The Antiangiogenic Properties of Adipose-Derived Mesenchymal Stem/Stromal Cells in Corneal Neovascularization in a Rabbit Model

The purpose was to study the anti-angiogenic effect of adipose-derived mesenchymal stem/stromal cells (ADMSCs) on experimentally induced corneal injuries. Corneal neovascularization (NV) was induced by incising and subsequently suturing the corneal surface in 32 New Zealand rabbits. Following suturing, the rabbits were randomly allocated into 2 groups, and received either phosphate-buffered saline (PBS) (control) or ADMSCs, both administered via three different routes. Digital images of the cornea were obtained two weeks post-incision to measure the area of neovascularized cornea. Tumor necrosis factor (TNF) was immunohistochemically assessed in the both groups. The corneal tissue was evaluated for vascular endothelial growth factor (VEGF). The extent of corneal NV in all eyes was assessed photographically by an independent observer. Fourteen days after the incisions, the degree of corneal NV was substantially decreased in the ADMSC-treated group (1.87 ± 0.9 mm2, 1.4 % ± 0.67 % of corneal surface) compared to the control and PBS-treated group (4.66 ± 1.74 mm2, 3.51 % ± 1.31 %, p < 0.001). ADMSCs significantly decreased injury-induced corneal NV in New Zealand rabbits two weeks post-treatment. This strategy has potential for use in the control of corneal NV in vivo.Â

The Antiangiogenic Properties of Adipose-Derived Mesenchymal Stem/Stromal Cells in Corneal Neovascularization in a Rabbit Model

The purpose was to study the anti-angiogenic effect of adipose-derived mesenchymal stem/stromal cells (ADMSCs) on experimentally induced corneal injuries. Corneal neovascularization (NV) was induced by incising and subsequently suturing the corneal surface in 32 New Zealand rabbits. Following suturing, the rabbits were randomly allocated into 2 groups, and received either phosphate-buffered saline (PBS) (control) or ADMSCs, both administered via three different routes. Digital images of the cornea were obtained two weeks post-incision to measure the area of neovascularized cornea. Tumor necrosis factor (TNF) was immunohistochemically assessed in the both groups. The corneal tissue was evaluated for vascular endothelial growth factor (VEGF). The extent of corneal NV in all eyes was assessed photographically by an independent observer. Fourteen days after the incisions, the degree of corneal NV was substantially decreased in the ADMSC-treated group (1.87 ± 0.9 mm2, 1.4 % ± 0.67 % of corneal surface) compared to the control and PBS-treated group (4.66 ± 1.74 mm2, 3.51 % ± 1.31 %, p < 0.001). ADMSCs significantly decreased injury-induced corneal NV in New Zealand rabbits two weeks post-treatment. This strategy has potential for use in the control of corneal NV in vivo.

Comparative study of clinicopathological and neuropathological investigation of brains of aged dogs with or without canine cognitive dysfunction syndrome (dementia)

