Case report: Treatment of constrictive epicarditis using the waffle procedure in a dog that had previously undergone a subtotal pericardiectomy.

A 10 year-old female spayed German Short-haired Pointer dog weighing 26.8 kg (59 lb) presented with a 2 week history of recurrent ascites. The dog had a 4 year history of idiopathic pericardial effusion causing sporadic episodes of cardiac tamponade and secondary ascites. A subtotal pericardiectomy was performed 3 months prior to presentation. The patient had done well for 2 months following this procedure, at which point the large-volume modified transudate ascites recurred, necessitating abdominocentesis every 10 days. Thoracic and abdominal computed tomography (CT) revealed no abdominal or vascular cause of ascites. Transthoracic echocardiography performed under general anesthesia showed constrictive epicarditis (visceral pericarditis) resulting in diastolic dysfunction and right-sided congestive heart failure. A sternotomy was performed for a pericardial waffle procedure or crosshatch pericardiotomy-scoring of crosshatched incisions into the thickened epicardium. Echocardiographic findings postoperatively were consistent with resolved constrictive epicarditis. At 8 months postoperatively, the dog was clinically normal and had only required one abdominocentesis one month after the waffle procedure. This case report describes the successful treatment of a dog with constrictive epicarditis using a novel surgical technique (waffle procedure) that has not yet been described in veterinary medicine.

Orthogonal IMiD-Degron Pairs Induce Selective Protein Degradation in Cells.

Immunomodulatory imide drugs (IMiDs) including thalidomide, lenalidomide, and pomalidomide, can be used to induce degradation of a protein of interest that is fused to a short zinc finger (ZF) degron motif. These IMiDs, however, also induce degradation of endogenous neosubstrates, including IKZF1 and IKZF3. To improve degradation selectivity, we took a bump-and-hole approach to design and screen bumped IMiD analogs against 8380 ZF mutants. This yielded a bumped IMiD analog that induces efficient degradation of a mutant ZF degron, while not affecting other cellular proteins, including IKZF1 and IKZF3. In proof-of-concept studies, this system was applied to induce efficient degradation of TRIM28, a disease-relevant protein with no known small molecule binders. We anticipate that this system will make a valuable addition to the current arsenal of degron systems for use in target validation.

Size regulation of the lateral organ initiation zone and its role in determining cotyledon number in conifers.

Introduction: Unlike monocots and dicots, many conifers, particularly Pinaceae, form three or more cotyledons. These are arranged in a whorl, or ring, at a particular distance from the embryo tip, with cotyledons evenly spaced within the ring. The number of cotyledons, nc, varies substantially within species, both in clonal cultures and in seed embryos. nc variability reflects embryo size variability, with larger diameter embryos having higher nc. Correcting for growth during embryo development, we extract values for the whorl radius at each nc. This radius, corresponding to the spatial pattern of cotyledon differentiation factors, varies over three-fold for the naturally observed range of nc. The current work focuses on factors in the patterning mechanism that could produce such a broad variability in whorl radius. Molecularly, work in Arabidopsis has shown that the initiation zone for leaf primordia occurs at a minimum between inhibitor zones of HD-ZIP III at the shoot apical meristem (SAM) tip and KANADI (KAN) encircling this farther from the tip. PIN1-auxin dynamics within this uninhibited ring form auxin maxima, specifying primordia initiation sites. A similar mechanism is indicated in conifer embryos by effects on cotyledon formation with overexpression of HD-ZIP III inhibitors and by interference with PIN1-auxin patterning. Methods: We develop a mathematical model for HD-ZIP III/KAN spatial localization and use this to characterize the molecular regulation that could generate (a) the three-fold whorl radius variation (and associated nc variability) observed in conifer cotyledon development, and (b) the HD-ZIP III and KAN shifts induced experimentally in conifer embryos and in Arabidopsis. Results: This quantitative framework indicates the sensitivity of mechanism components for positioning lateral organs closer to or farther from the tip. Positional shifting is most readily driven by changes to the extent of upstream (meristematic) patterning and changes in HD-ZIP III/KAN mutual inhibition, and less efficiently driven by changes in upstream dosage or the activation of HD-ZIP III. Sharper expression boundaries can also be more resistant to shifting than shallower expression boundaries. Discussion: The strong variability seen in conifer nc (commonly from 2 to 10) may reflect a freer variation in regulatory interactions, whereas monocot (nc = 1) and dicot (nc = 2) development may require tighter control of such variation. These results provide direction for future quantitative experiments on the positional control of lateral organ initiation, and consequently on plant phyllotaxy and architecture.

