CAR/CXCR5-T cell immunotherapy is safe and potentially efficacious in promoting sustained remission of SIV infection.

During chronic human immunodeficiency virus (HIV) or simian immunodeficiency virus (SIV) infection prior to AIDS progression, the vast majority of viral replication is concentrated within B cell follicles of secondary lymphoid tissues. We investigated whether infusion of T cells expressing an SIV-specific chimeric antigen receptor (CAR) and the follicular homing receptor, CXCR5, could successfully kill viral-RNA+ cells in targeted lymphoid follicles in SIV-infected rhesus macaques. In this study, CD4 and CD8 T cells from rhesus macaques were genetically modified to express antiviral CAR and CXCR5 moieties (generating CAR/CXCR5-T cells) and autologously infused into a chronically infected animal. At 2 days post-treatment, the CAR/CXCR5-T cells were located primarily in spleen and lymph nodes both inside and outside of lymphoid follicles. Few CAR/CXCR5-T cells were detected in the ileum, rectum, and lung, and no cells were detected in the bone marrow, liver, or brain. Within follicles, CAR/CXCR5-T cells were found in direct contact with SIV-viral RNA+ cells. We next infused CAR/CXCR5-T cells into ART-suppressed SIV-infected rhesus macaques, in which the animals were released from ART at the time of infusion. These CAR/CXCR5-T cells replicated in vivo within both the extrafollicular and follicular regions of lymph nodes and accumulated within lymphoid follicles. CAR/CXR5-T cell concentrations in follicles peaked during the first week post-infusion but declined to undetectable levels after 2 to 4 weeks. Overall, CAR/CXCR5-T cell-treated animals maintained lower viral loads and follicular viral RNA levels than untreated control animals, and no outstanding adverse reactions were noted. These findings indicate that CAR/CXCR5-T cell treatment is safe and holds promise as a future treatment for the durable remission of HIV.

Retrospective study of 20 cats surgically treated for insulinoma.

OBJECTIVE: To report the clinical signs, histopathology results, and prognostic factors for outcomes following excision for feline insulinoma (INS). STUDY DESIGN: Retrospective study. SAMPLE POPULATION: Twenty client-owned cats. METHODS: Medical records from 2006 to 2020 were reviewed by Veterinary Society of Surgical Oncology members for cats with hypoglycemia resulting from INS, with surgical excision and follow up. Clinical signs and histopathology results were summarized. Factors potentially related to disease-free interval (DFI), disease-related death (DRD), and overall survival time (OST) were analyzed with a Cox proportional hazards regression analysis. RESULTS: All cats were hypoglycemic on presentation with neurologic signs in 18 out of 20 and inappropriate insulin levels in 12/13. Excision of insulinomas resulted in immediate euglycemia or hyperglycemia in 18 cats. Eighteen cats survived to hospital discharge. The median time to death or last postoperative follow up was 664 days (range: 2-1205 days). Prognostic factors included age at presentation (for DFI); time to postoperative euglycemia (for DRD); preoperative and postoperative serum blood glucose concentrations; metastasis at the time of surgery (DFI and DRD), and histopathologic tumor invasion (for OST). The median OST for all cats was 863 days. The 1-, 2- and 3-year survival rates were 75%, 51%, and 10%, respectively. CONCLUSION: Excision of insulinoma resulted in euglycemia or hyperglycemia in most cats. Negative prognostic factors included young age, low serum glucose concentrations, metastasis at time of surgery, tumor invasion, and shorter time to euglycemia. CLINICAL SIGNIFICANCE: Surgical excision resulted in survival times comparable to those of canine INS.

Validation of the 0-10 verbal numeric scale for assessment of pruritus severity in dogs.

