@article{rivas_tan_shaverdian_nguyen_wouters_stern_li_2024, title={A novel ITGA2B double cytosine frameshift variant (c.1986_1987insCC) leads to Glanzmann's thrombasthenia in a cat}, volume={3}, ISSN={["1939-1676"]}, url={https://doi.org/10.1111/jvim.17030}, DOI={10.1111/jvim.17030}, abstractNote={AbstractBackgroundGlanzmann's thrombasthenia (GT) is a congenital platelet disorder affecting approximately 1:1 000 000 people globally and characterized by impaired platelet aggregation and clot retraction. Autosomal recessive, loss‐of‐function, variants in ITGA2B or ITGB3 of the αIIbβ3 receptor cause the disease in humans. A cat affected by Glanzmann's and macrothrombocytopenia was presented to the UC Davis VMTH.Hypothesis/ObjectivesSevere thrombopathia in this cat has an underlying genetic etiology.AnimalsA single affected patient, 2 age‐matched clinically healthy controls, and a geriatric population (n = 20) of normal cats.MethodsPhysical examination and clinical pathology tests were performed on the patient. Flow cytometry and platelet aggregometry analyses for patient phenotyping were performed. Patient and validation cohort gDNA samples were extracted for Sanger sequencing of a previously identified ITGA2B (c.1986delC) variant. Reverse transcriptase PCR was performed on patient and healthy control PRP samples to verify ITGA2B variant consequence.ResultsA novel c.1986_1987insCC autosomal recessive variant in ITGA2B was identified. This variant was absent in a population of 194 unrelated cats spanning 44 different breeds. Complete loss of ITGA2B transcript and protein expression was verified by RT‐PCR and flow cytometry, explaining the underlying etiology of GT, and likely macrothrombocytopenia, in this cat.Conclusions and Clinical ImportanceThis study emphasizes the role of precision medicine in cardiovascular disease of cats and identified yet another variant that may be of utility for screening in the feline population. This study provides a small‐volume, standardized, successful protocol for adequate platelet RNA isolation and subsequent molecular assessment of gene expression in cats.}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Rivas, Victor N. and Tan, Avalene W. K. and Shaverdian, Meg and Nguyen, Nghi P. and Wouters, Jalena R. and Stern, Joshua A. and Li, Ronald H. L.}, year={2024}, month={Mar} }
@article{rosati_jandrey_stern_nguyen_li_2024, title={Evaluation of clopidogrel response in healthy cats using a novel viscoelastic test and thromboelastography}, volume={11}, ISSN={["2297-1769"]}, url={https://europepmc.org/articles/PMC11217344}, DOI={10.3389/fvets.2024.1371781}, abstractNote={Introduction Cats with cardiomyopathy face an increased risk of arterial thromboembolism (ATE). Although clopidogrel is frequently utilized to mitigate this risk, feline responses to this therapy exhibit variability. This study evaluated 2 viscoelastic devices, thromboelastography (TEG) and Viscoelastic Coagulation Monitor (VCM), for monitoring clopidogrel in cats in comparison to light transmission aggregometry (LTA). Methods Twenty-eight healthy cats received clopidogrel for 7 days. Blood was collected at baseline and after treatment for analysis by TEG, VCM, and LTA. Results On LTA, maximum amplitude, slope, and area under the curve (AUC) significantly decreased after treatment ( p < 0.0001). On VCM, maximum clot firmness (MCF) significantly increased after treatment ( p = 0.002). On TEG, R-time significantly prolonged ( p = 0.024), while K and alpha angle significantly changed ( p = 0.0002 and p = 0.0014, respectively). There was a moderate negative correlation between TEG R-time and LTA AUC ( r = −0.39, p = 0.042). Eight cats were identified as non-responders to clopidogrel. Of the 8 non-responders, 6 (75%) had shortened R time after treatment. VCM appeared to be less discriminatory in identifying non-responders. Discussion LTA remained the gold standard of monitoring clopidogrel treatment in cats. Unexpected changes on VCM and TEG were likely related to high interindividual and assay variability and increased sensitivity of feline platelets. R-time on TEG may have potential utility for point-of-care monitoring of clopidogrel response in cats.}, journal={FRONTIERS IN VETERINARY SCIENCE}, author={Rosati, Tommaso and Jandrey, Karl E. and Stern, Joshua A. and Nguyen, Nghi and Li, Ronald H. L.}, year={2024}, month={Jun} }
@article{duler_visser_nguyen_johnson_stern_li_2024, title={Platelet hyperresponsiveness and increased platelet-neutrophil aggregates in dogs with myxomatous mitral valve disease and pulmonary hypertension}, volume={5}, ISSN={["1939-1676"]}, url={http://dx.doi.org/10.1111/jvim.17067}, DOI={10.1111/jvim.17067}, abstractNote={Abstract Background Pulmonary hypertension (PH) in dogs with myxomatous mitral valve disease (MMVD) is caused by increased pulmonary venous pressure. Thrombosis, vascular remodeling, and vasoconstriction mediated by platelets could exacerbate PH. Hypothesis Dogs with PH will exhibit a hypercoagulable state, characterized by increased platelet activation, platelet‐leukocyte, and platelet‐neutrophil aggregate formation. Animals Eleven dogs (≥3.5 kg) diagnosed with MMVD and PH and 10 dogs with MMVD lacking PH. Methods Prospective cohort ex vivo study. All dogs underwent echocardiographic examination, CBC, 3‐view thoracic radiographs, and heartworm antigen testing. Severity of PH and MMVD were assessed by echocardiography. Viscoelastic monitoring of coagulation was assessed using thromboelastography (TEG). Platelet activation and platelet‐leukocyte/platelet‐neutrophil interactions were assessed using flow cytometry. Plasma serotonin concentrations were measured by ELISA. Results Unstimulated platelets from dogs with MMVD and PH expressed more surface P‐selectin than MMVD controls ( P = .03). Platelets from dogs with MMVD and PH had persistent activation in response to agonists. The number of platelet‐leukocyte aggregates was higher in dogs with MMVD and PH compared with MMVD controls ( P = .01). Ex vivo stimulation of whole blood resulted in higher numbers of platelet‐neutrophil aggregates in dogs with MMVD and PH ( P = .01). Assessment of hypercoagulability based on TEG or plasma serotonin concentrations did not differ between groups. Conclusion and Clinical Importance Platelet hyperresponsiveness and increased platelet‐neutrophil interaction occur in dogs with MMVD and PH, suggesting that platelets play a role of in the pathogenesis of PH. Clinical benefits of antiplatelet drugs in dogs with MMVD and PH require further investigation.}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Duler, Laetitia and Visser, Lance and Nguyen, Nghi and Johnson, Lynelle R. and Stern, Joshua A. and Li, Ronald H. L.}, year={2024}, month={May} }
@article{philp_farrell_li_2023, title={Case report: Disseminated intravascular coagulation in a dog following treatment with melarsomine for Dirofilaria immitis}, volume={10}, ISSN={2297-1769}, url={http://dx.doi.org/10.3389/fvets.2023.1118798}, DOI={10.3389/fvets.2023.1118798}, abstractNote={Disseminated intravascular coagulation following melarsomine therapy for Dirofilaria immitis (D. immitis) is reported in a 9-year-old female intact pit bull-type dog. The dog had been diagnosed with D. immitis (antigen and microfilaria positive) and treated with imidacloprid, moxidectin, doxycycline and 3 doses of melarsomine over a 92-day period. Seven days after the third melarsomine injection, the patient was presented to her family veterinarian due to right pelvic limb swelling. Prothrombin and activated partial thromboplastin times were prolonged beyond the detectable range. Treatment included vitamin K1 and fresh frozen plasma (FFP) prior to referral to the authors' institution. At this time the patient remained coagulopathic. Further investigations included thoracic radiographs, abdominal ultrasound and an echocardiogram. The patient was administered multiple units of packed red blood cells and FFP, sildenafil, dexamethasone SP, aminocaproic acid and vitamin K1. Repeat CBC approximately 20 h after admission showed persistent anemia and thrombocytopenia. Despite ongoing administration of FFP, a repeat coagulation panel showed worsening of the coagulopathy with prothrombin time of 84.2s [reference interval (RI) 7.0–9.3s], activated partial thromboplastin time >140s (RI 10.4–12.9s) and fibrinogen <50 mg/dL (RI 109–311 mg/dL). Following discussion with the owners, the patient was euthanized. Necropsy was performed and confirmed heartworm infection with severe pulmonary arterial thrombosis, vascular remodeling, and intraluminal degenerate nematodes. Multifocal subcutaneous and organ hemorrhage was apparent. Although coagulopathy has been described in caval syndrome associated with heartworm disease and is listed as a potential side effect of melarsomine administration, this is the first report of documented disseminated intravascular coagulation following melarsomine treatment for D. immitis. Potential mechanisms for the coagulopathy are discussed and the case report highlights a rare, but serious complication of adulticide therapy.}, journal={Frontiers in Veterinary Science}, publisher={Frontiers Media SA}, author={Philp, Helen S. and Farrell, Kate S. and Li, Ronald H. L.}, year={2023}, month={Feb}, pages={1118798} }
@article{li_fabella_nguyen_kaplan_ontiveros_stern_2023, title={Circulating neutrophil extracellular traps in cats with hypertrophic cardiomyopathy and cardiogenic arterial thromboembolism}, volume={37}, ISSN={0891-6640 1939-1676}, url={http://dx.doi.org/10.1111/jvim.16676}, DOI={10.1111/jvim.16676}, abstractNote={AbstractBackgroundCats with hypertrophic cardiomyopathy (HCM) are at risk of cardiogenic arterial thromboembolism (CATE). Neutrophil extracellular traps (NETs) may be a potential biomarker and therapeutic target for cardiomyopathy in cats.Hypothesis/ObjectivesCharacterize NETs in cats with HCM or CATE. We hypothesized that circulating NETs assessed in the form of cell‐free DNA (cfDNA) and citrullinated histone H3 (citH3) are increased in cats with HCM and CATE and associated with reported predisposing factors for thrombus formation.AnimalsEighty‐five cats including client‐owned cats with HCM and CATE and staff‐ and student‐owned clinically healthy cats without HCM.MethodsAfter echocardiographic evaluations, NETs were measured as cfDNA and citH3.ResultsCats with CATE had significant increases in cfDNA (11.2 ng/μL; interquartile range [IQR], 8.1 to 29.6) compared to those without HCM (8.2 ng/μL; IQR, 5.7 to 11.7 μL; P = .01) and were responsible for 75% to 83% of cases with cfDNA fragments sized 100 to 2000 base pairs. Citrullinated histone 3, detected in 52% of cats with HCM (31.1 ng/mL; IQR, 16.9 to 29.8), was significantly lower than in those with CATE (48.2 ng/mL; IQR, 34.2 to 60.2; P = .007). The citH3 concentrations correlated significantly with reported risk factors of CATE, such as left atrial auricular velocity.Conclusions and Clinical ImportanceNeutrophil extracellualr traps, especially citH3, are increased in cats with HCM and CATE. They may serve as a novel therapeutic target and biomarker of thrombosis in cats with HCM.}, number={2}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Li, Ronald H. L. and Fabella, Arianne and Nguyen, Nghi and Kaplan, Joanna L. and Ontiveros, Eric and Stern, Joshua A.}, year={2023}, month={Mar}, pages={490–502} }