Early infection is an independent risk factor for increased mortality in patients with culture-confirmed infected pancreatic necrosis.

BACKGROUND: Mortality in infected pancreatic necrosis (IPN) is dynamic over the course of the disease, with type and timing of interventions as well as persistent organ failure being key determinants. The timing of infection onset and how it pertains to mortality is not well defined. OBJECTIVES: To determine the association between mortality and the development of early IPN. METHODS: International multicenter retrospective cohort study of patients with IPN, confirmed by a positive microbial culture from (peri) pancreatic collections. The association between timing of infection onset, timing of interventions and mortality were assessed using Cox regression analyses. RESULTS: A total of 743 patients from 19 centers across 3 continents with culture-confirmed IPN from 2000 to 2016 were evaluated, mortality rate was 20.9% (155/734). Early infection was associated with a higher mortality, when early infection occurred within the first 4 weeks from presentation with acute pancreatitis. After adjusting for comorbidity, advanced age, organ failure, enteral nutrition and parenteral nutrition, early infection (≤4 weeks) and early open surgery (≤4 weeks) were associated with increased mortality [HR: 2.45 (95% CI: 1.63-3.67), p < 0.001 and HR: 4.88 (95% CI: 1.70-13.98), p = 0.003, respectively]. There was no association between late open surgery, early or late minimally invasive surgery, early or late percutaneous drainage with mortality (p > 0.05). CONCLUSION: Early infection was associated with increased mortality, independent of interventions. Early surgery remains a strong predictor of excess mortality.

Cost analysis and outcomes of endoscopic, minimal access and open pancreatic necrosectomy.

OBJECTIVES: To assess both individual patient and institutional costs as well as outcomes in patients with pancreatic necrosis who underwent either endoscopic, minimal access or open pancreatic necrosectomy. These data can be used to evaluate clinical effectiveness with a view to informing local health care providers. SUMMARY BACKGROUND DATA: Intervention for infected pancreatic necrosis is associated with a high morbidity, mortality and long hospital stays. Minimal access surgical step-up approaches have been the gold standard of care, however endoscopic approaches are now offered preferentially. METHODS: All patients undergoing endoscopic (EN), minimal access retroperitoneal (MARPN) and open (OPN) necrosectomy at a single institution from April 2015-March 2017 were included. Patients were selected for intervention based on morphology and position of the necrosis and on clinical factors. Patient level costing systems were used to determine inpatient and outpatient costs. RESULTS: 86 patients were included: 38 underwent EN, 35 MARPN and 13 OPN. Pre-operative APACHEII was 6 vs 9 vs 9 (p=0.017) and CRP 107 vs 204 vs 278, (p=0.012), respectively. Post-operative stay was 19 days for EN vs. 41 for MARPN vs. 42 for OPN (p=0.007). Complications occurred in 68.4%, 68.6% and 46.2% (p=0.298) while mortality was 10.5%, 22.9% and 15.4% (p=0.379) respectively. Mean total cost was £31,364 for EN, £52,770 for MARPN (p=0.008) and £60,346 for OPN. Ward and critical care costs for EN were lower than for MARPN (ward: £9,430 vs. £14,033, p=0.024; critical care: £5,317 vs. £16,648, p=0.056).

Single port retroperitoneal pancreatic necrosectomy for the treatment of extra-pancreatic walled off necrotic collections.

