Assessment of misdiagnosis in small animal intensive care patients using the Modified Goldman criteria.

The postmortem examination can be used as a means of quality control for clinical diagnoses. A retrospective study on 300 dogs and cats that had been admitted to a small animal intensive care unit was performed comparing the clinical and postmortem findings, using the Modified Goldman criteria. All patient files were reevaluated for clinical diagnoses and all postmortem material was reevaluated for pathological diagnoses. After this, the Modified Goldman criteria were applied to score the discrepancies between them, and factors associated with the occurrence of an undiagnosed major unexpected finding were analyzed. The postmortem examination revealed additional findings in 65% of the cases. Major discrepancies, defined as those affecting treatment and possibly outcome of the patient, were present in 21.3% of the cases. The most frequently missed diagnoses detected at necropsy were pneumonia of various etiologies, meningitis/meningoencephalitis, myocarditis and generalized vasculitis. A shorter ICU stay was associated with increased odds of a major discrepancy. Conditions affecting the urinary or gastrointestinal system were negatively associated with major discrepancy.

Evidence of a sudden increase in α-chloralose poisoning in dogs and cats in the Netherlands between 2018 and 2021.

BACKGROUND: After changes in European Union biocide legislation, the Dutch Poisons Information Center observed a strong increase in information requests concerning dogs and cats exposed to α-chloralose. To investigate whether α-chloralose-based rodenticides are safe for non-professional use, additional information regarding poisoning scenarios and clinical course was collected. METHODS: Veterinarians reporting α-chloralose exposure over a 2.5-year period were contacted by mail for follow-up information concerning exposure scenario, product formulation, clinical course and treatment, and outcome. In total, information was collected for 96 dogs and 41 cats. RESULTS: Fifty-three of 96 dogs and 17 of 19 cats known to have been exposed to α-chloralose-based rodenticides developed signs of central nervous system (CNS) depression or sensory-induced CNS excitation. Mortality in dogs and cats following exposure was 1% and 18%, respectively. An additional 22 cats presented with clinical signs suggestive of α-chloralose poisoning, with a mortality of 5%. LIMITATIONS: Exposure to α-chloralose was not confirmed by biochemical analyses. CONCLUSION: Dogs and especially cats were at risk of poisoning from α-chloralose. If criteria such as acute toxicity and risk of (secondary) poisoning are applied during the approval of α-chloralose-based rodenticides, similar to anticoagulant-based rodenticides, it can be concluded that α-chloralose is also not safe for non-professional use.

Genomic Investigation of Two Acinetobacter baumannii Outbreaks in a Veterinary Intensive Care Unit in The Netherlands.

Acinetobacter baumannii is a nosocomial pathogen that frequently causes healthcare-acquired infections. The global spread of multidrug-resistant (MDR) strains with its ability to survive in the environment for extended periods imposes a pressing public health threat. Two MDR A. baumannii outbreaks occurred in 2012 and 2014 in a companion animal intensive care unit (caICU) in the Netherlands. Whole-genome sequencing (WGS) was performed on dog clinical isolates (n = 6), environmental isolates (n = 5), and human reference strains (n = 3) to investigate if the isolates of the two outbreaks were related. All clinical isolates shared identical resistance phenotypes displaying multidrug resistance. Multi-locus Sequence Typing (MLST) revealed that all clinical isolates belonged to sequence type ST2. The core genome MLST (cgMLST) results confirmed that the isolates of the two outbreaks were not related. Comparative genome analysis showed that the outbreak isolates contained different gene contents, including mobile genetic elements associated with antimicrobial resistance genes (ARGs). The time-measured phylogenetic reconstruction revealed that the outbreak isolates diverged approximately 30 years before 2014. Our study shows the importance of WGS analyses combined with molecular clock investigations to reduce transmission of MDR A. baumannii infections in companion animal clinics.

The Effect of the Preoperative Fasting Regimen on the Incidence of Gastro-Oesophageal Reflux in 90 Dogs.

