Platelets Rich Plasma Increases Antioxidant Defenses of Tenocytes via Nrf2 Signal Pathway.

Tendinopathies are common disabling conditions in equine and human athletes. The etiology is still unclear, although reactive oxygen species (ROS) and oxidative stress (OS) seem to play a crucial role. In addition, OS has been implicated in the failure of tendon lesion repair. Platelet-rich plasma (PRP) is rich in growth factors that promote tissue regeneration. This is a promising therapeutic approach in tendon injury. Moreover, growing evidence has been attributed to PRP antioxidant effects that can sustain tissue healing. In this study, the potential antioxidant effects of PRP in tenocytes exposed to oxidative stress were investigated. The results demonstrated that PRP reduces protein and lipid oxidative damage and protects tenocytes from OS-induced cell death. The results also showed that PRP was able to increase nuclear levels of redox-dependent transcription factor Nrf2 and to induce some antioxidant/phase II detoxifying enzymes (superoxide dismutase 2, catalase, heme oxygenase 1, NAD(P)H oxidoreductase quinone-1, glutamate cysteine ligase catalytic subunit and glutathione, S-transferase). Moreover, PRP also increased the enzymatic activity of catalase and glutathione S-transferase. In conclusion, this study suggests that PRP could activate various cellular signaling pathways, including the Nrf2 pathway, for the restoration of tenocyte homeostasis and to promote tendon regeneration and repair following tendon injuries.

Proteome Alterations in Equine Osteochondrotic Chondrocytes.

Osteochondrosis is a failure of the endochondral ossification that affects developing joints in humans and several animal species. It is a localized idiopathic joint disorder characterized by focal chondronecrosis and growing cartilage retention, which can lead to the formation of fissures, subchondral bone cysts, or intra-articular fragments. Osteochondrosis is a complex multifactorial disease associated with extracellular matrix alterations and failure in chondrocyte differentiation, mainly due to genetic, biochemical, and nutritional factors, as well as traumas. This study describes the main proteomic alterations occurring in chondrocytes isolated from osteochondrotic cartilage fragments. A comparative analysis performed on equine osteochondrotic and healthy chondrocytes showed 26 protein species as differentially represented. In particular, quantitative changes in the extracellular matrix, cytoskeletal and chaperone proteins, and in cell adhesion and signaling molecules were observed in osteochondrotic cells, compared to healthy controls. Functional group analysis annotated most of these proteins in "growth plate and cartilage development", while others were included in "glycolysis and gluconeogenesis", "positive regulation of protein import", "cell-cell adhesion mediator activity", and "mitochondrion nucleoid". These results may help to clarify some chondrocyte functional alterations that may play a significant role in determining the onset and progression of equine osteochondrosis and, being related, of human juvenile osteochondrosis.

Ultrasonographic findings in 38 horses with septic arthritis/tenosynovitis.

Septic arthritis/tenosynovitis in the horse can have life-threatening consequences. The purpose of this cross-sectional retrospective study was to describe ultrasound characteristics of septic arthritis/tenosynovitis in a group of horses. Diagnosis of septic arthritis/tenosynovitis was based on historical and clinical findings as well as the results of the synovial fluid analysis and/or positive synovial culture. Ultrasonographic findings recorded were degree of joint/sheath effusion, degree of synovial membrane thickening, echogenicity of the synovial fluid, and presence of hyperechogenic spots and fibrinous loculations. Ultrasonographic findings were tested for dependence on the cause of sepsis, time between admission and beginning of clinical signs, and the white blood cell counts in the synovial fluid. Thirty-eight horses with confirmed septic arthritis/tenosynovitis of 43 joints/sheaths were included. Degree of effusion was marked in 81.4% of cases, mild in 16.3%, and absent in 2.3%. Synovial thickening was mild in 30.9% of cases and moderate/severe in 69.1%. Synovial fluid was anechogenic in 45.2% of cases and echogenic in 54.8%. Hyperechogenic spots were identified in 32.5% of structures and fibrinous loculations in 64.3%. Relationships between the degree of synovial effusion, degree of the synovial thickening, presence of fibrinous loculations, and the time between admission and beginning of clinical signs were identified, as well as between the presence of fibrinous loculations and the cause of sepsis (P ≤ 0.05). Findings indicated that ultrasonographic findings of septic arthritis/tenosynovitis may vary in horses, and may be influenced by time between admission and beginning of clinical signs.

