Apelin-13 modulates the endometrial transcriptome of the domestic pig during implantation.

BACKGROUND: The peri-implantation period is a critical time during pregnancy that mostly defines the overall litter size. Most authors agree that the highest percentage of embryo mortality occurs during this time. Despite the brevity of the peri-implantation period, it is the most dynamic part of pregnancy in which the sequential and uninterrupted course of several processes is essential to the animal's reproductive success. Also then, the maternal uterine tissues undergo an intensive remodelling process, and their energy demand dramatically increases. It is believed that apelin, a member of the adipokine family, is involved in the control of female reproductive functions in response to the current metabolic state. The verified herein hypothesis assumed the modulatory effect of apelin on the endometrial tissue transcriptome on days 15 to 16 of gestation (beginning of implantation). RESULTS: The analysis of data obtained during RNA-seq (Illumina HiSeq2500) of endometrial slices treated and untreated with apelin (n = 4 per group) revealed changes in the expression of 68 genes (39 up-regulated and 29 down-regulated in the presence of apelin), assigned to 240 gene ontology terms. We also revealed changes in the frequency of alternative splicing events (397 cases), as well as single nucleotide variants (1,818 cases) in the presence of the adipokine. The identified genes were associated, among others, with the composition of the extracellular matrix, apoptosis, and angiogenesis. CONCLUSIONS: The obtained results indicate a potential role of apelin in the regulation of uterine tissue remodelling during the peri-implantation period.

Resistin in endocrine pancreas of sheep: Presence and expression related to different diets.

Resistin (RETN), a recently discovered adipokine, is a cysteine-rich and secretory protein produced by adipocytes. RETN has been detected in several tissues, including human and laboratory animals' pancreas, wherein impairs glucose tolerance and insulin (INS) action and causes INS resistance. This study aims to evaluate the presence and expression of RETN in the pancreas of 15 adult female sheep reared on Apennine pastures, which show a decrease in their nutritional value due to the drought stress linked to the increasing summer aridity. The sheep were divided into 3 groups according to the diet they were subjected to: maximum pasture flowering (MxF) group, maximum pasture dryness (MxD) group, and experimental (Exp) group which received a feed supplementation in addition to the MxD group feeding. Immunohistochemistry and immunofluorescence were performed on formalin-fixed and paraffin-embedded sections of the pancreas to detect the RETN presence and to evaluate the co-localization of RETN with both glucagon (GCG)- and INS-producing cells. In addition, the expression of the three molecules was evaluated also in relation to different diets. RETN was observed only in the endocrine pancreas, showing a wide distribution throughout the pancreatic islets with few negative cells and the RETN producing cells colocalized with both α cells and ß cells. No differences in distribution and immunostaining intensity of RETN, GCG and INS were observed among the three groups. Quantitative PCR showed the expression of RETN, GCG and INS in all tested samples. No significant differences were observed for RETN and GCG among all three groups of sheep. Instead, a high statistically significant expression of INS was detected in the MxF group with respect to the Exp and MxD groups. These results highlight the localization of RETN in GCG- and INS-secreting cells involved in glucose homeostasis suggesting a modulatory role for RETN. Furthermore, the RETN expression is not influenced by food supplementation and thus is not affected by diet.

Post-mortem ultrasonographic and computed tomographic features of the anatomical variations and acquired pathological bony changes of the lumbosacroiliac region in a mixed population of horses.

