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Kenney-Doig scale is considered the international standard method for classifying uterine biopsies in mares; however, its objectivity has been questioned by various studies. In the present study, we analysed the degree of agreement between two pathologists when assessing the same set of 201 uterine biopsies, obtaining a slight to moderate level of agreement (κ = .34/κ w  = .57). Subsequently, we developed a numerical scale based on the evaluation of histological parameters, including inflammation, fibrosis, glandular density and lymphatic lacunae. Partial scores were summed to obtain a fifth parameter called Summation. The correlation between both scales was demonstrated (p < .0001), and their combined use resulted in a notable increase in the degree of agreement between the two pathologists (κ = .53/κ w  = .67)., (© 2024 The Author(s). Reproduction in Domestic Animals published by Wiley‐VCH GmbH.)

Prostaglandins have many roles in the equine reproductive tract, including but not limited to luteolysis, luteal support, ovulation, transport through the uterine tube, uterine contraction, embryonic mobility, inflammation, and fibrosis. Altered secretion of inflammatory proteins are likely to disrupt the balance of endometrial function and could impair fertility. Our overall goal was to measure the expression of several prostaglandin- and inflammation-related genes in mares with different degrees of endometrial histological changes. Our hypothesis was that mares with neutrophilic and lymphocytic plasmocytic inflammation, fibrosis, or different biopsy grades would have altered concentrations of prostaglandin E2 (PGE2) and F2α (PGF2α), as well as altered expression of inflammation- and prostaglandin-related genes, compared to mares with minimal to no histological changes on biopsy evaluation. Forty-five endometrial biopsies from estrous mares were assessed by a reproductive pathologist for the degree of neutrophilic inflammation, lymphocytic and plasmocytic inflammation, and fibrosis, and a biopsy grade was assigned based on the Kenney-Doig system. A low-volume uterine lavage was collected from a subset of twenty-six mares prior to biopsy collection and was used to measure PGE2 and PGF2α concentrations via ELISA. Total RNA was extracted from biopsies and mRNA expression was evaluated for twenty-five genes of interest. A restricted maximum likelihood linear model was used to compare differences of mRNA expression, with a statistical significance set at P < 0.05. There was no difference in the abundance of PGE2 or PGF2α between any of the variables tested. Mares with endometrial biopsy grade I had lower expression of NF-kB, PTGS1 and HPGD compared to grade IIA or IIB (P < 0.05). Mares with neutrophilic inflammation had decreased expression of NF-kB, PTGS1, PTGER4, CBR1, mPGES2 and PTGIS compared to mares without inflammation. Mares with mild or minimal endometrial fibrosis had increased expression of mPGES2 and PTGIS, compared to mares with moderate endometrial fibrosis. In conclusion, several genes were identified to be differentially expressed in mares with histological changes compared to mares with no to minimal histological changes. The presence of inflammation and fibrosis may alter the concentration of prostaglandins in endometrial tissue, which could impair many of the uterine reproductive and immune functions during estrus, affecting early embryo survival., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

There is limited published data regarding cardiovascular disease in nondomestic felid populations. To address this knowledge gap, necropsy cases of tigers and lions with representative myocardial samples submitted to a diagnostic laboratory were histologically assessed with hematoxylin and eosin and Sirius red stains. A total of 32 submissions (15 tigers, 17 lions) were identified in a 4-year period. All tigers and lions had some degree of coronary artery lesions in the left ventricle and/or interventricular septum. Major findings included moderate to marked arteriosclerosis in 8 tigers (53%) and 4 lions (24%) and moderate to marked perivascular fibrosis in 10 tigers (67%) and 9 lions (53%). Moreover, 10 tigers (67%) and 8 lions (47%) had coronary artery lesions with variable degrees of perivascular cardiomyocyte degeneration and/or loss. To our knowledge, this is the first report describing coronary artery pathology in captive tigers and lions., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Equine endometrosis is a degenerative and predominantly fibrotic condition resulting from progressive and irreversible multifactorial causes that influence the endometrium of mare. Tissue remodeling in the equine endometrium occurs as part of the pathogenesis of endometrosis, a process characterized by a shift in extracellular matrix (ECM) components. The relationship between matrix metalloproteinases and their specific inhibitors is crucial for the remodeling process. Collagen play a significant role in maintaining a healthy uterus and may promote fibrotic processes. The aim of this study was to quantify endometrial collagen deposition using picrosirius 25 red (PSR) staining, and to evaluate gene expression of collagen type 2 (COL-2) and 3 (COL-3), matrix metalloproteinases 1 (MMP-1) and 2 (MMP-2), their tissue inhibitor (TIMP-2), and tumor necrosis factor (TNF-α) in the endometrium of mares with different grades of fibrosis. The samples (n = 34) were classified into three categories based on the frequency and distribution of fibrosis-related changes in the endometrium: Category I (healthy endometrium, n = 12), Category II (moderate fibrosis, n = 12), and Category III (severe fibrosis, n = 10). Collagen quantification demonstrate a substantial proportional increase (P < 0.0001) in collagen deposition across Category I (11.72 ± 1.39 %), Category II (17.76 ± 1.29 %), and Category III (24.15 ± 1.87 %). In transcript evaluations, higher COL-2 expression was found in Category II than in mares classified as Category I or III. MMP-1 showed increased transcript expression in Category II compared to Category III endometrial samples. Higher expression of MMP-2 was detected in Category III than in Category I and II. TIMP-2 showed lower mRNA expression in Category III vs Category I and II. However, TNF-α gene expression was higher in Category II than in Categories I and III. This study demonstrates that endometrial evaluation using PSR can play an important role in routine analyses for the detection and objective quantification of collagen in endometrial tissues. Additionally, this study demonstrated through gene expression analysis that MMP-1 may be linked to physiological endometrial remodeling. In contrast, MMP-2 could be associated with fibrogenesis in the endometrium, which is regulated by the inhibitor TIMP-2. Furthermore, COL-2 and TNF-α could be considered as biological markers involved in the progression endometrosis in mares. As such, the results of this study may contribute to the development of future antifibrotic therapies that aim to delay or even reverse the pathological remodeling of the extracellular matrix in the uterus, in addition to optimizing the diagnosis and prognosis of endometrial fibrosis in mares., Competing Interests: Declaration of competing interest None of the authors have any conflicts of interest to declare., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Myeloperoxidase is an enzyme released by neutrophils when neutrophil extracellular traps (NETs) are formed. Besides myeloperoxidase activity against pathogens, it was also linked to many diseases, including inflammatory and fibrotic ones. Endometrosis is a fibrotic disease of the mare endometrium, with a large impact on their fertility, where myeloperoxidase was shown to induce fibrosis. Noscapine is an alkaloid with a low toxicity, that has been studied as an anti-cancer drug, and more recently as an anti-fibrotic molecule. This work aims to evaluate noscapine inhibition of collagen type 1 (COL1) induced by myeloperoxidase in equine endometrial explants from follicular and mid-luteal phases, at 24 and 48 h of treatment. The transcription of collagen type 1 alpha 2 chain ( COL1A2 ), and COL1 protein relative abundance were evaluated by qPCR and Western blot, respectively. The treatment with myeloperoxidase increased COL1A2 mRNA transcription and COL1 protein, whereas noscapine was able to reduce this effect with respect to COL1A2 mRNA transcription, in a time/estrous cycle phase-dependent manner (in explants from the follicular phase, at 24 h of treatment). Our study indicates that noscapine is a promising drug to be considered as an anti-fibrotic molecule to prevent endometrosis development, making noscapine a strong candidate to be applied in future endometrosis therapies.