BACKGROUND: The pruritus Visual Analog Scale (pVAS) is currently the only validated tool for assessing canine pruritus. A verbal numeric scale (VNS) offers an alternative if clients are not present or able to complete the pVAS. HYPOTHESIS/OBJECTIVE: To validate the 0-10 VNS and evaluate its interchangeability with the pVAS. ANIMALS: 251 dogs were included in the study. MATERIALS AND METHODS: Dog owners were asked to assess their pet's pruritus level using the pVAS, the 0-10 VNS and the verbal rating categories (mild, moderate, severe) in one or two subsequent visits. To be valid, VNS must satisfy the following: correlate with pVAS (criterion validity); indicate reduction in itch score after an antipruritic intervention (construct validity); and have scores related to the verbal rating categories that are significantly different (content validity). VNS and pVAS interchangeability and the percentage of clients that preferred the VNS and pVAS also were evaluated. RESULTS: The VNS and pVAS correlation for visits 1 and 2 was good (rICC = 0.9) and excellent (rICC = 0.94). The VNS scores were significantly decreased after antipruritic interventions (P < 0.001). The VNS scores associated with the verbal rating scale categories (mild, moderate and severe) were significantly different (P < 0.001). The 95% limits of agreement for the pVAS and VAS were outside the limit of acceptability of ±2 (-2.0, 2.57). Most clients (69%) preferred the pVAS to the VNS. CONCLUSIONS AND CLINICAL RELEVANCE: The VNS is a valid scale to evaluate canine pruritus; however, the VNS and pVAS are not interchangeable.

Low levels of SIV-specific CD8+ T cells in germinal centers characterizes acute SIV infection.

CD8+ T cells play an important role in controlling of HIV and SIV infections. However, these cells are largely excluded from B cell follicles where HIV and SIV producing cells concentrate during chronic infection. It is not known, however, if antigen-specific CD8+ T cells are excluded gradually as pathogenesis progresses from early to chronic phase, or this phenomenon occurs from the beginning infection. In this study we determined that SIV-specific CD8+ T cells were largely excluded from follicles during early infection, we also found that within follicles, they were entirely absent in 60% of the germinal centers (GCs) examined. Furthermore, levels of SIV-specific CD8+ T cells in follicular but not extrafollicular areas significantly correlated inversely with levels of viral RNA+ cells. In addition, subsets of follicular SIV-specific CD8+ T cells were activated and proliferating and expressed the cytolytic protein perforin. These studies suggest that a paucity of SIV-specific CD8+ T cells in follicles and complete absence within GCs during early infection may set the stage for the establishment of persistent chronic infection.

DLA class II haplotypes show sex-specific associations with primary hypoadrenocorticism in Standard Poodle dogs.

Addison's disease (AD) is a life-threatening endocrine disorder that occurs spontaneously in both humans and dogs. Associations between MHC class II genes and AD have been shown in several human studies. Our goal was to identify MHC class II associations with AD in a large population of Standard Poodles, a breed highly predisposed to AD. We sequenced exon 2 of the class II genes DLA-DRB1, DLA-DQA1, and DLA-DQB1 in 110 affected and 101 unaffected Standard Poodles and tested for association with AD. After correcting for population structure, two haplotypes were found to confer risk of developing AD in a sex-specific manner: DLA-DRB1*015:01-DQA1*006:01-DQB1*023:01 in males (x2p = 0.03, OR 2.1) and DLA-DRB1*009:01-DQA1*001:01-DQB1*008:01:1 in females (x2p = 0.02, OR 8.43). Sex-specific associations have been previously described in human populations, but this is the first report of this kind in dogs. Consistent with findings in other studies, we found the DLA-DQA1*006:01 allele (x2p = 0.04) to be associated with AD in males independent of haplotype. In females, the haplotype DLA-DRB1*009:01-DQA1*001:01-DQB1*008:01:1 confers a very high risk for developing AD, although its frequency was rare (9 of 124 females) in our study population. Further studies are warranted to validate the findings of this exploratory dataset and to assess the usefulness of this haplotype as a risk marker for AD in female Standard Poodles. Our results highlight the importance of evaluating MHC class II disease associations in large populations, and accounting for both biological sex and population structure.

An in-home intervention of parent-implemented strategies to increase child vegetable intake: results from a non-randomized cluster-allocated community trial.