@article{walker_li_nguyen_jauregui_meurs_gagnon_stern_2023, title={Evaluation of autoantibodies to desmoglein-2 in dogs with and without cardiac disease}, volume={13}, ISSN={2045-2322}, url={http://dx.doi.org/10.1038/s41598-023-32081-x}, DOI={10.1038/s41598-023-32081-x}, abstractNote={AbstractAutoantibodies to desmoglein-2 have been associated with arrhythmogenic right ventricular cardiomyopathy (ARVC) in people. ARVC is a common disease in the Boxer dog. The role of anti-desmoglein-2 antibodies in Boxers with ARVC and correlation with disease status or severity is unknown. This prospective study is the first to evaluate dogs of various breeds and cardiac disease state for anti-desmoglein-2 antibodies. The sera of 46 dogs (10 ARVC Boxers, 9 healthy Boxers, 10 Doberman Pinschers with dilated cardiomyopathy, 10 dogs with myxomatous mitral valve disease, and 7 healthy non-Boxer dogs) were assessed for antibody presence and concentration via Western blotting and densitometry. Anti-desmoglein-2 antibodies were detected in all dogs. Autoantibody expression did not differ between study groups and there was no correlation with age or body weight. In dogs with cardiac disease, there was weak correlation with left ventricular dilation (r = 0.423, p = 0.020) but not left atrial size (r = 0.160, p = 0.407). In ARVC Boxers there was strong correlation with the complexity of ventricular arrhythmias (r = 0.841, p = 0.007) but not total number of ectopic beats (r = 0.383, p = 0.313). Anti-desmoglein-2 antibodies were not disease specific in the studied population of dogs. Correlation with some measures of disease severity requires further study with larger populations.}, number={1}, journal={Scientific Reports}, publisher={Springer Science and Business Media LLC}, author={Walker, Ashley L. and Li, Ronald H. L. and Nguyen, Nghi and Jauregui, Carina E. and Meurs, Kathryn M. and Gagnon, Allison L. and Stern, Joshua A.}, year={2023}, month={Mar} }
@article{rank_lynch_ruterbories_li_ueda_2023, title={Evaluation of thrombin generation in dogs administered clopidogrel}, volume={10}, ISSN={2297-1769}, url={http://dx.doi.org/10.3389/fvets.2023.1194242}, DOI={10.3389/fvets.2023.1194242}, abstractNote={IntroductionThe antiplatelet effect of clopidogrel can vary between patients. A modified thromboelastography (TEG) protocol (TEG-Platelet Mapping assay® [TEG-PM]) can be used for clopidogrel monitoring but is not widely available. Thrombin generation (TG) assays could offer a novel alternative. The main objective of this pilot study was to assess TG assay variables (lag time, peak, endogenous thrombin potential [ETP]) in dogs before and after 7 days of clopidogrel administration and compare with TEG-PM variables (maximum amplitude [MA]-ADP and percentage (%) inhibition).MethodsSix healthy mix-breed dogs were enrolled in this pilot study. Blood samples for platelet count, TG assays, and TEG-PM were obtained at two time points, corresponding to baseline, and after 7 days of clopidogrel administration (mean 2.3 +/− 0.3 mg/kg PO q24 hours). Data were then compared with a Student’s t-test.ResultsThere was no significant change in TG assay variables performed on platelet poor plasma after 7 days of clopidogrel administration: lag time (Day 1: 1.8 +/− 0.2 min, Day 7: 1.8 +/− 0.2 min, p = 0.42); peak (Day 1: 76 +/− 7 nM, Day 7: 72 +/− 10 nM, p = 0.49); and ETP (Day 1: 399 +/− 27 nM*min, Day 7: 392 +/− 32 nM*min; p = 0.49). There were significant changes in TEG MA-ADP (Day 1: 19 +/− 8 mm, Day 7: 9 +/− 6 mm, p = 0.04) and % inhibition (Day 1: 58 +/− 27, Day 7: 99 +/− 0.3, p = 0.02).DiscussionClopidogrel administration did not lead to changes in TG assay variables performed on platelet poor plasma samples, despite concomitant changes in TEG-PM variables consistent with platelet inhibition. Based on this pilot study, thrombin generation performed on platelet poor plasma may not be a useful antiplatelet monitoring tool in dogs.}, journal={Frontiers in Veterinary Science}, publisher={Frontiers Media SA}, author={Rank, Kaitlyn and Lynch, Alex M. and Ruterbories, Laura K. and Li, Ronald H. L. and Ueda, Yu}, year={2023}, month={Aug} }
@article{bannasch_oertle_vo_batcher_stern_kaplan_li_madden_christen_leeb_et al._2023, title={Naturally occurring canine laminopathy leading to a dilated and fibrosing cardiomyopathy in the Nova Scotia Duck Tolling Retriever}, volume={13}, ISSN={2045-2322}, url={http://dx.doi.org/10.1038/s41598-023-46601-2}, DOI={10.1038/s41598-023-46601-2}, abstractNote={AbstractDilated cardiomyopathy (DCM) is characterized by decreased systolic function and dilation of one or both ventricles, often leading to heart failure or sudden death. Two 10-month-old sibling Nova Scotia Duck Tolling Retrievers (NSDTR) died acutely with evidence of dilated cardiomyopathy with myocardial fibrosis. Association analysis using two cases and 35 controls identified three candidate regions homozygous in the two cases. Whole genome sequencing identified a frameshift deletion in the LMNA gene (NC_049228.1:g.41688530del, NP_001274080:p.(Asp576ThrfsTer124)). Three retrospectively identified NSDTRs with sudden death before 2 years of age and severe myocardial fibrosis were also homozygous for the deletion. One 5 year old with sudden death and myocardial fibrosis was heterozygous for the deletion. This variant was not identified in 722 dogs of other breeds, nor was it identified to be homozygous in 784 NSDTR. LMNA codes for lamin A/C proteins, which are type V intermediate filaments that provide structural support to the nuclear membrane. In humans, LMNA variants can cause DCM with sudden death as well as diseases of striated muscles, lipodystrophy, neuropathies, and accelerated aging disorders. This frameshift deletion is predicted to affect processing of prelamin A into lamin A. Pedigree analysis in the NSDTR and functional evaluation of heterozygotes is consistent with a predominantly recessive mode of inheritance and possibly low penetrance in heterozygotes in contrast to people, where most pathogenic LMNA variants are dominantly inherited.}, number={1}, journal={Scientific Reports}, publisher={Springer Science and Business Media LLC}, author={Bannasch, Danika L. and Oertle, Danielle T. and Vo, Julia and Batcher, Kevin L. and Stern, Joshua A. and Kaplan, Joanna L. and Li, Ronald H. L. and Madden, Indiana E. and Christen, Matthias and Leeb, Tosso and et al.}, year={2023}, month={Nov}, pages={19077} }
@article{shaverdian_li_2023, title={Preventing Cardiogenic Thromboembolism in Cats: Literature Gaps, Rational Recommendations, and Future Therapies}, volume={53}, ISSN={0195-5616}, url={http://dx.doi.org/10.1016/j.cvsm.2023.06.002}, DOI={10.1016/j.cvsm.2023.06.002}, abstractNote={Cardiogenic arterial thromboembolism (CATE) is a devastating complication in cats with cardiomyopathies with significant morbidity and mortality. Despite recent advances in the diagnosis and treatment of CATE, its recurrence and mortality remain high. This highlights the urgent need for a greater understanding of CATE pathophysiology so that novel diagnostic tests and therapeutics can be developed. This comprehensive review aims to summarize existing literature on pathophysiology, clinical diagnosis, and current recommendations on the prevention and treatment of CATE. It also identifies and describes knowledge gaps and research priorities in the roles of immunothrombosis and procoagulant platelets in the pathogenesis of CATE.}, number={6}, journal={Veterinary Clinics of North America: Small Animal Practice}, publisher={Elsevier BV}, author={Shaverdian, Meg and Li, Ronald H.L.}, year={2023}, month={Nov}, pages={1309–1323} }
@article{lo_li_georges_nguyen_chen_stuhlmann_oldach_rivas_fousse_harris_et al._2023, title={Synergistic inhibitory effects of clopidogrel and rivaroxaban on platelet function and platelet‐dependent thrombin generation in cats}, volume={37}, ISSN={0891-6640 1939-1676}, url={http://dx.doi.org/10.1111/jvim.16727}, DOI={10.1111/jvim.16727}, abstractNote={AbstractBackgroundDual antithrombotic treatment (DAT) with clopidogrel and rivaroxaban sometimes is prescribed to cats with hypertrophic cardiomyopathy at risk of thromboembolism. To date, no studies have evaluated their combined effects on platelet function.Objectives/HypothesisEvaluate the safety of DAT in healthy cats and compare, ex vivo, platelet‐dependent thrombin generation and agonist‐induced platelet activation and aggregation in cats treated with clopidogrel, rivaroxaban, or DAT. We hypothesized that DAT would safely modulate agonist‐induced platelet activation and aggregation more effectively than single agent treatment.AnimalsNine apparently healthy 1‐year‐old cats selected from a research colony.MethodsUnblinded, nonrandomized ex vivo cross‐over study. All cats received 7 days of rivaroxaban (0.6 ± 0.1 mg/kg PO), clopidogrel (4.7 ± 0.8 mg/kg PO), or DAT with defined washout periods between treatments. Before and after each treatment, adenosine diphosphate (ADP)‐ and thrombin‐induced platelet P‐selectin expression was evaluated using flow cytometry to assess platelet activation. Platelet‐dependent thrombin generation was measured by fluorescence assay. Platelet aggregation was assessed using whole blood impedance platelet aggregometry.ResultsNo cats exhibited adverse effects. Of the 3 treatments, only DAT significantly decreased the number of activated platelets (P = .002), modulated platelet activation in response to thrombin (P = .01), dampened thrombin generation potential (P = .01), and delayed maximum reaction velocity (P = .004) in thrombin generation. Like clopidogrel, DAT inhibited ADP‐mediated platelet aggregation. However, rivaroxaban alone resulted in increased aggregation and activation in response to ADP.Conclusion and Clinical ImportanceTreatment combining clopidogrel and rivaroxaban (DAT) safely decreases platelet activation, platelet response to agonists, and thrombin generation in feline platelets more effectively than monotherapy with either clopidogrel or rivaroxaban.}, number={4}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Lo, Sara T. and Li, Ronald H. L. and Georges, Catherine J. and Nguyen, Nghi and Chen, Cheyenne K. and Stuhlmann, Claire and Oldach, Maureen Sigmund and Rivas, Victor Noel and Fousse, Samantha and Harris, Samantha P. and et al.}, year={2023}, month={May}, pages={1390–1400} }