The Deepwater Horizon (DWH) oil spill profoundly impacted the health of bottlenose dolphins (Tursiops truncatus) in Barataria Bay, LA (BB). To comprehensively assess the cardiac health of dolphins living within the DWH oil spill footprint, techniques for in-water cardiac evaluation were refined with dolphins cared for by the U.S. Navy Marine Mammal Program in 2018 and applied to free-ranging bottlenose dolphins in BB (n = 34) and Sarasota Bay, Florida (SB) (n = 19), a non-oiled reference population. Cardiac auscultation detected systolic murmurs in the majority of dolphins from both sites (88% BB, 89% SB) and echocardiography showed most of the murmurs were innocent flow murmurs attributed to elevated blood flow velocity [1]. Telemetric six-lead electrocardiography detected arrhythmias in BB dolphins (43%) and SB dolphins (31%), all of which were considered low to moderate risk for adverse cardiac events. Echocardiography showed BB dolphins had thinner left ventricular walls, with significant differences in intraventricular septum thickness at the end of diastole (p = 0.002), and left ventricular posterior wall thickness at the end of diastole (p = 0.033). BB dolphins also had smaller left atrial size (p = 0.004), higher prevalence of tricuspid valve prolapse (p = 0.003), higher prevalence of tricuspid valve thickening (p = 0.033), and higher prevalence of aortic valve thickening (p = 0.008). Two dolphins in BB were diagnosed with pulmonary arterial hypertension based on Doppler echocardiography-derived estimates and supporting echocardiographic findings. Histopathology of dolphins who stranded within the DWH oil spill footprint showed a significantly higher prevalence of myocardial fibrosis (p = 0.003), regardless of age, compared to dolphins outside the oil spill footprint. In conclusion, there were substantial cardiac abnormalities identified in BB dolphins which may be related to DWH oil exposure, however, future work is needed to rule out other hypotheses and further elucidate the connection between oil exposure, pulmonary disease, and the observed cardiac abnormalities., Competing Interests: The authors have declared that no competing interests exist.

The objective of this pilot study was to examine the histologic effects associated with three known sclerosing agents and their ability to induce fibrosis in the subcutaneous space between the cervicocephalic air sac and skin. In the future, these drugs may prove useful in treating birds experiencing cervicocephalic diverticula rupture. The agents used were 1% polidocanol, absolute ethanol, and doxycycline hyclate. Twelve healthy adult chickens ( Gallus gallus domesticus ) were used in this study. The chickens were randomly allocated into three groups denoting day of euthanasia (day 4, 7, or 14). On day 0, all agents were injected (0.2 ml) subcutaneously, in a four-point grid fashion, in both the cervical and pectoral region of each bird. After euthanasia, the skin and subcutaneous tissues corresponding to the injection sites were harvested for histologic assessment. Tissue sections were assessed for fibrosis and lymphocytic and histiocytic inflammation. A scoring system was established to rank sclerosing agents by fibrosing and inflammatory ability. In the cervical region of chickens, 1% polidocanol induced the greatest inflammatory changes by day 7. Data suggest that doxycycline hyclate may produce the greatest cutaneous and subcutaneous fibrosis overall among all groups of birds. No adverse reactions were associated with any injection. Sterile saline produced the least amount of inflammation when assessed with the scoring system. Further investigation is needed to determine the safety of injections of larger volume with these chemicals and whether these findings can be extrapolated to birds with disease.

Fibrosis has also been recorded as a prominent pathological feature within wooden breast (WB) myopathy of broiler chickens. This study was conducted to evaluate the accumulation of fibril collagen, deposition of the extracellular matrix (ECM) components, and the underlying mechanism mediating the pathogenic fibrotic process in the pectoralis major (PM) muscle of WB-affected birds. Broiler chickens were categorized into the control and WB groups based on the evaluation of myopathic lesions. Results indicated that the total content and area of collagen in cross-sections of the PM muscle, as well as the augmented expression of collagen-I and fibronectin in the ECM, were greatly increased in birds with WB. Wooden breast myopathy upregulated expressions of transforming growth factor-beta (TGF-β) and the phosphorylation of Smad 2 and 3, thereby activating TGF-β-mediated Smad signaling pathway, which further enhanced the transcription of profibrotic mediators. In addition, regulators involved in collagen biosynthesis and cross-linking including prolyl 4-hydroxylase, lysyl oxidase, lysyl hydroxylase, and decorin were increased in the WB muscle. Finally, the expressions of both matrix metalloproteinases (MMP) and tissue inhibitor of metalloproteinases (TIMP) were increased in the WB muscle, which might be related with reduced ECM remodeling. Overall, WB myopathy induces severe fibrosis by enhancing ECM deposition and collagen cross-linking in the PM muscle of broiler chickens, possibly via the activation of TGF-β signaling and the dysregulation of the MMP and TIMP system., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

A 5-year-old Norwegian elkhound was referred due to an acute onset of lameness and persistent shoulder pain over a period of 3 weeks. Computed tomography demonstrated an enlarged, hypoattenuating right infraspinatus muscle with peripheral contrast enhancement and a nonenhancing center, without concurrent lesions in superficial structures or bones. The right infraspinatus muscle showed progressive atrophy on consecutive CT studies. The dog developed clinical symptoms compatible with fibrotic infraspinatus contracture 2 months after the initial presentation, and was treated with infraspinatus tenotomy. Histopathological diagnoses based on intraoperative biopsy samples were fibrotic muscle atrophy and muscle hypertrophy with regeneration., (© 2019 American College of Veterinary Radiology.)

Myocardial atrophy with fibrosis and fatty infiltration involving the cardiac conduction system is relatively unusual in horses. We herein report of such a case in a 13-year-old Arabian broodmare that had spontaneously died on a paddock. An autopsy revealed multifocal myocardial atrophy with concomitant fibrosis and fatty infiltration in both the ventricles and interventricular septum. The Purkinje fibres in the ventricles and interventricular septum were surrounded by thick fibrous or adipose tissues adjacent to atrophic myocardial cells. Myocardial fibrosis and fatty infiltration were likely secondary to myocardial atrophy, occurring as a pathological response triggered by the repair of muscular wall injury. However, there were no major vascular pathologies (e.g. atherosclerosis and arteriosclerosis); hence, the pathogenesis of myocardial atrophy was unclear. There was no evidence of myocardial atrophy ̵ induced pathologies such as infarct, ischaemic lesions, myocardial degeneration, myocarditis and endocarditis. However, such an unusual histopathological pattern may be associated with rapid clinical deterioration and death., (© 2020 The Authors. Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

There are age-related changes in testicular anatomy and physiology whereby there are modifications of sperm production and reproductive hormone functions. Effects of age on testicular microanatomy are well documented in humans, while there is limited understanding of these changes in dogs. The aim of this study was to evaluate age-related changes of seminiferous tubule morphology, interstitial fibrosis and spermatogenesis in dogs. Dogs (n = 32) were divided into four age groups: peripubertal (n = eight), relatively younger (n = seven), reproductively mature (n = seven) and relatively older (n = ten). Picrosirius Red stained sections were used for morphometrical analysis of testicular tissues, while the characteristics of seminiferous epithelium were assessed using a modified Johnsen scoring system for haematoxylin and eosin stained sections. Seminiferous epithelium and seminiferous tubule area increased from peripuberty to reproductive maturity, indicating there were changes during sexual maturation and subsequently there were decreases with further aging. There was a similar age-related trend for changes in seminiferous epithelium height with values being greatest in reproductively mature dogs; while there were no age-related differences in tubular diameter. Collagen content in the testicular interstitium gradually decreased from peripuberty to the age when dogs were reproductively mature and there were subsequent increases in relatively older dogs, thus, there was an association between the extent of testicular fibrosis and senescence. There was a decrease in spermatogenetic functions from relatively younger to older ages. Further investigations are warranted to establish mechanisms responsible for age-related changes of testicular morphology and related clinical implications., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper, (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Increased gene transcription of hypoxia-induced mediators of fibrosis in renal tissue has been identified in experimentally induced, ischemic chronic kidney disease (CKD)., Objective: To characterize hypoxia-induced profibrotic pathways in naturally occurring CKD in cats., Animals: Twelve client-owned cats with CKD and 8 healthy control cats., Methods: In this prospective, cross-sectional study, bilateral renal tissue samples were assessed histologically for inflammation, tubular atrophy, and fibrosis, and by reverse transcription-quantitative PCR for characterization of transcript levels of hypoxia-inducible factor-1α (HIF1A), matrix metalloproteinases-2 (MMP2), -7 (MMP7), and -9 (MMP9), tissue inhibitor of metalloproteinase-1 (TIMP1), transforming growth factor-β1 (TGFB1), and vascular endothelial growth factor-A (VEGFA). Linear mixed models were used to compare gene transcription between diseased and healthy kidneys, and to examine the association between transcript levels and serum creatinine concentration for all cats, and between transcript levels and histologic scores of diseased kidneys., Results: Kidneys from cats with CKD had significantly higher transcript levels of HIF1A, MMP2, MMP7, MMP9, TIMP1, and TGFB1 (all P < .001), and lower levels of VEGFA (P = .006) than those from control cats. Transcript levels of MMP7 (P = .05) and TIMP1 (P = .005) were positively associated with serum creatinine in cats with CKD, but not in control cats. In diseased kidneys, transcript levels of MMP2 (P = .002), MMP7 (P = .02), and TIMP1 (P = .02) were positively, whereas those of VEGFA (P = .003) were negatively, associated with histologic score severity., Conclusion and Clinical Significance: Evaluation of the expression of the corresponding proteins in larger populations could identify therapeutic targets and/or biomarkers of tubulointerstitial fibrosis in cats., (© 2020 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.)