Single Port Retroperitoneal Pancreatic Necrosectomy (SPRPN), a novel method to debride extra-pancreatic necrosis after failed conventional treatment, was undertaken in 7 patients with a median collection diameter of 98 x 85 x 124mm, with resolution at a median of 42 days and post-operative median stay of 47 days.

Understanding and Measuring Long-Term Outcomes of Fingertip and Nail Bed Injuries and Treatments.

There are many outcome measures to choose from when caring for or studying fingertip and nail bed trauma and treatments. This article outlines general outcome measures principles as well as guidelines on choosing, implementing, and interpreting specific tools for these injuries. It also presents recent results from the literature for many of these measures, which can help learners, educators, and researchers by providing a clinical knowledge base and aiding study design.

Control of skeletal morphogenesis by the Hippo-YAP/TAZ pathway.

The Hippo-YAP/TAZ pathway is an important regulator of tissue growth, but can also control cell fate or tissue morphogenesis. Here, we investigate the function of the Hippo pathway during the development of cartilage, which forms the majority of the skeleton. Previously, YAP was proposed to inhibit skeletal size by repressing chondrocyte proliferation and differentiation. We find that, in vitro, Yap/Taz double knockout impairs murine chondrocyte proliferation, whereas constitutively nuclear nls-YAP5SA accelerates proliferation, in line with the canonical role of this pathway in most tissues. However, in vivo, cartilage-specific knockout of Yap/Taz does not prevent chondrocyte proliferation, differentiation or skeletal growth, but rather results in various skeletal deformities including cleft palate. Cartilage-specific expression of nls-YAP5SA or knockout of Lats1/2 do not increase cartilage growth, but instead lead to catastrophic malformations resembling chondrodysplasia or achondrogenesis. Physiological YAP target genes in cartilage include Ctgf, Cyr61 and several matrix remodelling enzymes. Thus, YAP/TAZ activity controls chondrocyte proliferation in vitro, possibly reflecting a regenerative response, but is dispensable for chondrocyte proliferation in vivo, and instead functions to control cartilage morphogenesis via regulation of the extracellular matrix.

MDH1 and MPP7 Regulate Autophagy in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is driven by metabolic changes in pancreatic cells caused by oncogenic mutations and dysregulation of p53. PDAC cell lines and PDAC-derived xenografts grow as a result of altered metabolic pathways, changes in stroma, and autophagy. Selective targeting and inhibition of one of these may open avenues for the development of new therapeutic strategies. In this study, we performed a genome-wide siRNA screen in a PDAC cell line using endogenous autophagy as a readout and identified several regulators of autophagy that were required for autophagy-dependent PDAC cell survival. Validation of two promising candidates, MPP7 (MAGUK p55 subfamily member 7, a scaffolding protein involved in cell-cell contacts) and MDH1 (cytosolic Malate dehydrogenase 1), revealed their role in early stages of autophagy during autophagosome formation. MPP7 was involved in the activation of YAP1 (a transcriptional coactivator in the Hippo pathway), which in turn promoted autophagy, whereas MDH1 was required for maintenance of the levels of the essential autophagy initiator serine-threonine kinase ULK1, and increased in the activity upon induction of autophagy. Our results provide a possible explanation for how autophagy is regulated by MPP7 and MDH1, which adds to our understanding of autophagy regulation in PDAC. SIGNIFICANCE: This study identifies and characterizes MPP7 and MDH1 as novel regulators of autophagy, which is thought to be responsible for pancreatic cancer cell survival.

Identification and Validation of Novel Autophagy Regulators Using an Endogenous Readout siGENOME Screen.

Autophagy is a highly regulated process, and its deregulation can contribute to various diseases, including cancer, immune diseases, and neurodegenerative disorders. Here we describe the design, protocol, and analysis of an imaging-based high-throughput screen with an endogenous autophagy readout. The screen uses a genome-wide siRNA library to identify autophagy regulators in mammalian cells.

A systematic review and meta-analysis of metal versus plastic stents for drainage of pancreatic fluid collections: metal stents are advantageous.

