Blood Vessel Topography of the Feet in Selected Species of Birds of Prey and Owls.

Birds of prey and owls are susceptible to diseases of and traumatic injuries to their feet, which regularly require surgical intervention. A precise knowledge of the blood vessel topography is essential for a targeted therapy. Therefore, the metatarsal and digital vasculature was examined in eight species of birds of prey and owls. The study included contrast micro-computed tomography scans and anatomical dissections after intravascular injection of colored latex. In all examined species, the dorsal metatarsal arteries provided the main supply to the foot and their branching pattern and number differed between species. They continued distally as digital arteries. All examined species showed a basic pattern of four collaterally located digital blood vessels per toe: a prominent artery and small vein on one side and a small artery and prominent vein on the other side. Digital veins united to form common digital veins, most of which joined into a superficial, medially located metatarsal vein. This vein provided the main drainage of the foot. The detailed visualization of the topography of pedal blood vessels will help veterinary surgeons during surgical procedures. In addition, differences in the plantar arterial arch between hawks and falcons were discussed regarding their possible influence on the prevalence of pododermatitis (bumblefoot).

Historic Horse Family Displaying Malformations of the Cervicothoracic Junction and Their Connection to Modern German Warmblood Horses.

Malformations of the equine cervicothoracic junction affect the C6 and C7 cervical vertebrae, the T1 thoracic vertebra and in variable extent the first and second sternal ribs. To date, the clinical impact of this malformation, its prevalence and mode of inheritance in equine populations are not yet determined. We examined five skeletons for signs of malformation of the cervicothoracic junction, including three skeletons from widely used Thoroughbred stallions affected with the malformation and two skeletons serving as a comparison. The three affected historical horses were the Thoroughbred stallions Der Loewe XX, Birkhahn XX and their common great grandsire Dark Ronald XX. Malformations of C6 and C7 showed a large variation between the three stallions, as Dark Ronald XX, Der Loewe XX and Birkhahn XX were affected uni-laterally at C6 and C7, uni-laterally at C6 and bi-laterally at C6 and C7, respectively, with varying grades. In order to evaluate whether or not these malformations are incidental, we took a random sample of 20 living German Warmblood horses, which are distant descendants of these stallions. This sample consisted of ten controls and ten horses with malformations of C6/C7. Blood proportions of the historical sires in the modern Warmblood horses ranged from 0.10 to 6.25%. The contribution to inbreeding in each individual horse of our selected horse group by those sires was expressed as a percentage of the total inbreeding coefficient and ranged from 0.01 to 17.96%, demonstrating their influence on the modern Warmblood. In the present study, we were able to describe the variability of the malformation of C6/C7 within a horse family including historic and modern horses. Additionally, we detected variations appearing in connection with malformations of the cervicothoracic junction that have not been described in the literature yet. This is the first time that the malformations of C6 and C7 have been described within a familial context, providing hints on inheritance in Thoroughbreds and Warmbloods. It is worthwhile to carry out further studies in a larger setting to gain more comprehensive insights into the inheritance of the malformation and the role of important ancestors.

Alpha2 Antagonist Vatinoxan Does Not Abolish the Preconditioning Effect of Dexmedetomidine on Experimental Ischaemia-Reperfusion Injury in the Equine Small Intestine.

Pharmacological preconditioning with dexmedetomidine has been shown to ameliorate intestinal ischaemia reperfusion injury in different species, including horses. However, it remains unknown if this effect is related to alpha2 adrenoreceptor activity. Therefore, the aim of this study was to determine the effect of dexmedetomidine preconditioning with and without the administration of the peripheral alpha2 antagonist vatinoxan. This prospective randomized experimental trial included 12 horses equally divided between two treatment groups. Horses in group Dex received a bolus of dexmedetomidine followed by a continuous rate infusion (CRI), while group DexV additionally received vatinoxan as bolus and CRI. A median laparotomy was performed under general anaesthesia, and jejunal ischaemia was applied for 90 min, followed by 30 min of reperfusion. Mucosal damage was evaluated in full thickness biopsies by use of a semiquantitative mucosal injury score and by determining the apoptotic cell counts with immunohistochemical staining for cleaved caspase-3 and TUNEL. Comparisons between the groups and time points were performed using non-parametric tests (p < 0.05). During pre-ischaemia and ischaemia, no differences could be found in mucosal injury between the groups. After reperfusion, group DexV showed lower mucosal injury scores compared to group Dex. The apoptotic cell counts did not differ between the groups. In conclusion, antagonizing the peripheral alpha2 adrenoreceptors did not negatively affect dexmedetomidine preconditioning.