Equine endometrosis is a multifactorial chronic degenerative condition, considered to be one of a major causes of equine infertility. The formation of periglandular fibrosis seems to be linked to chronic inflammation of the mare endometrium in a paracrine way and in a response to numerous forms of inflammatory stimuli elicit the net deposition of extracellular matrix (ECM) around the endometrial glands and stroma. We hypothesized some of these stimuli, such as monocyte chemoattractant protein-1 (MCP-1), interleukin-6 (IL-6), and hyaluronan synthases (HASs), may share the nuclear factor-κB (NF-κB) dependent activation pathway. This study aimed to determine whether mRNA expression of MCP-1, IL-6, HASs, and proteins of canonical (RelA/NK-κβ1) and noncanonical (NK-κβ2) signaling pathways for NF-kB would change in subsequent categories of endometrosis during the estrous cycle. The expression of selected genes was established in mare endometrium (n = 80; Kenney and Doig categories I, IIA, IIB, III), obtained in the follicular phase (FLP) and mid-luteal phase (MLP). The high expression of RelA mRNA was observed in III, whereas of NK-κβ1 and NK-κβ2 also in IIA, and IIA and IIB, respectively. The expression of MCP-1 mRNA occurred constantly, regardless of the category, whereas IL-6 mRNA was low in IIA, IIB, and III. The expression of HAS 1 was high in IIA and HAS 3 in IIA, IIB, and III. All those changes were observed in FLP, but not MLP. Our results suggest that NF-κB may be involved in progression of the chronic degenerative condition of the mare endometrium, on both canonical and noncanonical pathways. The most important changes in target genes expression were observed only in FLP, which may suggest the hormone-dependent activation of the NF-κB-dependent fibrosis pathway., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interests regarding the publication of this paper., (Copyright © 2020. Published by Elsevier Inc.)

Diabetic human patients have increased risk of heart failure compared to healthy subjects. The underlying mechanisms for this are not fully understood, and to help develop improved treatment strategies, well-characterized animal models are essential. To investigate cardiac dysfunction in diabetes, this study evaluated myocardial changes in 10 aging rhesus monkeys with and without diabetes. Based on evaluation of plasma glycosylated hemoglobin and glucose, 7 of 10 rhesus macaques had diabetes for a minimum of 11 months, while 3 of 10 were categorized as nondiabetic. A detailed histological examination of formalin-fixed left ventricular myocardial samples was followed by a semiquantitative evaluation of myocardial fibrosis and fat infiltration; digital quantifications of myocardial collagen, lipofuscin, and nuclear area fractions; and measurements of cardiomyocyte diameter. Histological myocardial evaluation revealed the presence of lipofuscin; large nuclei; interstitial, replacement, and vascular fibrosis; adipocyte infiltration; and vacuolar degeneration with atrophy of cardiomyocytes and fibrosis. However, there were no differences between groups for semiquantitative fat infiltration, fibrosis, cardiomyocyte size, collagen, or nuclear and lipofuscin area fraction. Lipofuscin area fraction correlated with plasma insulin, triglyceride, total cholesterol, and high-density lipoprotein cholesterol concentrations. In conclusion, myocardial pathological changes were found in left ventricular myocardium in aged rhesus macaques, independent of the stage of diabetes. The duration of diabetes might have been too short to cause differences between groups.

Background: Thoracolumbar myelopathies associated with spinal cord and vertebral column lesions, with a similar clinical phenotype, but different underlying etiologies, occur in pugs., Objectives: To further characterize the clinical and neuropathological characteristics of pugs with longstanding thoracolumbar myelopathy., Animals: Thirty client-owned pure-bred pugs with a history of more than a month of ataxia and paresis of the pelvic limbs, suggesting a myelopathy localized to the thoracolumbar spinal cord, were included in the study., Methods: Prospective clinicopathological study. Included pugs underwent a complete neurological examination and gross and histopathologic postmortem studies with focus on the spinal cord. Computed tomography (n = 18), magnetic resonance imaging (n = 17), and cerebrospinal fluid analysis (n = 27) were performed before or immediately after death., Results: Twenty male and 10 female pugs had a median age at clinical onset of 84 months (interquartile range, 66-96). Affected pugs presented with a progressive clinical course and 80% were incontinent. There was circumferential meningeal fibrosis with concomitant focal, malacic, destruction of the neuroparenchyma in the thoracolumbar spinal cord in 24/30 pugs. Vertebral lesions accompanied the focal spinal cord lesion, and there was lympho-histiocytic inflammation associated or not to the parenchymal lesion in 43% of the pugs., Conclusions and Clinical Importance: Meningeal fibrosis with associated focal spinal cord destruction and neighboring vertebral column lesions were common findings in pugs with long-standing thoracolumbar myelopathy., (© 2020 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.)

Cardiovascular disorders and predominantly idiopathic myocardial fibrosis are frequently associated with mortality among zoo-housed chimpanzees ( Pan troglodytes ). Formalin-fixed whole hearts of deceased chimpanzees housed in zoos ( n = 33) and an African sanctuary ( n = 2) underwent detailed macroscopic and histopathologic examination using a standardized protocol. Archived histological slides from the hearts of 23 additional African sanctuary-housed chimpanzees were also examined. Myocardial fibrosis (MF) was identified in 30 of 33 (91%) of the zoo-housed chimpanzees but none of the 25 sanctuary-housed chimpanzees. MF was shown to be characterized by both interstitial and replacement fibrosis. Immunophenotyping demonstrated that the fibrotic lesions were accompanied by the increased presence of macrophages, alpha smooth muscle actin-positive myofibroblasts, and a minimal to mild T-cell-dominant leukocyte infiltration. There was no convincing evidence of cardiotropic viral infection or suggestion that diabetes mellitus or vitamin E or selenium deficiency were associated with the presence of the lesion. However, serum vitamin D concentrations among zoo-housed chimpanzees were found to be lower in seasons of low ultraviolet light levels.

Myocarditis can cause death or permanent heart damage. As epidemiologic and etiopathologic data for canine myocarditis are lacking, we performed a retrospective study using nucleic acid extracted from archived (2007 to 2015) tissues from myocarditis cases and control dogs without myocardial lesions. Heart tissue from pediatric/juvenile and adult dogs was tested with a comprehensive panel of conventional and real-time polymerase chain reaction (PCR) assays targeting recognized agents of canine myocarditis based on a literature review and informed by the comparative epidemiology of human myocarditis. The PCR screen, which included canine parvovirus 2 (CPV-2), canine distemper virus, canine herpesvirus, Borrelia spp , West Nile virus, adenovirus, parainfluenza virus, pneumovirus, respiratory coronavirus, influenza virus, Bartonella spp , Rickettsia spp , Mycoplasma spp , and Neospora caninum , did not detect agents in 35 of 66 cases (53%; 95% confidence interval [CI], 41%-65%) and was frequently negative in adults (21/26); by comparison, agents were not detected in 27 of 57 controls (47%; 95% CI, 35%-60%). Canine distemper virus, herpesvirus, adenovirus, coronavirus, parainfluenza virus, Mycoplasma haemocanis , and N. caninum were occasionally detected in both cases and controls; thus, PCR detection was not considered to indicate causation. We previously reported that CPV-2 continues to be associated with myocarditis in young dogs despite widespread vaccination; in adults, CPV-2 was detected in 2 of 26 cases and 4 of 22 controls. As several agents were similarly detected in cases and controls, it is unclear if these are cardiopathogenic, incidental, or latent. West Nile virus was detected at the analytic limit in 1 adult case. We did not detect Borrelia spp , Bartonella spp , Rickettsia spp , or influenza A virus in the myocarditis cases. These data demonstrate the limitations of current targeted diagnostic tests and the need for additional research to identify unknown agents and develop testing strategies for canine myocarditis.