BACKGROUND: The use of fully covered metal stents (FCSEMS) and specifically designed lumen apposing metal stents for transmural drainage of pancreatic fluid collections has become widespread. A systematic review published in 2015 did not support the routine use of metal stents for drainage of pancreatic fluid collections. However, recent studies have shown conflicting data; therefore a systematic review and meta-analysis was performed. METHOD: We conducted a database search for original comparative studies between plastic and metal stents. The random effects model was used to calculate pooled risk ratios (RR) with 95% confidence intervals (CI). Outcomes analysed were clinical success, adverse events and requirement of further intervention. RESULTS: The search identified 936 studies, 7 studies with 681 (340 metal, 341 plastic) patients met inclusion criteria and were included in the meta-analysis. Clinical success was achieved in 93.8% versus 86.2% in the metal and plastic groups, respectively, RR 1.08 [95% CI 1.02-1.14]; p = 0.009. Adverse events were reduced for metal stents when compared with plastic (10.2% vs. 25.0%), RR 0.42 [95% CI 0.22-0.81]; p = 0.010. Metal stent usage reduced bleeding (2.8% vs. 7.9%), RR 0.37; [95% CI 0.18-0.75]; p = 0.006. Further intervention was required in 12.4% of patients in the metal stent group versus 26.7% for plastic stents, RR 0.54; [95% CI 0.22-1.29]; p = 0.165. CONCLUSIONS: The use of metal stents for drainage of pancreatic fluid collections is associated with improved clinical success, fewer adverse events and reduced bleeding compared to plastic stents.

A genome-wide RNAi screen identifies the SMC5/6 complex as a non-redundant regulator of a Topo2a-dependent G2 arrest.

The Topo2a-dependent arrest is associated with faithful segregation of sister chromatids and has been identified as dysfunctional in numerous tumour cell lines. This genome-protecting pathway is poorly understood and its characterization is of significant interest, potentially offering interventional opportunities in relation to synthetic lethal behaviours in arrest-defective tumours. Using the catalytic Topo2a inhibitor ICRF193, we have performed a genome-wide siRNA screen in arrest-competent, non-transformed cells, to identify genes essential for this arrest mechanism. In addition, we have counter-screened several DNA-damaging agents and demonstrate that the Topo2a-dependent arrest is genetically distinct from DNA damage checkpoints. We identify the components of the SMC5/6 complex, including the activity of the E3 SUMO ligase NSE2, as non-redundant players that control the timing of the Topo2a-dependent arrest in G2. We have independently verified the NSE2 requirement in fibroblasts from patients with germline mutations that cause severely reduced levels of NSE2. Through imaging Topo2a-dependent G2 arrested cells, an increased interaction between Topo2a and NSE2 is observed at PML bodies, which are known SUMOylation hotspots. We demonstrate that Topo2a is SUMOylated in an ICRF193-dependent manner by NSE2 at a novel non-canonical site (K1520) and that K1520 sumoylation is required for chromosome segregation but not the G2 arrest.

The Dynamics of TGF-ß Signaling Are Dictated by Receptor Trafficking via the ESCRT Machinery.

Signal transduction pathways stimulated by secreted growth factors are tightly regulated at multiple levels between the cell surface and the nucleus. The trafficking of cell surface receptors is emerging as a key step for regulating appropriate cellular responses, with perturbations in this process contributing to human diseases, including cancer. For receptors recognizing ligands of the transforming growth factor ß (TGF-ß) family, little is known about how trafficking is regulated or how this shapes signaling dynamics. Here, using whole genome small interfering RNA (siRNA) screens, we have identified the ESCRT (endosomal sorting complex required for transport) machinery as a crucial determinant of signal duration. Downregulation of ESCRT components increases the outputs of TGF-ß signaling and sensitizes cells to low doses of ligand in their microenvironment. This sensitization drives an epithelial-to-mesenchymal transition (EMT) in response to low doses of ligand, and we demonstrate a link between downregulation of the ESCRT machinery and cancer survival.