@article{delaforcade_bacek_blais_boyd_brainard_chan_cortellini_goggs_hoareau_koenigshof_et al._2022, title={2022 Update of the Consensus on the Rational Use of Antithrombotics and Thrombolytics in Veterinary Critical Care (CURATIVE) Domain 1‐ Defining populations at risk}, volume={32}, ISSN={1479-3261 1476-4431}, url={http://dx.doi.org/10.1111/vec.13204}, DOI={10.1111/vec.13204}, abstractNote={AbstractObjectivesTo expand the number of conditions and interventions explored for their associations with thrombosis in the veterinary literature and to provide the basis for prescribing recommendations.DesignA population exposure comparison outcome format was used to represent patient, exposure, comparison, and outcome. Population Exposure Comparison Outcome questions were distributed to worksheet authors who performed comprehensive searches, summarized the evidence, and created guideline recommendations that were reviewed by domain chairs. The revised guidelines then underwent the Delphi survey process to reach consensus on the final guidelines. Diseases evaluated in this iteration included heartworm disease (dogs and cats), immune‐mediated hemolytic anemia (cats), protein‐losing nephropathy (cats), protein‐losing enteropathy (dogs and cats), sepsis (cats), hyperadrenocorticism (cats), liver disease (dogs), congenital portosystemic shunts (dogs and cats) and the following interventions: IV catheters (dogs and cats), arterial catheters (dogs and cats), vascular access ports (dogs and cats), extracorporeal circuits (dogs and cats) and transvenous pacemakers (dogs and cats).ResultsOf the diseases evaluated in this iteration, a high risk for thrombosis was defined as heartworm disease or protein‐losing enteropathy. Low risk for thrombosis was defined as dogs with liver disease, cats with immune‐mediated hemolytic anemia, protein‐losing nephropathy, sepsis, or hyperadrenocorticism.ConclusionsAssociations with thrombosis are outlined for various conditions and interventions and provide the basis for management recommendations. Numerous knowledge gaps were identified that represent opportunities for future studies.}, number={3}, journal={Journal of Veterinary Emergency and Critical Care}, publisher={Wiley}, author={deLaforcade, Armelle and Bacek, Lenore and Blais, Marie‐Claude and Boyd, Corrin and Brainard, Benjamin M and Chan, Daniel L. and Cortellini, Stefano and Goggs, Robert and Hoareau, Guillaume L and Koenigshof, Amy and et al.}, year={2022}, month={May}, pages={289–314} }
@article{fowler_bolton_rossmeisl_arendse_vernau_li_parker_2022, title={Clinical, Diagnostic, and Imaging Findings in Three Juvenile Dogs With Paraspinal Hyperesthesia or Myelopathy as a Consequence of Hemophilia A: A Case Report}, volume={9}, ISSN={2297-1769}, url={http://dx.doi.org/10.3389/fvets.2022.871029}, DOI={10.3389/fvets.2022.871029}, abstractNote={Three juvenile dogs presented with an acute onset of paraspinal hyperesthesia and/or neurologic deficits. These dogs underwent anesthesia for MRI and additional diagnostics. The thoracolumbar MRI in Dog 1 revealed an accumulation of T2-weighted (T2W) hyperintense, T1-weighted (T1W) iso- to hyperintense, contrast enhancing extradural material. The differential diagnoses were meningitis with secondary hemorrhage or empyema or late subacute hemorrhage. The initial cervical MRI in Dog 2 revealed T1W meningeal contrast enhancement suspected to be secondary to meningitis. A repeat MRI following neurologic decline after CSF sampling revealed a large area of T2W and T1W hyperintensity between fascial planes of the cervical musculature as well as T2W iso- to hyperintense and T1W iso- to hypointense extradural material at the level of C1 consistent with hemorrhage. The cervical MRI in Dog 3 revealed T2W hyperintense and T1W iso- to hypointense extradural compressive material consistent with hemorrhage. Dogs 1 and 2 underwent CSF sampling and developed complications, including subcutaneous hematoma and vertebral canal hemorrhage. Dog 3 underwent surgical decompression, which revealed a compressive extradural hematoma. In each case, a hemophilia panel including factor VIII concentration confirmed the diagnosis of hemophilia A. Dog 1 had a resolution of clinical signs for ~5 months before being euthanized from gastrointestinal hemorrhage. Dog 2 was euthanized due to neurologic decompensation following CSF sampling. Dog 3 did well for 2 weeks after surgery but was then lost to follow-up. This case series provides information on clinical signs, MRI findings, and outcome in 3 juvenile dogs with hemophilia A that developed neurologic deficits or paraspinal hyperesthesia secondary to spontaneous or iatrogenic vertebral canal hemorrhage. Hemophilia A should be considered as a differential in any young dog presenting with an acute onset of hyperesthesia with or without neurologic deficits. This diagnosis should be prioritized in young male dogs that have other evidence of hemorrhage on physical exam.}, journal={Frontiers in Veterinary Science}, publisher={Frontiers Media SA}, author={Fowler, Kayla M. and Bolton, Timothy A. and Rossmeisl, John H. and Arendse, Avril U. and Vernau, Karen M. and Li, Ronald H. L. and Parker, Rell L.}, year={2022}, month={Apr}, pages={871029} }
@article{tan_li_ueda_stern_hussain_haginoya_sharpe_gunther-harrington_epstein_nguyen_2022, title={Platelet Priming and Activation in Naturally Occurring Thermal Burn Injuries and Wildfire Smoke Exposure Is Associated With Intracardiac Thrombosis and Spontaneous Echocardiographic Contrast in Feline Survivors}, volume={9}, ISSN={2297-1769}, url={http://dx.doi.org/10.3389/fvets.2022.892377}, DOI={10.3389/fvets.2022.892377}, abstractNote={Wildfires pose a major health risk for humans, wildlife, and domestic animals. We previously discovered pathophysiologic parallels between domestic cats with naturally occurring smoke inhalation and thermal burn injuries and human beings with similar injuries; these were characterized by transient myocardial thickening, cardiac troponin I elevation and formation of intracardiac thrombosis. While the underlying mechanisms remain unclear, results from murine models suggest that platelet priming and activation may contribute to a global hypercoagulable state and thrombosis. Herein, we evaluated and compared the degree of platelet activation, platelet response to physiologic agonists and levels of platelet-derived microvesicles (PDMV) in 29 cats with naturally occurring wildfire thermal injuries (WF), 21 clinically healthy cats with subclinical hypertrophic cardiomyopathy (HCM) and 11 healthy cats without HCM (CC). We also quantified and compared circulating PDMVs in WF cats to CC cats. In addition, we examined the association between thrombotic events, severity of burn injuries, myocardial changes, and the degree of platelet activation in cats exposed to wildfires. Flow cytometric detection of platelet surface P-selectin expression showed that WF cats had increased platelet response to adenosine diphosphate (ADP) and thrombin compared to the two control groups indicating the presence of primed platelets in circulation. In addition, cats in the WF group had increased circulating levels of PDMV, characterized by increased phosphatidylserine on the external leaflet. Cats in the WF group with documented intracardiac thrombosis had elevated platelet activation and platelet priming in the presence of ADP. While high dose arachidonic acid (AA) mostly resulted in platelet inhibition, persistent response to AA was noted among cats in the WF group with intracardiac thrombosis. Univariate and multiple logistic regression analyses demonstrated that increased platelet response to AA was independently associated with thrombotic events. This is the first study reporting the significant association between platelet priming and intracardiac thrombosis in domestic cats with naturally occurring wildfire-related injuries and smoke inhalation. Further studies are required to delineate additional mechanisms between inflammation and thrombosis, especially regarding platelet primers and the cyclooxygenase pathway.One Sentence SummaryPlatelet activation and shedding of platelet-derived microvesicles due to platelet priming is present following naturally occurring wildfire smoke exposure and thermal burn injuries in a population of domestic cats.}, journal={Frontiers in Veterinary Science}, publisher={Frontiers Media SA}, author={Tan, Avalene W. K. and Li, Ronald H. L. and Ueda, Yu and Stern, Joshua A. and Hussain, Mehrab and Haginoya, Satoshi and Sharpe, Ashely N. and Gunther-Harrington, Catherine T. and Epstein, Steven E. and Nguyen, Nghi}, year={2022}, month={Jul} }
@article{ueda_li_nguyen_ontiveros_kovacs_oldach_vernau_court_stern_2021, title={A genetic polymorphism in P2RY(1) impacts response to clopidogrel in cats with hypertrophic cardiomyopathy}, volume={11}, ISSN={["2045-2322"]}, url={https://europepmc.org/articles/PMC8206363}, DOI={10.1038/s41598-021-91372-3}, abstractNote={AbstractClopidogrel is converted to its active metabolite by cytochrome P450 isoenzymes and irreversibly inhibits platelet activation by antagonizing the adenosine-diphosphate (ADP) receptor. It is frequently used in cats with hypertrophic cardiomyopathy (HCM) to prevent thromboembolic complications. However, significant interpatient variability of the response to clopidogrel therapy has been suspected. In this study, we assessed the impact of single nucleotide polymorphisms (SNPs) within ADP receptor (P2RY1, P2RY12) and cytochrome P450 isoenzyme (CYP2C41) genes on platelet inhibition by clopidogrel administration in cats with HCM. Forty-nine cats completed the study, and blood samples were obtained before and after clopidogrel therapy to assess the degree of platelet inhibition based on flow cytometry and whole blood platelet aggregometry. Plasma concentrations of clopidogrel metabolites were measured after the last dose of clopidogrel. Whole blood platelet aggregometry revealed a significant reduction of platelet inhibition by clopidogrel in cats with the P2RY1:A236G and the P2RY12:V34I variants. The association with the P2RY1:A236G variant and clopidogrel resistance remained significant after adjustment for multiple comparisons. This study demonstrated that a genetic polymorphism in the P2RY1 gene altered response to clopidogrel therapy and suggests that clinicians may consider alternative or additional thromboprophylactic therapy in cats with the P2RY1:A236G variant.}, number={1}, journal={SCIENTIFIC REPORTS}, author={Ueda, Yu and Li, Ronald H. L. and Nguyen, Nghi and Ontiveros, Eric S. and Kovacs, Samantha L. and Oldach, Maureen S. and Vernau, Karen M. and Court, Michael H. and Stern, Joshua A.}, year={2021}, month={Jun} }