Previous work demonstrated renal fibrosis 70 days after a single unilateral in vivo renal ischemic event, but changes associated with a single episode of renal ischemia past this time are unknown. In this study, we evaluated renal function and structural changes 6 months after a 90-minute in vivo unilateral renal ischemic event. Six adult female cats underwent unilateral renal ischemia and renal function was followed for 6 months, at which time the kidneys were evaluated by histology and histomorphometry. Over time, there was a significant reduction in the glomerular filtration rate and an elevation of serum creatinine of 31% and 42%, respectively. All cats had tubulointerstitial lesions characterized by segmental interstitial inflammation, tubular atrophy, and interstitial fibrosis. Unlike short-term studies, ischemic kidneys had variable numbers of obsolescent glomeruli, consistent with the development of atubular glomeruli and subsequent ischemic glomerulosclerosis. Chronic changes associated with acute renal ischemia may include loss of function and glomerulosclerosis.

Benign stricture is an uncommon cause of chronic small intestinal obstruction in the cat. The purpose of this retrospective case series was to describe the ultrasonographic features, histopathological findings, and clinical presentation in a group of cats with benign small intestinal stricture. Inclusion criteria were cats presenting during the period 2010-2017, and that had ultrasonography and small intestinal stricture confirmed at surgery. For each cat, clinical data and ultrasonographic findings were retrieved from the medical record, and histopathology, where available, was reviewed. Eight cats met the inclusion criteria. The location of strictures was duodenum (1/8), mid- to distal jejunum (4/8), and ileum (3/8). Ultrasonographic findings included gastric distension (8/8) and generalized (3/8) or segmental (5/8) intestinal dilation consistent with mechanical obstruction. Ingesta did not propagate beyond the strictured segment. Wall thickening was mild to moderate (3-6 mm). Normal wall layering was disrupted in all cats. Strictures were predominantly hypoechoic (7/8) and associated with hyperechoic peri-intestinal mesentery (6/8). Annular strictures (5/8) were less than 15 mm in length whereas long-segment strictures (3/8) were greater than 15 mm in length. Histopathology showed transmural disease with fibrosis and inflammation (8/8), often (6/8) extending into the bordering mesentery. The mucosa was the most severely affected layer and epithelial injury accompanied the mucosal fibrosis/inflammation. Clinical presentation reflected delayed diagnosis of chronic bowel obstruction with debilitation (8/8), marked weight loss (8/8), and prerenal azotemia (5/8). Benign fibrostenotic stricture should be considered a differential diagnosis in debilitated young cats presenting with chronic bowel disease and ultrasonographic features of intestinal obstruction., (© 2019 American College of Veterinary Radiology.)

Objective: To evaluate tissue levels, safety, and efficacy of topical ophthalmic 0.5% and 1% pirfenidone in decreasing subconjunctival fibrosis., Animal Studied: Twelve normal beagle dogs PROCEDURES: A 5 × 1 mm diameter silicone disk was implanted subconjunctivally in one eye, and then dogs were treated with topical 0.5% pirfenidone (n = 9) in artificial tears or artificial tears alone (n = 3) for 28 days. To evaluate tissue drug levels, a single sample of tears, conjunctiva, and aqueous humor was collected 30 (n = 3), 90 (n = 3), and 180 min (n = 3) following administration of the last drop of pirfenidone, respectively. Fibrous capsule thickness and staining for Ki67 and fibroblast activation protein alpha (FAPα) were evaluated histologically. After a 2-week washout, the experiment was repeated in the opposite eye and using 1% pirfenidone., Results: Treatment with pirfenidone resulted in thinner fibrous capsules and decreased staining for FAPα with no adverse effects. The implant in one dog treated with pirfenidone extruded. There was no difference in tissue levels, capsular thickness, or staining for Ki67 or FAPα between dogs treated with 0.5% or 1% pirfenidone., Conclusions: Pirfenidone may decrease fibrosis following glaucoma shunt surgery and can potentially be used indefinitely due to minimal side effects., (© 2019 American College of Veterinary Ophthalmologists.)

The changes associated with condemned lungs in cattle with chronic pleural lesions of the caudal lobes were characterized by histology and immunohistochemistry (IHC). Fibroproliferative pleural lesions were microscopically confirmed. Occasionally, the pleural lesions also included adipose, chondroid, and osseous metaplasia that were covered by mesothelial cells, mostly in the absence of inflammation. Other lungs also showed fibrosis in the subpleural interstitium and interlobular septa. In both condemned and noncondemned lungs, immunoreactivity to Wilms tumor 1 (WT1) was normally observed on surface mesothelial cells but not on the submesothelial fibroblasts and myofibroblasts. Conversely, the myofibroblasts beneath the pleura, but not the mesothelial cells, showed immunoreactivity to alpha smooth muscle actin and calponin. However, in the lungs with myofibroblastic foci in the pleura, the proliferated cells maintained WT1 immunoreactivity similar to those of some metaplastic cells. These findings may reflect the plasticity of mesothelial cells in vivo.

Background: Pulmonary hemorrhage is a rare cause of death in horses. Hemorrhage within the respiratory tract has many causes, including mycosis of the guttural pouch, invasive procedures causing serious trauma to nasal conchae, or lung biopsy. We report on a rare case of a fatal pulmonary hemorrhage in a horse after a severe cough during bronchoalveolar lavage. To the best of our knowledge, this is the first report of spontaneous hemorrhage in a horse during bronchoalveolar lavage., Case Presentation: A 21-year-old mare which belonged to the didactic herd of The Faculty of Veterinary Medicine underwent BAL procedure for training purposes. Clinical examination prior to the procedure did not reveal any abnormalities and the horse had been classified as healthy. The horse was sedated with 0.01 mg/kg of detomidine and 0.01 mg/kg of butorphanol. The silicon BAL catheter was passed through the nasal passage into the trachea and then into the bronchus. Before catheter was wedged, the mare began to cough heavily and massive haemorrhage from mouth and nostrils occurred. Despite fluid therapy, shock occurred within 15 min and the mare was euthanized. Upon necropsy, site of hemorrhage was identified in the left lobar caudal bronchi, from a large blood vessel running directly beneath the bronchial wall. Upon histology, a chronic lympho-plasmocytic inflammatory process in left bronchi was identified. Moreover, Masson's trichrome staining revealed severe, perivascular fibrosis., Conclusion: Although BAL is a relatively safe procedure, and such complications should be treated as extremely rare, this case indicates that, in some individuals with specific subclinical problems, even mild physical force such as a cough can lead to rupture of the artery.

Our recent Communications Biology research article revealed the genomic drivers and therapeutic vulnerabilities of sea turtle fibropapillomatosis tumors. Fibropapillomatosis is a debilitating tumorous disease afflicting populations of green sea turtles globally. While a virus is involved in the development of this disease, it is increasingly understood that the key trigger is linked to anthropogenic disturbances of the environment. The specific environmental co-trigger(s) has yet to be functionally confirmed. Here we outline the next steps required to advance our understanding of this enigmatic disease, to enable us to more effectively clinically combat it and to ultimately tackle its environmental co-trigger to halt and hopefully reverse the spread of fibropapillomatosis., Competing Interests: The authors declare no competing interests.

Background: Chronic intestinal pseudo-obstruction (CIPO) is a rare clinical syndrome in veterinary medicine characterized by severe intestinal dysmotility without evidence of mechanical occlusion of the intestinal lumen. The exact pathogenesis of CIPO is unknown., Case Presentation: A 1-year-old male Miniature Bull Terrier dog was presented with a history of chronic weight loss, regurgitation, lethargy, vomiting and diarrhea. The dog was submitted for exploratory laparotomy. A full thickness intestinal biopsy was taken and a CIPO was suspected. The clinical condition deteriorated and the dog was euthanized. At gross examination the small intestine was severely dilated. Histologically severe fibrosis of the submucosa and severe atrophy of the tunica muscularis were present in small intestine and colon. Immunohistochemical examination with a panel of antibodies for gastro-intestinal neuromuscular disease-associated antigens revealed a severely reduced expression of alpha-smooth muscle actin in the tunica muscularis., Conclusions: This case report describes the gross, histological and immunohistochemical findings of CIPO affecting a 1-year-old Miniature Bull Terrier; on the basis of these findings a myopathic form of CIPO is hypothesized in this case.