ß-Hydroxybutyrate Effects on Bovine Caruncular Epithelial Cells: A Model for Investigating the Peri-Implantation Period Disruption in Ketotic Dairy Cows.

Ketosis is a metabolic disorder arising from a negative energy balance (NEB). It is characterized by high ß-Hydroxybutyrate (BHBA) blood levels and associated with reduced fertility in dairy cows. To investigate the impact of BHBA on bovine caruncular epithelial cells (BCEC) in vitro, these cells were stimulated with different concentrations of BHBA. Cell metabolism and motility were examined using an MTT assay and Live-cell imaging. RT-qPCR was used to examine mRNA expressions of TNF, IL6, RELA, prostaglandin E2 synthase (PTGES2) and receptor (PTGER2) as well as integrin subunits ITGAV, ITGA6, ITGB1 and ITGB3. Stimulation with 1.8 and 2.4 mM of BHBA negatively affected cell metabolism and motility. TNF showed increased mRNA expression related to rising BHBA concentrations. IL6, RELA, ITGAV, ITGA6, ITGB1 and ITGB3 as well as PTGER2 showed no changes in mRNA expression. Stimulation with 0.6 and 1.2 mM of BHBA significantly increased the mRNA expression of PTGES2. This does not indicate a negative effect on reproductive performance because low BHBA concentrations are found in steady-state conditions. However, the results of the study show negative effects of high BHBA concentrations on the function of BCECs as well as an inflammatory response. This could negatively affect the feto-maternal communication during the peri-implantation period in ketotic dairy cows.

ß-Hydroxybutyrate affects cell physiological parameters, inflammatory markers and hormone receptor expression in bovine endometrial gland cells in vitro.

INTRODUCTION: After calving, dairy cows are commonly affected by negative energy balance (NEB), indicated by high ß-Hydroxybutyrate (BHBA) blood levels. These are associated with subfertility frequently related to uterine inflammation. Since this could compromise functionality of endometrial glands that are essential for proper embryo implantation in sheep, we investigated effects of BHBA on bovine endometrial gland cells (BEGC) in vitro. MATERIAL AND METHODS: BEGC were stimulated with different concentrations of BHBA over different periods. Cell metabolism and motility were examined by MTT-assay and Live-cell-imaging. The mRNA expression of the receptors for estrogen (ESR1, ESR2), progesterone (PR) and IFNτ (IFNAR1, IFNAR2), and the inflammatory cytokines TNFα and IL-6 was determined by RT-qPCR. Protein expression for PR and ESR1 was analyzed by semiquantitative Western Blot. RESULTS: BEGC metabolism was significantly decreased after stimulation with 1.2, 1.8 and 2.4 mM BHBA over 24 and 36 h. Cell motility was significantly reduced by 1.8 and 2.4 mM BHBA already after 11 h. After 24 h stimulation, the ESR1 mRNA expression was significantly increased in BEGC stimulated with 0.6 mM BHBA. PR and TNFα mRNA expressions were increased in cells stimulated with 2.4 mM BHBA. Protein expression of ESR1 and PR was not altered. DISCUSSION: Treatment with BHBA leads to restriction of BEGC metabolism and motility, and increased expression of TNFα, ESR1 and PR in vitro. This could explain how increased BHBA blood levels might compromise functionality of uterine glands in vivo and thus could contribute to compromised reproductive success of cows suffering from NEB.

Obesity impacts hypoxia adaptation of the lung.