Canine cognitive dysfunction syndrome (CCDS) is a progressive neurodegenerative disease, that is highly prevalent in old dogs but critically underreported. CCDS models early Alzheimer disease. To date, no specific multi-targeted protocol exists for the diagnosis of the disease and many key aspects of the syndrome are not elucidated. The aim of this study was firstly to comprehensively quantify Aβ42 and Aβ40 peptides in blood and CSF and to investigate their contribution in the improvement of CCDS individual’s characterization. Secondly, through the histopathological and immunohistochemical investigation of these brains of interest, we aim to validate the above measurements as prominent biomarkers of CCDS, as pathology is considered as the “gold standard” for the credibility rating of candidate biomarkers. A total of 61 dogs were assigned to four groups: i) Young (YG) ii) Middle-aged (MA) iii) Cognitively Unimpaired and Aged (CU) iv) Cognitively Impaired and Aged (CI). Animal’s cognitive status was assessed using Canine Cognitive Assessment Scale modified by the Companion Animal Welfare Education Center. Based on the scale score the CI group was further subdivided in dogs with Mild Cognitive Impairment (CI-MCI) and in dogs with Severe Cognitive Impairment (CI-SCI). Direct and indirect Aβ markers were determined in plasma fractions (total-TP, free-FP, bound to plasma components-CP) and CSF using AΒtest assays (Araclon Biotech). The dogs of the study died either naturally or were euthanized due to debilitating disorders and their brains were collected. The brains were routinely fixed, and samples were collected from: 1) prefrontal cortex 2) frontal lobe and caudate nucleus 3) parietal lobe 4) temporal lobe, piriform lobe and diencephalon 5) temporal lobe and hippocampus and 6) occipital lobe. Histopathological and immunohistochemical (Aβ40, Αβ42, p-tauSer396, ApoE4, GFAP) examination of paraffin wax-embedded brain sections were performed. A semiquantitative scoring system was used for the evaluation of the immunohistochemical findings. ΤP42 and CP42 levels were significantly higher in CI-MCI dogs when compared to CU and CI-SCI dogs. CSFAβ42 levels in YG were the lowest and were higher in CU than in CI dogs. CSF Aβ42 levels were correlated only with CPAβ42 and the CPAβ40. TPAβ42/40 allowed discrimination between CI-MCI and CU aged dogs. Positive labelling for Aβ40, Αβ42, p-tau and ApoE4 was detected in the brain of CU and CI aged dogs. In CU and MCI dogs, the fraction ΤΡ42 proved to be the most sensitive for the prediction of the Aβ42 brain depositions, the fraction ΤP42/40 was the most appropriate for the detection of early p-tau depostitions and the fraction CP42 was the most useful biomarker for the prediction of the overall of the p-tau depositions. We conclude that the role of p-tau protein on CCDS should be reconsidered. TP42/40 could generate, in the earliest phase of the symptoms, a new recruitment strategy for efficient preselection in the population of interest. The determination of a combination of CSF and plasma markers (TP42/40, ΤP42, CP42, CP42/40 and CSF42) on a large scale could serve a comprehensive role in a manifold approach of the disease, especially in the early stagesΤο σύνδρομο γνωστικής δυσλειτουργίας του σκύλου (CCDS) είναι ένα προοδευτικά εξελισσόμενο νευροεκφυλιστικό νόσημα, το οποία εάν και είναι συχνό στους παρήλικους σκύλους, είναι σημαντικά υποδιαγνωσμένο. Το CCDS έχει πολλές ομοιότητες με το πρώιμο στάδιο της νόσου Alzheimer. Μέχρι σήμερα, δεν υπάρχει ειδικό πολυπαραμετρικό πρωτόκολλο για τη διάγνωση της νόσου και πολλές βασικές πτυχές του συνδρόμου δεν έχουν διασαφηνιστεί. Ο στόχος αυτής της μελέτης ήταν: πρώτον, ο ποσοτικός προσδιορισμός των πεπτιδίων Αβ42 και Αβ40, στο αίμα και το εγκεφαλονωτιαίο υγρό (ΕΝΥ) και η διερεύνηση της συμβολής αυτών στη διάγνωση σκύλων με CCDS. Δεύτερον, η επιβεβαίωση της κλινικής διάγνωσης καθώς και η επικύρωση της αξιοπιστίας των παραπάνω παραμέτρων ως βιοδείκτες του CCDS, μέσω της ιστοπαθολογικής και ανοσοϊστοχημικής διερεύνησης των εγκεφάλων των σκύλων της μελέτης, αφού, η παθολογοανατομική διερεύνηση αποτελεί τη μέθοδο εκλογής για την αξιολόγηση της αξιοπιστίας ενός βιοδείκτη. Στη μελέτη συμμετείχαν 61 σκύλοι χωρισμένοι σε τέσσερις ομάδες: i) Νεαροί (Υoung-YG, 0-4 ετών) ii) Μέσης Ηλικίας (Middle Aged, MA, 4-8 ετών) iii) Γνωστικά υγιείς παρήλικοι (Cognitive unimpaired-CU, >8 ετών) iv) Παρήλικοι με γνωστικές διαταραχές (Cognitive Impaired-CI, >8 ετών). Για την αξιολόγηση του γνωστικού προφίλ των παρήλικων σκύλων χρησιμοποιήθηκε βαθμολογική κλίμακα με τη μορφή ερωτηματολογίου που πρέπει να απαντάται από τον κηδεμόνα του κάθε ζώου. Με βάση τη βαθμολογία της κλίμακας, η ομάδα των CI παρήλικων σκύλων διακρίθηκε σε δύο υποομάδες: 1) Σκύλοι με ήπια γνωστική διαταραχή (Mild Cognitive Impairment, MCI) 2) Σκύλοι με σοβαρή γνωστική διαταραχή (Severe Cognitive Impairment, SCI). Tα επίπεδα των Αβ40 και Αβ42 προσδιορίστηκαν σε τρία κλάσματα του πλάσματος (ολικό στο πλάσμα-ΤΡ, ελεύθερο στο πλάσμα-FΡ, δεσμευμένο σε συστατικά του πλάσματος-CP) και στο ΕΝΥ χρησιμοποιώντας δύο επικυρωμένες δοκιμές (AΒtest, Araclon Biotech). Από τους σκύλους της μελέτης συλλέχθηκαν οι εγκέφαλοι. Οι εγκέφαλοι μονιμοποιούνταν σε διάλυμα φορμόλης 10% και στη συνέχεια, από κάθε εγκέφαλο λαμβάνονταν ιστοτεμάχια από όλες τις ανατομικές περιοχές ενδιαφέροντος. Ακολούθως, σε τομές παραφίνης έγινε ιστοπαθολογική και ανοσοϊστοχημική (Αβ40, Αβ42, p-tauSer396, ApoE4 και GFAP) διερεύνηση. Τα επίπεδα των ΤP42 και CP42 ήταν σημαντικά υψηλότερα στους ΜCI σκύλους σε σύγκριση με τις άλλες ομάδες των παρήλικων σκύλων. Τα επίπεδα του Αβ42 στο ΕΝΥ συσχετίστηκαν μόνο με το κλάσμα CP42 και το CP40. Ο ΤΡ42/40 επέτρεψε τη διάκριση των CU από τους MCI σκύλους. Στους CU και ΜCI σκύλους, ο ΤΡ42 ήταν ο πιο ευαίσθητος για να τεθεί υποψία παρουσίας εναποθέσεων του Αβ42 στον εγκεφάλο. Ο ΤΡ42/40 ήταν ο καταλληλότερος για την ανίχνευση πρώιμων εναποθέσεων της p-tau και ο CP42 ήταν ο πιο χρήσιμος για την εκτίμηση της συνολικής εναπόθεσης της p-tau πρωτεΐνης. Συμπερασματικά, ο ρόλος της p-tau στην παθογένεια του CCDS θα πρέπει να επανεξεταστεί. Ο βιοδείκτης TP42/40 μπορεί να δημιουργήσει, μια νέα στρατηγική εντόπισης των παρήλικων σκύλων που εμφανίζουν περισσότερες πιθανότητες να νοσήσουν, ανεξαρτήτως της διακριτικής ικανότητας του κηδεμόνα τους. Η συνδυαστική αξιολόγηση μιας ομάδας βιοδεικτών ( TP42/40, ΤP42, CP42, CP42/40 στο πλάσμα και του Aβ42 στο ΕΝΥ ) σε μεγάλη κλίμακα θα μπορούσε να αποτελέσει χρήσιμο εργαλείο για την πρώιμη διάγνωση και τον ακριβή ορισμό των αρχικών σταδίων του CCDS