Objective: To explore the impact of equine corneal fibroblast (ECF) to myofibroblast (ECM) differentiation by altering the expression of the Smad genes either individually or in combination. Specifically, we sought to examine the ECF differentiation after (a) silencing of Smad2, 3, and 4 profibrotic genes individually and (b) overexpression of antifibrotic Smad7 gene and in a combination with pro- and antifibrotic Smad genes., Methods: Equine corneal fibroblast primary cultures were generated as previously described. ECFs were transfected with individual plasmids which silenced gene expression of either Smad2, 3, or 4 or in combination with a plasmid overexpressing Smad7 using Lipofectamine 2000™ or Lipofectamine BLOCK-iT™. Smad-transfected clones were then exposed to TGF-β1 to induce differentiation to myofibroblasts. Immunofluorescence and qRT-PCR techniques quantified levels of ECF differentiation to ECM by measuring alpha smooth muscle actin, a known marker of ECM transdifferentiation., Results: Silencing of individual Smad2, 3, or 4 genes or overexpression of Smad7 showed significant inhibition of ECF transdifferentiation (73-83% reduction). Silencing of Smad2 showed the greatest inhibition of ECF transdifferentiation in (a) and was therefore utilized for the combination gene transfer testing. The combination gene transfer consisting of Smad7 overexpression and Smad2 silencing attenuated ECF differentiation significantly; however, the level was not significant compared to the overexpression of Smad7 individually., Conclusions: Using gene transfer technology involving profibrotic Smad silencing, antifibrotic Smad overexpression or its combination is a novel strategy to control TGF-β1-mediated fibrosis in equine fibroblasts. Combination gene therapy was not better than single gene therapy in this study., (© 2017 American College of Veterinary Ophthalmologists.)

Renal fibrosis is characterized by glomerulosclerosis and tubulointerstitial fibrosis and its pathogenesis is associated with the activity of mesenchymal cells (fibroblasts), being essentially characterized by a process of excessive accumulation resulting from the deposition of extracellular matrix components. The aim of this study was to characterize the morphological presentation of chronic and fibrotic lesions in the glomerular, tubular, interstitial, and vascular compartments in feline CKD, as well as the possible participation of myofibroblasts in renal fibrotic processes in this species. Cat kidneys were collected and processed according to the conventional techniques for light microscopy, circular polarization, immunohistochemistry, and electron microscopy. Fibrotic alterations were present in all compartments analyzed. The main findings in the glomerular compartment were different degrees of glomerular sclerosis, synechia formation, Bowman's capsule calcification, in addition to glomerular basement membrane thickening and pericapsular fibrosis. The tubulointerstitial compartment had intense tubular degeneration and the immunostaining in tubular cells for mesenchymal cell markers demonstrated the possibility of mesenchymal epithelial transition and consequent involvement of myofibroblasts in the development of interstitial tubule damage. Infiltration of inflammatory cells, added to vessel thickening and fibrosis, demonstrated the severity and role of inflammation in the development and perpetuation of damage. Thus, we may conclude that fibrotic lesions play a relevant role in feline CKD and the mechanism of perpetuation of these lesions need further elucidation regarding the origin and participation of myofibroblasts and consequent mesenchymal epithelial transition in this species., (© 2017 Wiley Periodicals, Inc.)

Environmental exposure and health risk upon engineered nanomaterials are increasingly concerned. The family of mesoporous carbon nanomaterials (MCNs) is a rising star in nanotechnology for multidisciplinary research with versatile applications in electronics, energy and gas storage, and biomedicine. Meanwhile, there is mounting concern on their environmental health risks due to the growing production and usage of MCNs. The lung is the primary site for particle invasion under environmental exposure to nanomaterials. Here, we studied the comprehensive toxicological profile of MCNs in the lung under the scenario of moderate environmental exposure. It was found that at a low concentration of 10μg/mL MCNs induced biophysical inhibition of natural pulmonary surfactant. Moreover, MCNs at similar concentrations reduced viability of J774A.1 macrophages and lung epithelial A549 cells. Incubating with nature pulmonary surfactant effectively reduced the cytotoxicity of MCNs. Regarding the pro-inflammatory responses, MCNs activated macrophages in vitro, and stimulated lung inflammation in mice after inhalation exposure, associated with lung fibrosis. Moreover, we found that the size of MCNs played a significant role in regulating cytotoxicity and pro-inflammatory potential of this nanomaterial. In general, larger MCNs induced more pronounced cytotoxic and pro-inflammatory effects than their smaller counterparts. Our results provided valuable information on the toxicological profile and environmental health risks of MCNs, and suggested that fine-tuning the size of MCNs could be a practical precautionary design strategy to increase safety and biocompatibility of this nanomaterial., (Copyright © 2017. Published by Elsevier B.V.)

Perinatal parvoviral infection causes necrotizing myocarditis in puppies, which results in acute high mortality or progressive cardiac injury. While widespread vaccination has dramatically curtailed the epidemic of canine parvoviral myocarditis, we hypothesized that canine parvovirus 2 (CPV-2) myocardial infection is an underrecognized cause of myocarditis, cardiac damage, and/or repair by fibrosis in young dogs. In this retrospective study, DNA was extracted from formalin-fixed, paraffin-embedded tissues from 40 cases and 41 control dogs under 2 years of age from 2007 to 2015. Cases had a diagnosis of myocardial necrosis, inflammation, or fibrosis, while age-matched controls lacked myocardial lesions. Conventional polymerase chain reaction (PCR) and sequencing targeting the VP1 to VP2 region detected CPV-2 in 12 of 40 cases (30%; 95% confidence interval [CI], 18%-45%) and 2 of 41 controls (5%; 95% CI, 0.1%-16%). Detection of CPV-2 DNA in the myocardium was significantly associated with myocardial lesions ( P = .003). Reverse transcription quantitative PCR amplifying VP2 identified viral messenger RNA in 12 of 12 PCR-positive cases and 2 of 2 controls. PCR results were confirmed by in situ hybridization, which identified parvoviral DNA in cardiomyocytes and occasionally macrophages of juvenile and young adult dogs (median age 61 days). Myocardial CPV-2 was identified in juveniles with minimal myocarditis and CPV-2 enteritis, which may indicate a longer window of cardiac susceptibility to myocarditis than previously reported. CPV-2 was also detected in dogs with severe myocardial fibrosis with in situ hybridization signal localized to cardiomyocytes, suggesting prior myocardial damage by CPV-2. Despite the frequency of vaccination, these findings suggest that CPV-2 remains an important cause of myocardial damage in dogs.

Fibroblasts are the structural base of mammary breast tissues. TGF-β1 can regulate the fibrotic process; however, it remains unclear whether TGF-β1 influences the susceptibility of fibroblasts to bacteria. Staphylococcus aureus (S. aureus) is a major bacterium in both chronic and subclinical mastitis in lactating cows that acts by invading host cells. To better understand the function of TGF-β1 in bovine mammary fibroblasts' (BMFBs) susceptibility to bacteria as well as the mechanisms involved, a primary BMFB model was established by treating cells with TGF-β1 followed by infection with S. aureus. The results revealed that the adhesion and invasion of S. aureus into BMFBs was significantly increased after cells were treated with 5 ng/ml TGF-β1 for 12 h. Moreover, TGF-β1 can increase Collagen I and α-SMA expression via activation of ERK signaling. However, the increased adhesion and invasion of S. aureus can be blocked by specific antibodies against either Collagen I or α-SMA, indicating that the increased adhesion and invasion are dependent on TGF-β1-induced upregulation of both Collagen I and α-SMA. Using PD98059, an ERK inhibitor, could also decrease the adhesion and invasion of S. aureus. These results indicate that TGF-β1 could promote S. aureus adhesion to and invasion into BMFBs by increasing Collagen I and α-SMA expression and may provide a novel target for controlling bovine mastitis., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Objective: In the context of a larger study on morphological changes of deep sole ulcers, claws with Rusterholz ulcer (RU) were investigated to describe the varied pathological changes of the RU., Material and Methods: Hind limbs of 55 cows displaying external signs of complicated sole ulcers were examined externally and internally. To examine the samples internally, a sagittal section was performed. Furthermore, the material was examined histologically and after bone maceration. A total of 59 claws of 112 digits with 120 deep sole ulcers displayed a complicated RU., Results: The most frequent findings were: fibrosis of the heel cushion, partly in conjunction with cartilaginous/osseous areas; osteomyelitis of the tuberculum flexorium, resulting in osteolysis of the macerated bones, often together with peripheral periosteal exostoses; avulsion fracture of the tuberculum flexorium in severely deformed claws; arthritis in the distal joint, frequently in conjunction with subchondral osteomyelitis, with the macerated bone showing osteolyses near the fossae synoviales and in the periphery of the joint surface as well as periosteal exostoses on the insertion point of the joint capsule. Changes in the bursa podotrochlearis, the common digital flexor tendon sheath and the pastern joint occurred less frequently., Conclusions and Clinical Relevance: Fibrosis and ossification of the heel cushion were classified as metaplasia, because these processes were considered an adaption to non-physiological strain of the heel. As the infection of the RU spreads, the tuberculum flexorium may play a central role. In advanced cases, an inflammation-related weakening of the bone tissue can lead to a pathological fracture with avulsion of the deep digital flexor tendon. Secondary infection of the distal joint may occur subsequent to any avulsion of the tendon in combination with opening of the joint cavity. More frequently, the infection is likely to have expanded from the insertion point of the joint capsule to the subchondral bone to ultimately infest the joint cavity. Changes in the bursa podotrochlearis, the common digital flexor tendon sheath and the pastern joint are assumed to develop as a consequence of arthritis in the distal joint.