Obesity is mostly associated with adverse health consequences, but may also elicit favorable effects under chronic conditions. This "obesity paradox" is under debate for pulmonary diseases. As confounding factors complicate conclusions from human studies, this study used a controlled animal model combining diet-induced obesity and chronic hypoxia as a model for pulmonary hypertension and chronic obstructive pulmonary disease. Male C57BL/6 mice were fed control or high-fat diet for 30 wk, and half of the animals were exposed to chronic hypoxia (13% O2) for 3 wk. Hypoxia induced right ventricular hypertrophy, thickening of pulmonary arterial and capillary walls, higher lung volumes, and increased hemoglobin concentrations irrespective of the body weight. In contrast, lung proteomes differed substantially between lean- and obese-hypoxic mice. Many of the observed changes were linked to vascular and extracellular matrix (ECM) proteins. In lean-hypoxic animals, circulating platelets were reduced and abundances of various clotting-related proteins were altered, indicating a hypercoagulable phenotype. Moreover, the septal ECM composition was changed, and airspaces were significantly distended pointing to lung hyperinflation. These differences were mostly absent in the obese-hypoxic group. However, the obesity-hypoxia combination induced the lowest blood CO2 concentrations, indicating hyperventilation for sufficient oxygen supply. Moreover, endothelial surface areas were increased in obese-hypoxic mice. Thus, obesity exerts differential effects on lung adaptation to hypoxia, which paradoxically include not only adverse but also rather protective changes. These differences have a molecular basis in the lung proteome and may influence the pathogenesis of lung diseases. This should be taken into account for future individualized prevention and therapy.NEW & NOTEWORTHY An "obesity paradox" is discussed for pulmonary diseases. By linking lung proteome analyses to pulmonary structure and function, we demonstrate that diet-induced obesity affects lung adaptation to chronic hypoxia in various ways. The observed changes include not only adverse but also protective effects and are associated with altered abundances of vascular and extracellular matrix proteins. These results highlight the existence of relevant differences in individuals with obesity that may influence the pathogenesis of lung diseases.

Hypoxia signaling in the equine small intestine: Expression and distribution of hypoxia inducible factors during experimental ischemia.

Introduction: Hypoxia inducible factors (HIF) are widely researched in human medicine for their role in different disease processes. The aim of this study was to investigate the expression and distribution of HIF in experimental small intestinal ischemia in the horse. Methods: In 14 horses under general anesthesia, segmental jejunal ischemia with 90% reduction in blood flow was induced. The horses were randomly divided into two groups of seven horses, one subjected to ischemic postconditioning (IPoC) by delayed reperfusion, and a control group (group C) undergoing undelayed reperfusion. Intestinal samples were taken pre-ischemia, after ischemia and after reperfusion. Following immunohistochemical staining for HIF1α and -2α, the immunoreactivity pattern in the small intestine was evaluated by light microscopy, and the mucosal enterocyte and muscularis staining were semi-quantitatively scored. Additionally, mucosal HIF1α protein levels were determined by an Enzyme Linked Immunosorbent Assay (ELISA), and mRNA levels of HIF1α and its target genes by a two-step real-time Reverse Transcriptase Polymerase Chain Reaction. Statistical comparison was performed between the groups and time points using parametric and non-parametric tests (p < 0.05). Results: All cell types exhibited cytoplasmic and nuclear immunoreactivity for HIF1α. After reperfusion, the cytoplasmic staining of the crypt and villus enterocytes as well as the villus nuclear staining significantly increased, whereas the perinuclear granules in the crypts decreased. The protein levels showed a significant decrease in group C at reperfusion, with lower HIF1α levels in group C compared to group IPoC during ischemia and reperfusion. No other group differences could be detected. In the HIF2α stained slides, mild to moderate cytoplasmic staining yet no nuclear immunoreactivity of the enterocytes was observed, and no significant changes over time were noted. Discussion: the changes in HIF1α immunoreactivity pattern and expression over time suggest that this transcription factor plays a role in the intestinal response to ischemia in horses. However, the current study could not identify an effect of IPoC on HIF distribution or expression.

Ultrastructural and Immunohistochemical Characterization of Maternal Myofibroblasts in the Bovine Placenta around Parturition.

