FEELnc : a tool for long non-coding RNA annotation and its application to the dog transcriptome

Whole transcriptome sequencing (RNA-seq) has become a standard for cataloguing and monitoring RNA populations. One of the main bottlenecks, however, is to correctly identify the different classes of RNAs among the plethora of reconstructed transcripts, particularly those that will be translated (mRNAs) from the class of long non-coding RNAs (lncRNAs). Here, we present FEELnc (FlExible Extraction of LncRNAs), an alignment-free program that accurately annotates lncRNAs based on a Random Forest model trained with general features such as multi k-mer frequencies and relaxed open reading frames. Benchmarking versus five state-of-the-art tools shows that FEELnc achieves similar or better classification performance on GENCODE and NONCODE data sets. The program also provides specific modules that enable the user to fine-tune classification accuracy, to formalize the annotation of lncRNA classes and to identify lncRNAs even in the absence of a training set of non-coding RNAs. We used FEELnc on a real data set comprising 20 canine RNA-seq samples produced by the European LUPA consortium to substantially expand the canine genome annotation to include 10 374 novel lncRNAs and 58 640 mRNA transcripts. FEELnc moves beyond conventional coding potential classifiers by providing a standardized and complete solution for annotating lncRNAs and is freely available at https://github.com/tderrien/FEELnc.Peer reviewe

Genetic and Functional Analysis of Congenital Portosystemic Shunts in Dogs

The general aim of this thesis was to gain further insight into the pathogenesis of congenital portosystemic shunts (CPSS), elucidate mechanisms involved in the pathophysiology of CPSS, and explore predictors of recovery after surgical ligation of the shunt. For intrahepatic portosystemic shunts (IHPSS), the genetic background of the different anatomical locations is likely to be identical as they are observed within one litter of Irish Wolfhounds. In this thesis, the anatomical distribution of extrahepatic portosystemic shunt (EHPSS) subtypes was retrospectively analysed in 135 dogs with a single EHPSS. It was demonstrated that both EHPSS subtypes, portocaval and porto-azygous shunts, coexist in all studied dog breeds (apart from the Pug), suggesting that the two subtypes are variants of the same inherited disorder. These findings indicate that for studies into the genetic background of both IHPSS and EHPSS no distinction is required considering subtypes. Subsequently, a micro-array study was performed on liver tissue of 47 CPSS dogs to unravel genes involved in the pathogenesis of CPSS.By comparing expression levels in both shunt types with the levels observed in healthy liver tissue, a list of only 26 genes that were differentially expressed for one of the two types of shunt was found. Follow-up experiments including qPCR, immunohistochemistry and Western blots, resulted in two possible candidate genes, or their regulating pathways, causing CPSS in dogs; namely VCAM1 for EHPSS and WEE1 for IHPSS. Another aspect of this study was to gain further insight into the pathophysiology of one of the characteristics shared in hepatic biopsies of both IHPSS and EHPSS; hepatic lipid accumulation. A 12-fold increase of lipid content was observed in shunt biopsies when compared to normal liver tissue using Oil-red-O staining. Liver organoids from CPSS dogs and healthy controls were incubated with palmitic- and oleic-acid, and lipid accumulation was quantified. All organoids showed a similar increase in triacylglycerols after FFA enrichment indicating that lipid accumulation in the presence of portosystemic shunting is not caused by primary genetic defects. Taken together, lipid accumulation was observed in all shunt types and occurs secondary to portosystemic shunting. In the final part of the thesis, a predictive model was proposed, based on pre-operative plasma albumin and intra-operative mRNA expression levels of hepatic gene products (DHDH, ERLEC1, and LYSMD2) as predictors of recovery from portosystemic shunting after surgical ligation of the shunt. If surgery would be performed on dogs with a predicted probability >50% only, this model resulted in a sensitivity to correctly predicted recovery of 77% and a specificity of 90%. Furthermore, this model seems to have good discriminating abilities between recovered and non-recovered dogs. In conclusion, extensive genomic research based on the different aetiology and the shared physiology of EHPSS and IHPSS has resulted in novel insights in inherited congenital portosystemic shunts in dogs

Chronische Diarrhoe beim Hund

Die chronische Diarrhoe kann bei Hunden durch zahlreiche Erkrankungen bedingt sein. Die diagnostische Aufarbeitung dieser Patienten ist häufig langwierig und kostenintensiv, und die Prognose für chronische Enteropathien im Allgemeinen ist schwierig vorauszusehen. Da differenzialdiagnostisch aber auch gut kontrollierbare Erkrankungen in Betracht kommen, sollten die Besitzer immer zu einer Diagnostik ermutigt werden