@article{lo_walker_georges_li_stern_2021, title={Dual therapy with clopidogrel and rivaroxaban in cats with thromboembolic disease}, volume={24}, ISSN={1098-612X 1532-2750}, url={http://dx.doi.org/10.1177/1098612x211013736}, DOI={10.1177/1098612x211013736}, abstractNote={Objectives Feline arterial thromboembolism (ATE), an often devastating outcome, was recently shown to affect 11.3% of cats with hypertrophic cardiomyopathy over 10 years. Current American College of Veterinary Internal Medicine guidelines recommend the use of clopidogrel in cats at risk for ATE, with addition of a factor Xa inhibitor in very high risk or post-ATE cases. To date, no studies have examined the safety or efficacy of this combined antithrombotic therapy. This retrospective case series aimed to assess the frequency and type of adverse events that occurred in cats prescribed dual clopidogrel and rivaroxaban therapy. Secondary aims were to evaluate indications for dual therapy and clinical outcome. Methods The study included 32 cats prescribed clopidogrel (18.75 mg PO q24h) and rivaroxaban (2.5 mg PO q24h) on an outpatient basis over a 5-year period. Results Cats were prescribed dual therapy for at least one of the following: ATE event (n = 18), presence of an intracardiac thrombi (n = 17) or presence of spontaneous echocardiographic contrast (SEC) (n = 16). Five cats experienced adverse effects that could be attributed to medications, a median of 13 days from initiation (epistaxis, hematemesis, hematochezia or hematuria). No cat required hospitalization as a result of these events. Median survival time from onset of therapy was 257 days (interquartile range [IQR] = 38–497) for all cats, 502 days (IQR = 171–663) for ATE cats, 725 days (IQR = 133–856) for cats with an ATE to two or more limbs and 301 days (IQR = 221–431) for cats with only one limb affected. Recurrence rate of ATE while on dual therapy was 16.7%; no cat newly developed an ATE while on dual therapy. Conclusions and relevance Dual antithrombotic therapy with clopidogrel and rivaroxaban resulted in a low reported incidence of adverse events. Cats placed on dual therapy for an ATE event experienced a low rate of recurrence and effective thromboprophylaxis was achieved in cats with intracardiac thrombi or SEC. }, number={4}, journal={Journal of Feline Medicine and Surgery}, publisher={SAGE Publications}, author={Lo, Sara T and Walker, Ashley L and Georges, Catherine J and Li, Ronald HL and Stern, Joshua A}, year={2021}, month={May}, pages={277–283} }
@article{li_hommel_nguyen_2021, title={Lipopolysaccharide-Activated Canine Platelets Upregulate High Mobility Group Box-1 via Toll-Like Receptor 4}, volume={8}, url={https://europepmc.org/articles/PMC8255672}, DOI={10.3389/fvets.2021.674678}, abstractNote={High mobility group box-1 (HMGB1) and the toll-like receptor 4 (TLR4) axis is a key mediator of inflammation. Platelet-derived high mobility group box-1 (HMGB1) may also play a critical role in sepsis-mediated thrombosis resulting in complications like disseminated intravascular coagulation and multiple organ failure. While elevated levels of HMGB1 have been documented in humans and dogs with systemic inflammatory response syndrome and sepsis, a better understanding of how platelet agonists and lipopolysaccharide (LPS) mediate platelet HMGB1 expression would open doors to novel therapies for sepsis-mediated thrombosis. Herein, we sought to determine if canine platelets express HMGB1 in the presence or absence of LPS and agonists (ADP or thrombin) and if surface expression of HMGB1 is dependent on platelet TLR4. Canine platelets were unstimulated (resting) or activated with thrombin or adenosine diphosphate (ADP) in the presence or absence of Escherichia coli LPS prior to flow cytometric and western blot analyses for HMGB1 expression. We also treated canine platelets with or without TLR4 function blocking antibody or its isotype control. We discovered that while thrombin upregulated both surface and cellular HMGB1 expression, LPS-mediated activation in the presence of ADP priming led to upregulation of surface HMGB1 expression. This expression was found to be most prominent in platelets that had undergone alpha-granule secretion. Inhibition of TLR4 attenuated LPS-induced HMGB1 expression indicating that exteriorization of HMGB1 may be dependent on the non-genomic pathway of platelet TLR4. Our findings indicate that upregulation of platelet-derived HMGB1 occurs as a result of thrombin or TLR4-mediated activation in dogs. Future studies should explore the translational implication of platelet-derived HMGB1 as novel therapeutic targets in humans and dogs with sepsis.}, journal={Frontiers in Veterinary Science Experimental and Diagnostic Pathology}, author={Li, R.H.L. and Hommel, C.T. and Nguyen, N.}, year={2021}, month={Jun}, pages={674678} }
@article{lyons_buckley_harvey_2021, title={Mining the 99 Lives Cat Genome Sequencing Consortium database implicates genes and variants for the Ticked locus in domestic cats (Felis catus)}, volume={52}, url={https://europepmc.org/articles/PMC8252059}, DOI={10.1111/age.13059}, abstractNote={SummaryTabby patterns of fur coats are defining characteristics in wild and domestic felids. Historically, three autosomal alleles at one locus (Tabby): Abyssinian (Ta; a.k.a. ticked), mackerel (Tm; a.k.a. striped) and blotched (tb; a.k.a. classic, blotched) were thought to control these patterns in domestic cats and their breeds. Currently, at least three loci influence cat tabby markings, two of which are designated Tabby and Ticked. The Tabby locus is laeverin (LVRN) and affects the mackerel and blotched patterns. The unidentified gene for the Ticked locus on cat chromosome B1 was suggested to control the presence or absence of the ticked pattern (Tabby – Abyssinian (Ta; a.k.a. ticked). The cat reference genome (Cinnamon, the Abyssinian) has the ticked phenotype and the variant dataset and coat phenotypes from the 99 Lives Cat Genome Consortium (195 cats) were used to identify candidate genes and variants associated with the Ticked locus. Two strategies were used to find the Ticked allele(s), one considered Cinnamon with the reference allele or heterozygous (Strategy A) and the other considered Cinnamon as having the variant allele or heterozygous (Strategy B). For Strategy A, two variants in Dickkopf Wnt Signaling Pathway Inhibitor 4 (DKK4), a p.Cys63Tyr (B1:41621481, c.188G>A) and a less common p.Ala18Val (B1:42620835, c.53C>T) variant are suggested as two alleles influencing the Ticked phenotype. Bioinformatic and molecular modeling analysis suggests that these changes disrupt a key disulfide bond in the Dkk4 cysteine‐rich domain 1 or Dkk4 signal peptide cleavage respectively. All coding variants were excluded as Ticked alleles using Strategy B.}, number={3}, journal={Animal genetics}, author={Lyons, LA and Buckley, RM and Harvey, RJ}, year={2021}, pages={321—332} }
@article{genova_nonnis_maffioli_tedeschi_strillacci_carisetti_sironi_cupaioli_di nanni_mezzelani_et al._2021, title={Multi-omic analyses in Abyssinian cats with primary renal amyloid deposits}, volume={11}, ISSN={2045-2322}, url={http://dx.doi.org/10.1038/s41598-021-87168-0}, DOI={10.1038/s41598-021-87168-0}, abstractNote={AbstractThe amyloidoses constitute a group of diseases occurring in humans and animals that are characterized by abnormal deposits of aggregated proteins in organs, affecting their structure and function. In the Abyssinian cat breed, a familial form of renal amyloidosis has been described. In this study, multi-omics analyses were applied and integrated to explore some aspects of the unknown pathogenetic processes in cats. Whole-genome sequences of two affected Abyssinians and 195 controls of other breeds (part of the 99 Lives initiative) were screened to prioritize potential disease-associated variants. Proteome and miRNAome from formalin-fixed paraffin-embedded kidney specimens of fully necropsied Abyssinian cats, three affected and three non-amyloidosis-affected were characterized. While the trigger of the disorder remains unclear, overall, (i) 35,960 genomic variants were detected; (ii) 215 and 56 proteins were identified as exclusive or overexpressed in the affected and control kidneys, respectively; (iii) 60 miRNAs were differentially expressed, 20 of which are newly described. With omics data integration, the general conclusions are: (i) the familial amyloid renal form in Abyssinians is not a simple monogenic trait; (ii) amyloid deposition is not triggered by mutated amyloidogenic proteins but is a mix of proteins codified by wild-type genes; (iii) the form is biochemically classifiable as AA amyloidosis.}, number={1}, journal={Scientific Reports}, publisher={Springer Science and Business Media LLC}, author={Genova, Francesca and Nonnis, Simona and Maffioli, Elisa and Tedeschi, Gabriella and Strillacci, Maria Giuseppina and Carisetti, Michela and Sironi, Giuseppe and Cupaioli, Francesca Anna and Di Nanni, Noemi and Mezzelani, Alessandra and et al.}, year={2021}, month={Apr} }
@article{li_nguyen_stern_duler_2021, title={Neutrophil extracellular traps in feline cardiogenic arterial thrombi: a pilot study}, volume={24}, ISSN={1098-612X 1532-2750}, url={http://dx.doi.org/10.1177/1098612x211044986}, DOI={10.1177/1098612x211044986}, abstractNote={Objectives The aim of this study was to investigate the spatial distribution of neutrophil extracellular traps (NETs) in cardiogenic arterial thromboembolism (CATE). Specifically, we aimed to examine the related structural features of NETs in feline arterial thrombi in relation to their arterial locations. Methods Paraffin-embedded aortic bifurcations from nine cats with hypertrophic cardiomyopathy (four with CATE and five without) were deparaffinized, and NETs were identified by immunodetection based on colocalization of cell-free DNA, citrullinated histone H3 and neutrophil elastase. The distribution of NETs in thrombi within the aortic bifurcations and common iliac arteries (CIAs) was compared based on their proximity to the descending aorta (proximal, mid, distal). Ten random fields per section were captured at × 10 and × 20 magnification for each section of the clot and analyzed. Results The distributions of NETs in thrombi within the aortic bifurcation and CIAs were found to differ in relation to their assigned zones (proximal, mid, distal; P = 0.04); NETs were concentrated mostly in the proximal region in the aortic bifurcations (47.56%, interquartile range [IQR] 14.07–77.95) and CIAs (44.69%, IQR 24.65–85.28), compared with the distal regions (2.69%, IQR 0.10–50.04 [P = 0.027]; 7.08%, IQR 1.27–59.33 [P = 0.02]). Conclusions and relevance The variation in NET distribution within arterial thrombi may shed light on the pathogenesis of thrombus growth. This may be due to possible neutrophil entrapment or variations in shear stress. }, number={6}, journal={Journal of Feline Medicine and Surgery}, publisher={SAGE Publications}, author={Li, Ronald HL and Nguyen, Nghi and Stern, Joshua A and Duler, Laetitia M}, year={2021}, month={Sep}, pages={580–586} }