Myofibroblasts are contractile cells that exhibit features of both fibroblasts and smooth muscle cells. In the synepitheliochorial placenta of the cow myofibroblasts are found in the maternal stroma. However, a deeper understanding of the structure and function of the stromal myofibroblasts in the developed bovine placenta is still missing. Thus, immunohistochemical and ultrastructural analyses in bovine term placentomes, compared to non-pregnant caruncle samples, were conducted. To investigate functional aspects, contractility of placentomal caruncle slices was assessed in an in vitro contraction assay. Additionally, a three-dimensional reconstruction of a bovine placental myofibroblast was created. Immunofluorescent staining revealed a characteristic pattern, including cytoplasmic expression of α-smooth muscle actin, strong perinuclear signal for the intermediate filament vimentin and nuclear progesterone receptor staining. Ultrastructurally, stress fibers, extended cisternae of the rough endoplasmic reticulum and perinuclear intermediate filaments were observed. Moreover, in vitro stimulation with angiotensin-II, but not with prostaglandin F2α, induced contraction of placental caruncle tissue. Altogether, these results indicate that progesterone-responsive myofibroblasts represent a mesenchymal phenotype that is involved in the contractile properties of bovine placental stroma. Therefore, the present findings suggest a potential involvement of myofibroblasts in post-partum events of cattle, i.e., expulsion of fetal membranes and uterine involution.

Influence of Von Willebrand Disease (VWD) and pregnancy on the expression of angiogenic factors in the porcine female reproductive tract.

Von Willebrand Disease (VWD) is a heritable disorder caused by defects of the Von Willebrand Factor (VWF), leading to deficiencies in coagulation and also angiogenesis. Women affected by VWD frequently show bleeding concerning the reproductive tract and may present with increased rates of miscarriages. We used a porcine model representing VWD type 1 and type 3 as well as the wildtype. Samples were obtained from the reproductive tract of non-pregnant sows and sows pregnant at time of placentation. Relative expression of the genes CALR, CCN2, CXCL8, ECE1, EDN1, F8, IGFBP7, and LGALS3 was analyzed. CCN2 and FVIII proteins were additionally analyzed using immunohistochemistry. In uterus and ovary significant upregulation of CCN2 was seen in non-pregnant pigs affected by VWD. This might be caused by the higher VEGFA-levels in these pigs and could have an influence angiogenesis. During pregnancy, CCN2 expression increased in wildtype pig uteri but hardly changed in those of pregnant pigs affected by VWD, presumably because the expression level in the latter pigs already was significantly increased before pregnancy. F8 expression was significantly reduced in uterus and ovary of VWD-affected pigs. VWF is known to protect FVIII from decomposition and a lack of VWF leads to lower levels of FVIII. Our results suggest that a reduced F8 expression primarily might contribute to those reduced FVIII levels in VWD-affected pigs. Additional significant results involving the pregnant pigs were detected for CALR, EDN1, and LGALS3. These genes are promising candidates for more detailed future studies.

Oral Supplementation with the Polyamine Spermidine Affects Hepatic but Not Pulmonary Lipid Metabolism in Lean but Not Obese Mice.

The polyamine spermidine is discussed as a caloric restriction mimetic and therapeutic option for obesity and related comorbidities. This study tested oral spermidine supplementation with regard to the systemic, hepatic and pulmonary lipid metabolism under different diet conditions. Male C57BL/6 mice were fed a purified control (CD), high sucrose (HSD) or high fat (HFD) diet with (-S) or without spermidine for 30 weeks. In CD-fed mice, spermidine decreased body and adipose tissue weights and reduced hepatic lipid content. The HSD induced hepatic lipid synthesis and accumulation and hypercholesterolemia. This was not affected by spermidine supplementation, but body weight and blood glucose were lower in HSD-S compared to HSD. HFD-fed mice showed higher body and fat depot weights, prediabetes, hypercholesterolemia and severe liver steatosis, which were not altered by spermidine. Within the liver, spermidine diminished hepatic expression of lipogenic transcription factors SREBF1 and 2 under HSD and HFD and affected the expression of other lipid-related enzymes. In contrast, diet and spermidine exerted only minor effects on pulmonary parameters. Thus, oral spermidine supplementation affects lipid metabolism in a diet-dependent manner, with significant reductions in body fat and weight under physiological nutrition and positive effects on weight and blood glucose under high sucrose intake, but no impact on dietary fat-related parameters.