Objective: To describe a novel technique for ameliorating cerebrospinal fluid flow obstruction secondary to pia-arachnoid fibrosis in dogs and report outcome., Study Design: Descriptive report and retrospective case series., Animals: Dogs with cerebrospinal fluid (CSF) flow obstruction (n = 7)., Methods: Medical records were searched for dogs that had a subarachnoid-subarachnoid shunt placed for treatment of CSF flow obstruction. Data collected included age, sex, breed, clinical signs and duration of signs prior to examination, neurologic status and localization prior to surgery, pre-surgical diagnostics, surgical technique, histopathology, postoperative neurologic examination, time to discharge from hospital, and outcome., Results: All dogs were diagnosed at surgery with a fibrotic adhesion between the arachnoid and pia mater. A subarachnoid shunting tube was implanted to allow CSF flow across the lesion site. Five dogs showed improvement of clinical signs, 3 of which showed complete recovery and 2 of which showed improvement without resolution of all clinical signs. Two dogs showed no change at 7 and 24 months postoperatively., Conclusion: Bridging a region of pia-arachnoid fibrosis with a tube placed in the subarachnoid space can ameliorate or prevent progression of associated clinical signs., (© 2017 The American College of Veterinary Surgeons.)

The association between pancreatic disorder and abdominal fat necrosis in cattle remains unclear. The pancreases of 29 slaughtered cattle with or without fat necrosis were collected to investigate pathological changes. Japanese Black (JB) cattle were classified into the FN group (with abdominal fat necrosis; n=9) and N group (without fat necrosis; n=5). The pancreases were also collected from 15 Holstein Friesian (HF) cows. All JB cattle showed high body condition scores. Regarding the pathological findings, fatty pancreas which involves adipocyte infiltration into the pancreas and fat necrosis (saponification) were observed in 25 and 27 cases, respectively. Immunohistochemical staining with anti-Iba-1 antibody showed large numbers of macrophages surrounding the saponified fat in the pancreas. CD3-positive T cells were significantly more common in the pancreas of both the FN and N groups compared with the HF group (P<0.05). Furthermore, fibrosis in the pancreas exhibited a correlative tendency with the formation of necrotic fat mass in the peritoneal cavity (P<0.1). These results indicate that obesity leads to increased severity of pancreatic disorder, including fatty pancreas and pancreatitis. The pathological lesions in the pancreas may play a key role in abdominal fat necrosis through the inflammatory process.

Vertebral column lordosis, kyphosis and scoliosis (LKS) can result in downgrading of farmed Chinook salmon Oncorhynchus tshawytscha in New Zealand. No cause of LKS has been identified. Radiography and histology were used to quantify LKS and perivertebral fibrosis in 27 fish with LKS visible at harvest and 30 visually normal fish from 3 New Zealand farms. Radiographic LKS was present in all 27 fish with LKS and in 18 of 30 fish without visible LKS. Quantification of the radiographic severity revealed significantly higher radiographic severity scores in fish with visible LKS (mean ± SD = 5.89 ± 2.41) than in fish with no visible, but radiographic LKS (1.44 ± 0.86, p < 0.001). The most frequent histological finding was unilateral perivertebral fibrosis that often extended into the horizontal septum and adjacent myomeres resulting in separation or loss of myocytes. Fibrosis was visible in all fish with LKS and in 12 of 30 fish without visible LKS. Fibrosis scores were higher in fish with visible LKS (3.32 ± 1.71) than in fish without visible LKS (0.35 ± 0.57, p < 0.001). The radiographic LKS severity scores were significantly correlated to the fibrosis scores (R2 = 0.59 p < 0.001) in the fish. Histology of other tissues revealed multifocal inflammation within muscle, peripheral connective tissues and myocardium which were considered most likely incidental in these fish. In this study, LKS was consistently and significantly associated with perivertebral fibrosis, suggesting that perivertebral fibrosis is an important process in the development of LKS. Further research to determine the cause of the fibrosis is required.

An 8-year-old spayed female Munsterlander was evaluated for a chronic low grade fever and a two month history of exercise intolerance. On physical examination, tachycardia and a grade II/VI right systolic heart murmur were detected. Echocardiography revealed marked thickening of the atrial and ventricular walls with mixed echogenicity and concentric hypertrophy of the left and right ventricles and equivocal systolic dysfunction. Serum cardiac troponin I level was markedly elevated. Endomyocardial biopsy was attempted; however, the patient arrested during the procedure and resuscitation was unsuccessful. Post-mortem examination revealed severe, chronic atrial and ventricular eosinophilic myocarditis associated with marked interstitial fibrosis. Serological testing, histopathology and immunohistochemistry staining did not reveal an underlying infectious agent or neoplasm. To our knowledge, this is the first reported case of primary eosinophilic myocarditis in the absence of a peripheral eosinophilia and multi-organ eosinophilic inflammation in a dog., (Published by Elsevier B.V.)

The cytokine transforming growth factor beta 1 (TGF-β1) has been widely implicated in the development and progression of renal fibrosis in chronic kidney disease (CKD) in humans and in experimental models. The aims of this study were to assess the association between urinary active TGF-β1 and (a) development of CKD in a cross-sectional study, (b) deterioration of renal function over 1 year in a longitudinal study, and (c) renal histopathological parameters in cats. A human active TGF-β1 ELISA was validated for use in feline urine. Cross-sectional analysis revealed no significant difference in urinary active TGF-β1:creatinine ratio (aTGF-β1:UCr) between groups with differing renal function. Longitudinally, non-azotaemic cats that developed CKD demonstrated a significant (P = 0.028) increase in aTGF-β1:UCr approximately 6 months before the development of azotaemia, which remained elevated (P = 0.046) at diagnosis (approximately 12 months prior, 8.4 pg/mg; approximately 6 months prior, 22.2 pg/mg; at CKD diagnosis, 24.6 pg/mg). In the histopathology study, aTGF-β1:UCr was significantly higher in cats with moderate (P = 0.02) and diffuse (P = 0.005) renal fibrosis than in cats without fibrosis. Cats with moderate renal inflammation had significantly higher urinary active aTGF-β1 concentrations than cats with mild (P = 0.035) or no inflammatory change (P = 0.004). The parameter aTGF-β1:UCr was independently associated with Log urine protein:creatinine ratio in a multivariable analysis of clinicopathological parameters and interstitial fibrosis score in a multivariable analysis of histopathological features. These results suggest that urinary aTGF-β1 reflects the severity of renal pathology. Increases in urinary aTGF-β1 followed longitudinally in individual cats may indicate the development of CKD., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The abnormal proliferation of bovine mammary fibroblasts (BMFBs) impairs mammary gland development and lactation. Severe manifestations develop into breast fibrosis, leading to the culling of cows and causing serious losses to the dairy industry. Transforming growth factor β1 (TGF-β1) is an important modulator of cell proliferation and extracellular matrix formation; however, limited information is available on BMFBs. In this study, a convenient and stable culture method for BMFBs was established. Treatment with 5 ng/mL of TGF-β1 significantly promoted the proliferation of BMFBs and accelerated the cell cycle. TGF-β1 stimulation for up to 12 h significantly increased the relative ERK1/2 mRNA expression and enhanced the protein expression of p-ERK1/2 and cyclin D1. Conversely, the ERK1/2 inhibitor PD98059 blocked these TGF-β1 effects. Further exploration using a mouse model showed that TGF-β1 significantly increased the proportion of fibroblasts and accelerating the cell transition from the G1 to G2/M phases. In addition, TGF-β1 enhanced the expression of fibrosis markers, α-SMA and I Collagen, which could be blocked efficiently by the PD98059 in mouse mammary gland. Finally, immunofluorescence analysis confirmed that TGF-β1 promoted fibroblast proliferation in healthy dairy cows after normal long-term dietary corn straw roughage supplementation. It is suggested that the diet may promote mammary fibroblast proliferation by raising the level of TGF-β1. Our study provides new insights into how nutrition causes undesirable changes in mammary gland structure., (© 2016 International Federation for Cell Biology.)