Cytotoxicity of local anaesthetics and protective effects of platelet rich plasma on equine tenocytes: An in vitro study.

Local anaesthetics (LAs) can have detrimental effects on rat, bovine, canine, and human tendon tissues and cells. Currently, there has been no available data on the impact of these drugs on equine tenocytes. Even if LA injection for managing painful tendon conditions in horses is limited, it is usually used via intra-articular, intrasynovial, perineural, and intrathecal as well as for lameness examinations. In this in vitro study, the cytotoxic effects of LAs, including lidocaine, mepivacaine, and bupivacaine on equine tenocytes, in the presence and absence of platelet rich plasma (PRP), were investigated. PRP accelerates tissue healing and can exert cytoprotective effects on different cell types exposed to different stressful conditions, including drugs. Results indicated that the exposure to LAs significantly reduced tenocytes viability in dose- and time-dependent manners while PRP was able to counteract their cytotoxic effects. Furthermore, microscopy and flow cytometry analyses revealed apoptosis and necrosis in equine tenocytes exposed to these drugs, that were both reduced when PRP was in the medium. These findings highlight the importance of considering the tenocyte toxicity associated with intrathecal and intraneural LA injections, as they might affect tenocytes or reduce the efficacy of associated therapies. Moreover, this study also highlights the protective effects of PRP, which could make LA injections safer.

Adult-onset cyclic esotropia: a case report.

PURPOSE: This case report presents the diagnosis and management of an adult patient with cyclic esotropia, a rare and poorly understood form of strabismus, that is characterized by alternating periods of manifest strabismus and orthotropia. Few cases of adult-onset cyclic strabismus have been reported. The etiology, pathogenesis, and treatment for this condition are reviewed. CASE REPORT: A 37-year-old man with high unilateral myopia and anisometropic amblyopia in his left eye developed cyclic esotropia 2 months after a scleral buckle procedure for a retinal detachment. A 48-hour cycle that consisted of a 24-hour period of orthotropia followed by a 24-hour period of constant left esotropia was present. Magnetic resonance imaging, electromyography, electroencephalography, and ocular and orbital echography were unremarkable, as was a neurological assessment that included fatigue and edrophonium testing. The patient was successfully managed with muscle surgery. CONCLUSIONS: The etiology and pathogenesis of adult-onset cyclic esotropia remain unknown. Cyclic esotropia should be considered in the differential diagnosis when an individual presents with a history of intermittent strabismus that is present some days and not others.

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.

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.

Animal models of Soft Tissue Sarcoma for alternative anticancer therapy studies: characterization of the A-72 Canine Cell Line.

Canine Soft Tissue Sarcoma (STS) cell line A-72 has been largely employed for antiviral and antiproliferative studies. However, there are few information on their characteristics. Our aim was to evaluate A-72 expression level of genes and proteins involved in the innate immune response and cell cycle, their ability to respond to infective stressors and their possible use as a cellular model for anti-cancer studies in human and animal medicine. For this purpose, we evaluated the basal expression of immune-related, cell cycle and DNA repair genes on this cell line and tumoral tissues. A-72 ability to respond to a wild-type strain of Salmonella typhimurium was assessed. S. typhimurium showed ability to penetrate A-72 causing pro-inflammatory response accompanied by a decrease of cell viability. IL10 and IL18 genes were not expressed in A-72 while CXCL8, NOS2, CXCR4 and PTEN were highly expressed in all samples and TP53 was slightly expressed, as shown in human STS. Our results outline the ability of A-72 to respond to a bacterial agent by modifying the expression of important genes involved in innate immune response and provide a useful model for in vitro evaluation of new therapeutic approaches that could be translated into the human oncology.