This study aimed to investigate the effect of three different preoperative fasting regimens on the incidence of gastro-oesophageal reflux (GOR) in dogs under general anaesthesia. Ninety dogs undergoing non-abdominal and non-thoracic elective surgery were included in the study and equally allocated to three groups. Dogs received canned food providing half the daily resting energy requirements (RER) 3 h prior to premedication (group 3H), a quarter of the daily RER 3 h before premedication (group 3Q), and half the daily RER 12 h before premedication (group 12H). The animals were premedicated with acepromazine and pethidine, anaesthesia was induced with propofol and maintained with isoflurane vaporised in oxygen. Oesophageal pH was monitored throughout anaesthesia. Demographic and surgery-related parameters were not different among groups. The incidence of GOR was 11/30 in group 3H (36.7%), 9/30 in group 3Q (30.0%) and 5/30 in group 12H (16.7%), which was not statistically different (p = 0.262). Reduction of the amount of the preoperative meal from half to a quarter of the daily RER did not reduce the incidence of GOR but resulted in a lower oesophageal pH (p = 0.003). The results of this study suggest that the administration of a meal 3 h before anaesthesia does not have any beneficial effect in the reduction of GOR incidence in dogs compared to the administration of a meal 12 h before anaesthesia.

A novel insertion technique for urinary catheters in female dogs with the use of a guidewire.

BACKGROUND: Complicated transurethral urinary bladder catheterization in female dogs is a commonly encountered urologic problem, often causing inadvertent trauma to the lower urinary tract and breach in aseptic technique. KEY CONCEPTS: A novel method for transurethral insertion of urinary catheters in female dogs is described. The main variation from current procedures involves the application of the Seldinger technique with the use of an introduction catheter that is specifically designed to feed into the urethra, and the use of a guidewire to replace the introducer with an indwelling balloon catheter. SIGNIFICANCE: The technique presented offers a step-by-step approach that could improve success in urinary catheter placement. This modification may facilitate catheter insertion, be less traumatic, and offer better control of asepsis, especially with challenging urethral catheterization. The technique also offers an easy method to replace the catheter. Application of the technique may reduce catheter-associated urinary tract infections as it addresses certain infection-related risk factors. Prospective validation studies are needed to support its advantages over current urinary catheter placement techniques.

Genes Involved in Energy Metabolism Are Differentially Expressed During the Day-Night Cycle in Murine Retinal Pigment Epithelium.

Purpose: The functional interaction between photoreceptors and retinal pigment epithelium (RPE) cells is essential for vision. Phagocytosis of photoreceptor outer segments (POSs) by the RPE follows a circadian pattern; however, it remains unknown whether other RPE processes follow a daily rhythm. Therefore, our aim was to identify RPE processes following a daily rhythm. Methods: Murine RPE was isolated at Zeitgeber time (ZT) 0, 2, 4, 9, 14, and 19 (n = 5 per time point), after which RNA was isolated and sequenced. Genes with a significant difference in expression between time points (P < 0.05) were subjected to EnrichR pathway analysis to identify daily rhythmic processes. Results: Pathway enrichment revealed 13 significantly enriched KEGG pathways (P < 0.01), including the metabolic pathway (P = 0.002821). Analysis of the metabolic pathway differentially expressed genes revealed that genes involved in adenosine triphosphate production, glycolysis, glycogenolysis, and glycerophospholipid were low at ZT0 (light onset) and high at ZT19 (night). Genes involved in fatty acid degradation and cholesterol synthesis were high at light onset and low at night. Conclusions: Our transcriptome data suggest that the highest energy demand of RPE cells is at night, whereas POS phagocytosis and degradation take place in the morning. Furthermore, we identified genes involved in fatty acid and glycerophospholipid synthesis that are upregulated at night, possibly playing a role in generating building blocks for membrane synthesis.

Gene expressions of de novo hepatic lipogenesis in feline hepatic lipidosis.