BACKGROUND: Less than 2% of children in the U.S., ages 9-13, meet the minimum dietary recommendations for vegetable intake. The home setting provides potential opportunities to promote dietary behavior change among children, yet limited trials exist with child vegetable intake as a primary outcome. Strategies to increase vegetable intake grounded in behavioral economics are no/low cost and may be easily implemented in the home by parents. METHODS: This non-randomized, controlled study tested whether an intervention of parent-led strategies informed by behavioral economics and implemented within a series of 6 weekly parent-child vegetable cooking skills classes, improved dietary outcomes of a diverse sample of low-income children (ages 9-12) more than the vegetable cooking skills classes alone. The primary outcomes were total vegetable intake, dietary quality (HEI scores), total energy intake, vegetable liking, variety of vegetables tried, child BMI-z score, and home availability of vegetables. Outcome measures were collected at baseline, immediate post-treatment, 6 and 12 months follow-up. Mixed model regression analyses with fixed independent effects (treatment condition, time point and treatment condition x time interaction) were used to compare outcomes between treatment conditions. RESULTS: A total of 103 parent/child pairs (intervention = 49, control = 54) were enrolled and 91 (intervention = 44, control = 47) completed the weekly cooking skills program. The intervention did not improve child total vegetable intake. Intervention children increased dark green vegetable intake from immediate post-treatment to 12 months. The number of vegetables children tried increased and mean vegetable liking decreased over time for both control and intervention children. CONCLUSIONS: Findings from this study suggest that the strategies and the manner in which they were implemented may not be effective in low-income populations. The burden of implementing a number of strategies with potentially higher food costs may have constrained the ability of families in the current study to use the strategies as intended. TRIAL REGISTRATION: This trial has been retrospectively registered at : # NCT03641521 on August 21, 2018.

Canine Pruritus Visual Analog Scale: how does it capture owners' perception of their pet's itching level?

BACKGROUND: Accurate measurement of pruritus severity is difficult in veterinary medicine. OBJECTIVES: To determine how the changes in Pruritus Visual Analog Scale (PVAS) scores at follow-up visits agree with the owners' perceptions of improvement of their pet's pruritus. ANIMALS: One hundred and ninety two pruritic dogs were included in the prospective study and 196 in the retrospective study. METHODS AND MATERIALS: Owners were randomly assigned into five groups and PVAS scores were recorded during two consecutive visits. Group A: previous scores were shown before completing the PVAS; Group B: PVAS was completed then owners were shown previous scores and asked to repeat the PVAS; Group C: PVAS was completed as reported previously; Group D: PVAS and a 0-10 verbal scale (VS) were completed. Retrospectively, PVAS scores were analysed during at least three consecutive visits. The average percentage and kappa agreements were calculated for all groups. In addition, PVAS and VS scores were compared in Group D. RESULTS: The average percentage and kappa agreements were higher in groups A (96%; 0.81), B [before (80%; 0.54), after (82%; 0.59) previous score] and D (85%; 0.47). Group C (79%; 0.37) had the lowest agreement. PVAS and VS scores were not significantly different (P = 0.56) in Group D. The average percentage and kappa agreements for the retrospective study were 50.8% and 0.25. The highest values (63%; 0.355) were noted at 30-60 day visit intervals. CONCLUSIONS AND CLINICAL IMPORTANCE: Showing owners previous scores could improve how PVAS captures the owner's perception of their dog's itching level.

Once daily oral extended-release hydrocodone as analgesia following tibial plateau leveling osteotomy in dogs.

OBJECTIVE: To determine the efficacy of oral extended-release hydrocodone compared with oral firocoxib for analgesia following tibial plateau leveling osteotomy surgery in dogs in a hospital setting. STUDY DESIGN: Prospective, randomized, blinded, clinical trial. ANIMALS: Thirty-six client-owned dogs weighing 25-43 kg with unilateral hindlimb lameness and cranial cruciate ligament rupture. METHODS: Dogs were assigned to 1 of 2 groups (n = 18/group). Group 1 received hydrocodone 3 mg/kg orally every 24 hours, and group 2 received firocoxib 5 mg/kg orally every 24 hours. Both hydrocodone and firocoxib (according to group assignment) were provided as preemptive analgesia 10 hours before induction to anesthesia and then every 24 hours for the remainder of the study period The level of analgesia was compared between treatments on the basis of a modified Glasgow Composite Pain Score (mGCPS) in each dog, the number of dogs requiring rescue analgesia (hydromorphone 0.05 mg/kg subcutaneously), pressure platform stance data, and number of adverse events. RESULTS: Nine of 18 dogs that received hydrocodone and 2/18 dogs that received firocoxib had an mGCPS ≥6 (P = .02). Two dogs had an mGCPS ≥6 three times, and 1 had an mGCPS ≥6 two times; all 3 of these dogs were in the hydrocodone group. Average postoperative peak pressure placed on the affected limb was lower in dogs that received hydrocodone (P = .01). Regurgitation and decreased appetite were more common in the dogs that received hydrocodone. CONCLUSION: Dogs that were treated with hydrocodone exhibited higher pain scores and lower limb function than dogs treated with firocoxib under the conditions of our study. CLINICAL SIGNIFICANCE: Our results do not provide evidence to justify the administration of extended-release hydrocodone at 3 mg/kg orally every 24 hours rather than firocoxib at 5 mg/kg orally every 24 hours in dogs undergoing tibial plateau leveling osteotomy.