Chronic kidney disease (CKD) is the most common metabolic disease of domesticated cats, with most affected cats being geriatric (>12 years of age). The prevalence of CKD in cats exceeds that observed in dogs, and the frequency of the diagnosis of CKD in cats has increased in recent decades. Typical histologic features include interstitial inflammation, tubular atrophy, and fibrosis with secondary glomerulosclerosis. In contrast to people and dogs, primary glomerulopathies with marked proteinuria are remarkably rare findings in cats. Although a variety of primary renal diseases have been implicated, the disease is idiopathic in most cats. Tubulointerstitial changes, including fibrosis, are present in the early stages of feline CKD and become more severe in advanced disease. A variety of factors-including aging, ischemia, comorbid conditions, phosphorus overload, and routine vaccinations-have been implicated as factors that could contribute to the initiation of this disease in affected cats. Factors that are related to progression of established CKD, which occurs in some but not all cats, include dietary phosphorus intake, magnitude of proteinuria, and anemia. Renal fibrosis, a common histologic feature of aged feline kidneys, interferes with the normal relationship between peritubular capillaries and renal tubules. Experimentally, renal ischemia results in morphologic changes similar to those observed in spontaneous CKD. Renal hypoxia, perhaps episodic, may play a role in the initiation and progression of this disease., (© The Author(s) 2016.)

Cardiorenal syndrome involves disease and dysfunction of the heart that leads to progressive renal dysfunction. This study investigated the relationship between cardiac and renal disease in 91 aged chimpanzees at the Alamogordo Primate Facility by evaluation of the medical histories, metabolic parameters, functional measurements of the cardiovascular system, clinical pathology, and histopathology focused on the heart and kidney. Cardiac fibrosis was the most frequent microscopic finding in 82 of 91 animals (90%), followed by glomerulosclerosis with tubulointerstitial fibrosis in 63 of 91 (69%). Cardiac fibrosis with attendant glomerulosclerosis and tubulointerstitial fibrosis was observed in 58 of 91 animals (63%); there was a statistically significant association between the 2 conditions. As the severity of cardiac fibrosis increased, there was corresponding increase in severity of glomerulosclerosis with tubulointerstitial fibrosis. Altered metabolic, cardiovascular, and clinical pathology parameters indicative of heart and kidney failure were commonly associated with the moderate to severe microscopic changes, and concurrent heart and kidney failure were considered the cause of death. The constellation of findings in the chimpanzees were similar to cardiorenal syndrome in humans., (© The Author(s) 2016.)

Objectives: This study aimed to determine the association of cardiac fibrosis with the galectin-3 (Gal-3) expression, a fibrosis marker in the myocardium and to compare plasma Gal-3 levels in normal and degenerative mitral valve disease (DMVD) dogs., Animals: Studies of muscle expression and plasma levels of Gal-3 were performed in separate groups of dogs. The tissue study was performed on cardiac tissues collected from 22 dogs. The plasma study was performed on 46 client-owned dogs., Methods: Papillary muscle and left ventricular (LV) wall obtained from 10 normal and 12 DMVD dogs were stained with Masson trichrome and Gal-3 immunohistochemistry to determine fibrosis areas and Gal-3 expression. Plasma samples were collected from 19 normal and 27 DMVD dogs for Gal-3 measurement by ELISA., Results: Percentage of fibrosis was higher in papillary muscle and LV wall of DMVD dogs (66.13 ± 5.58%; 52.98 ± 8.45%) than in normal dogs (35.40 ± 8.46%; 27.41 ± 7.91%; p < 0.0001). Gal-3 was higher in papillary muscle and LV wall of DMVD dogs (27.95 ± 6.94%; 17.25 ± 8.76%) than in normal dogs (1.08 ± 0.67%; 0.52 ± 0.42%; p < 0.0001). Fibrosis areas correlated strongly with the Gal-3 expression (r = 0.821, p < 0.0001). Plasma Gal-3 levels were increased in DMVD dogs (1.50; 0.87-2.36 ng/mL) compared to normal dogs (0.42; 0.27-0.63 ng/mL; p < 0.0001)., Conclusions: Gal-3 expression in cardiac muscle was associated with cardiac fibrosis and was higher in DMVD dogs than in normal dogs. DMVD dogs had higher plasma Gal-3 concentrations than normal dogs. Tissue Gal-3 is a candidate of fibrosis biomarker in DMVD; however, further investigation of associations between plasma Gal-3 and myocardial fibrosis is necessary., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Objectives: To evaluate gross and histopathologic lesions in Irish wolfhounds (IWs) with atrial fibrillation (AF) and/or primary dilated cardiomyopathy (DCM) in different stages of disease., Methods: Twenty-six formalin-fixed IW hearts were studied. Clinical diagnosis was based upon results of their most recent cardiovascular examinations including electrocardiography and echocardiography and categorized as normal (n = 4); preclinical (asymptomatic) DCM with AF (n = 6); DCM with congestive heart failure and AF (n = 4); AF with left ventricular reverse remodeling after DCM diagnosis (n = 3); AF without DCM (n = 7); and DCM with sinus rhythm (n = 2). All hearts were evaluated by one pathologist (HA) blinded to the clinical diagnosis., Results: Ten of 15 DCM hearts showed mild to moderate multifocal myocardial fibrosis with variable diffuse adipocyte infiltration within the left and right ventricular myocardium. In five DCM hearts, there were no histopathological findings identified. Right atrial appendages from AF dogs with and without DCM had significantly more myocardial fibrosis and adipocyte infiltration compared with normal hearts and compared to left atrial appendages., Conclusions: Gross and histological findings in the ventricular myocardium of IWs with clinical diagnosis of DCM were variable; in some dogs, histopathology was normal. In IWs, the etiology of DCM might be different from that in other breeds with conditions causing functional impairment rather than evident histological changes. Right and left atrial appendages from IWs with AF displayed substantial pathology (interstitial fibrosis and adipocytes) most prevalent in the right atrial appendages which may be correlated to the pathogenesis of AF. These preliminary findings merit further study., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The objectives of this study were to define the acute and chronic effects of 1-hour unilateral in vivo renal ischemia on renal function and histology in cats. Twenty-one adult purpose-bred research cats were anesthetized, and 1 kidney underwent renal artery and vein occlusion for 1 hour. Serum creatinine and urea concentrations, urine protein:creatinine ratio, urine-specific gravity, glomerular filtration rate, hematocrit, platelet concentration and function, and white blood cell count were measured at baseline and variable time points after ischemia. Renal histopathology was evaluated on days 3, 6, 12, 21, 42, and 70 postischemia; changes in smooth muscle actin and interstitial collagen were examined. Following ischemia, whole animal glomerular filtration rate was significantly reduced (57% of baseline on day 6; P < .05). At the early time points, the ischemic kidneys exhibited severe acute epithelial necrosis accompanied by evidence of regeneration of tubules predominantly within the corticomedullary junction. At later periods, postischemic kidneys had evidence of tubular atrophy and interstitial inflammation with significantly more smooth muscle actin and interstitial collagen staining and interstitial fibrosis when compared with the contralateral control kidneys. This study characterizes the course of ischemic acute kidney injury in cats and demonstrates that ischemic acute kidney injury triggers chronic fibrosis, interstitial inflammation, and tubular atrophy in feline kidneys. These late changes are typical of those observed in cats with naturally occurring chronic kidney disease., (© The Author(s) 2015.)

Although Chlamydia causes disease of the urethra and prostate of male koalas, its impact on the testis and epididymis has not been examined. This study describes chronic-active and granulomatous orchitis and epididymitis with interstitial fibrosis associated with infection by Chlamydia pecorum in 2 of 18 adult male koalas being euthanized at a koala hospital, 8 of which also had chlamydial prostatitis. By immunohistochemistry and transmission electron microscopy, chlamydial inclusions were demonstrated within Sertoli cells directly associated with mild inflammation surrounding intact seminiferous and epididymal tubules, marked pyogranulomatous inflammation around disrupted tubules, replacement of tubules by interstitial fibrosis, and aspermia. The presence of C. pecorum but not Chlamydia pneumoniae was detected by quantitative polymerase chain reaction of formalin-fixed tissues of the left and right testes and right epididymis in 1 animal. This is the first report of orchitis and epididymitis in a koala infected with C. pecorum., (© The Author(s) 2015.)