@article{li_tan_ueda_stern_hussain_haginoya_sharpe_gunther-harrington_epstein_nguyen_2021, title={Platelet Priming Following Naturally Occurring Thermal Burn Injuries And Wildfire Smoke Exposure is Associated with Intracardiac Thrombosis in Feline Survivors}, url={https://doi.org/10.21203/rs.3.rs-778724/v1}, DOI={10.21203/rs.3.rs-778724/v1}, abstractNote={Abstract
Wildfires pose a major health risk for humans, wildlife, and domestic animals. We previously discovered pathophysiologic parallels between domestic cats with naturally occurring smoke inhalation and thermal burn injuries and human beings with similar injuries; these were characterized by transient myocardial thickening, cardiac troponin I elevation and formation of intracardiac thrombosis. While the underlying mechanisms remain unclear, results from murine models suggest that platelet priming and activation may contribute to a global hypercoagulable state and thrombosis. Herein, we evaluated and compared the degree of platelet activation, platelet response to physiologic agonists and levels of platelet-derived microvesicles (PDMV) in 29 cats with naturally occurring wildfire thermal injuries, 21 clinically healthy cats with subclinical hypertrophic cardiomyopathy (HCM) and 11 healthy cats without HCM (CC). We also quantified and compared circulating PDMVs in WF cats to CC cats. In addition, we examined the association between thrombotic events, severity of burn injuries, myocardial changes, and the degree of platelet activation in cats exposed to wildfires. Flow cytometric detection of platelet surface P-selectin expression showed that cats in the wildfire group (WF) had increased platelet response to adenosine diphosphate (ADP) and thrombin compared to the 2 control groups indicating the presence of primed platelets in circulation. In addition, cats in the WF group had increased circulating levels of PDMV, characterized by increased phosphatidylserine on the external leaflet. Cats in the WF group with documented intracardiac thrombosis had elevated platelet activation and platelet priming in the presence of ADP or thrombin. While high dose arachidonic acid (AA) mostly resulted in platelet inhibition, persistent platelet activation in response to AA was noted among cats in the WF group with intracardiac thrombosis. Univariate and multiple logistic regression analyses demonstrated that increased platelet response to AA was independently associated with thrombotic events. This is the first study reporting the significant association between platelet priming and intracardiac thrombosis in domestic cats with naturally occurring wildfire-related injuries and smoke inhalation. Further studies are required to delineate additional mechanisms between inflammation and thrombosis, especially regarding platelet primers and the cyclooxygenase pathway.}, publisher={Research Square}, author={Li, Ronald and Tan, Avalene and Ueda, Yu and Stern, Joshua and Hussain, Mehrab and Haginoya, Satoshi and Sharpe, Ashley and Gunther-Harrington, Catherine and Epstein, Steven and Nguyen, Nghi}, year={2021} }
@article{yu_creighton_buckley_lyons_li_2020, title={A Deletion in GDF7 is Associated with a Heritable Forebrain Commissural Malformation Concurrent with Ventriculomegaly and Interhemispheric Cysts in Cats}, volume={11}, ISSN={2073-4425}, url={http://dx.doi.org/10.3390/genes11060672}, DOI={10.3390/genes11060672}, abstractNote={An inherited neurologic syndrome in a family of mixed-breed Oriental cats has been characterized as forebrain commissural malformation, concurrent with ventriculomegaly and interhemispheric cysts. However, the genetic basis for this autosomal recessive syndrome in cats is unknown. Forty-three cats were genotyped on the Illumina Infinium Feline 63K iSelect DNA Array and used for analyses. Genome-wide association studies, including a sib-transmission disequilibrium test and a case-control association analysis, and homozygosity mapping, identified a critical region on cat chromosome A3. Short-read whole genome sequencing was completed for a cat trio segregating with the syndrome. A homozygous 7 bp deletion in growth differentiation factor 7 (GDF7) (c.221_227delGCCGCGC [p.Arg74Profs]) was identified in affected cats, by comparison to the 99 Lives Cat variant dataset, validated using Sanger sequencing and genotyped by fragment analyses. This variant was not identified in 192 unaffected cats in the 99 Lives dataset. The variant segregated concordantly in an extended pedigree. In mice, GDF7 mRNA is expressed within the roof plate when commissural axons initiate ventrally-directed growth. This finding emphasized the importance of GDF7 in the neurodevelopmental process in the mammalian brain. A genetic test can be developed for use by cat breeders to eradicate this variant.}, number={6}, journal={Genes}, publisher={MDPI AG}, author={Yu, Yoshihiko and Creighton, Erica K. and Buckley, Reuben M. and Lyons, Leslie A. and Li, Ronald}, year={2020}, month={Jun}, pages={672} }
@article{li_nguyen_rosati_jandrey_2020, title={Assessment of P2Y12 Inhibition by Clopidogrel in Feline Platelets Using Flow Cytometry Quantification of Vasodilator-Stimulated Phosphoprotein Phosphorylation}, volume={7}, ISSN={2297-1769}, url={http://dx.doi.org/10.3389/fvets.2020.00267}, DOI={10.3389/fvets.2020.00267}, abstractNote={The primary objective of this study was to evaluate a novel flow cytometry-based assay of quantifying platelet phosphorylation of vasodilator-stimulated phosphoprotein (P-VASP) in cats that received clopidogrel treatment. Eight healthy cats received 18.75 mg PO q24h of clopidogrel for 7 days. Prior to and after clopidogrel treatment, blood was collected for ADP-induced light transmission aggregometry (LTA) and P-VASP measurement by flow cytometry. Flow cytometry measurement of P-VASP levels was used to derive platelet reactivity index (PRI) before and after clopidogrel treatment. Based on P-VASP and LTA findings, platelet response to ADP was significantly attenuated after 7 days of clopidogrel treatment. By eliciting the competing platelet pathways of P2Y12 and cAMP using ADP and PGE1, respectively, ADP had no effect on P-VASP levels following clopidogrel treatment (p = 0.94). Clopidogrel also significantly decreased PRI from 28.84 ± 28.52% to 1.69 ± 12.39% (p = 0.0078). PRI on day 8 correlated moderately with the degree of slope inhibition on LTA (r = −0.4, p = 0.4). Flow cytometry analysis of P-VASP is effective at monitoring the inhibitory effects of clopidogrel on feline platelets.}, journal={Frontiers in Veterinary Science}, publisher={Frontiers Media SA}, author={Li, Ronald H. L. and Nguyen, Nghi and Rosati, Tommaso and Jandrey, Karl}, year={2020}, month={May}, pages={267} }
@article{sharpe_gunther-harrington_epstein_li_stern_2020, title={Cats with thermal burn injuries from California wildfires show echocardiographic evidence of myocardial thickening and intracardiac thrombi}, volume={10}, ISSN={2045-2322}, url={http://dx.doi.org/10.1038/s41598-020-59497-z}, DOI={10.1038/s41598-020-59497-z}, abstractNote={AbstractRecent increases in the prevalence and severity of wildfires in some regions have resulted in an increased frequency of veterinary burn patients. Few studies exist regarding diagnostics and management of burn wounds in veterinary patients and current knowledge is extrapolated from human literature and research models. Post-burn cardiac injury is a common finding and predictor of mortality in human patients and echocardiography is an important tool in monitoring response to therapy and predicting outcome. We describe the notable findings from cats naturally exposed to California wildfires in 2017 and 2018. Domestic cats (n = 51) sustaining burn injuries from the Tubbs (2017) and Camp (2018) wildfires were prospectively enrolled and serial echocardiograms and cardiac troponin I evaluations were performed. Echocardiograms of affected cats revealed a high prevalence of myocardial thickening (18/51) and spontaneous echocardiographic contrast and thrombi formation (16/51). Forty-two cats survived to discharge and 6 died or were euthanized due to a possible cardiac cause. For the first time, we describe cardiovascular and coagulation effects of thermal burn and smoke inhalation in cats. Further studies in veterinary burn victims are warranted and serve as a translational research opportunity for uncovering novel disease mechanisms and therapies.}, number={1}, journal={Scientific Reports}, publisher={Springer Science and Business Media LLC}, author={Sharpe, Ashley N. and Gunther-Harrington, Catherine T. and Epstein, Steven E. and Li, Ronald H. L. and Stern, Joshua A.}, year={2020}, month={Feb}, pages={2648} }
@article{duler_nguyen_ontiveros_li_2020, title={Identification of Neutrophil Extracellular Traps in Paraffin-Embedded Feline Arterial Thrombi using Immunofluorescence Microscopy}, volume={3}, ISSN={1940-087X}, url={http://dx.doi.org/10.3791/60834}, DOI={10.3791/60834}, abstractNote={Neutrophil extracellular traps (NETs), composed of cell-free DNA (cfDNA) and proteins like histones and neutrophil elastase (NE), are released by neutrophils in response to systemic inflammation or pathogens. Although NETs have previously been shown to augment clot formation and inhibit fibrinolysis in humans and dogs, the role of NETs in cats with cardiogenic arterial thromboembolism (CATE), a life-threatening complication secondary to hypertrophic cardiomyopathy, is unknown. A standardized method to identify and quantify NETs in cardiogenic arterial thrombi in cats will advance our understanding of their pathological role in CATE. Here, we describe a technique to identify NETs in formaldehyde-fixed and paraffin-embedded thrombi within the aortic bifurcation, extracted during necropsy. Following deparaffinization with xylene, aortic sections underwent indirect heat-induced antigen retrieval. Sections were then blocked, permeabilized, and ex vivo NETs were identified by colocalization of cell-free DNA (cfDNA), citrullinated histone H3 (citH3), and neutrophil elastase (NE) using immunofluorescence microscopy. To optimize the immunodetection of NETs in thrombi, autofluorescence of tissue elements was limited by using an autofluorescence quenching process prior to microscopy. This technique could be a useful tool to study NETs and thrombosis in other species and offers new insights into the pathophysiology of this complex condition.}, number={157}, journal={Journal of Visualized Experiments}, publisher={MyJove Corporation}, author={Duler, Laetitia and Nguyen, Nghi and Ontiveros, Eric and Li, Ronald H. L.}, year={2020}, month={Mar} }