OBJECTIVES: The aim of this study was to evaluate if de novo hepatic lipid synthesis contributes to fatty acid overload in the liver of cats with feline hepatic lipidosis (FHL). METHODS: Lipogenic gene expression of peroxisome proliferator-activated receptor-alpha (PPAR-α), peroxisome proliferator-activated receptor-gamma (PPAR-γ), fatty acid synthase (FASN) and sterol regulatory element-binding factor (SREBF1) were evaluated using quantitative RT-PCR in liver tissue of six cats with FHL and compared with the liver tissue of eight healthy cats. RESULTS: In liver tissue, PPAR-α, PPAR-γ and FASN mRNA expression levels were not significantly different (P >0.12, P >0.89 and P >0.5, respectively) in the FHL group compared with the control group. SREBF1 gene expression was downregulated around 10-fold in the FHL group vs the control group (P = 0.039). CONCLUSIONS AND RELEVANCE: The downregulation of SREBF1 in the liver tissue of cats with FHL does not support the hypothesis that de novo lipogenesis in the liver is an important pathway of fatty acid accumulation in FHL.

Innovative application of an implantable venous access system in the portal vein: technique, results and complications in three dogs.

BACKGROUND: Vascular access port (VAP) systems are widely used in human medicine to provide long-term venous access. However, in veterinary medicine the use of VAP systems is not common practice and publications on their potential applications have been limited. A VAP system was used as part of an experimental study on liver regeneration and implanted in the canine portal vein to create direct access to the portal venous circulation of the liver. The aim of the present study is to describe the surgical technique, its use, and the complications of a VAP system in three research dogs. RESULTS: The VAP system was successfully used for the intraoperative measurement of portal blood pressure, the administration of cell suspensions, and the collection of portal venous blood samples. Long-term complications consisted of dislocation of the VAP system in one dog (2 months after implantation) and thrombus formation at the catheter tip in two dogs (3 months after implantation). Both complications prevented further use of the VAP but had no adverse clinical implications. CONCLUSIONS: This pilot study suggests that the VAP system is an effective and safe technique to obtain long term access to the portal venous system in dogs. However, complications with port detachment and thrombosis may limit long term use of VAPs in the portal system of dogs.

Immunohistochemical characterisation of the hepatic stem cell niche in feline hepatic lipidosis: a preliminary morphological study.

OBJECTIVES: The aim of this study was to describe the cellular and stromal components of the hepatic progenitor cell niche in feline hepatic lipidosis (FHL). METHODS: Immunohistochemical staining for the progenitor/bile duct marker (K19), activated Kupffer cells (MAC387), myofibroblasts (alpha-smooth muscle actin [α-SMA]) and the extracellular matrix component laminin were used on seven liver biopsies of cats with FHL and three healthy cats. Double immunofluorescence stainings were performed to investigate co-localisation of different cell types in the hepatic progenitor cell (HPC) niche. RESULTS: HPCs, Kupffer cells, myofibroblasts and laminin deposition were observed in the liver samples of FHL, although with variability in the expression and positivity of the different immunostainings between different samples. When compared with the unaffected cats where K19 positivity and minimal α-SMA and laminin positivity were seen mainly in the portal area, in the majority of FHL samples K19 and α-SMA-positive cells and laminin positivity were seen also in the periportal and parenchymatous area. MAC387-positive cells were present throughout the parenchyma. CONCLUSIONS AND RELEVANCE: This is a preliminary morphological study to describe the activation and co-localisation of components of the HPC niche in FHL. Although the HPC niche in FHL resembles that described in hepatopathies in dogs and in feline lymphocytic cholangitis, the expression of K19, α-SMA, MAC387 and lamin is more variable in FHL, and a common pattern of activation could not be established. Nevertheless, when HPCs were activated, a spatial association between HPCs and their niche could be demonstrated.

Negative pressure therapy versus passive open abdominal drainage for the treatment of septic peritonitis in dogs: A randomized, prospective study.