In Switzerland, dicrocoeliasis is regarded as the most significant parasitic infection of llamas and alpacas. Fasciola hepatica infestation is also a problem but less common. The aim of the present retrospective study was to evaluate the lungs of New World camelids (NWCs) for evidence of arterial hypertension in association with liver changes due to liver fluke infestation. The lungs of 20 llamas and 20 alpacas with liver fluke infestation were histologically evaluated. The hematoxylin and eosin and van Gieson (VG)-elastica stains as well as immunohistology for the expression of α-smooth muscle actin (α-SMA) were used to visualize the structures of arterial walls. Parasitology of fecal matter (11 llamas and 17 alpacas) confirmed that most of these animals were infested with both Dicrocoelium dendriticum and other gastrointestinal parasites. In most cases (10/12 llamas, 4/6 alpacas), liver enzyme activity in serum was elevated. Histologically, arteries in the lungs of 9 of 20 llamas (45%) and 3 of 20 alpacas (15%) showed severe intimal and adventitial and slight to moderate medial thickening, which was confirmed with α-SMA and VG-elastica staining. All animals exhibited typical liver changes, such as fibrosis and biliary hyperplasia, in association with the presence of liver flukes. This study shows that liver flukes can induce proliferative changes in lung arteries in NWCs that resemble those seen with pulmonary arterial hypertension due to liver parasites in humans. However, the degree of liver fluke infestation was not correlated with the extent of liver damage, or with the amount of thoracic or abdominal effusion or pulmonary arterial changes., (© The Author(s) 2015.)

A 7-year-old female boxer dog died suddenly without any clinical signs. It was suspected that the dog had arrhythmogenic right ventricular cardiomyopathy (ARVC) due to ventricular premature complexes and ventricular tachycardia at 3 years of age. The final diagnosis of ARVC was confirmed by histological characteristics, such as loss of cardiocytes and fibrofatty replacement, occurring in the right and left ventricular walls. In the cardiocytes, non-lipid vacuoles were observed. Cardiac fibrosis and intimal thickening of the small arteries occurred without fatty replacement in the inner muscle layer including the papillary muscles of the left ventricular wall. This paper describes the pathomorphological details of an ARVC case with coincidental cardiac fibrosis in the inner muscle layer of the left ventricular wall.

We report coincident sternal segment dislocation and focally extensive right ventricular epicardial fibrosis observed during routine histopathology evaluation of C57BL/6N mice as part of a high throughput phenotyping screen conducted between 4 and 16 weeks of age. This retrospective case series study was conducted to determine whether cardiac fibrosis was a pathological consequence of sternal segment dislocation. We identified sternal segment dislocation in 51 of the total 1103 mice (4.6%) analyzed at 16 weeks of age. Males were more frequently affected. In all cases but 2, the dislocation occurred at the fourth intersternebral joint. In 42 of the 51 cases (82.4%), the dislocation was encased by regenerative cartilaginous callus that protruded internally into the thoracic cavity (intrathoracic callus) and/or externally to the outer aspect of the sternum (extrathoracic callus). Displacement of dislocated ends of the sternum into the thoracic cavity was present in 19 of 51 cases (36.5%). Coincident minimal or mild right ventricular epicardial and subepicardial fibrosis was observed in 22 of the 51 cases (43%) but was not observed in any of the mice in the absence of sternal segment dislocation. Our data suggest that right ventricular fibrosis was likely caused by direct injury of the right ventricle by the dislocated ends of the sternum and/or by intrathoracic callus that develops post dislocation. Potential pathogenesis for the sternal and cardiac lesions and their implication for the interpretation of phenotypes in mouse models of cardiopulmonary and skeletal disease are discussed., (© The Author(s) 2014.)

Mares that fail to become pregnant after repeated breeding, without showing typical signs of clinical endometritis, should be suspected of subclinical endometritis (SE). Contact with infectious agents results in altered synthesis and secretion of inflammatory mediators, including cytokines and arachidonic acid metabolites, and disturbs endometrial functional balance. To address the hypothesis that SE affects the immune endocrine status of the equine endometrium, spontaneous secretion of prostaglandin E(2) (PGE(2)), prostaglandin F(2α) (PGF(2α)), 6-keto-PGF(1α )(a metabolite of prostacyclin I(2)), leukotriene B(4) (LTB(4)), and leukotriene C(4) (LTC(4)) was examined. In addition, secretion of these factors was examined relative to the grade of inflammation, fibrosis, and estrous cycle stage. Eighty-two warmblood mares, of known breeding history, were enrolled in this study. On the basis of histopathologic assessment, mares were classified as suffering from first-grade SE, second-grade SE, or being healthy. The grade of fibrosis and the infiltration of endometrial tissue with polymorphonuclear leukocytes were examined by routine hematoxylin-eosin staining. In mares suffering from SE, the secretion profiles of PGE(2), 6-keto-PGF(1α), LTB(4), and LTC(4) were changed compared to mares that did not suffer from endometritis. The secretion of PGE(2) and 6-keto-PGF1α was increased, whereas that of LTB(4) and LTC(4) was decreased. Secretion of 6-keto-PGF(1α) was increased in first- and second-grade SE (P < 0.01). The concentration of PGI(2) metabolite was increased only in inflamed endometrium, independently of the inflammation grade, but was not affected by fibrosis. Prostaglandin E(2) secretion was increased in second-grade SE (P < 0.05). The secretion of LTB(4) decreased in both first- and second-grade SE (P < 0.05), whereas secretion of LTC(4) was decreased only in second-grade SE (P < 0.05). Fibrosis did not change the secretion profile of PGE(2), PGF(2α), and 6-keto-PGF(1α) during the course of SE. However, the secretion profile of LTB(4) was affected during the course of fibrosis. Evident divergences between PGE(2) and PGF(2α) profiles and in PGE2:PGF(2α) ratios in the control versus SE mares observed during the course of diestrus contribute to shortened or prolonged interestrous intervals observed clinically in SE mares., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Objectives: To evaluate associations between N-terminal procollagen type III (PIIINP), a serum biomarker of collagen biosynthesis, and myocardial fibrosis in dogs with naturally-occurring myxomatous mitral valve disease (MMVD)., Animals: Twenty-two dogs with echocardiographically-confirmed MMVD were prospectively recruited from a hospital population. All died as a result of MMVD and their hearts were available for post mortem examination., Methods: Echocardiographic measurements and serum PIIINP concentrations were obtained from all dogs prior to death or euthanasia. Serum PIIINP concentrations (μg/mL) were measured using a validated commercially available radioimmunoassay. Myocardial tissue samples were collected post mortem and myocardial fibrosis was scored. The average fibrosis score for all cardiac sites in the heart was designated the global fibrosis score (GFS). The average fibrosis score for all papillary muscle sites was designated the papillary fibrosis score (PFS). Univariate and multivariate linear regression analyses were used separately to evaluate associations between GFS and PFS, respectively, and PIIINP and echocardiographic variables., Results: Left ventricular end-diastolic diameter normalized for body weight (LVEDDN) and PIIINP were weakly independently positively associated with both GFS and PFS. LVEDDN and PIIINP were weakly negatively correlated., Conclusions: Both LVEDDN and serum PIIINP increase with increasing fibrosis score, although these relationships were not strong enough to be clinically useful. Although LVEDDN and PIIINP were positively correlated with fibrosis, PIIINP decreased with increasing LVEDDN, suggesting a complex interplay between fibrosis and remodeling in MMVD., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Cattle grazing turnips or other brassica forage crops occasionally develop hepatogenous photosensitization. In New Zealand, cases of bovine photosensitization associated with such crops frequently occur during late summer and fall, and this coincides with the facial eczema (sporidesmin toxicosis) "season." Clinical chemistry findings in acute photosensitization cases associated with both brassica and facial eczema include marked serum elevations in γ-glutamyl transferase and glutamate dehydrogenase activities. Distinctive bile duct lesions of "subacute" turnip photosensitization in 2 cows, comprising microscopic cholangiectasis with concentric periductal fibrosis of small bile ducts, and a spectrum of changes from bile duct necrosis progressing to obliterative sclerosis are described. These bile duct lesions are compared with those in 3 cases of "subacute" facial eczema in adult cows, where medium-sized and larger ducts tend to be involved and bile duct hyperplasia and portal fibrosis are more prominent, often leading to bridging between neighboring portal triads., (© The Author(s) 2013.)