Pooled human bone marrow-derived mesenchymal stromal cells with defined trophic factors cargo promote dermal wound healing in diabetic rats by improved vascularization and dynamic recruitment of M2-like macrophages.

Human Mesenchymal Stromal Cells (hMSCs) are a promising source for cell-based therapies. Yet, transition to phase III and IV clinical trials is remarkably slow. To mitigate donor variabilities and to obtain robust and valid clinical data, we aimed first to develop a manufacturing concept balancing large-scale production of pooled hMSCs in a minimal expansion period, and second to test them for key manufacture and efficacy indicators in the clinically highly relevant indication wound healing. Our novel clinical-scale manufacturing concept is comprised of six single donor hMSCs master cell banks that are pooled to a working cell bank from which an extrapolated number of 70,000 clinical doses of 1x106 hMSCs/cm2 wound size can be manufactured within only three passages. The pooled hMSC batches showed high stability of key manufacture indicators such as morphology, immune phenotype, proliferation, scratch wound healing, chemotactic migration and angiogenic support. Repeated topical hMSCs administration significantly accelerated the wound healing in a diabetic rat model by delivering a defined growth factor cargo (specifically BDNF, EGF, G-CSF, HGF, IL-1α, IL-6, LIF, osteopontin, VEGF-A, FGF-2, TGF-ß, PGE-2 and IDO after priming) at the specific stages of wound repair, namely inflammation, proliferation and remodeling. Specifically, the hMSCs mediated epidermal and dermal maturation and collagen formation, improved vascularization, and promoted cell infiltration. Kinetic analyses revealed transient presence of hMSCs until day (d)4, and the dynamic recruitment of macrophages infiltrating from the wound edges (d3) and basis (d9), eventually progressing to the apical wound on d11. In the wounds, the hMSCs mediated M2-like macrophage polarization starting at d4, peaking at d9 and then decreasing to d11. Our study establishes a standardized, scalable and pooled hMSC therapeutic, delivering a defined cargo of trophic factors, which is efficacious in diabetic wound healing by improving vascularization and dynamic recruitment of M2-like macrophages. This decision-making study now enables the validation of pooled hMSCs as treatment for impaired wound healing in large randomized clinical trials.

Low Flow versus No Flow: Ischaemia Reperfusion Injury Following Different Experimental Models in the Equine Small Intestine.

In experimental studies investigating strangulating intestinal lesions in horses, different ischaemia models have been used with diverging results. Therefore, the aim was to comparatively describe ischaemia reperfusion injury (IRI) in a low flow (LF) and no flow (NF) model. Under general anaesthesia, 120 min of jejunal ischaemia followed by 120 min of reperfusion was induced in 14 warmbloods. During ischaemia, blood flow was reduced by 80% (LF, n = 7) or by 100% (NF, n = 7). Intestinal blood flow and oxygen saturation were measured by Laser Doppler fluxmetry and spectrophotometry. Clinical, histological, immunohistochemical and Ussing chamber analyses were performed on intestinal samples collected hourly. Tissue oxygen saturation was significantly lower in NF ischaemia. The LF group exhibited high variability in oxygen saturation and mucosal damage. Histologically, more haemorrhage was found in the LF group at all time points. Cleaved-caspase-3 and calprotectin-stained cells increased during reperfusion in both groups. After NF ischaemia, the tissue conductance was significantly higher during reperfusion. These results aid in the selection of suitable experimental models for future studies. Although the LF model has been suggested to be more representative for clinical strangulating small intestinal disease, the NF model produced more consistent IRI.

Connexin43 represents an important regulator for Sertoli cell morphology, Sertoli cell nuclear ultrastructure, and Sertoli cell maturation.