BACKGROUND: Transrectal ultrasonography is the best technique for evaluating the ventral aspect of the lumbosacral and sacroiliac regions yet this diagnostic technique does not always lead to a final diagnosis of back pain in horses. OBJECTIVES: To describe anatomical variations and acquired pathological bony changes (APBCs) in the lumbosacral and sacroiliac regions detected by ultrasonography (US) and computed tomography (CT) examinations on specimens. We hypothesised that age, body mass, previous use and anatomical variations may be correlated with the presence and/or severity of APBCs. STUDY DESIGN: Descriptive cadaver study. METHODS: Lumbosacroiliac specimens were obtained from 51 horses that died or were euthanised for reasons other than the study and underwent US and CT examinations post-mortem. RESULTS: Forty-two specimens were analysed. The most prevalent lumbosacral disc morphology was type 2 (21/42), and protrusions were found in 15/42 specimens. Abnormal echogenicity of the L5-L6 and lumbosacral discs was detected in 11/42 and 30/42 specimens, respectively. Abnormalities in the size of the L5-L6 disc were found in 10/42 specimens and correlated with promontorium localisation (Cramér's V coefficient [V] = 0.42) and lumbosacral disc morphology (type 1: V = 0.41; type 5: V = 0.69). The most prevalent orientation of the L6 spinous process (SP) was convergent (24/42). The promontoria were mostly located between L6 and S1 (36/42). Lumbosacral spondylosis was detected in 24/42 specimens and spondylolisthesis in one. Age was associated with the severity of APBCs in the sacroiliac joints, lumbosacral intertransverse joints, articular process joints, spondylosis and L6 or S1 bone plate irregularities. Lumbosacral spondylosis or L6 extremitas caudalis irregularity was associated with lumbosacral angulation, variability in the size of the L5-L6 disc and localisation of the promontorium, L6 SP orientation and L6 extremitas caudalis irregularity with abnormal echogenicity of the LS disc. MAIN LIMITATIONS: Lack of information on clinical signs. CONCLUSIONS: High variability and prevalence of the anatomical variations and APBCs in the lumbosacroiliac region were observed, and correlations between some anatomical variations and APBCs and between APBCs and age were reported.

Omics Technologies in Veterinary Medicine: Literature Review and Perspectives in Transfusion Medicine.

Background: Omics technologies represent a new analytical approach that allows a full cellular readout through the simultaneous analysis of thousands of molecules. The application of such technologies represents a flourishing field of research in human medicine, especially in transfusion medicine, while their application in veterinary medicine still needs to be developed. Summary: Omics technologies, especially proteomics, metabolomics, and lipidomics, are currently applied in several fields of human medicine. In transfusion medicine, the creation and integration of multiomics datasets have uncovered intricate molecular pathways occurring within blood bags during storage. In particular, the research has been directed toward the study of storage lesions (SLs), i.e., those biochemical and structural changes that red blood cells (RBCs) undergo during hypothermic storage, their causes, and the development of new strategies to prevent them. However, due to their challenges to perform and high costs, these technologies are hardly accessible to veterinary research, where their application dates back only to the last few years and thus a great deal of progress still needs to be made. As regards veterinary medicine, there are only a few studies that have focused mainly on fields such as oncology, nutrition, cardiology, and nephrology. Other studies have suggested omics datasets that provide important insights for future comparative investigations between human and nonhuman species. Regarding the study of storage lesions and, more generally, the veterinary transfusion field, there is a marked lack of available omics data and results with relevance for clinical practice. Key Messages: The use of omics technologies in human medicine is well established and has led to promising results in blood transfusion and related practices knowledge. Transfusion practice is a burgeoning field in veterinary medicine, but, to date, there are no species-specific procedures and techniques for the collection and storage of blood units and those validated in the human species are univocally pursued. Multiomics analysis of the species-specific RBCs' biological characteristics could provide promising results both from a comparative perspective, by increasing our understanding of species suitable to be used as animal models, and in a strictly veterinary view, by contributing to the development of animal-targeted procedures.

Presence, Tissue Localization, and Gene Expression of the Adiponectin Receptor 1 in Testis and Accessory Glands of Male Rams during the Non-Breeding Season.

Adiponectin (ADIPOQ) is a member adipocytokines, and its actions are supported by two receptors, ADIPOQ receptor 1 and -2, respectively (ADIPOR1 and -R2). Our study was performed to evaluate the ADIPOR1 presence and location and its gene expression in reproductive tissues of the male ram, during its non-breading season. The different portions of the male ram reproductive system (testis, epididymis, seminal vesicle, ampoule vas deferens, bulb-urethral gland) were collected in a slaughterhouse. Immunohistochemistry showed ADIPOR1 positive signals in the cytoplasm of all the glandular epithelial cells, with a location near the nucleus; in the testes, the positive reaction was evidenced in the cytoplasm in the basal portion of the germinal epithelial cells. The immune reaction intensity was highest (p < 0.001) in the prostate and seminal vesicles glands than that of other parts of the ram reproductive tract. RT-qPCR detected the ADIPOR1 transcript in the testes, epididymis, vas deferens, bulbourethral glands, seminal vesicles, and prostate; the expression levels were high (p < 0.01) in the prostate and low (p < 0.01) in the testis, epididymis, and bulbourethral glands. The present results evidenced the possible ADIPOQ/ADIPOR1 system's role in regulating the testicular activity of male rams during the non-breading season.