Proteome Scale-Protein Turnover Analysis Using High Resolution Mass Spectrometric Data from Stable-Isotope Labeled Plants.

Protein turnover is an important aspect of the regulation of cellular processes for organisms when responding to developmental or environmental cues. The measurement of protein turnover in plants, in contrast to that of rapidly growing unicellular organismal cultures, is made more complicated by the high degree of amino acid recycling, resulting in significant transient isotope incorporation distributions that must be dealt with computationally for high throughput analysis to be practical. An algorithm in R, ProteinTurnover, was developed to calculate protein turnover with transient stable isotope incorporation distributions in a high throughput automated manner using high resolution MS and MS/MS proteomic analysis of stable isotopically labeled plant material. ProteinTurnover extracts isotopic distribution information from raw MS data for peptides identified by MS/MS from data sets of either isotopic label dilution or incorporation experiments. Variable isotopic incorporation distributions were modeled using binomial and beta-binomial distributions to deconvolute the natural abundance, newly synthesized/partial-labeled, and fully labeled peptide distributions. Maximum likelihood estimation was performed to calculate the distribution abundance proportion of old and newly synthesized peptides. The half-life or turnover rate of each peptide was calculated from changes in the distribution abundance proportions using nonlinear regression. We applied ProteinTurnover to obtain half-lives of proteins from enriched soluble and membrane fractions from Arabidopsis roots.

Genome-wide analysis reveals selection for important traits in domestic horse breeds.

Intense selective pressures applied over short evolutionary time have resulted in homogeneity within, but substantial variation among, horse breeds. Utilizing this population structure, 744 individuals from 33 breeds, and a 54,000 SNP genotyping array, breed-specific targets of selection were identified using an F(ST)-based statistic calculated in 500-kb windows across the genome. A 5.5-Mb region of ECA18, in which the myostatin (MSTN) gene was centered, contained the highest signature of selection in both the Paint and Quarter Horse. Gene sequencing and histological analysis of gluteal muscle biopsies showed a promoter variant and intronic SNP of MSTN were each significantly associated with higher Type 2B and lower Type 1 muscle fiber proportions in the Quarter Horse, demonstrating a functional consequence of selection at this locus. Signatures of selection on ECA23 in all gaited breeds in the sample led to the identification of a shared, 186-kb haplotype including two doublesex related mab transcription factor genes (DMRT2 and 3). The recent identification of a DMRT3 mutation within this haplotype, which appears necessary for the ability to perform alternative gaits, provides further evidence for selection at this locus. Finally, putative loci for the determination of size were identified in the draft breeds and the Miniature horse on ECA11, as well as when signatures of selection surrounding candidate genes at other loci were examined. This work provides further evidence of the importance of MSTN in racing breeds, provides strong evidence for selection upon gait and size, and illustrates the potential for population-based techniques to find genomic regions driving important phenotypes in the modern horse.

Genome-wide association study implicates testis-sperm specific FKBP6 as a susceptibility locus for impaired acrosome reaction in stallions.