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.

Radiographic findings and anatomical variations of the caudal cervical area in horses with neck pain and ataxia: case-control study on 116 horses.

BACKGROUND: Abnormalities of the ventral lamina of the sixth cervical vertebra (AVL-C6) are thought to exert abnormal stress on the articular process joints (APJs) of the cervicothoracic junction. The aim of the study was to investigate the association between AVL-C6 and radiographic findings in the caudal cervical area and between clinical signs of neck pain and ataxia and radiographic findings. METHODS: Medical records of horses subjected to cervical radiography were reviewed. Horses were classified into those with neck pain (group C), those with ataxia (group A) and healthy horses (group H). Presence of AVL-C6 and increased size, dysplasia, remodelling, fragmentation and osteochondral fragment at the APJs (C5-T1) were recorded. Univariable logistic regression analysis was performed to identify the associations between explanatory and dependent variables. Variables with P<0.2 were included in the multivariable analysis. RESULTS: One hundred and sixteen horses were included (44 in group C, 29 in group A, 66 in group H); 24 of 116 horses had radiographic AVL-C6. Age, AVL-C6 and overall/C6-C7 increase in size remained in the final models. CONCLUSIONS: The presence of AVL-C6 and moderate/severe increase in size of the caudal cervical APJs increase the odds of showing neck pain and, if severe, ataxia.

3D Biomimetic Porous Titanium (Ti6Al4V ELI) Scaffolds for Large Bone Critical Defect Reconstruction: An Experimental Study in Sheep.

The main goal in the treatment of large bone defects is to guarantee a rapid loading of the affected limb. In this paper, the authors proposed a new reconstructive technique that proved to be suitable to reach this purpose through the use of a custom-made biomimetic porous titanium scaffold. An in vivo study was undertaken where a complete critical defect was experimentally created in the diaphysis of the right tibia of twelve sheep and replaced with a five-centimeter porous scaffold of electron beam melting (EBM)-sintered titanium alloy (EBM group n = 6) or a porous hydroxyapatite scaffold (CONTROL group, n = 6). After surgery, the sheep were allowed to move freely in the barns. The outcome was monitored for up to 12 months by periodical X-ray and clinical examination. All animals in the CONTROL group were euthanized for humane reasons within the first month after surgery due to the onset of plate bending due to mechanical overload. Nine months after surgery, X-ray imaging showed the complete integration of the titanium implant in the tibia diaphysis and remodeling of the periosteal callus, with a well-defined cortical bone. At 12 months, sheep were euthanized, and the tibia were harvested and subjected to histological analysis. This showed bone tissue formations with bone trabeculae bridging titanium trabeculae, evidencing an optimal tissue-metal interaction. Our results show that EBM-sintered titanium devices, if used to repair critical bone defects in a large animal model, can guarantee immediate body weight-bearing, a rapid functional recovery, and a good osseointegration. The porous hydroxyapatite scaffolds proved to be not suitable in this model of large bone defect due to their known poor mechanical properties.

Reproductive Traits of an Invasive Alien Population of Grey Squirrel (Sciurus carolinensis) in Central Italy.

The reproductive cycle of an invasive alien Italian grey squirrel population was studied to understand its adaptation and limit its spread, in order to conserve the autochthonous red squirrel. Female and male genital traits were evaluated throughout the reproductive cycle, including the ovary, uterus, testicle, epididymis, seminiferous tubule morphometry, and germinative epithelium histology. Moreover, individual female fecundity was determined by counting uterine scars. Ovary width and uterus weight, length, and width reached their highest values in the luteal and pregnancy phases. On conducting a histological evaluation of the testicular germinal epithelium, four morphotypes related to the different reproductive phases of the male squirrels were identified: immature, pubertal, spermatogenesis, and regressive. Testicle and epididymis weights and seminiferous tubule diameters reached their largest values during spermatogenesis. Uterine scar analysis showed that 69% of the females had given birth to one or two litters, while 31% had no uterine scars. Litters were larger in the first breeding period than in the second; annual fecundity was 4.52 ± 1.88 uterine scars/female. Umbrian grey squirrels have adapted to their non-native range, showing two annual mating periods at times similar to those in their native range, and high reproductive success.