Post-Mortem Computed Tomographic Features of the Most Caudal Lumbar Vertebrae, Anatomical Variations and Acquired Osseous Pathological Changes, in a Mixed Population of Horses.

The radiographic, ultrasonographic, and scintigraphic findings of horses with thoracolumbosacral pain have been previously reported. In this study, the computed tomographic appearance of anatomical variations and pathological changes of the equine caudal lumbar region through a post-mortem examination were investigated. A total of 40 horses that had died or were submitted for euthanasia, for reasons unrelated to the study, were included in the study. From all the specimens, the modified vertebral system was adopted to evaluate and describe the four most caudal lumbar vertebrae, which were numbered from a caudal reference point (lumbosacral junction), with the segment number designated within parentheses (i.e., L(i)-L(iv)). Contact of the spinous processes was detected in 21 specimens (54%) and fusion in 6 specimens (15%). Lumbar spondylosis was seen in 17 specimens (42.5%), more commonly on the lateral aspect or on both ventral and lateral aspects in 12 specimens (71%). The presence of spondylosis was found more commonly in older horses (p < 0.001). There was no difference in bony density in specimens with spondylosis or spinous processes contact compared to specimens without. The highest prevalence of bony changes was found at L(ii)-L(i) intertransverse joints in 28 specimens (97%) on the left and in 22 specimens (96%) on the right side. Spondylolisthesis and partial fusion of the L(ii)-L(i) vertebral disc were found in association with degenerative pathologies. This study showed a high frequency of several anatomical variations and acquired osseous pathological changes in the most caudal lumbar vertebrae via a CT examination.

Leptin system is not affected by different diets in the abomasum of the sheep reared in semi-natural pastures of the Central Apennines.

The growing summer drought stress is affecting the nutritional value of pastures, no longer sufficient to support the nutritional status of sheep in extensive rearing. Adipokines affect organ and tissue functionality can be useful to evaluate animal welfare and prompt an improvement in the management of the grazing animals. Leptin (Lep) is an adipokine mainly produced by adipose tissue that regulates food intake by an anorexigenic action. Lep has also been detected in the human and rat gastrointestinal tract, where it regulates the rate of gastric emptying. In this study, Lep system was evaluated in the abomasum of 15 adult sheep reared on Apennine pastures and subjected to different diets. Until the maximum pasture flowering (MxF group), the sheep fed on fresh forage; from that moment until the maximum pasture dryness (MxD group), the experimental group (Exp group) received a feed supplementation in addition to MxD group feeding. The Lep system was investigated in the abomasum samples by immunohistochemistry (IHC) and RT-qPCR. Double-label localisation of Lep and leptin receptor (LepR) with neuroendocrine hormones was conducted to distinguish the gland cell types. The analysis performed revealed the presence of Lep and LepR in the chief and neuroendocrine cells of the fundic glands of the abomasum. RT-qPCR evidenced the transcript for Lep and LepR also identifying the long isoform (LepRb). No significant differences were observed among the three groups of sheep subjected to different diets. The abundant immunostaining observed in the fundic glands suggests that the Lep intervenes in the regulation of abomasum in sheep with a similar pattern to monogastric species while long term food supplementation seems do not influence the local function of the Lep system. A better understanding of the gastrointestinal system can contribute to improving sheep management and optimising the sustainability of livestock production.

Presence and localization of apelin and its cognate receptor in canine testes using immunohistochemical and RT-PCR techniques.