Efficacy of a Dietary Polyherbal Formula on the Performance and Gut Health in Broiler Chicks after Experimental Infection with Eimeria spp.

One-hundred and fifty, one-day-old Ross-308 female chicks were randomly allocated to five equal treatments: NCONTR negative control—not challenged; PCONTR positive control—challenged; PHERB1 and PHERB2 diets were supplemented with phytogenic formula (1 and 2 g/kg feed, respectively)—challenged; PSALIN diet was supplemented with salinomycin (60 mg/kg feed)—challenged. Challenge was made by oral inoculation with 3.5 × 104 E. acervulina, 7.0 × 103 E. maxima and 5.0 × 103 E. tenella oocysts, at 14 days of age. One week post inoculation, bloody diarrhea, oocysts numbers, and intestinal lesions were evaluated, along with intestinal microbiota, viscosity, and pH of digesta, and histopathology. PHERB2 had a comparable (p ≤ 0.001) growth performance and feed conversion ratio to PSALIN. PHERB1 and PHERB2 had similar (p ≤ 0.001) oocyst counts to PSALIN and lower than PCONTROL. PHERB2 and PSALIN had lower (p ≤ 0.001) jejunal, ileal, and cecal lesion scores compared to PCONTR. PHERB1 and PHERB2 had higher (p ≤ 0.001) jejunal and cecal lactobacilli and lower (p ≤ 0.001) coliform counts compared to other treatments. PCONTR had lower (p ≤ 0.001) jejunum villus height, height to crypt ratio, and villus goblet cells. Breast and thigh meat resistance to oxidation was improved (p ≤ 0.001) in PHERB1 and PHERB2 compared to the PCONTR. The polyherbal formula exerted a substantial improvement on growth performance and intestinal health of the Eimeria-challenged birds

Effects of Enriched-in-Oleuropein Olive Leaf Extract Dietary Supplementation on Egg Quality and Antioxidant Parameters in Laying Hens