OBJECTIVE: To compare passive open abdominal drainage (POAD) and negative-pressure abdominal drainage (NPAD) using the ABThera™ system in the treatment of septic peritonitis. STUDY DESIGN: Randomized prospective clinical trial. ANIMALS: Dogs (n = 16) with septic peritonitis. METHODS: Dogs with septic peritonitis were randomly assigned to one of two treatment protocols: NPAD versus POAD. Anesthesia time, operating time, duration of drainage, costs, survival, and complications were compared between techniques. Hematological and biochemical parameters in blood and abdominal fluid, and histopathological findings of omentum and abdominal wall tissue samples were compared between NPAD and POAD at time of initial surgery and at time of closure. RESULTS: Overall survival was 81%. Treatment costs, anesthesia and operating time, drainage time, survival, and postoperative complications were similar between techniques. Loss of total plasma protein and decreased inflammation-related factors in abdominal fluid at time of closure were noted in all patients. Neutrophilic inflammation was greater in abdominal wall samples after NPAD. POAD patients showed discomfort during bandage changes and had frequent leakage of abdominal fluid outside of the bandage. CONCLUSION: NPAD is an effective alternative to POAD for treatment of septic peritonitis, based on costs and survival. NPAD resulted in less abdominal fluid leakage, and evidence of superior healing on histological evaluation of abdominal tissues.

Sex specific differences in hepatic and plasma lipid profiles in healthy cats pre and post spaying and neutering: relationship with feline hepatic lipidosis.

BACKGROUND: A link between lipid metabolism and disease has been recognized in cats. Since hepatic lipidosis is a frequent disorder in cats, the aim of the current study was to evaluate liver and plasma lipid dimorphism in healthy cats and the effects of gonadectomy on lipid profiling. From six female and six male cats plasma and liver lipid profiles before and after spaying/neutering were assessed and compared to five cats (three neutered male and two spayed female) diagnosed with hepatic lipidosis. RESULTS: Intact female cats had a significantly lower level of plasma triacylglycerides (TAG) and a higher liver level of the long chain polyunsaturated fatty acid arachidonic acid (AA) compared to their neutered state. Both male and female cats with lipidosis had a higher liver, but not plasma TAG level and an increased level of plasma and liver sphingomyelin compared to the healthy cats. CONCLUSION: Although lipid dimorphism in healthy cats resembles that of other species, intact female cats show differences in metabolic configuration that could predispose them to develop hepatic lipidosis. The increased sphingomyelin levels in cats with lipidosis could suggest a potential role in the pathogenesis of hepatic lipidosis in cats.

SUCNR1-mediated chemotaxis of macrophages aggravates obesity-induced inflammation and diabetes.

AIMS/HYPOTHESIS: Obesity induces macrophages to drive inflammation in adipose tissue, a crucial step towards the development of type 2 diabetes. The tricarboxylic acid (TCA) cycle intermediate succinate is released from cells under metabolic stress and has recently emerged as a metabolic signal induced by proinflammatory stimuli. We therefore investigated whether succinate receptor 1 (SUCNR1) could play a role in the development of adipose tissue inflammation and type 2 diabetes. METHODS: Succinate levels were determined in human plasma samples from individuals with type 2 diabetes and non-diabetic participants. Succinate release from adipose tissue explants was studied. Sucnr1 -/- and wild-type (WT) littermate mice were fed a high-fat diet (HFD) or low-fat diet (LFD) for 16 weeks. Serum metabolic variables, adipose tissue inflammation, macrophage migration and glucose tolerance were determined. RESULTS: We show that hypoxia and hyperglycaemia independently drive the release of succinate from mouse adipose tissue (17-fold and up to 18-fold, respectively) and that plasma levels of succinate were higher in participants with type 2 diabetes compared with non-diabetic individuals (+53%; p < 0.01). Sucnr1 -/- mice had significantly reduced numbers of macrophages (0.56 ± 0.07 vs 0.92 ± 0.15 F4/80 cells/adipocytes, p < 0.05) and crown-like structures (0.06 ± 0.02 vs 0.14 ± 0.02, CLS/adipocytes p < 0.01) in adipose tissue and significantly improved glucose tolerance (p < 0.001) compared with WT mice fed an HFD, despite similarly increased body weights. Consistently, macrophages from Sucnr1 -/- mice showed reduced chemotaxis towards medium collected from apoptotic and hypoxic adipocytes (-59%; p < 0.05). CONCLUSIONS/INTERPRETATION: Our results reveal that activation of SUCNR1 in macrophages is important for both infiltration and inflammation of adipose tissue in obesity, and suggest that SUCNR1 is a promising therapeutic target in obesity-induced type 2 diabetes. DATA AVAILABILITY: The dataset generated and analysed during the current study is available in GEO with the accession number GSE64104, www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE64104 .