The Sertoli cell (SC)-specific knockout (KO) of connexin43 (Cx43) was shown to be an effector of multiple histological changes in tubular morphology, resulting in germ cell loss through to a Sertoli-cell-only (SCO) phenotype and vacuolated seminiferous tubules containing SC-clusters. Our present study focused on the effects of Cx43 loss on SC ultrastructure. Using serial block-face scanning electron microscopy (SBF-SEM), we could confirm previous results. Ultrastructural analysis of Sertoli cell nuclei (SCN) revealed that these appear in clusters with a phenotype resembling immature/proliferating SCs in KO mice. Surprisingly, SCs of fertile wild type (WT) mice contained SCN with a predominantly smooth surface instead of deep indentations of the nuclear envelope, suggesting that these indentations do not correlate with germ cell support or spermatogenesis. SBF-SEM facilitated the precise examination of clustered SCs. Even if the exact maturation state of mutant SCs remained unclear, our study could detect indications of cellular senescence as well as immaturity, emphasising that Cx43 affects SC maturation. Moreover, Sudan III staining and transmission electron microscopy (TEM) demonstrated an altered lipid metabolism in SCs of Cx43 deficient mice.

Number of Primordial Follicles in Juvenile Ringed Seals (Pusa hispida) from the Gulf of Bothnia and West Greenland.

Primordial follicles are important for the reproduction cycle and, therefore, also for the survival of the whole population of a species. Mammals have a large pool of primordial follicles, and it is thought that this pool represents the total number of oocytes. The aim of the present study was to determine the total primordial follicle number of juvenile ringed seals (Pusa hispida) from the Gulf of Bothnia and Greenland. Overall, 52 ovaries from two ringed seal populations (West Greenland (N = 6), Gulf of Bothnia, region in the Baltic Sea (N = 46)) were examined. All ovaries were cut into 2 mm thick slices and every slice was embedded in paraffin. Out of each tissue block, a 5 µm thick section was cut and stained with haematoxylin-eosin. The mean volume of the follicles and the total volume of primordial follicles per ovary were estimated by stereology and used to calculate the total estimated number of primordial follicles. The median of the total estimated number of primordial follicles seemed to be higher in Baltic individuals than in Greenland individuals (Gulf of Bothnia = 565,657; Greenland Sea = 122,475). This widens the total range of primordial follicles in ringed seals overall and might bear some potential for discussions regarding the influence of endocrine disruptors and environmental influences depending on different regions/populations and their exposure to various factors. Thus, this study aims to provide basic reference data of the number and mean volume of ringed seal primordial follicles.

The effect of ischaemic postconditioning on mucosal integrity and function in equine jejunal ischaemia.

BACKGROUND: Ischaemic postconditioning (IPoC) has been shown to ameliorate ischaemia reperfusion injury in different species and tissues. OBJECTIVES: To assess the feasibility of IPoC in equine small intestinal ischaemia and to assess its effect on histomorphology, electrophysiology and paracellular permeability. STUDY DESIGN: Randomised in vivo experiment. METHODS: Experimental jejunal ischaemia was induced for 90 min in horses under general anaesthesia. In the control group (C; n = 7), the jejunum was reperfused without further intervention. In the postconditioning group (IPoC; n = 7), reocclusion was implemented following release of ischaemia by clamping the mesenteric vessels in three cycles of 30 seconds. This was followed by 120 minutes of reperfusion in both groups. Intestinal microperfusion and oxygenation was measured during IPoC using spectrophotometry and Doppler flowmetry. Histomorphology and histomorphometry of the intestinal mucosa were assessed. Furthermore, electrophysiological variables and unidirectional flux rates of 3 H-mannitol were determined in Ussing chambers. Western blot analysis was performed to determine the tight junction protein levels of claudin-1, claudin-2 and occludin in the intestinal mucosa. Comparisons between the groups and time points were performed using a two-way repeated measures analysis of variance (ANOVA) or non-parametric statistical tests for the ordinal and not normally distributed data (significance P < .05). RESULTS: IPoC significantly reduced intestinal microperfusion during all clamping cycles yet affected oxygen saturation only during the first cycle. After reperfusion, Group IPoC showed significantly less mucosal villus denudation (mean difference 21.5%, P = .02) and decreased mucosal-to-serosal flux rates (mean difference 15.2 nM/cm2 /h, P = .007) compared to Group C. There were no significant differences between the groups for the other tested variables. MAIN LIMITATIONS: Small sample size, long-term effects were not investigated. CONCLUSIONS: Following IPoC, the intestinal mucosa demonstrated significantly less villus denudation and paracellular permeability compared to the untreated control group, possibly indicating a protective effect of IPoC on ischaemia reperfusion injury.