Protective effects of platelet-rich plasma against lidocaine cytotoxicity on canine articular chondrocytes.

BACKGROUND: Lidocaine (LD) is one of the most commonly used local anesthetics for performing arthroscopic surgery and managing of osteoarthritic pain in both human and veterinary medicine. However, over the last years, several studies have focused on the chondrotoxic effects of LD. In order to ensure that intra-articular lidocaine is safe to use, treatments aimed at mitigating chondrocyte death have recently been investigated. The aim of this study is to evaluate the possible protective effects of platelet-rich plasma (PRP) against LD cytotoxicity on canine articular chondrocytes. RESULTS: Articular canine chondrocytes, were exposed to 1% or 1.8% LD alone or in co-presence with 10% PRP for 30 min. In order to evaluate the effects of PRP pre-treatments, experiments were carried out on cells cultured in serum-free medium-or in medium supplemented with 10% PRP or 10% fetal bovine serum. Cell viability was evaluated by methyl thiazolyl tetrazolium assay and cell apoptosis was analyzed by flow cytometry using annexin V-fluorescein isothiocyanate/propidium iodide. The results showed that LD significantly reduced canine chondrocytes viability, probably due to apoptosis induction. Pre-treatment or the co-presence of PRP in the media restored the number of viable chondrocytes. The PRP also seemed to protect the cells from LD-induced apoptosis. CONCLUSIONS: Pre-treatments and/or the simultaneous administration of PRP reduced LD-induced cytotoxicity in canine chondrocytes. Further in vivo studies are required to determine whether PRP can be used as a save protective treatment for dogs receiving intra-articular LD injections.

Heart rate and blood pressure variations after transvascular patent ductus arteriosus occlusion in dogs.

The objective of the study was to retrospectively analyse the cardiovascular effects that occurs following the transvascular occlusion of patent ductus arteriosus in dogs. Sixteen anaesthesia records were included. Variables were recorded at the time of placing the arterial introducer, occlusion of the ductus, and from 5 to 60min thereafter, including, among the other, heart rate, systolic, diastolic and mean arterial blood pressure. The maximal percentage variation of the aforementioned physiological parameters within 60min of occlusion, compared with the values recorded at the introducer placing, was calculated. The time at which maximal variation occurred was also computed. Correlations between maximal percentage variation of physiological parameters and the diameter of the ductus and systolic and diastolic flow velocity through it were evaluated with linear regression analysis. Heart rate decreased after occlusion of the ductus with a mean maximal percentage variation of 41.0±14.8% after 21.2±13.7min. Mean and diastolic arterial blood pressure increased after occlusion with a mean maximal percentage variation of 30.6±18.1 and 55.4±27.1% after 19.6±12.1 and 15.7±10.8min, respectively. Mean arterial blood pressure variation had a significant and moderate inverse correlation with diastolic and systolic flow velocity through the ductus. Transvascular patent ductus arteriosus occlusion in anaesthetised dogs causes a significant reduction in heart rate and an increase in diastolic and mean blood arterial pressure within 20min of closure of the ductus.

Intraperitoneal injection of microencapsulated Sertoli cells restores muscle morphology and performance in dystrophic mice.