Impaired acrosomal reaction (IAR) of sperm causes male subfertility in humans and animals. Despite compelling evidence about the genetic control over acrosome biogenesis and function, the genomics of IAR is as yet poorly understood, providing no molecular tools for diagnostics. Here we conducted Equine SNP50 Beadchip genotyping and GWAS using 7 IAR-affected and 37 control Thoroughbred stallions. A significant (P<6.75E-08) genotype-phenotype association was found in horse chromosome 13 in FK506 binding protein 6 (FKBP6). The gene belongs to the immunophilins FKBP family known to be involved in meiosis, calcium homeostasis, clathrin-coated vesicles, and membrane fusions. Direct sequencing of FKBP6 exons in cases and controls identified SNPs g.11040315G>A and g.11040379C>A (p.166H>N) in exon 4 that were significantly associated with the IAR phenotype both in the GWAS cohort (n = 44) and in a large multi-breed cohort of 265 horses. All IAR stallions were homozygous for the A-alleles, while this genotype was found only in 2% of controls. The equine FKBP6 was exclusively expressed in testis and sperm and had 5 different transcripts, of which 4 were novel. The expression of this gene in AC/AG heterozygous controls was monoallelic, and we observed a tendency for FKBP6 up-regulation in IAR stallions compared to controls. Because exon 4 SNPs had no effect on the protein structure, it is likely that FKBP6 relates to the IAR phenotype via regulatory or modifying functions. In conclusion, FKBP6 was considered a susceptibility gene of incomplete penetrance for IAR in stallions and a candidate gene for male subfertility in mammals. FKBP6 genotyping is recommended for the detection of IAR-susceptible individuals among potential breeding stallions. Successful use of sperm as a source of DNA and RNA propagates non-invasive sample procurement for fertility genomics in animals and humans.

Clustering analysis of lipoprotein profiles to identify subtypes of hypertriglyceridemia in Miniature Schnauzers.

BACKGROUND: Hypertriglyceridemia (HTG) is prevalent in Miniature Schnauzers, predisposing them to life-threatening diseases. Varied responses to management strategies suggest the possibility of multiple subtypes. HYPOTHESIS/OBJECTIVE: To identify and characterize HTG subtypes in Miniature Schnauzers through cluster analysis of lipoprotein profiles. We hypothesize that multiple phenotypes of primary HTG exist in this breed. ANIMALS: Twenty Miniature Schnauzers with normal serum triglyceride concentration (NTG), 25 with primary HTG, and 5 with secondary HTG. METHODS: Cross-sectional study using archived samples. Lipoprotein profiles, generated using continuous lipoprotein density profiling, were clustered with hierarchical cluster analysis. Clinical data (age, sex, body condition score, and dietary fat content) was compared between clusters. RESULTS: Six clusters were identified. Dogs with primary HTG were dispersed among 4 clusters. One cluster showed the highest intensities for triglyceride-rich lipoprotein (TRL) and low-density lipoprotein (LDL) fractions and also included 4 dogs with secondary HTG. Two clusters had moderately high TRL fraction intensities and low-to-intermediate LDL intensities. The fourth cluster had high LDL but variable TRL fraction intensities with equal numbers of NTG and mild HTG dogs. The final 2 clusters comprised only NTG dogs with low TRL intensities and low-to-intermediate LDL intensities. The clusters did not appear to be driven by differences in the clinical data. CONCLUSIONS AND CLINICAL IMPORTANCE: The results of this study support a spectrum of lipoprotein phenotypes within Miniature Schnauzers that cannot be predicted by triglyceride concentration alone. Lipoprotein profiling might be useful to determine if subtypes have different origins, clinical consequences, and response to treatment.

Effect of Holstein genotype on ex-vivo interleukin-1ß response to lipopolysaccharide (LPS), lipoteichoic acid (LTA) and heat-killed Gram-negative and Gram-positive bacteria.