Hypoxic Processes Induce Complement Activation via Classical Pathway in Porcine Neuroretinas.

Considering the fact that many retinal diseases are yet to be cured, the pathomechanisms of these multifactorial diseases need to be investigated in more detail. Among others, oxidative stress and hypoxia are pathomechanisms that take place in retinal diseases, such as glaucoma, age-related macular degeneration, or diabetic retinopathy. In consideration of these diseases, it is also evidenced that the immune system, including the complement system and its activation, plays an important role. Suitable models to investigate neuroretinal diseases are organ cultures of porcine retina. Based on an established model, the role of the complement system was studied after the induction of oxidative stress or hypoxia. Both stressors led to a loss of retinal ganglion cells (RGCs) accompanied by apoptosis. Hypoxia activated the complement system as noted by higher C3+ and MAC+ cell numbers. In this model, activation of the complement cascade occurred via the classical pathway and the number of C1q+ microglia was increased. In oxidative stressed retinas, the complement system had no consideration, but strong inflammation took place, with elevated TNF, IL6, and IL8 mRNA expression levels. Together, this study shows that hypoxia and oxidative stress induce different mechanisms in the porcine retina inducing either the immune response or an inflammation. Our findings support the thesis that the immune system is involved in the development of retinal diseases. Furthermore, this study is evidence that both approaches seem suitable models to investigate undergoing pathomechanisms of several neuroretinal diseases.

A double deletion prevents replication of the pestivirus bovine viral diarrhea virus in the placenta of pregnant heifers.

In contrast to wild type bovine viral diarhea virus (BVDV) specific double deletion mutants are not able to establish persistent infection upon infection of a pregnant heifer. Our data shows that this finding results from a defect in transfer of the virus from the mother animal to the fetus. Pregnant heifers were inoculated with such a double deletion mutant or the parental wild type virus and slaughtered pairwise on days 6, 9, 10 and 13 post infection. Viral RNA was detected via qRT-PCR and RNAscope analyses in maternal tissues for both viruses from day 6 p.i. on. However, the double deletion mutant was not detected in placenta and was only found in samples from animals infected with the wild type virus. Similarly, high levels of wild type viral RNA were present in fetal tissues whereas the genome of the double deletion mutant was not detected supporting the hypothesis of a specific inhibition of mutant virus replication in the placenta. We compared the induction of gene expression upon infection of placenta derived cell lines with wild type and mutant virus via gene array analysis. Genes important for the innate immune response were strongly upregulated by the mutant virus compared to the wild type in caruncle epithelial cells that establish the cell layer on the maternal side at the maternal-fetal interface in the placenta. Also, trophoblasts which can be found on the fetal side of the interface showed significant induction of gene expression upon infection with the mutant virus although with lower complexity. Growth curves recorded in both cell lines revealed a general reduction of virus replication in caruncular epithelial cells compared to the trophoblasts. Compared to the wild type virus this effect was dramtic for the mutant virus that reached only a TCID50 of 1.0 at 72 hours post infection.

SPI2 T3SS effectors facilitate enterocyte apical to basolateral transmigration of Salmonella-containing vacuoles in vivo.