Duchenne muscular dystrophy (DMD) is a genetic disease characterized by progressive muscle degeneration leading to impaired locomotion, respiratory failure and premature death. In DMD patients, inflammatory events secondary to dystrophin mutation play a major role in the progression of the pathology. Sertoli cells (SeC) have been largely used to protect xenogeneic engraftments or induce trophic effects thanks to their ability to secrete trophic, antiinflammatory, and immunomodulatory factors. Here we have purified SeC from specific pathogen-free (SPF)-certified neonatal pigs, and embedded them into clinical grade alginate microcapsules. We show that a single intraperitoneal injection of microencapsulated SPF SeC (SeC-MC) in an experimental model of DMD can rescue muscle morphology and performance in the absence of pharmacologic immunosuppressive treatments. Once i.p. injected, SeC-MC act as a drug delivery system that modulates the inflammatory response in muscle tissue, and upregulates the expression of the dystrophin paralogue, utrophin in muscles through systemic release of heregulin-ß1, thus promoting sarcolemma stability. Analyses performed five months after single injection show high biocompatibility and long-term efficacy of SeC-MC. Our results might open new avenues for the treatment of patients with DMD and related diseases.

MODELLI CHIMICI DEL VIVENTE LE ORIGINI DEL CENCETTO DI «MACCHINA CHIMICA¼.

The concept of "chemical machine" is currently very much in use in studies of the physiology of living bodies especially at the level of their intimate structure. Its appearance, however, has a long history that begins as early as the sixteenth century, when it became apparent that the living body was the abode of complex and fundamental chemical processes. When this awareness met the equally widespread idea that living bodies could be described as "machinery", the problem emerged of reconciling the two approaches, which gave origin to the concept of "chemical machine."

Effects of intraperitoneal injection of microencapsulated Sertoli cells on chronic and presymptomatic dystrophic mice.

We report data about the effects of intraperitoneal (i.p.) injection of specific pathogen-free (SPF) porcine Sertoli cells (SeC) encapsulated into clinical grade alginate-based microcapsules (SeC-MC) on muscles of chronic and presymptomatic dystrophic, mdx mice. Mdx mouse is the best characterized animal model of Duchenne muscular dystrophy (DMD), an X-linked lethal myopathy due to mutation in the gene of dystrophin, which is crucial for myofiber integrity during muscle contraction. Our data show that three weeks after i.p. injection of SeC-MC significantly reduced adipose and fibrous tissue deposition, reduced macrophage infiltrate, and reduced numbers of damaged myofibers are found in muscles of 12-month-old mdx mice, which reproduce chronic DMD conditions. Compared with muscles of mock-treated mdx mice muscles of SeC-MC-treated mice show upregulation of the dystrophin paralogue, utrophin which is localized to the periphery of myofibers. Moreover, our data show that i.p. injection of SeC-MC into presymptomatic, 2-week-old mdx mice, although not fully preventing myofiber degeneration, results in protection against myofiber necrosis and muscle inflammation. Extensive discussion of these data can be found in Ref. [1].

Radiographic evaluation of changes in the proximal phalanx of Thoroughbreds in race training.

OBJECTIVE: To evaluate changes in the cortical bone of the proximal phalanx of the fore-limbs of Thoroughbreds in response to training. ANIMALS: Twenty-seven 2-year-old Thoroughbreds (20 females, 2 males, and 5 geldings). PROCEDURES: Horses were principally in training for races in a straight line and in a clockwise direction. Lateromedial and dorsopalmar radiographic views of each metacarpophalangeal joint were obtained before the horses started training and 1 year after starting exercise and racing. Width of the dorsal, palmar, lateral, and medial cortex and the width and thickness of the medulla were measured. Ratios (rather than absolute values) were used to remove the effect of differences in bone size among horses. RESULTS: 10 horses were lost from the study. Radiographs were obtained for 17 horses 1 year after starting training (9 horses raced in a clockwise direction, and 8 raced in clockwise and counterclockwise directions). There was no difference between the cortical bone in the right and left forelimbs at the start of the study. After training for 1 year, the palmar cortex in the right forelimb was significantly thicker than that in the left forelimb. CONCLUSIONS AND CLINICAL RELEVANCE: The strain patterns, biomechanics of rapid exercise, and type of training most probably determined differences in the adaptive responses of the proximal phalanx. The data reported here can be used in the evaluation of weight-bearing distribution along the proximal phalanx and evaluation of the relationship between exercise and bone remodelling of the proximal phalanx.