Nonpharmacological Treatment Strategies for the Management of Canine Chronic Inflammatory Enteropathy-A Narrative Review

Chronic inflammatory enteropathy (CIE) refers to a heterogeneous group of idiopathic diseases of the dog characterised by persistent gastrointestinal (GI) clinical signs. If conventional dietary treatment alone would be unsuccessful, management of CIE is traditionally attained by the use of pharmaceuticals, such as antibiotics and immunosuppressive drugs. While being rather effective, however, these drugs are endowed with side effects, which may impact negatively on the animal's quality of life. Therefore, novel, safe and effective therapies for CIE are highly sought after. As gut microbiota imbalances are often associated with GI disorders, a compelling rationale exists for the use of nonpharmacological methods of microbial manipulation in CIE, such as faecal microbiota transplantation and administration of pre-, pro-, syn- and postbiotics. In addition to providing direct health benefits to the host via a gentle modulation of the intestinal microbiota composition and function, these treatments may also possess immunomodulatory and epithelial barrier-enhancing actions. Likewise, intestinal barrier integrity, along with mucosal inflammation, are deemed to be two chief therapeutic targets of mesenchymal stem cells and selected vegetable-derived bioactive compounds. Although pioneering studies have revealed encouraging findings regarding the use of novel treatment agents in CIE, a larger body of research is needed to address fully their mode of action, efficacy and safety

Nonpharmacological Treatment Strategies for the Management of Canine Chronic Inflammatory Enteropathy-A Narrative Review

Chronic inflammatory enteropathy (CIE) refers to a heterogeneous group of idiopathic diseases of the dog characterised by persistent gastrointestinal (GI) clinical signs. If conventional dietary treatment alone would be unsuccessful, management of CIE is traditionally attained by the use of pharmaceuticals, such as antibiotics and immunosuppressive drugs. While being rather effective, however, these drugs are endowed with side effects, which may impact negatively on the animal's quality of life. Therefore, novel, safe and effective therapies for CIE are highly sought after. As gut microbiota imbalances are often associated with GI disorders, a compelling rationale exists for the use of nonpharmacological methods of microbial manipulation in CIE, such as faecal microbiota transplantation and administration of pre-, pro-, syn- and postbiotics. In addition to providing direct health benefits to the host via a gentle modulation of the intestinal microbiota composition and function, these treatments may also possess immunomodulatory and epithelial barrier-enhancing actions. Likewise, intestinal barrier integrity, along with mucosal inflammation, are deemed to be two chief therapeutic targets of mesenchymal stem cells and selected vegetable-derived bioactive compounds. Although pioneering studies have revealed encouraging findings regarding the use of novel treatment agents in CIE, a larger body of research is needed to address fully their mode of action, efficacy and safety

Influence of macronutrient composition of commercial diets on circulating leptin and adiponectin concentrations in overweight dogs

Leptin and adiponectin play important roles in obesity-related inflammation and comorbidities. Previous research suggests that alterations in dietary macronutrient composition can modify circulating leptin and adiponectin concentrations in people, but limited research on this subject has been performed in dogs. This study investigated the effects of commercial high protein (HP), high fat (HF) and high carbohydrate medium protein (HCMP) diets on baseline (T-1 ) concentrations, post-prandial peak concentrations and total release in a ten-hour time span of leptin and adiponectin in dogs, when compared to a maintenance high carbohydrate low protein (HCLP) diet. Thirty-six overweight dogs were fed the HCLP diet in a one-week control period, after which the animals were assigned to one of three groups. In three four-week periods, each group was fed all test diets in a different sequence. At the last day of each period, blood was sampled at one hour before feeding (T-1 ) and at three (T3 ), six (T6 ) and nine (T9 ) hours after feeding. Feeding caused peak leptin concentrations at T6 and T9 (p < .001). No significant post-prandial change in adiponectin concentrations was found (p = .056). The HP diet resulted in lower leptin peak concentrations (p = .004) and AUCT-1-T9 (p = .01), but none of the diets influenced baseline leptin concentrations (p = .273). Baseline adiponectin concentrations were lower for the HF diet (p = .018) and HCMP (p < .001), and the HP, HF and HCMP AUCT-1-T9 (p < .001) were lower compared with the HCLP diet. Female dogs had lower adiponectin baseline concentrations (p = .041) and AUCT-1-T9 (p = .023) than male dogs. In conclusion, the HP diet was associated with the lowest post-prandial peak leptin concentration and the least decrease in adiponectin release, suggesting that a HP diet may improve immune-metabolic health and post-prandial satiety in overweight dogs