Apelin, a member of the adipokine family, is a novel endogenous peptide which regulates the male reproductive system of mammals by interacting with a specific receptor. Recent studies have highlighted that apelin may play a role in the regulation of reproduction by reducing testosterone production and inhibiting LH secretion. To the best of our knowledge, there is no available data on the presence of the apelin and its receptor in canine testes. Therefore, the aim of this study was to reveal the presence of apelin and evaluate its distribution in the canine testes using immunohistochemical and RT-PCR techniques. For this purpose, five fertile and healthy male dogs were subjected to elective orchiectomy. The immunohistochemical reaction revealed the presence of apelin and its receptor in the canine testes. Apelin was localized in spermatids and spermatozoa with a positive signal in the "acrosomal bodies". As regards the apelin receptor, a positive immunoreaction was detected in the cytoplasm of the cells localized near to the basal membrane of the seminiferous tubules and in the cytoplasm of Leydig cells. The RT-PCR analysis showed the presence of transcripts for apelin and apelin receptor in all of the samples under study. A 35kDa band confirmed apelin receptor protein expression in all of the samples analysed. In conclusion, the paracrine and endocrine role of apelin and its cognate receptor on male reproduction reported in humans and laboratory animals could also be hypothesized in dogs.

Age and sex correlate with bony changes and anatomic variations of the lumbosacroiliac region of the vertebral column in a mixed population of horses.

OBJECTIVE: To provide a postmortem description of anatomic variations and changes of the lumbosacroiliac region in horses. The authors hypothesized that lesion severity would increase with age and body weight and correlate to anatomic variations. SAMPLES: Lumbosacroiliac vertebral specimens from 38 horses (mean age, 16 years; range, 5 to 30 years) that died or were euthanized for reasons unrelated to the study between November 2019 and October 2021. PROCEDURES: The lumbosacroiliac region of the vertebral column was removed from each cadaver. After dissection, disarticulation, and boiling, the anatomic specimens were examined for anatomic variations and osseous changes of the articular process joints (APJs), intertransverse joints (ITJs), and sacroiliac joints (SIJs). The lengths of L6-S1 intertransverse articular surfaces were measured and their ratios calculated. Descriptive statistics were obtained, and the χ2 test was used to assess differences in anatomic variations and abnormal changes of the APJs, ITJs, and SIJs. RESULTS: The most common anatomic variation was a sacrum-like shape of the transverse processes of L6 (29/38 [76%]) and converging orientation of dorsal spinous process of L6 (33/38 [87%]). The highest prevalence of bony changes was detected at L5-L6 (right, 34/38 [89%]; left, 33/38 [87%]) and L6-S1 APJs (right, 38/38 [100%]; left, 37/38 [97%]) and at SIJs (right, 32/38 [86%]; left, 31/38 [82%]). The shape of L6 transverse processes differed between breed (P = .01) and was associated with presence of L4-L5 ITJs (P < .01). CLINICAL RELEVANCE: Age and sex were associated with changes of the sacral dorsal spinous processes, ITJs, and APJs. The clinical significance of these findings could not be confirmed based on the study limitations.

Metabolic milieu and localization of ovarian leptin and receptor in queens under different reproduction phases.

Metabolic profile and leptin presence and expression of receptor in the ovary of 8 queens were analyzed. All queens were assessed for nutritional state and underwent ovariectomy in accordance with Italian law for the control of stray cats and dogs. Three queens were at early gestation, three prepubescent and two non-pregnant. All animals appeared healthy but underweight. Haemogram and biochemical profile did not show to be of prognostic value. On microscopy, granulosa cells of secondary follicles tested positive to leptin in pregnant queens, while testing negative in non-pregnant ones. Muscular layer of vessels of the ovary in non-pregnant queens tested positive to receptor, but negative in those found pregnant. Results suggest that leptin is produced and receptor expressed in the different tissues of the ovary in underweight queens. Additionally, the expression of receptor did not necessarily imply local leptin production and clearly displayed to vary according to the reproduction phase. Taken altogether, these findings allow to conclude that the expression of receptor can be interpreted as the pillar bridging ovary tissues as target sites of circulating leptin produced elsewhere. Arguably, ovary removal may affect hormonal cross-talk at the systemic level, likely affecting different systemic functions, including the nutritional status.

Pharmacokinetics of cannabidiol following single oral and oral transmucosal administration in dogs.