@article{cogné_latypova_senaratne_martin_koboldt_kellaris_fievet_le meur_caldari_debray_et al._2020, title={Mutations in the Kinesin-2 Motor KIF3B Cause an Autosomal-Dominant Ciliopathy}, volume={106}, ISSN={0002-9297}, url={http://dx.doi.org/10.1016/j.ajhg.2020.04.005}, DOI={10.1016/j.ajhg.2020.04.005}, abstractNote={Kinesin-2 enables ciliary assembly and maintenance as an anterograde intraflagellar transport (IFT) motor. Molecular motor activity is driven by a heterotrimeric complex comprised of KIF3A and KIF3B or KIF3C plus one non-motor subunit, KIFAP3. Using exome sequencing, we identified heterozygous KIF3B variants in two unrelated families with hallmark ciliopathy phenotypes. In the first family, the proband presents with hepatic fibrosis, retinitis pigmentosa, and postaxial polydactyly; he harbors a de novo c.748G>C (p.Glu250Gln) variant affecting the kinesin motor domain encoded by KIF3B. The second family is a six-generation pedigree affected predominantly by retinitis pigmentosa. Affected individuals carry a heterozygous c.1568T>C (p.Leu523Pro) KIF3B variant segregating in an autosomal-dominant pattern. We observed a significant increase in primary cilia length in vitro in the context of either of the two mutations while variant KIF3B proteins retained stability indistinguishable from wild type. Furthermore, we tested the effects of KIF3B mutant mRNA expression in the developing zebrafish retina. In the presence of either missense variant, rhodopsin was sequestered to the photoreceptor rod inner segment layer with a concomitant increase in photoreceptor cilia length. Notably, impaired rhodopsin trafficking is also characteristic of recessive KIF3B models as exemplified by an early-onset, autosomal-recessive, progressive retinal degeneration in Bengal cats; we identified a c.1000G>A (p.Ala334Thr) KIF3B variant by genome-wide association study and whole-genome sequencing. Together, our genetic, cell-based, and in vivo modeling data delineate an autosomal-dominant syndromic retinal ciliopathy in humans and suggest that multiple KIF3B pathomechanisms can impair kinesin-driven ciliary transport in the photoreceptor.}, number={6}, journal={The American Journal of Human Genetics}, publisher={Elsevier BV}, author={Cogné, Benjamin and Latypova, Xenia and Senaratne, Lokuliyanage Dona Samudita and Martin, Ludovic and Koboldt, Daniel C. and Kellaris, Georgios and Fievet, Lorraine and Le Meur, Guylène and Caldari, Dominique and Debray, Dominique and et al.}, year={2020}, month={Jun}, pages={893–904} }
@article{li_ontiveros_nguyen_stern_lee_hardy_2020, title={Precision medicine identifies a pathogenic variant of the ITGA2B gene responsible for Glanzmann's thrombasthenia in a cat}, volume={34}, ISSN={0891-6640 1939-1676}, url={http://dx.doi.org/10.1111/jvim.15886}, DOI={10.1111/jvim.15886}, abstractNote={AbstractBackgroundA nonpedigreed male cat presented with epistaxis, severe bladder hemorrhage, and secondary urethral obstruction after cystocentesis.ObjectivesTo characterize the phenotype of a cat with bleeding diathesis and use a precision medicine approach to identify the molecular genetic defect by whole genome sequencing.MethodsAdenosine diphosphate (ADP) and arachidonic acid (AA)‐induced whole blood platelet aggregometry was performed in the affected cat and a healthy cat. Platelet activation, measured by P‐selectin expression, and surface integrin subunit β3 expression were evaluated by flow cytometry in the affected cat and healthy control. Total integrin subunit αIIb expression was assessed by western blot. Whole genome sequencing at 30× coverage was used to identify genetic variants that segregated in the affected cat compared to 194 cats from the 99 Lives Sequencing Consortium.ResultsPlatelet aggregometry identified significant impairment in platelet aggregation in response to ADP and AA compared to the control cat. Targeted protein expression analyses by flow cytometry and immunoblot analysis determined that the surface expression and total expression of the integrin, αIIbβ3, was absent. Whole genome sequencing identified a homozygous c.1986delC frameshift variant in the integrin subunit αIIb (ITGA2B) gene that was not detected in the control population. The p.Pro662fs (ITGA2B P662X) variant terminates translation of the protein at the extracellular domain of the integrin prematurely, which is predicted to affect expression of the β3 unit.Conclusions and Clinical ImportanceThis novel ITGA2B variant and the associated phenotype closely resemble Glanzmann's thrombasthenia, which has never been reported in cats.}, number={6}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Li, Ronald H. L. and Ontiveros, Eric and Nguyen, Nghi and Stern, Joshua A. and Lee, Elizabeth and Hardy, Brian T.}, year={2020}, month={Sep}, pages={2438–2446} }
@article{buckley_gandolfi_creighton_pyne_bouhan_leroy_senter_gobble_abitbol_lyons_et al._2020, title={Werewolf, There Wolf: Variants in Hairless Associated with Hypotrichia and Roaning in the Lykoi Cat Breed}, volume={11}, ISSN={2073-4425}, url={http://dx.doi.org/10.3390/genes11060682}, DOI={10.3390/genes11060682}, abstractNote={A variety of cat breeds have been developed via novelty selection on aesthetic, dermatological traits, such as coat colors and fur types. A recently developed breed, the lykoi (a.k.a. werewolf cat), was bred from cats with a sparse hair coat with roaning, implying full color and all white hairs. The lykoi phenotype is a form of hypotrichia, presenting as a significant reduction in the average numbers of follicles per hair follicle group as compared to domestic shorthair cats, a mild to severe perifollicular to mural lymphocytic infiltration in 77% of observed hair follicle groups, and the follicles are often miniaturized, dilated, and dysplastic. Whole genome sequencing was conducted on a single lykoi cat that was a cross between two independently ascertained lineages. Comparison to the 99 Lives dataset of 194 non-lykoi cats suggested two variants in the cat homolog for Hairless (HR) (HR lysine demethylase and nuclear receptor corepressor) as candidate causal gene variants. The lykoi cat was a compound heterozygote for two loss of function variants in HR, an exon 3 c.1255_1256dupGT (chrB1:36040783), which should produce a stop codon at amino acid 420 (p.Gln420Serfs*100) and, an exon 18 c.3389insGACA (chrB1:36051555), which should produce a stop codon at amino acid position 1130 (p.Ser1130Argfs*29). Ascertainment of 14 additional cats from founder lineages from Canada, France and different areas of the USA identified four additional loss of function HR variants likely causing the highly similar phenotypic hair coat across the diverse cats. The novel variants in HR for cat hypotrichia can now be established between minor differences in the phenotypic presentations.}, number={6}, journal={Genes}, publisher={MDPI AG}, author={Buckley, Reuben M. and Gandolfi, Barbara and Creighton, Erica K. and Pyne, Connor A. and Bouhan, Delia M. and LeRoy, Michelle L. and Senter, David A. and Gobble, Johnny R. and Abitbol, Marie and Lyons, Leslie A. and et al.}, year={2020}, month={Jun}, pages={682} }
@article{li_nguyen_tablin_2019, title={Canine platelets express functional Toll-like receptor-4: lipopolysaccharide-triggered platelet activation is dependent on adenosine diphosphate and thromboxane A2 in dogs}, volume={15}, ISSN={1746-6148}, url={http://dx.doi.org/10.1186/s12917-019-1997-3}, DOI={10.1186/s12917-019-1997-3}, abstractNote={Functional Toll-like receptor 4 (TLR4) has been characterized in human and murine platelets indicating that platelets play a role in inflammation and hemostasis during sepsis. It is unclear whether canine platelets could express functional TLR4 by responding to its ligand, lipopolysaccharide (LPS). We sought to determine if dogs express functional TLR4 and if LPS-induced platelet activation requires co-stimulation with ADP or thromboxane A2 (TxA2). Canine platelets were unstimulated (resting) or activated with thrombin or ADP prior to flow cytometric or microscopic analyses for TLR4 expression. We treated resting or ADP-primed platelets with LPS in the absence or presence of acetylsalicylic acid (ASA) and inhibited TLR4 with function blocking antibody or LPS from Rhodobacter sphaeroides (LPS-RS).We discovered that dog platelets have variable TLR4 expression, which was upregulated following thrombin or ADP activation. LPS augmented P-selectin expression and thromboxane B2 secretion in ADP-primed platelets via TLR4. Inhibition of cyclooxygenase by ASA attenuated LPS-mediated P-selectin expression demonstrating that TLR4 signaling in platelets is partially dependent on TxA2 pathway.Expression of functional TLR4 on canine platelets may contribute to hypercoagulability in clinical septic dogs. Cyclooxygenase and TxA2 pathways in TLR4-mediated platelet activation may present novel therapeutic targets in dogs with sepsis.}, number={1}, journal={BMC Veterinary Research}, publisher={Springer Science and Business Media LLC}, author={Li, Ronald H. L. and Nguyen, Nghi and Tablin, Fern}, year={2019}, month={Jul}, pages={245} }
@article{jaffey_reading_giger_abdulmalik_buckley_johnstone_lyons_li_2019, title={Clinical, metabolic, and genetic characterization of hereditary methemoglobinemia caused by cytochrome b5 reductase deficiency in cats}, volume={33}, ISSN={0891-6640 1939-1676}, url={http://dx.doi.org/10.1111/jvim.15637}, DOI={10.1111/jvim.15637}, abstractNote={AbstractTwo non‐pedigreed male castrated cats had persistent cyanosis over a 3‐year observation period. Clinical cardiopulmonary evaluations did not reveal abnormalities, but the blood remained dark after exposure to air. Erythrocytic methemoglobin concentrations were high (~40% of hemoglobin) and cytochrome b5 reductase (CYB5R) activities in erythrocytes were low (≤15% of control). One cat remained intolerant of exertion, and the other cat developed anemia and died due to an unidentified comorbidity. Whole‐genome sequencing revealed a homozygous c.625G>A missense variant (B4:137967506) and a c.232‐1G>C splice acceptor variant (B4:137970815) in CYB5R3, respectively, which were absent in 193 unaffected additional cats. The p.Gly209Ser missense variant likely disrupts a nicotinamide adenine dinucleotide (NADH)‐binding domain, while the splicing error occurs at the acceptor site for exon 4, which likely affects downstream translation of the protein. The 2 novel CYB5R3 variants were associated with methemoglobinemia using clinical, biochemical, genomics, and in silico protein studies. The variant prevalence is unknown in the cat population.}, number={6}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Jaffey, Jared A. and Reading, N. Scott and Giger, Urs and Abdulmalik, Osheiza and Buckley, Ruben M. and Johnstone, Sophie and Lyons, Leslie A. and Li, Ronald}, year={2019}, month={Oct}, pages={2725–2731} }