Lipid Therapy for Intoxications.

This review discusses the use of intravenous lipid emulsion (ILE) in the treatment of intoxications with lipophilic agents in veterinary medicine. Despite growing scientific evidence that ILE has merit in the treatment of certain poisonings, there is still uncertainty on the optimal composition of the lipid emulsion, the dosing, the mechanism of action, and the efficacy. Therefore, a critical view of the clinician on the applicability of this modality in intoxications is still warranted. The use of ILE therapy is advocated as an antidote in cases of intoxications with some lipophilic substances.

Evaluation of a virtual pet visit system with live video streaming of patient images over the Internet in a companion animal intensive care unit in the Netherlands.

OBJECTIVE: To evaluate the impact of a virtual pet visit system ("TelePet" System, TPS) on owners and staff of a companion animal ICU. DESIGN: Longitudinal interventional study (2010-2013). SETTING: Companion animal ICU at a university veterinary medical teaching hospital. STUDY POPULATIONS: Pet owners, ICU technicians. INTERVENTIONS: The introduction of the TPS, with live video streaming of patient images over the Internet, in a companion animal ICU. MEASUREMENTS AND MAIN RESULTS: Pet owners experienced TPS as a valuable extra service. Most TPS users (72.4%) experienced less anxiety and felt less need (40.4% of TPS users) to visit their hospitalized pet in person. Most users (83.5%) shared TPS access with their family. The introduction of the TPS did not improve overall owner satisfaction, except for the score on "quality of medical treatment." Seven of 26 indicators of owner satisfaction were awarded higher scores by TPS users than by TPS nonusers in the survey after the introduction of the system. However, the lack of randomization of owners might have influenced findings. The enthusiasm of the ICU technicians for the system was tempered by the negative feedback from a small number of owners. Nevertheless they recognized the value of the system for owners. The system was user friendly and ICU staff and TPS users experienced few technical problems. CONCLUSIONS: As veterinary healthcare is moving toward a more client-centered approach, a virtual pet visit system, such as TPS, is a relatively simple application that may improve the well-being of most owners during the hospitalization of their pet.

Utstein-style guidelines on uniform reporting of in-hospital cardiopulmonary resuscitation in dogs and cats. A RECOVER statement.

OBJECTIVE: To provide recommendations for reviewing and reporting clinical in-hospital cardiopulmonary resuscitation (CPR) events in dogs and cats and to establish nonambiguous operational definitions for CPR terminology. DESIGN: Consensus guidelines. SETTING: International, academia, referral practice, general practice, and human medicine. METHODS: An international veterinary Utstein task force was convened in April 2013 in San Francisco to determine the scope of the project, the variables to be reported, their definitions, and a reporting template. Factors that were essential for meaningful data reporting and were amenable to accurate collection (ie, core variables) and additional variables useful for research projects and hypothesis generation (ie, supplemental variables) were defined. Consensus on each item was either achieved during that meeting or during the subsequent online modified Delphi process and dialogue between task force members. RESULTS: Variables were defined and categorized as hospital, animal, event (arrest), and outcome variables. This report recommends a template for standardized reporting of veterinary in-hospital CPR studies involving dogs or cats. Core elements include the suspected cause(s) and location of arrest, first rhythm identified, the occurrence of return of spontaneous circulation (ROSC) of more than 30 seconds (any ROSC) or more than 20 minutes (sustained ROSC), survival to discharge, and functional capacity at discharge. If CPR is discontinued or the patient is euthanized by owner request, a reason is reported. The task force suggests a case report form to be used for individual resuscitation events. CONCLUSIONS: The availability of these veterinary small animal CPR reporting guidelines will encourage and facilitate high-quality veterinary CPR research, improve data comparison between studies and across study sites, and serve as the foundation for veterinary CPR registries.