Introduction: In the last few years, different formulations containing cannabidiol (CBD) were tested with regard to its efficacy on chronic pain, refractory epilepsy, anxiety, aggressive behavior and atopic dermatitis in dogs. CBD is generally administered orally, but its low bioavailability, probably due to a first-pass metabolism, represents a great limitation. The aim of this study was to evaluate if CBD bioavailability increases after oral transmucosal administration (OTM) compared to oral treatment. Methods: Twelve dogs diagnosed with mild chronic pain were enrolled in the study and treated once orally or OTM (6 dogs/group) with a pure CBD in oil formulation at a dosing rate of 1 mg/kg b.w. At prefixed time points, blood samples were collected to define CBD plasma concentrations vs. time profiles, and the main pharmacokinetics parameters were obtained by non-compartmental model. Results: CBD Cmax, Tmax, terminal half-life and AUC0 - t were 206.77 ± 167 and 200.33 ± 158.33 ng/mL, 2.17 ± 0.98 and 1.92 ± 1.11 h, 2.67 ± 0.53 and 2.62 ± 0.64 h, 647.51 ± 453.17, and 536.05 ± 370.21 h*ng/mL, following oral and OTM administration, respectively. No significant difference in pharmacokinetic parameters were observed between treatments. Discussion: The OTM administration did not increase cannabidiol bioavailability compared to oral treatment. The almost perfectly superimposable mean plasma concentrations of cannabidiol following the two treatments suggests that CBD is not able to be adsorbed by the oral mucosa or that its absorption is very scarce, and that CBD is swallowed and absorbed in the gastrointestinal tract.

The Apelinergic System Immuno-Detection in the Abomasum and Duodenum of Sheep Grazing on Semi-Natural Pasture.

Apelin (APLN) is an adipokine mainly produced by adipose tissue and related to an individual's nutritional status as well as digestive apparatus functions. In this work, APLN and its receptor (APLNR) were investigated, by immunohistochemistry, in the abomasum and duodenum of 15 Comisana × Appenninica adult sheep reared in a semi-natural pasture. Organ samples were collected after maximum pasture flowering (M × F group) and after maximum pasture dryness (M × D group); the experimental group (E × p group) received a feed supplementation of 600 grams/day/head of barley and corn in addition to M × D group feeding. APLN and APLNR were identified in the lining epithelium and the fundic gland chief cells of the abomasum. APLNR was observed in the lining epithelium, in the crypts and the serotonin secreting cells of the duodenum. Similar reactivity was observed between the M × F and E × p groups, while the M × D group showed a lower intensity of immunostaining for both APLN and APLNR in all positive structures but the duodenal serotonin neuroendocrine cells. Hence, our findings show that the E × p group presents a picture quite overlapped with M × F and suggest that food supplementation has a maintaining effect on the apelinergic system expression in the investigated digestive tracts of the sheep.

Effects of Obesity on Adiponectin System Skin Expression in Dogs: A Comparative Study.

Obesity is an important health issue in dogs since it influences a plethora of associated pathologies, including dermatological disorders. Considering the scarcity of information in pets, this work aimed to evaluate the localization and expression of adiponectin (ADIPOQ) and its two receptors (ADIPOR1 and ADIPOR2) in the skin of 10 obese dogs, compared with serum ADIPOQ level. Through immunohistochemistry, ADIPOQ and ADIPOR2 were observed in the adipose tissue, sweat and sebaceous glands, endothelium, and some connective cells. Both receptors were observed in the epidermis and the hair follicles, other than in the sweat and sebaceous glands. Real-time PCR evidenced that the ADIPOQ and ADIPOR2 transcripts were expressed 5.4-fold (p < 0.01) and 2.3-fold less (p < 0.01), respectively, in obese than in normal weight dogs, while ADIPOR1 expression did not change. Obese dogs showed lower serum ADIPOQ levels than the normal weight group. Accordingly, ADIPOQ and ADIPOR2 expression in the skin appear negatively correlated with obesity in the same way as the serum ADIPOQ level. These findings evidence that ADIPOQ system changes in the skin of obese dogs and suggest that the ADIPOQ effect on the skin is at least in part regulated by the reduced expression of ADIPOR2.