@article{goggs_bacek_bianco_koenigshof_li_2019, title={Consensus on the Rational Use of Antithrombotics in Veterinary Critical Care (CURATIVE): Domain 2—Defining rational therapeutic usage}, volume={29}, ISSN={1479-3261 1476-4431}, url={http://dx.doi.org/10.1111/vec.12791}, DOI={10.1111/vec.12791}, abstractNote={AbstractObjectivesTo systematically review available evidence to determine when small animals at risk of thrombosis should be treated with antiplatelet agents and anticoagulants, which antiplatelet and anticoagulant agents are most effective, and when multimodal therapy is indicated.DesignStandardized, systematic evaluation of the literature, categorization of relevant articles according to level of evidence (LOE) and quality (Good, Fair, or Poor), and development of consensus on conclusions via a Delphi‐style survey for application of the concepts to clinical practice. Draft recommendations were presented at 2 international veterinary conferences and made available for community assessment, review, and comment prior to final revisions and publication.SettingsAcademic and referral veterinary medical centers.ResultsDatabases searched included Medline via PubMed and CAB abstracts. Twelve Population Intervention Comparison Outcome questions were devised and generated corresponding worksheets investigating indications for use of antithrombotic drugs in small animals. Seventy‐eight studies were reviewed in detail. Most studies assessed were experimentally controlled laboratory studies in companion animals (56 LOE 3) with smaller numbers of LOE 2 (1), LOE 4 (5), LOE 5 (6), and LOE 6 (4) studies assessed. Only 5 randomized controlled clinical trials were identified (LOE 1, Good–Fair). The 12 worksheets generated 21 guidelines with 17 guideline statements that were refined during 3 rounds of Delphi surveys. A high degree of consensus was reached across all guideline recommendations during the Delphi process.ConclusionsOverall, systematic evidence evaluations generated 2 strong recommendations, 19 weak recommendations (formulated as suggestions), 9 situations where the evidence was insufficient to make strong recommendations, and 8 situations where no relevant evidence was retrieved to aid guideline generation. Numerous significant knowledge gaps were highlighted by the evidence reviews undertaken, indicating the need for substantial additional research in this field.}, number={1}, journal={Journal of Veterinary Emergency and Critical Care}, publisher={Wiley}, author={Goggs, Robert and Bacek, Lenore and Bianco, Domenico and Koenigshof, Amy and Li, Ronald H. L.}, year={2019}, month={Jan}, pages={49–59} }
@article{goggs_jeffery_levine_li_2019, title={Neutrophil-Extracellular Traps, Cell-Free DNA, and Immunothrombosis in Companion Animals: A Review}, volume={57}, ISSN={0300-9858 1544-2217}, url={http://dx.doi.org/10.1177/0300985819861721}, DOI={10.1177/0300985819861721}, abstractNote={Immunothrombosis is a potentially beneficial physiological process that aids innate immunity and host defense against pathogen invasion. However, this process can also be damaging when it occurs to excess or in critical blood vessels. Formation of extracellular traps by leukocytes, particularly neutrophils, is central to our understanding of immunothrombosis. In addition to degranulation and phagocytosis, extracellular traps are the third mechanism by which neutrophils combat potential pathogens. These traps consist of extracellular DNA decorated with bactericidal cellular proteins, including elastase, myeloperoxidase, and cathepsins. Neutrophils can release these structures as part of a controlled cell-death process or via a process termed vital NETosis that enables the cells to extrude DNA but remain viable. There is accumulating evidence that NETosis occurs in companion animals, including dogs, horses, and cats, and that it actively contributes to pathogenesis. Numerous studies have been published detailing various methods for identification and quantification of extracellular trap formation, including cell-free DNA, measurements of histones and proteins such as high-mobility group box–1, and techniques involving microscopy and flow cytometry. Here, we outline the present understanding of these phenomena and the mechanisms of extracellular trap formation. We critically review the data regarding measurement of NETosis in companion animals, summarize the existing literature on NETosis in veterinary species, and speculate on what therapeutic options these insights might present to clinicians in the future.}, number={1}, journal={Veterinary Pathology}, publisher={SAGE Publications}, author={Goggs, Robert and Jeffery, Unity and LeVine, Dana N. and Li, Ronald H. L.}, year={2019}, month={Jul}, pages={6–23} }
@article{li_tablin_2018, title={In Vitro Canine Neutrophil Extracellular Trap Formation: Dynamic and Quantitative Analysis by Fluorescence Microscopy}, volume={8}, ISSN={1940-087X}, url={http://dx.doi.org/10.3791/58083}, DOI={10.3791/58083}, abstractNote={In response to invading pathogens, neutrophils release neutrophil extracellular traps (NETs), which are extracellular networks of DNA decorated with histones and antimicrobial proteins. Excessive NET formation (NETosis) and citH3 release during sepsis is associated with multiple organ dysfunction and mortality in mice and humans but its implications in dogs are unknown. Herein, we describe a technique to isolate canine neutrophils from whole blood for observation and quantification of NETosis. Leukocyte-rich plasma, generated by dextran sedimentation, is separated by commercially available density gradient separation media and granulocytes collected for cell count and viability testing. To observe real-time NETosis in live neutrophils, cell permeant and cell impermeant fluorescent nucleic acid stains are added to neutrophils activated either by lipopolysaccharide (LPS) or phorbol 12-myristate 13-acetate (PMA). Changes in nuclear morphology and NET formation are observed over time by fluorescence microscopy. In vitro NETosis is further characterized by co-colocalization of cell-free DNA (cfDNA), myeloperoxidase (MPO) and citrullinated histone H3 (citH3) using a modified double-immunolabelling protocol. To objectively quantify NET formation and citH3 expression using fluorescence microscopy, NETs and citH3-positive cells are quantified in a blinded manner using available software. This technique is a specific assay to evaluate the in vitro capacity of canine neutrophils to undergo NETosis.}, number={138}, journal={Journal of Visualized Experiments}, publisher={MyJove Corporation}, author={Li, Ronald H.L. and Tablin, Fern}, year={2018}, month={Aug} }
@article{li_tablin_2018, title={A Comparative Review of Neutrophil Extracellular Traps in Sepsis}, volume={5}, ISSN={2297-1769}, url={http://dx.doi.org/10.3389/fvets.2018.00291}, DOI={10.3389/fvets.2018.00291}, abstractNote={Sepsis is the leading cause of critical illness and mortality in human beings and animals. Neutrophils are the primary effector cells of innate immunity during sepsis. Besides degranulation and phagocytosis, neutrophils also release neutrophil extracellular traps (NETs), composed of cell-free DNA, histones, and antimicrobial proteins. Although NETs have protective roles in the initial stages of sepsis, excessive NET formation has been found to induce thrombosis and multiple organ failure in murine sepsis models. Since the discovery of NETs nearly a decade ago, many investigators have identified NETs in various species. However, many questions remain regarding the exact mechanisms and fate of neutrophils following NET formation. In humans and mice, platelet-neutrophil interactions via direct binding or soluble mediators seem to play an important role in mediating NET formation during sepsis. Preliminary data suggest that these interactions may be species dependent. Regardless of these differences, there is increasing evidence in human and veterinary medicine suggesting that NETs play a crucial role in the pathogenesis of intravascular thrombosis and multiple organ failure in sepsis. Because the outcome of sepsis is highly dependent on early recognition and intervention, detection of NETs or NET components can aid in the diagnosis of sepsis in humans and veterinary species. In addition, the use of novel therapies such as deoxyribonuclease and non-anticoagulant heparin to target NET components shows promising results in murine septic models. Much work is needed in translating these NET-targeting therapies to clinical practice.}, journal={Frontiers in Veterinary Science}, publisher={Frontiers Media SA}, author={Li, Ronald H. L. and Tablin, Fern}, year={2018}, month={Nov}, pages={291} }
@article{li_johnson_kohen_tablin_2018, title={A novel approach to identifying and quantifying neutrophil extracellular trap formation in septic dogs using immunofluorescence microscopy}, volume={14}, ISSN={1746-6148}, url={http://dx.doi.org/10.1186/s12917-018-1523-z}, DOI={10.1186/s12917-018-1523-z}, abstractNote={Canine neutrophils release neutrophil extracellular traps (NETs) in response to lipopolysaccharide but NETs from clinical septic dogs had not been identified. The primary aim is to describe the methodology of identifying and quantifying neutrophil extracellular traps (NETs) in cytology samples of septic foci in dogs with sepsis using immunofluorescence microscopy. Cytology samples including endotracheal tracheal wash (ETW), bronchoalveolar lavage (BAL), abdominal and pleural effusion collected from 5 dogs (3 septic, 2 non-septic) were fixed, permeabilized and stained for myeloperoxidase (MPO), citrullinated histone H3 (citH3) and cell-free DNA (cfDNA). Fluorescence microscopy was used to identify and quantify NETs in 10 random views at 40× magnification. NETs were identified based on co-localization of MPO, citH3 and cfDNA. NETs were quantified as a ratio (number of NETs: number of neutrophils). Neutrophils were identified based on cytoplasmic MPO, cellular diameter and nuclear morphology.NETs were identified and quantified in all cytology samples collected from septic dogs. A small number of NETs was documented in one dog with sterile chronic bronchitis. No NETs were found in sterile abdominal effusion collected from one dog with congestive heart failure.Immunofluorescence microscopy could be a useful tool for the study of NETs in dogs with clinical sepsis.}, number={1}, journal={BMC Veterinary Research}, publisher={Springer Science and Business Media LLC}, author={Li, Ronald H. L. and Johnson, Lynelle R. and Kohen, Casey and Tablin, Fern}, year={2018}, month={Jun}, pages={210} }