Effects of Holstein genotype on interleukin-1ß response were assessed by ex-vivo stimulation of whole blood with lipopolysaccharide (LPS), lipoteichoic acid (LTA), or sonicated, heat-killed Gram-negative or Gram-positive bacteria. Holstein genotypes were unselected Holsteins (UH, n = 14) not subjected to selection pressures since the mid-1960s and contemporary Holsteins (CH, n = 13). Milk yield of UH and CH cows differ by more than 4500 kg/lactation. Whole blood was mixed with 0.01 µg LPS, 10 µg LTA or 2.5 × 106 CFU of sonicated, heat-killed E. coli, K. pneumoniae, S. marcescens, S. aureus, S. dysgalactiae, or S. uberis per mL of blood and incubated (4 h, 37 °C). Plasma IL-1ß was quantified by ELISA and log10-transformed concentrations analyzed with a multivariate linear mixed effects model. Responses to bacteria were greater than responses to LPS or LTA. Responses to LPS, LTA and the Gram-negative stimulants were greater in UH than in CH cows while responses to the Gram-positive bacteria did not differ between Holstein genotypes. In both genotypes, strong correlations were detected among IL-1ß responses to the Gram-negative stimulants and to LTA. There were strong correlations among IL-1ß responses to the Gram-positive bacteria in CH cows but only between S. aureus and S. dysgalactiae in UH cows. The IL-1ß response to S. uberis was highly correlated with responses to all of the Gram-negative stimulants in CH cows but only with E. coli in the UH cows. The reduced immune response could make contemporary cows more susceptible to infection by Gram-negative bacteria. Results confirm selection practices since the mid-1960s have altered immune response in the Holstein, at least to Gram-negative bacteria, and validate the need for additional studies to further evaluate the impacts of these selection practices on immune function in contemporary Holsteins.

Prevalence and Predictors of Radiographically Apparent Upper Urinary Tract Urolithiasis in Eight Dog Breeds Predisposed to Calcium Oxalate Urolithiasis and Mixed Breed Dogs.

Data on upper urinary tract (UUT) uroliths in dogs are important to understanding their etiology. The aim of this retrospective case-control study was to determine the prevalence and identify predictors of radiographically apparent UUT uroliths in dog breeds at increased risk for calcium oxalate uroliths (CaOx risk breeds) and mixed breed dogs. Radiologist reports of three-view abdominal radiographs were reviewed from 251 purebred dogs of 8 CaOx risk breeds and 68 mixed breed dogs. UUT uroliths were more common in CaOx risk breeds than mixed breed dogs (23% versus 6%, respectively; OR = 4.8, 95% confidence interval [CI] 1.7−18.9, p < 0.001). UUT uroliths were more common in dogs with lower urinary tract (LUT) uroliths (predominantly calcium-containing) than those without (41% versus 5%, respectively; OR = 13.6, 95% CI 6.3−33.1, p < 0.001), and LUT uroliths predicted the presence of UUT uroliths in the multivariable regression (OR = 6.5, 95% CI 2.8−16.7, p < 0.001). Increasing age (p < 0.001) and lower body weight (p = 0.0016) were also predictors of UUT urolith presence in the multivariable regression. The high prevalence of UUT uroliths in dogs with LUT uroliths supports a shared mechanism for their formation.

Retrospective evaluation of fluid production at the time of thoracostomy tube removal following elective and emergency surgery in dogs (2010-2017): 185 cases.

OBJECTIVE: To report the rate of fluid production at the time of removal of thoracostomy tubes placed intraoperatively and to determine the association of this rate with specific patient factors, surgical factors, or clinical diagnosis. The secondary objective was to determine whether identification of pleural effusion within 2 weeks of thoracostomy tube removal was associated with the same variables. DESIGN: Retrospective study. SETTING: University teaching hospital. ANIMALS: One hundred eighty-five client-owned dogs with thoracostomy tubes placed intraoperatively between January 2010 and March 2017. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Thoracostomy tubes were removed at a median fluid production of 0.09 mL/kg/h (range, 0-7.0 m L/kg/h). Median fluid production at the time of thoracostomy tube removal was significantly higher in dogs with preoperative pleural effusion compared to dogs without preoperative pleural effusion (0.21 vs 0.05 mL/kg/h; P = 0.0001) and in dogs that had a median sternotomy compared to dogs that had a lateral thoracotomy (0.14 vs 0.09 mL/kg/h; P = 0.04). Of the 169 dogs available for follow-up, 12 (7.1%) had pleural effusion within 2 weeks of removal of the thoracostomy tube. Detection of pleural effusion during the follow-up period was significantly associated with the presence of preoperative pleural effusion (P = 0.0019) and the diagnosis (P = 0.01). A greater proportion of dogs with a lung lobe torsion (4/9, 44.4%) and idiopathic chylothorax (2/7, 28.5%) had pleural effusion within 2 weeks compared to other diagnoses. Reintervention was performed in 4.7% of dogs. CONCLUSIONS: Thoracostomy tubes were removed at pleural fluid production rates that frequently exceeded current veterinary guidelines. However, the fluid production rate at the time of thoracostomy tube removal was not associated with the detection of pleural effusion within 2 weeks of thoracostomy tube removal, and the overall need for reintervention following thoracostomy tube removal was low (4.7%).