Effects of Orexin B on Swine Granulosa and Endothelial Cells.

In addition to the well-known central modulatory role of orexins, we recently demonstrated a peripheral involvement in swine granulosa cells for orexin A and in adipose tissue for orexin B (OXB). The aim of present research was to verify immunolocalization of OXB and its potential role in modulating the main features of swine granulosa cells. In particular, we explored the effects on granulosa cell proliferation (through the incorporation of bromodeoxyuridine), cell metabolic activity (as indirect evaluation by the assessment of ATP), steroidogenic activity (by immunoenzymatic examination) and redox status (evaluating the production of superoxide anion by means of the WST test, production of nitric oxide through the use of the Griess test and the non-enzymatic reducing power by FRAP test). Our data point out that OXB does not modify granulosa cell growth, steroidogenesis and superoxide anion generation. On the contrary, the peptide stimulates (p < 0.05) nitric oxide output and non-enzymatic reducing power. Since new vessel growth is crucial for ovarian follicle development, a further aim of this study was to explore the expression of prepro-orexin and the effects of OXB on swine aortic endothelial cells. We found that the peptide is ineffective in modulating cell growth, while it inhibits redox status parameters. In addition, we demonstrated a stimulatory effect on angiogenesis evaluated in fibrin gel angiogenesis assay. Taken together, OXB appears to be potentially involved in the modulation of redox status in granulosa and endothelial cells and we could argue an involvement of the peptide in the follicular angiogenic events.

Effects of Dietary Polyphenols from Olive Mill Waste Waters on Inflammatory and Apoptotic Effectors in Rabbit Ovary.

The aim of this study was to evaluate the effect of dietary polyphenols on the expression of the effectors involved in inflammation and apoptosis in rabbit ovary. New Zealand White female rabbits were fed a basal control diet (CTR), or the same diet supplemented with a polyphenolic concentrate (POL, 282.4 mg/kg) obtained from olive mill waste waters. The follicle counts and the relative mRNA (RT-qPCR) and protein (immunohistochemistry) expression of the effectors involved in inflammation (cyclooxygenase-2; interleukin-1beta; tumor necrosis factor-alpha, TNFA) and apoptosis (BCL2-associated X protein, BAX), detected in the ovaries of both groups, were examined. The POL diet increased the primary and total follicles number. Cyclooxygenase-2 gene expression was higher (p < 0.05) in the POL group than in the CTR group, whereas BAX was lower (p < 0.05) in POL than CTR. Immunohistochemistry revealed the presence of all the proteins examined, with weaker (p < 0.05) COX2 and BAX signals in POL. No differences between the CTR and POL groups were observed for IL1B and TNFA gene and protein expression. These preliminary findings show that dietary polyphenols modulate inflammatory and apoptotic activities in rabbit ovary, regulating cyclooxygenase-2 and BAX expression, thus suggesting a functional involvement of these dietary compounds in mammalian reproduction.

Effects of Dietary Zn/Se and α-Tocopherol Supplementation on Metabolic Milieu, Haemogram and Semen Traits of Breeding Stallions.

Trace element status and metabolic milieu are sometimes overlooked in common veterinary clinical practice across animal species. The evaluation of requirements of trace elements, in fact, may be useful to prevent the perturbation of tissue-specific metabolic impair. In particular, essential trace elements in the diet play key roles within sub-cellular metabolic patterns with macro effects at the systemic level, like blood cell stability and semen quality. This effect was studied in breeding stallions, in which semen quality and haemogram are important for reproduction. A case-control feeding trial involved 40 stallions (age: 8-21 years; body weight, BW: 510-531 kg) of one stud centre, allotted to two experimental groups (n = 20 control, CON vs. n = 20 supplemented, SUPPL100), following a matched-pairs approach based on age. Supplemented stallions (SUPPL100) received a mixed mineral and vitamin supplement of Zn/Se and α-tocopherol (α-TOH) (100 g/day stallion) to compound feed, fed as control diet to horses of the control group (CON). Horses resulted deficient in circulating α-TOH and Zn at the start, though clinically healthy. After supplementation, different plasmatic levels of α-TOH, Zn and Se were found between groups. Circulating basophils (BASO) and mean cell haemoglobin concentration (MCHC) were affected by the dietary treatment (p < 0.05). Plasmatic Se affected monocyte count, haematocrit, mean cell volume and mean cell haemoglobin concentration. Semen traits were not affected by the dietary treatment per se, except for mobile/progressive sperm cells (%) of stallions aged > 13 years marginal circulating levels of α-TOH (p = 0.04). Ameliorating the micromineral status showed to improve the haemogram of stallions in view of circulating levels of Cu. Semen quality appeared to be strongly dependent on animal effects.