No up-regulation of the phosphatidylethanolamine N-methyltransferase pathway and choline production by sex hormones in cats.

BACKGROUND: Feline hepatic lipidosis (FHL) is a common cholestatic disease affecting cats of any breed, age and sex. Both choline deficiency and low hepatic phosphatidylethanolamine N-methyltransferase (PEMT) activity are associated with hepatic lipidosis (HL) in humans, mice and rats. The PEMT expression is known to be upregulated by oestrogens, protecting the females in these species from the development of HL when exposed to choline deficient diets. The aim of the present study was to evaluate the influence of sex hormones on choline synthesis via the PEMT pathway in healthy male and female cats before and after spaying/neutering, when fed a diet with recommended dietary choline content. RESULTS: From six female and six male cats PEMT activity was assayed directly in liver biopsies taken before and after spaying/neutering, and assessed indirectly by analyses of PEMT-specific hepatic phosphatidylcholine (PC) species and plasma choline levels. Hepatic PEMT activity did not differ between intact female and male cats and no changes upon spaying/neutering were observed. Likewise, no significant differences in liver PC content and PEMT-specific polyunsaturated PC species were found between the sexes and before or after spaying/neutering. CONCLUSION: These results suggest that choline synthesis in cats differs from what is observed in humans, mice and rats. The lack of evident influence of sex hormones on the PEMT pathway makes it unlikely that spaying/neutering predisposes cats for HL by causing PC deficiency as suggested in other species.

Intellectual disability and bleeding diathesis due to deficient CMP--sialic acid transport.

OBJECTIVE: To identify the underlying genetic defect in a patient with intellectual disability, seizures, ataxia, macrothrombocytopenia, renal and cardiac involvement, and abnormal protein glycosylation. METHODS: Genetic studies involved homozygosity mapping by 250K single nucleotide polymorphism array and SLC35A1 sequencing. Functional studies included biochemical assays for N-glycosylation and mucin-type O-glycosylation and SLC35A1-encoded cytidine 5'-monophosphosialic acid (CMP-sialic acid) transport after heterologous expression in yeast. RESULTS: We performed biochemical analysis and found combined N- and O-glycosylation abnormalities and specific reduction in sialylation in this patient. Homozygosity mapping revealed homozygosity for the CMP-sialic acid transporter SLC35A1. Mutation analysis identified a homozygous c.303G > C (p.Gln101His) missense mutation that was heterozygous in both parents. Functional analysis of mutant SLC35A1 showed normal Golgi localization but 50% reduction in transport activity of CMP-sialic acid in vitro. CONCLUSION: We confirm an autosomal recessive, generalized sialylation defect due to mutations in SLC35A1. The primary neurologic presentation consisting of ataxia, intellectual disability, and seizures, in combination with bleeding diathesis and proteinuria, is discriminative from a previous case described with deficient sialic acid transporter. Our study underlines the importance of sialylation for normal CNS development and regular organ function.

Evaluation of a veterinary triage list modified from a human five-point triage system in 485 dogs and cats.