The objective of the present study was to evaluate the effects of an olive leaf extract obtained with an up-to-date laboratory method, when supplemented at different levels in laying hens’ diets, on egg quality, egg yolk antioxidant parameters, fatty acid content, and liver pathology characteristics. Thus, 96 laying hens of the ISA-Brown breed were allocated to 48 experimental cages with two hens in each cage, resulting in 12 replicates per treatment. Treatments were: T1 (Control: basal diet); T2 (1% olive leaf extract); T3 (2.5% olive leaf extract); T4 (Positive control: 0.1% encapsulated oregano oil). Eggshell weight and thickness were improved in all treatments compared to the control, with T2 being significantly higher till the end of the experiment (p < 0.001). Egg yolk MDA content was lower for the T2 and T4 groups, while total phenol content and Haugh units were greater in the T2. The most improved fatty acid profile was the one of T3 yolks. The α-tocopherol yolk content was higher in all groups compared to T1. No effect was observed on cholesterol content at any treatment. Based on the findings, it can be inferred that the inclusion of olive leaf extract at a concentration of 1% in the diet leads to enhancements in specific egg quality attributes, accompanied by an augmentation of the antioxidant capacity

Effect of an Herbal Mixture of Oregano, Garlic, Sage and Rock Samphire Extracts in Combination with Tributyrin on Growth Performance, Intestinal Microbiota and Morphology, and Meat Quality in Broilers

The present study investigated the effects of two feed additives, the first containing an herbal mixture of oregano, garlic, sage, and rock samphire extracts and the second containing tributyrin (glyceryl tributyrate) when fed to broiler chickens. A total of 360 one-day-old chicks were randomly allocated to four treatments (6 replicate pens of 15 chicks). One treatment served as the unsupplemented control, whereas the feeds of the other three treatments were supplemented either with the herbal additive (3 g/kg), the tributyrin additive (1 g/kg), or both additives. The duration of the trial was 37 days. Data were collected on growth performance, intestinal microbiota and morphology, and some meat quality parameters. The combined supplementation improved (p p Escherichia coli, total Enterobacteriaceae, and Clostridium spp. compared to the other treatments. Fecal coccidial oocyst counts were also reduced (p p < 0.05) breast and thigh meat resistance to oxidation. In conclusion, the combined dietary supplementation with the examined feed additives could be utilized to improve the performance and intestinal health of broiler chickens

A phytobiotic extract, in an aqueous or in a cyclodextrin encapsulated form, added in diet affects meat oxidation, cellular responses and intestinal morphometry and microbiota of broilers

The present trial examined the effects of diet supplementation with an extract including Greek oregano, garlic, rock samphire, and camelina, administered either in aqueous form or encapsulated in cyclodextrin, on broiler chickens. The duration of the trial was 35 days. Mixed broiler chicks (Ross-308, 120 individuals, 1 day old) were randomly allocated to one of three groups, each with four replicates. Control group A (CONTROL) was fed a basal diet consisting of maize and soybean. The diet of the AQORGCC and CDORGCC groups was further supplemented with aqueous and cyclodextrin-encapsulated herbal extracts, respectively. Levels of lipid and protein oxidation were determined in breast and thigh meat samples. Furthermore, to address cellular stress and signaling responses, the expression patterns of heat shock proteins (Hsp60, Hsp70, and Hsp90), mitogen-activated protein kinases (P38 and P44/42 MAPKs), and apoptotic-related proteins (Bcl-2/Bad ratio) were investigated in breast and thigh tissues using Western blot analysis. The intestinal morphometry of the duodenum, jejunum, and ileum was also assessed. To investigate ileal and cecal bacterial community diversity, 16S rRNA gene high-throughput amplicon sequencing on the V3–V4 hypervariable region was performed. The results showed that the herbal extract in cyclodextrin delayed meat lipid oxidation. According to the protein expression patterns, the formulated diets elicited tissue-specific cellular responses. Compared with the CONTROL group, dietary supplementation with the encapsulated form resulted in significant Hsp induction and MAPK activation, whereas, in the group whose diet was supplemented with the aqueous form, the expression of most of the examined proteins decreased or was maintained at a constant level. Villus height and lamina propria width were mostly affected by the aqueous herbal extract, whereas the number of goblet cells remained unchanged among the groups. Firmicutes, Proteobacteria, and Bacteroidota were the major phyla in mean relative abundance in all diets in both cecal and ileal samples. Alpha-diversity indices highlighted higher species richness and diversity in the cecum than in the ileum, as well as in chicks treated with the aqueous extract of the herbal mixture, but only in the cecum. Cecal beta-diversity differed between the cyclodextrin and the CONTROL groups, while ileal beta-diversity varied only between the aqueous-treated group and the CONTROL group. In conclusion, the dietary mixtures of herbal extracts (particularly those encapsulated in cyclodextrin) improved protein and lipid oxidation and increased the number of beneficial lactic acid-producing bacteria in the cecum, whereas the aqueous herbal extract mostly affected bacterial activity in the proximal part of the chicken intestine. Similarly, intestinal morphometry in the duodenum, jejunum, and ileum was mostly affected by the aqueous herbal extract, which seems to inhibit proteins associated with stress signaling in meat