Behavioral and Physiologic Effects of Dirty Bedding Exposure in Female ICR Mice.

Exposure of sentinel mice to dirty bedding is commonly used in health monitoring programs to screen colonies for clinical and subclinical disease. Despite the potential stressors present in dirty bedding, including but not limited to microorganisms, pheromones, and ammonia, it is unknown whether sentinel mice exposed to soiled bedding experience stress. In this study, select behavioral and physiologic changes associated with stress were assessed in female ICR mice exposed to dirty bedding. Behavioral parameters included evaluation in the home cage and selected behavioral tests; physiologic measurements included neutrophil:lymphocyte ratio and weight. Mice in the acute group were exposed for 24 h whereas mice in the chronic group were exposed for 4 wk. Mice in the chronic group exposed to dirty bedding weighed less at days 21 and 28 than did control mice. Chronic mice exposed to dirty bedding also exhibited decreased net weight gain over the entire study period as compared with control mice. No significant differences were detected in the other behavioral and physiologic parameters measured. These results indicate that dirty bedding exposure may affect sentinel mice, but further investigation is needed to determine the specific mechanism(s) behind the weight difference.

HIV-Specific CAR T Cells with CD28 or 4-1BB Signaling Domains Are Phenotypically and Functionally Distinct and Effective at Suppressing HIV and Simian Immunodeficiency Virus.

Despite mounting a robust antiviral CD8 T cell response to HIV infection, most infected individuals are unable to control HIV viral load without antiretroviral therapy (ART). Chimeric Ag receptor (CAR) T cell treatment is under intensive investigation as an alternative therapy for ART-free remission of chronic HIV infection. However, achieving durable remission of HIV will require a successful balance between CAR T cell effector function and persistence. CAR T cells with CD28 costimulatory domains have robust effector function but limited persistence in vivo, whereas CAR T cells with 4-1BB costimulatory domains present a more undifferentiated phenotype and greater in vivo persistence. We compared the in vitro phenotype and function of rhesus macaque and human CAR T cells that contained either the CD28 or 4-1BB costimulatory domain; both constructs also included CARs that are bispecific for gp120 of HIV or SIV and the CXCR5 moiety to promote in vivo homing of CAR/CXCR5 T cells to B cell follicles. Cells were transduced using a gammaretroviral vector and evaluated using flow cytometry. 4-1BB-CAR/CXCR5 T cells were phenotypically distinct from CD28-CAR/CXCR5 T cells and showed increased expression of CAR and CD95. Importantly, both CD28- and 4-1BB-CAR/CXCR5 T cells retained equal capacity to recognize and suppress SIV in vitro. These studies provide new insights into rhesus macaque and human 4-1BB- and CD28-bearing CAR T cells.

Evaluation of coated platelets, a subset of highly procoagulant platelets, in healthy dogs and dogs with neoplasia.

OBJECTIVE: To determine if dogs with neoplasia produce more coated platelets, a subpopulation of activated platelets generated by dual stimulation with thrombin and convulxin, a glycoprotein VI agonist, than healthy control dogs. ANIMALS: Client-owned dogs diagnosed with lymphoma (n = 19) or solid tumors (14) and healthy control dogs (14). PROCEDURES: Platelets were stimulated ex vivo with thrombin and convulxin. Flow cytometry was used to quantify the percentage of coated platelets based on high levels of surface fibrinogen. To compare the percentage of coated platelets between the three groups, an ANOVA was performed followed by pairwise 95% confidence intervals (CI) adjusted for multiple comparisons using Tukey's method. RESULTS: We observed a greater mean percentage of coated platelets in dogs with solid tumors, compared with healthy control dogs, by 10.9 percentage points (95% CI: -1.0, 22.8), and a mean percentage of coated platelets in dogs with lymphoma that was less than healthy control dogs by 0.3 percentage points (95% CI: -11.4, 10.8). CLINICAL RELEVANCE: This study provides the first data-based evidence that dogs with solid tumors may have a greater mean coated platelet percentage when compared with healthy control dogs, although there is overlap between groups. Further studies are needed investigating coated platelets in specific subsets of neoplasia and investigating additional mechanisms of hypercoagulability in dogs with neoplasia.