OBJECTIVES: To devise a veterinary triage list (VTL) and to determine whether the application of this VTL results in more accurate categorization of emergency patients compared with intuitive triage. DESIGN: Prospective and retrospective observational study. SETTING: Private veterinary emergency clinic. ANIMALS: Four hundred and eighty-five client-owned dogs and cats. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: A VTL was composed using a human triage system and data from medical records of the study group. Target waiting times were prospectively determined using intuition by veterinary nurses (TWT-N).  Target waiting times were subsequently determined retrospectively by the use of the VTL (TWT-VTL).  Both TWT-N and TWT-VTL were compared against target waiting times determined by a review team (TWT-R), which was considered the gold standard. TWT categories included 0, 15, 30-60, and 120 minutes, and were associated with triage categories red, orange, yellow, and green, respectively. Differences in agreement were tested for significance. One hundred and eighty-five dogs and 300 cats fulfilled the inclusion criteria. TWT-N and TWT-R agreed on 30 cases of 67 (44.8%) in triage category red and 22 of 89 (24.7%) in category orange. TWT-VTL and TWT-R agreed on 64 cases of 67 (95.5%) in category red and 75 of 89 (84.3%) in category orange. Agreement between TWT-VTL and TWT-R (Pearson's R = 0.848) was significantly greater (P < 0.001) than agreement between TWT-N and TWT-R (Pearson's R = 0.519). CONCLUSIONS: Intuitive triage performed by veterinary nurses showed significantly less correlation with TWT-R than triage performed with the VTL. A short physical examination in all emergency patients appears to be essential in recognizing critical disease. The use of a standardized VTL can help to categorize veterinary emergency patients.

The succinate receptor as a novel therapeutic target for oxidative and metabolic stress-related conditions.

The succinate receptor (also known as GPR91) is a G protein-coupled receptor that is closely related to the family of P2Y purinoreceptors. It is expressed in a variety of tissues, including blood cells, adipose tissue, the liver, retina, and kidney. In these tissues, this receptor and its ligand succinate have recently emerged as novel mediators in local stress situations, including ischemia, hypoxia, toxicity, and hyperglycemia. Amongst others, the succinate receptor is involved in recruitment of immune cells to transplanted tissues. Moreover, it was shown to play a key role in the development of diabetic retinopathy. However, most prominently, the role of locally increased succinate levels and succinate receptor activation in the kidney, stimulating the systemic and local renin-angiotensin system, starts to unfold: the succinate receptor is a key mediator in the development of hypertension and possibly fibrosis in diabetes mellitus and metabolic syndrome. This makes the succinate receptor a promising drug target to counteract or prevent cardiovascular and fibrotic defects in these expanding disorders. Recent development of SUCNR1-specific antagonists opens novel possibilities for research in models for these disorders and may eventually provide novel opportunities for the treatment of patients.

Novel role for proteinase-activated receptor 2 (PAR2) in membrane trafficking of proteinase-activated receptor 4 (PAR4).

Proteinase-activated receptors 4 (PAR(4)) is a class A G protein-coupled receptor (GPCR) recognized through the ability of serine proteases such as thrombin and trypsin to mediate receptor activation. Due to the irreversible nature of activation, a fresh supply of receptor is required to be mobilized to the cell surface for responsiveness to agonist to be sustained. Unlike other PAR subtypes, the mechanisms regulating receptor trafficking of PAR(4) remain unknown. Here, we report novel features of the intracellular trafficking of PAR(4) to the plasma membrane. PAR(4) was poorly expressed at the plasma membrane and largely retained in the endoplasmic reticulum (ER) in a complex with the COPI protein subunit ß-COP1. Analysis of the PAR(4) protein sequence identified an arginine-based (RXR) ER retention sequence located within intracellular loop-2 (R(183)AR → A(183)AA), mutation of which allowed efficient membrane delivery of PAR(4). Interestingly, co-expression with PAR(2) facilitated plasma membrane delivery of PAR(4), an effect produced through disruption of ß-COP1 binding and facilitation of interaction with the chaperone protein 14-3-3ζ. Intermolecular FRET studies confirmed heterodimerization between PAR(2) and PAR(4). PAR(2) also enhanced glycosylation of PAR(4) and activation of PAR(4) signaling. Our results identify a novel regulatory role for PAR(2) in the anterograde traffic of PAR(4). PAR(2) was shown to both facilitate and abrogate protein interactions with PAR(4), impacting upon receptor localization and cell signal transduction. This work is likely to impact markedly upon the understanding of the receptor pharmacology of PAR(4) in normal physiology and disease.