Salmonella pathogenicity island (SPI) 2 type three secretion system (T3SS)-mediated effector molecules facilitate bacterial survival in phagocytes but their role in the intestinal epithelium in vivo remains ill-defined. Using our neonatal murine infection model in combination with SPI2 reporter technology and RNA-Seq of sorted primary enterocytes, we demonstrate expression of SPI2 effector molecules by intraepithelial Salmonella Typhimurium (S. Typhimurium). Contrary to expectation, immunostaining revealed that infection with SPI2 T3SS-mutants resulted in significantly enlarged intraepithelial Salmonella-containing vacuoles (SCV) with altered cellular positioning, suggesting impaired apical to basolateral transmigration. Also, infection with isogenic tagged S. Typhimurium strains revealed a reduced spread of intraepithelial SPI2 T3SS mutant S. Typhimurium to systemic body sites. These results suggest that SPI2 T3SS effector molecules contribute to enterocyte apical to basolateral transmigration of the SCV during the early stage of the infection.

Comparative morphometric study of the mimic facial muscles of brachycephalic and dolichocephalic dogs.

Excessive breeding of dogs has led to a disadvantageous morphology in some breeds, for example extreme brachycephaly, which is responsible for many health issues. We hypothesize that alterations of the mimic muscular system are present in brachycephalic dogs and could contribute to behavioural problems due to a restricted mimic display. Therefore, the purpose of this paper was to compare the mimic muscular systems of brachycephalic with dolichocephalic dogs. Mimic muscles were measured and set in ratio to measurements of the head and calculated indices. When bringing the length of the muscles m(x) in proportion to the length of the head, highly significant differences (p < .0001) and significant difference (p < .05) were found in all but two of the examined muscles. Calculations of the m(x) divided by the cranial index and the square index showed significant differences for all muscles. For example, the musculus (m.) levator nasolabialis was morphologically different from the one of dolichocephalic dogs. Muscle fibres of the m. levator nasolabialis were localized in the fold over the nasal bridge of brachycephalic dogs. The raphe of the m. orbicularis oris was not always apparent in brachycephalic dogs. The proportions of the muscle lengths and the length of the eye slots to the size of the skull have shifted considerably in brachycephalic dogs. We conclude that many alterations contribute to the strong shift in the proportions of the head of brachycephalic dogs versus that of dolichocephalic dogs. Our findings suggest that brachycephalic dogs have reduced mimic skills that can lead to ambiguous communication.

Studies on the weight of the gastrointestinal tract, digesta composition and occurrence of gastro- and enteroliths in adult domesticated ostriches fed different diets.

Comminuting the ingested material in the stomach and fermentation in the large intestine of ostriches, allows an efficient utilization of fiber-rich feedstuffs. The entire gastrointestinal tract (GIT) of 61 adult ostriches (both sexes; av. age of 15 mo), which had previously been fed a ration consisting of either haylage and pelleted compound feed (HP) or haylage, corn silage and pelleted compound feed (HCP), was the subject of the present investigations. The weight of the different compartments of the GIT was measured. The digesta was differentiated into inorganic and organic substances. Wet sieving was used to separate the collected stones (>19 mm), small stones (1 mm), and sand (<1 mm). Ostriches fed the HCP diet had a significantly higher empty gizzard weight (3,435 g) compared to those fed the HP diet (3,064 g). Additionally, the weight of the empty cecum (left and right parts) was increased (P < 0.05) for ostriches fed the HCP diet (107 and 122 g, respectively) in comparison to those fed the HP diet (93.4 and 108 g, respectively). The weight of pure digesta in the gizzard and left or right cecum for ostriches fed the HP diet was higher (1,640, 448, and 471 g, respectively) compared to those fed the HCP diet (P < 0.05). The contents of crude ash and HCl-insoluble ash in the digesta of all the GIT compartments were higher for ostriches fed the HP diet in comparison to those fed the HCP diet (P < 0.05). Independent of the type of the offered diet, the large stones occurred only in the proventriculus and gizzard (2.71 and 4.76%, respectively), while sand dominated in the distal colon (30.3%). The high proportion of stones in the gizzard form the "mechanical equipment" which enables the animals to grind basic feed such as corn silage or haylage, and these are almost completely excreted as sand. Continuous stone replacement for ostriches is necessary but the amount mostly depends on the type of feed.