and CTNNA1).

Delineation of signalling pathway leading to antioxidant-dependent inhibition of dexamethasone-mediated muscle cell death.

The molecular mechanism of the cell death-promoting effect of dexamethasone (Dex) was studied during myogenesis (10 days) in L6 muscle cells by making use of several indices such as cell viability (protein synthesis, mitochondrial respiration), mortality (DNA fragmentation, chromatin condensation, structural modifications) and immunocytochemical studies [hydrogen peroxide, m-calpain (calpain 2)]. Dex initially (2 nM) stimulated protein synthesis (P < 0.001), but a further increase (20 nM) did not stimulate, whereas a higher dose (200 nM) inhibited formation of cellular proteins (P < 0.001). The latter, apparently, resulted from impaired cell viability (P < 0.001). From the day 4, structural changes featuring cell death were observed. Antioxidants [sodium ascorbate (ASC), catalase (CAT) or N-acetyl-L-cysteine (NAC)] as well as the inhibition of transcription and translation by actinomycin D abrogated Dex-induced cell death (P < 0.001). Using a fluorescent probe (DCFH-DA) we directly corroborated the working hypothesis of the mediating role of H2O2 in the reduction of cell viability by the excess of glucocorticoids. We also found that tPKC, PLCgamma, PLA2 were required to induce Dex-dependent cell death since inactivation of tPKC by H7 completely abolished the cytotoxic effect of Dex, while the blockade of PLCgamma and PLA2 by U 73122 partially abolished the effect. Cell death was triggered by Ca2+ influx necessary to activate m-calpain since it was reversed by the calcium chelator EGTA or m-calpain inhibitor ALLN but not EDTA nor ALLM. However, cell viability impaired by Ca2+ ionophore A 23187 (P < 0.001) was neither reversed by EGTA, nor EDTA, nor caspase-3 blocker--Ac DEVD CHO, nor ALLN, nor antioxidants--ASC, NAC, CAT. Specific caspase-3 inhibitor Ac DEVD CHO also did not rescue cells from Dex-induced cell death (P < 0.001), in contrast to m-calpain inhibitor--ALLN. Taken together, these findings suggest that reactive oxygen species inhibit protein synthesis and amplify m-calpain-dependent proteolysis. The events that led to the death of L6 muscle cells most likely resulted from Dex-mediated repression of antioxidative defences on the genomic level.