Leptin System in Obese Dog Skin: A Pilot Study.

Obesity predisposes to several health problems including skin diseases. However, information on the relationship between obesity and skin disorders in pets is very scarce. Leptin (LEP) is mainly produced by adipose tissue and has a prominent role in skin biology. This study evaluated the LEP system in the skin of obese dogs compared to normal-weight animals. The investigation was carried out on 10 obese (Obese group) and 10 normal-weight (Normal-weight group) dogs through Real-time PCR and immunohistochemistry. Cells of skin associated immune system were also evaluated. No differences were evidenced between the two groups as well as skin inflammation. LEP differences were no significant, while LEPR transcript appeared 10-fold higher in obesedogs than in normal-weight ones. Immunostaining for both molecules was observed in several skin structures such as the epidermis, hair follicles, and glands. No differences appeared in the skin associated immune system composition. This study is a preliminary report showing that LEP system changes in obese dog skin. The increased LEPR expression observed in the obese group suggests that the receptor plays a modulating role in the system control. However, the exact role of LEPin the skin under obesity conditions needs further elucidation.

Seasonal Expression of NGF and Its Cognate Receptors in the Ovaries of Grey Squirrels (Sciurus carolinensis).

The grey squirrel is an invasive alien species that seriously threatens the conservation of the native red squirrel species. With the aim of characterizing the reproductive physiology of this species due to its great reproductive success, the function of the ovarian nerve growth factor (NGF) system was analyzed in a grey squirrel population living in central Italy. During the breeding and nonbreeding seasons, the ovarian presence, distribution, and gene expression of NGF, neurotrophic tyrosine kinase receptor 1 (NTRK1), and nerve growth factor receptor (NGFR), as well as NGF plasma concentrations, were evaluated in female grey squirrels. NGF was found in the luteal cells and in the thecal and granulosa cells of follicles, while NTRK1 and NGFR were only observed in follicular thecal and granulosa cells. NGF and NGFR transcripts were almost two-fold greater during the breeding season, while no seasonal differences were observed in NTRK1 gene expression. During the breeding season, NGFR was more expressed than NTRK1. Moreover, no changes were observed in NGF plasma levels during the reproductive cycle. The NGF system seems to be involved in regulating the ovarian cycle mainly via local modulation of NGF/NGFR, thus playing a role in the reproductive physiology of this grey squirrel population.

Influence of Different Feed Physical Forms on Mandibular Gland in Growing Pigs.

A study was performed on the mandibular gland obtained from growing pigs enrolled in a wide research project aiming to test the effects of different feed physical forms on animal health, production and welfare. We used 48 pigs fed for four weeks with different dietary treatments based on different grinding intensities and compactions of the same diet, namely coarsely ground meal (CM), finely ground pelleted (FP) and coarsely ground pelleted (CP) diets. Samples were analyzed by conventional histochemistry to identify the glycohistochemical profile and by immunohistochemistry to localize aquaporin 5, apelin and apelin receptor. Statistical elaborations were performed using the Stats R-package, version 3.5.3. Pig mandibular gland adenomere increased both the quantity and acidity of produced glycoconjugates from CM to FP and CP diets. This probably calls forth higher watery saliva, thus promoting a better feed softening facilitating the amalgamation of the bolus. Mandibular gland increased aquaporin 5 positivity in the CP diet, supporting the hypothesis of an augmented demand for water. Based on apelin/receptor localization, it was hypothesized that in pig mandibular gland the apelinergic system likely performs an endocrine control on the demilunes activity and a paracrine control on ducts, facilitating the production of a more fluid saliva.