@article{ueda_li_tablin_ontiveros_stern_2018, title={Nonsynonymous single nucleotide polymorphisms in candidate genes P2RY1, P2RY12 and CYP2C19 for clopidogrel efficacy in cats}, volume={49}, ISSN={0268-9146 1365-2052}, url={http://dx.doi.org/10.1111/age.12666}, DOI={10.1111/age.12666}, abstractNote={Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.}, number={4}, journal={Animal Genetics}, publisher={Wiley}, author={Ueda, Yu and Li, Ronald Hak Long and Tablin, Fern and Ontiveros, Eric S. and Stern, Joshua A.}, year={2018}, month={May}, pages={356–357} }
@article{li_ng_tablin_2017, title={Lipopolysaccharide-induced neutrophil extracellular trap formation in canine neutrophils is dependent on histone H3 citrullination by peptidylarginine deiminase}, volume={193-194}, ISSN={0165-2427}, url={http://dx.doi.org/10.1016/j.vetimm.2017.10.002}, DOI={10.1016/j.vetimm.2017.10.002}, abstractNote={Neutrophils release neutrophil extracellular traps (NETs), which are extracellular chromatin decorated with histones and antimicrobial proteins. Although known for antimicrobial properties, overzealous production of NETs (NETosis) may lead to cytotoxicity and multiple organ failure in sepsis. Pathogen-induced NETosis has been extensively studied in mice but its importance in dogs remains largely unknown. This study sought to characterize in vitro NETosis induced by E.coli LPS, including assessing the role of peptidylarginine deiminase (PAD) in canine NETosis. Neutrophils (1×106 cells/ml) from healthy dogs were isolated and treated with 100μg/ml LPS, 100nM phorbol 12-myristate 13-acetate (PMA), or buffer for either 90 or 180min. NETs were assessed using fluorescence microscopy of living neutrophils and immunofluorescent microscopy. Supernatant and cellular debris were purified and cell-free DNA was quantified by spectrophotometry. The role of PAD was assessed by treating LPS- and PMA-activated neutrophils with 50, 100 or 200μM of the PAD inhibitor, Cl-amidine. In vitro NETosis was characterized by co-localization of cell-free DNA, citrullinated histone H3, and myeloperoxidase. LPS stimulation resulted in intracellular citrullination of histone H3. Compared to PMA chemically-induced NETosis, LPS resulted in smaller NETs with less extracellular citrullinated histone H3. Cl-amidine decreased citrullination of histones and NET production in either LPS- or PMA-stimulated neutrophils demonstrating that neutrophil PAD is essential for these cellular processes.}, journal={Veterinary Immunology and Immunopathology}, publisher={Elsevier BV}, author={Li, Ronald H.L. and Ng, Geena and Tablin, Fern}, year={2017}, month={Dec}, pages={29–37} }
@article{li_chan_2016, title={Evaluation of platelet function using multiple electrode platelet aggregometry in dogs with septic peritonitis}, volume={26}, ISSN={1479-3261 1476-4431}, url={http://dx.doi.org/10.1111/vec.12508}, DOI={10.1111/vec.12508}, abstractNote={AbstractObjectiveTo assess platelet function via multiple electrode platelet aggregometry (MEPA) in dogs with septic peritonitis and in healthy dogs. The secondary aim was to determine if there is prognostic significance to changes in platelet function observed in septic dogs.DesignProspective, observational cohort study conducted from January 2012 to March 2014.SettingUniversity teaching hospital.AnimalsTwenty dogs with septic peritonitis and 23 healthy dogs.InterventionsNone.Measurements and Main ResultsMEPA using arachidonic acid, adenosine diphosphate, and collagen (COL) as agonists was measured within 24 hours of diagnosis of sepsis. Compared to healthy dogs, platelet aggregation was reduced in dogs with septic peritonitis for all agonists (P < 0.01). Overall mortality rate was 40%. MEPA in response to COL was significantly reduced in nonsurvivors compared to survivors (P = 0.019). Using receiver‐operating characteristic curve statistics, a COL‐activated MEPA less than 43.5 aggregation units had a sensitivity and specificity of 85.7% and 90.9%, respectively, for predicting nonsurvival in dogs with septic peritonitis.ConclusionsCirculating platelets from dogs with septic peritonitis have diminished aggregation in response to multiple platelet agonists. MEPA may serve as an assessment tool for illness severity in this patient population.}, number={5}, journal={Journal of Veterinary Emergency and Critical Care}, publisher={Wiley}, author={Li, Ronald H.L. and Chan, Daniel L.}, year={2016}, month={Jul}, pages={630–638} }
@article{li_stern_ho_tablin_harris_2016, title={Platelet Activation and Clopidogrel Effects on ADP-Induced Platelet Activation in Cats with or without the A31P Mutation in MYBPC3}, volume={30}, ISSN={0891-6640 1939-1676}, url={http://dx.doi.org/10.1111/jvim.14568}, DOI={10.1111/jvim.14568}, abstractNote={BackgroundClopidogrel is commonly prescribed to cats with perceived increased risk of thromboembolic events, but little information exists regarding its antiplatelet effects.ObjectiveTo determine effects of clopidogrel on platelet responsiveness in cats with or without the A31P mutation in the MYBPC3 gene. A secondary aim was to characterize variability in feline platelet responses to clopidogrel.AnimalsFourteen healthy cats from a Maine Coon/outbred mixed Domestic cat colony: 8 cats homozygous for A31P mutation in the MYPBC3 gene and 6 wild‐type cats without the A31P mutation.MethodsEx vivo study. All cats received clopidogrel (18.75 mg PO q24h) for 14 days. Before and after clopidogrel treatment, adenosine diphosphate (ADP)‐induced P‐selectin expression was evaluated. ADP‐ and thrombin‐induced platelet aggregation was measured by optical aggregometry (OA). Platelet pVASP and ADP receptor response index (ARRI) were measured by Western blot analysis.ResultsPlatelet activation from cats with the A31P mutation was significantly (P = .0095) increased [35.55% (18.58–48.55) to 58.90% (24.85–69.90)], in response to ADP. Clopidogrel treatment attenuated ADP‐induced P‐selectin expression and platelet aggregation. ADP‐ and PGE1‐treated platelets had a similar level of pVASP as PGE1‐treated platelets after clopidogrel treatment. Clopidogrel administration resulted in significantly lower ARRI [24.13% (12.46–35.50) to 11.30% (−7.383 to 23.27)] (P = .017). Two of 13 cats were nonresponders based on OA and flow cytometry.Conclusion and Clinical ImportanceClopidogrel is effective at attenuating platelet activation and aggregation in some cats. Cats with A31P mutation had increased platelet activation relative to the variable response seen in wild‐type cats.}, number={5}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Li, R.H.L. and Stern, J.A. and Ho, V. and Tablin, F. and Harris, S.P.}, year={2016}, month={Sep}, pages={1619–1629} }
@article{mugford_li_humm_2013, title={Acute kidney injury in dogs and cats 1. Pathogenesis and diagnosis}, volume={35}, ISSN={0263-841X 2042-7689}, url={http://dx.doi.org/10.1136/inp.f2868}, DOI={10.1136/inp.f2868}, abstractNote={Acute kidney injury (AKI), an abrupt loss of renal function, is a commonly encountered emergency in small animal practice. This article, the first of two on AKI, reviews the pathophysiology and diagnosis of the condition in dogs and cats. The second article, to be published in the June issue of In Practice, will cover the treatment, recovery and prognosis associated with AKI.}, number={5}, journal={In Practice}, publisher={Wiley}, author={Mugford, Adam and Li, Ronald and Humm, Karen}, year={2013}, month={May}, pages={253–264} }
@article{li_mugford_humm_2013, title={Acute kidney injury in dogs and cats 2. Management, treatment and outcome}, volume={35}, ISSN={0263-841X 2042-7689}, url={http://dx.doi.org/10.1136/inp.f3640}, DOI={10.1136/inp.f3640}, abstractNote={Acute kidney injury (AKI) is a commonly encountered emergency in small animal practice. Dogs and cats with AKI are mostly presented in the maintenance phase of the disease, by which point renal function has been severely compromised and clinical signs are apparent. The first article in this two‐part series, which was published in the May issue of In Practice discussed the pathogenesis and diagnosis of AKI. This article considers both specific therapy and general supportive treatment in dogs and cats.}, number={6}, journal={In Practice}, publisher={Wiley}, author={Li, Ronald and Mugford, Adam and Humm, Karen}, year={2013}, month={Jun}, pages={302–316} }
@article{cruz_li_kenney_monteith_2006, title={Effects of indwelling nasogastric intubation on gastric emptying of a liquid marker in horses}, volume={67}, ISSN={0002-9645}, url={http://dx.doi.org/10.2460/ajvr.67.7.1100}, DOI={10.2460/ajvr.67.7.1100}, abstractNote={Abstract
Objective—To determine the effects of indwelling nasogastric intubation on the gastric emptying rate of liquid in horses.
Animals—6 healthy horses.
Procedures—Horses were assigned to treatment and control groups in a prospective randomized crossover study with a washout period of at least 4 weeks between trials. Acetaminophen (20 mg/kg) diluted in 1 L of distilled water was administered via nasogastric tube at time points of 0, 12, 30, 48, and 72 hours to evaluate the liquid-phase gastric emptying rate. In control horses, nasogastric tubes were removed after administration of acetaminophen. In horses receiving treatment, the tube was left indwelling and maintained for 72 hours. A 10-mL sample of blood was collected from a jugular vein immediately before and 20, 40, 60, 80, 100, 120, and 180 minutes after acetaminophen administration. Serum acetaminophen concentrations were measured by use of a colorimetric method.
Results—Peak serum acetaminophen concentration was significantly higher in the control group (38.11 μg/mL) than in the treatment group (29.09 μg/mL), and the time required to reach peak serum acetaminophen concentration was significantly shorter in the control group (22.79 minutes) than in the treatment group (35.95 minutes).
Conclusions and Clinical Relevance—Results indicated that indwelling nasogastric intubation has a delaying effect on the gastric emptying rate of liquids. Veterinarians should consider the potential for delayed gastric emptying when placing and maintaining an indwelling nasogastric tube for an extended period of time after surgery. Repeated nasogastric intubation may be better than maintenance of an indwelling tube in horses with ileus.}, number={7}, journal={American Journal of Veterinary Research}, publisher={American Veterinary Medical Association (AVMA)}, author={Cruz, Antonio M. and Li, Ronald and Kenney, Dan G. and Monteith, Gabrielle}, year={2006}, month={Jul}, pages={1100–1104} }
@article{crus_cote_mcdonell_geor_wilson_monteith_li_2006, title={Postoperative effects of anesthesia and surgery on resting energy expenditure in horses as measured by indirect calorimetry}, volume={70}, url={https://www.ncbi.nlm.nih.gov/pmc/articles/pmid/17042377/?tool=EBI}, number={4}, journal={Canadian Journal of Veterinary Research}, author={Crus, A.M. and Cote, N. and McDonell, W. and Geor, R.J. and Wilson, B.A. and Monteith, G. and Li, R.}, year={2006}, month={Oct}, pages={257–262} }