Effect of an Herbal Mixture of Oregano, Garlic, Sage and Rock Samphire Extracts in Combination with Tributyrin on Growth Performance, Intestinal Microbiota and Morphology, and Meat Quality in Broilers

The present study investigated the effects of two feed additives, the first containing an herbal mixture of oregano, garlic, sage, and rock samphire extracts and the second containing tributyrin (glyceryl tributyrate) when fed to broiler chickens. A total of 360 one-day-old chicks were randomly allocated to four treatments (6 replicate pens of 15 chicks). One treatment served as the unsupplemented control, whereas the feeds of the other three treatments were supplemented either with the herbal additive (3 g/kg), the tributyrin additive (1 g/kg), or both additives. The duration of the trial was 37 days. Data were collected on growth performance, intestinal microbiota and morphology, and some meat quality parameters. The combined supplementation improved (p &lt; 0.05) weigh gain, feed conversion ratio, and the European Efficiency Factor. In the cecum, the combined supplementation lowered (p &lt; 0.05) the microbial populations of aerobes, anaerobes, Escherichia coli, total Enterobacteriaceae, and Clostridium spp. compared to the other treatments. Fecal coccidial oocyst counts were also reduced (p &lt; 0.01) by the combined supplementation. The herbal mixture supplementation improved (p &lt; 0.05) breast and thigh meat resistance to oxidation. In conclusion, the combined dietary supplementation with the examined feed additives could be utilized to improve the performance and intestinal health of broiler chickens

Administration of a Multi-Genus Synbiotic to Broilers: Effects on Gut Health, Microbial Composition and Performance

In recent years, the applicability of prebiotics, probiotics and their mixtures, defined as synbiotics, in poultry production has received considerable attention. Following the increasing regulation of antibiotic use, these nutraceuticals are seen as an alternative way to sustain production efficiency and resistance to pathogens and stressors by modulating birds’ gut health. The aim of this study was to evaluate the benefits provided under field conditions by administering the multi-species synbiotic PoultryStar® sol to broilers in drinking water. To this purpose, three Ross 308 broiler flocks, representing separate progenies of a breeder flock which was treated with the same synbiotic, were housed in separate farms, divided into treatment and control groups, and followed throughout the productive cycle. Synbiotic administration was shown to improve gut health even in absence of a challenge, with limited changes in terms of macroscopic intestinal lesions and more overt differences related to histopathological scores and villi length. Synbiotic-fed chickens performed consistently better in terms of body weight gain, feed conversion ratio and survivability. Lastly, the evaluation of the caecal microbiome through next-generation sequencing highlighted the effects of synbiotic supplementation on the composition of the bacterial population, the implications of which will, however, require further studies to be better comprehended

Quantitative proteomics of cerebrospinal fluid using tandem mass tags in dogs with recurrent epileptic seizures

This prospective study included four dog groups (group A: healthy dogs, groups B: dogs with idiopathic epilepsy under antiepileptic medication (AEM), C: idiopathic epilepsy dogs without AEM administration, D: dogs with structural epilepsy). The purpose of the study was to compare the proteomic profile among the four groups. Samples were analyzed by a quantitative Tandem Mass Tags approach using a Q-Exactive-Plus mass-spectrometer. Identification and relative quantification were performed using Proteome Discoverer, and data were analyzed using R. Gene ontology terms were analyzed based on Canis lupus familiaris database. Data are available via ProteomeXchange with identifier PXD018893. Eighteen proteins were statistically significant among the four groups (P &lt; 0.05). MMP2 and EFEMP2 appeared down-regulated whereas HP and APO-A1 were up-regulated (groups B, D). CLEC3B and PEBP4 were up-regulated whereas APO-A1 was down-regulated (group C). IGLL1 was down-regulated (groups B, C) and up-regulated (group D). EFEMP2 was the only protein detected among the four groups and PEBP4 was significantly different among the epileptic dogs. Western blot and SPARCL immunoassay were used to quantify HP abundance change, validating proteomic analysis. Both, showed good correlation with HP levels identified through proteomic analysis (r = 0.712 and r = 0.703, respectively)