Measuring the turnover rates of Arabidopsis proteins using deuterium oxide: an auxin signaling case study.

Rapid environmental responses in plants rely on endogenous signaling mechanisms, which in many cases are mediated by changes in protein turnover rates. It is therefore necessary to develop methods for measuring protein dynamics that monitor large sets of plant proteins to begin to apply a systems biology approach to the study of plant behavior. The use of stable isotope labeling strategies that are adaptable to proteomic methods is particularly attractive for this purpose. Here, we explore one example of such methods that is particularly suitable for plants at the seedling stage, where measurement of amino acid and protein turnover rates is accomplished using a heavy water labeling strategy. The method is backed by microarray evaluation to define its feasibility for specific experimental approaches, and the CULLIN-ASSOCIATED AND NEDDYLATION DISSOCIATED 1 (CAND1) and TRANSPORT INHIBITOR RESPONSE 1 (TIR1) proteins are used to illustrate the potential utility in understanding hormonal signaling regulation. These studies provide insight not only into the potential utility of the method, but also address possible areas of concern regarding the use of heavy water labeling during plant growth. These considerations suggest a prescription for specific experimental designs that minimize interference resulting from the induction of treatment-specific gene expression in the results obtained.

Public Views of Dairy Calf Welfare and Dairy Consumption Habits of American Youth and Adults.

The primary objective of this study was to explore views of dairy calf welfare and dairy product consumption habits among youth and adults. The secondary objective was to explore views of dairy calf welfare and dairy product consumption habits among a subset of parent-child pairs. Participants 5-17 years of age (n = 463) and 18 years old or greater (n = 1,310) completed an in-person survey at the Minnesota State Fair (St. Paul, MN, USA) in summer 2018. A subset of these data was comprised of parent-child pairs (n = 188). The survey was administered via Qualtrics using iPads and included multiple-choice questions about demographics and calf welfare, an open-ended question on "what dairy calves need to have a good life," and multiple-choice questions about participants' consumption of dairy products and nondairy alternative products. Content analysis was used for responses to the open-ended question, and concepts to describe dairy calf welfare views were identified. Fisher's exact test and Cohen's Kappa were used to investigate the relationships between parent-child pair responses about dairy calf welfare. In addition to these methods, prevalence-adjusted and bias-adjusted kappa (PABAK) were used to investigate the relationships between parent-child pair responses about consumption habits. The median age of all youth participants was 11 years and 61% were female, 82% were urban residents, and 63% did not have prior experience handling agricultural animals but 83% had visited a farm in the past. Most youth participants (94.4%) indicated that they consumed dairy products, while 47.1% consumed nondairy alternatives products. Median age range of all adult participants was 45-54 years, 65% were female, 82% urban residents, and 81% did not have prior experience handling agricultural animals but 63% had visited a farm in the past. Most adult participants (94%) indicated that they consumed dairy products and 47% indicated that they consumed nondairy alternative products. In response to "what dairy calves need to have a good life," youth and adults most commonly focused on issues related to biological functioning (82 and 70% of youth and adults mentioning this concept, respectively), followed by natural living (44 and 50%, respectively), humane care (30 and 20%, respectively), and affective states (5% of both youth and adults). For the natural living concept of animal welfare, parent and child responses were slightly associated (Kappa = 0.19; P = 0.01; overall agreement = 61%). Almost all participants reported consuming dairy products, therefore, the agreement is high between parents and children because in most households (90%), both parents and children consume dairy products. However, child consumption was observed to be lower (75%, 9/12) when parents do not consume dairy than when parents do consume dairy (95%, 167/176), leading to a Kappa of 0.20 (P = 0.006, PABAK = 0.81) and a slight association between parents and children. The results suggest that biological functioning is highly valued by the public and views of parents and their children related to natural living in dairy calves are slightly associated.