Use of an intravenous sclerosing foam (3% sodium tetradecyl sulfate) for treatment of orbital varix in a dog.

A 6-month-old intact male Standard Dachshund was referred for evaluation of a soft tissue swelling above the right eye. Examination of the right eye revealed mild lateral deviation of the globe, normal vision, and a dorsonasal soft tissue swelling. Examination of the posterior segment was normal. Dual-phase computed tomography angiography was consistent with an orbital varix of the angularis oculi and right dorsal external ophthalmic veins with no evidence of arterial involvement. Treatment involved fluoroscopically guided coil embolization of the venous outflow with nine platinum microcoils, followed by sclerotherapy of the varix using 1.5 mL of 3% sodium tetradecyl sulfate foam. Moderate-to-marked swelling was noted at the treatment site in the weeks following therapy, which gradually resolved. At final reexamination 3 months post-therapy, complete sclerosis and resolution of the orbital varix were documented. To the authors' knowledge, this is the first reported case involving the use of a sclerotic agent for successful treatment of a venous malformation in a dog.

Viability of Hand and Wrist Photogoniometry.

BACKGROUND: No goniometric technique is both maximally convenient and completely accurate, although photogoniometry (ie, picture taking to facilitate digital angle measurement) shows promise in this regard. Our purpose was to test the feasibility and reliability of a photogoniometric protocol designed to measure wrist and digit range of motion in general. METHODS: Two independent observers examined a sample of joints in both normal and abnormal hands according to a photogoniometric protocol. Interrater and intrarater correlation were calculated, and these measurements were compared with measurements made by a third independent examiner with a manual goniometer. RESULTS: The photo-based measurements were reliable within and between observers; however, only a minority of these measurements were in agreement with manually collected values. CONCLUSIONS: At present, photogoniometry is not an acceptable alternative to manual goniometry for determining wrist and digit range of motion in general. Joint-specific photogoniometry should be the subject of future study, as should relevant imaging and software technology.

Multiomic Analysis of the UV-Induced DNA Damage Response.

In order to facilitate the identification of factors and pathways in the cellular response to UV-induced DNA damage, several descriptive proteomic screens and a functional genomics screen were performed in parallel. Numerous factors could be identified with high confidence when the screen results were superimposed and interpreted together, incorporating biological knowledge. A searchable database, bioLOGIC, which provides access to relevant information about a protein or process of interest, was established to host the results and facilitate data mining. Besides uncovering roles in the DNA damage response for numerous proteins and complexes, including Integrator, Cohesin, PHF3, ASC-1, SCAF4, SCAF8, and SCAF11, we uncovered a role for the poorly studied, melanoma-associated serine/threonine kinase 19 (STK19). Besides effectively uncovering relevant factors, the multiomic approach also provides a systems-wide overview of the diverse cellular processes connected to the transcription-related DNA damage response.

Inhibition of fatty acid desaturation is detrimental to cancer cell survival in metabolically compromised environments.

BACKGROUND: Enhanced macromolecule biosynthesis is integral to growth and proliferation of cancer cells. Lipid biosynthesis has been predicted to be an essential process in cancer cells. However, it is unclear which enzymes within this pathway offer the best selectivity for cancer cells and could be suitable therapeutic targets. RESULTS: Using functional genomics, we identified stearoyl-CoA desaturase (SCD), an enzyme that controls synthesis of unsaturated fatty acids, as essential in breast and prostate cancer cells. SCD inhibition altered cellular lipid composition and impeded cell viability in the absence of exogenous lipids. SCD inhibition also altered cardiolipin composition, leading to the release of cytochrome C and induction of apoptosis. Furthermore, SCD was required for the generation of poly-unsaturated lipids in cancer cells grown in spheroid cultures, which resemble those found in tumour tissue. We also found that SCD mRNA and protein expression is elevated in human breast cancers and predicts poor survival in high-grade tumours. Finally, silencing of SCD in prostate orthografts efficiently blocked tumour growth and significantly increased animal survival. CONCLUSIONS: Our data implicate lipid desaturation as an essential process for cancer cell survival and suggest that targeting SCD could efficiently limit tumour expansion, especially under the metabolically compromised conditions of the tumour microenvironment.