Redox unbalance in the hyperthyroid cat : a comparison with healthy and non-thyroidal diseased cats

BACKGROUND: Feline hyperthyroidism, the most common endocrinopathy in older cats, provides a spontaneous model for human thyrotoxicosis. Human thyrotoxicosis is associated with redox unbalance, which may result in organ damage. The redox status of hyperthyroid cats is largely unknown. The aims of the present study were to compare the redox status of cats with hyperthyroidism with that of healthy cats and cats with chronic non-thyroidal illness. RESULTS: Forty cats with untreated hyperthyroidism (group H), 45 chronically ill cats with non-thyroidal illness (group I), and 39 healthy cats (group C) were recruited for this observational cross-sectional study. All cats were screened for redox status markers. Determinable reactive oxygen metabolites (d-ROMs) were used as oxidative stress markers. Antioxidant status was determined using the OXY-Adsorbent test to quantify the plasma barrier to oxidation. The Oxidative Stress index (OSi) was calculated as the ratio of d-ROMs and OXY-Adsorbent test values. Data were compared by ANOVA with Tukey's multiple comparisons post-hoc test. The dROMs of group H (193 ± 47 CarrU) were significantly higher (p &lt; 0.001) than those of the healthy cats (103 ± 17 CarrU). The OXY-Adsorbent test results in group H (265 ± 68 μmol HClO/ml) were significantly lower than those in healthy cats (390 ± 83 μmol HClO/ml; p &lt; 0.01) and chronically ill cats (306 ± 45 μmol HClO/ml, p &lt; 0.05). Moreover, the Osi value in group H (0.8 ± 0.2 CarrU/μmol HClO/ml) was significantly higher (p &lt; 0.001) than that of the healthy cats (0.3 ± 0.1 CarrU/μmol HClO/ml). CONCLUSIONS: As described in humans with hyperthyroidism, feline hyperthyroidism is associated with redox unbalance. Free radical production is increased in hyperthyroid cats and their antioxidant depletion seems to be more severe than in cats with non-thyroidal illnesses. Our results support the rationale for a clinical trial investigating the potential positive effects of antioxidant supplementation to cats with hyperthyroidism

Intestinal Organoids—Current and Future Applications

Recent technical advances in the stem cell field have enabled the in vitro generation of complex structures resembling whole organs termed organoids. Most of these approaches employ culture systems that allow stem cell-derived or tissue progenitor cells to self-organize into three-dimensional (3D)-structures. Since organoids can be grown from different species (human, mouse, cat, dog), organs (intestine, kidney, brain, liver), and from patient-derived induced pluripotent stem cells, they create significant prospects for modelling development and diseases, for toxicology and drug discovery studies, and in the field of regenerative medicine. Here, we report on intestinal stem cells, organoid culture, organoid disease modeling, transplantation, specifically covering the current and future uses of this exciting new insight model to the field of veterinary medicine

Generation of Differentiating and Long-Living Intestinal Organoids Reflecting the Cellular Diversity of Canine Intestine

Functional intestinal disorders constitute major, potentially lethal health problems in humans. Consequently, research focuses on elucidating the underlying pathobiological mechanisms and establishing therapeutic strategies. In this context, intestinal organoids have emerged as a potent in vitro model as they faithfully recapitulate the structure and function of the intestinal segment they represent. Interestingly, human-like intestinal diseases also affect dogs, making canine intestinal organoids a promising tool for canine and comparative research. Therefore, we generated organoids from canine duodenum, jejunum and colon, and focused on simultaneous long-term expansion and cell differentiation to maximize applicability. Following their establishment, canine intestinal organoids were grown under various culture conditions and then analyzed with respect to cell viability/apoptosis and multi-lineage differentiation by transcription profiling, proliferation assay, cell staining, and transmission electron microscopy. Standard expansion medium supported long-term expansion of organoids irrespective of their origin, but inhibited cell differentiation. Conversely, transfer of organoids to differentiation medium promoted goblet cell and enteroendocrine cell development, but simultaneously induced apoptosis. Unimpeded stem cell renewal and concurrent differentiation was achieved by culturing organoids in the presence of tyrosine kinase ligands. Our findings unambiguously highlight the characteristic cellular diversity of canine duodenum, jejunum and colon as fundamental prerequisite for accurate in vitro modelling