Evaluation of a feline-specific multiplex, bead-based assay for detection of cytokines, chemokines, growth factors, and other immunologically active proteins in serum and plasma samples from cats.

OBJECTIVE To evaluate a feline-specific multiplex, bead-based assay system for detection of recombinant and native proteins in serum samples and in EDTA-treated and heparinized plasma samples. SAMPLE Serum samples and EDTA-treated and heparinized plasma samples from 30 sick cats and 9 healthy client-owned cats and heparinized whole blood samples from 5 healthy purpose-bred cats. PROCEDURES Ability of the assay system to detect 19 recombinant and native immunologically active proteins in plasma and serum samples from healthy and purpose-bred cats was evaluated via spike-and-recovery tests, assessments of inter- and intra-assay variation, linearity results, and leukocyte stimulation. Effects of various concentrations of heparin and serum matrix solution on percentages of analytes recovered were also evaluated. Analyte concentrations in samples from healthy and sick cats were measured and compared between groups. RESULTS Percentages of analytes recovered were unsatisfactory for most assays. Serum and heparinized plasma samples yielded better recovery results than did EDTA-treated plasma samples. Use of serum matrix solution did not improve results. Use of heparin concentrations greater than the recommended range affected the results. Linearity of results was difficult to assess because of the poor recovery. For the analytes that were recovered sufficiently for assessment, linearity appeared to be reasonable despite the limited detection. CONCLUSIONS AND CLINICAL RELEVANCE Poor percentages of analytes recovered and adverse effects of sample protein matrix limited the usefulness of the multiplex, bead-based assay system for measurement of immunologically active proteins in solutions with high protein content; however, recovery results were fairly linear, potentially allowing evaluation of feline plasma or serum samples with high analyte concentrations.

Acetyl-CoA synthetase 2 promotes acetate utilization and maintains cancer cell growth under metabolic stress.

A functional genomics study revealed that the activity of acetyl-CoA synthetase 2 (ACSS2) contributes to cancer cell growth under low-oxygen and lipid-depleted conditions. Comparative metabolomics and lipidomics demonstrated that acetate is used as a nutritional source by cancer cells in an ACSS2-dependent manner, and supplied a significant fraction of the carbon within the fatty acid and phospholipid pools. ACSS2 expression is upregulated under metabolically stressed conditions and ACSS2 silencing reduced the growth of tumor xenografts. ACSS2 exhibits copy-number gain in human breast tumors, and ACSS2 expression correlates with disease progression. These results signify a critical role for acetate consumption in the production of lipid biomass within the harsh tumor microenvironment.

Reduced NF1 expression confers resistance to EGFR inhibition in lung cancer.

Activating mutations in the EGF receptor (EGFR) are associated with clinical responsiveness to EGFR tyrosine kinase inhibitors (TKI), such as erlotinib and gefitinib. However, resistance eventually arises, often due to a second EGFR mutation, most commonly T790M. Through a genome-wide siRNA screen in a human lung cancer cell line and analyses of murine mutant EGFR-driven lung adenocarcinomas, we found that erlotinib resistance was associated with reduced expression of neurofibromin, the RAS GTPase-activating protein encoded by the NF1 gene. Erlotinib failed to fully inhibit RAS-ERK signaling when neurofibromin levels were reduced. Treatment of neurofibromin-deficient lung cancers with a MAP-ERK kinase (MEK) inhibitor restored sensitivity to erlotinib. Low levels of NF1 expression were associated with primary and acquired resistance of lung adenocarcinomas to EGFR TKIs in patients. These findings identify a subgroup of patients with EGFR-mutant lung adenocarcinoma who might benefit from combination therapy with EGFR and MEK inhibitors.