Effect of a mindfulness exercise on stress in veterinary students performing surgery.

OBJECTIVE: To determine students' stress while performing surgery and evaluate the ability of a mindfulness intervention to reduce this stress. STUDY DESIGN: Quasi-experimental design. SAMPLE POPULATION: Eighteen fourth-year DVM program students (n = 9 student/group). METHODS: Utilizing a quasi-experimental design, students were randomly assigned to a control or treatment group. The treatment group performed a 5-minute breathing (mindfulness) exercise immediately prior to performing surgery. Each student provided 3 samples of saliva, at time 0, at 10 minutes before surgery, and at 10 minutes after surgery. Students' salivary cortisol and α-amylase levels were compared between groups. Students' self-reported mood measures were also correlated to levels of salivary biomarkers. RESULTS: Cortisol and α-amylase levels of students in both groups greatly exceeded normative reference groups (>90th percentile) prior to surgery and diminished to average levels (50th-60th percentile) after surgery but did not differ between groups at any time point. Immediately prior to surgery when stress values were likely to peak, salivary α-amylase levels decreased approximately 30 U/L units for students in the treatment group compared with an increase of approximately 10 U/L units for students in the control group. Students in the treatment group reported being more calm (mean [M] 2.67, SD 1.03, d = 0.75) and relaxed (M 2.33, SD 1.51, d = 0.90) than students in the control group (M 3.44, SD 1.01 and M 3.44, SD 0.88, respectively). CONCLUSION: This study provides some evidence that the mindfulness intervention temporarily decreased stress levels and improved students' sense of calmness and relaxation immediately before operating on a live animal. CLINICAL IMPACT: Students who are experiencing less stress may be less likely to commit a medical error and negatively impact animal health. This study, the first of its kind in veterinary surgery, may serve as a model for related future studies.

Evaluation of Fourth-Year Veterinary Students' Client Communication Skills: Recommendations for Scaffolded Instruction and Practice.

Effective client communication is important for success in veterinary practice. The purpose of this project was to describe one approach to communication training and explore fourth-year veterinary students' communication skills through an evaluation of their interactions with clients during a general practice rotation. Two raters coded 20 random videotaped interactions simultaneously to assess students' communication, including their ability to initiate the session, incorporate open-ended questions, listen reflectively, express empathy, incorporate appropriate nonverbal communication, and attend to organization and sequencing. We provide baseline data that will guide future instruction in client communication. Results showed that students' communication skills require development. Half of the students sampled excelled at open-ended inquiry (n=10), and 40% (n=8) excelled at nonverbal communication. Students needed improvement on greeting clients by name and introducing themselves and their role (n=15), reflective listening (n=18), empathy (n=17), and organization and sequencing (n=18). These findings suggest that more focused instruction and practice is necessary in maintaining an organized structure, reflective listening, and empathy to create a relationship-centered approach to care.

Mapping the Energetic Epitope of an Antibody/Interleukin-23 Interaction with Hydrogen/Deuterium Exchange, Fast Photochemical Oxidation of Proteins Mass Spectrometry, and Alanine Shave Mutagenesis.

Epitope mapping the specific residues of an antibody/antigen interaction can be used to support mechanistic interpretation, antibody optimization, and epitope novelty assessment. Thus, there is a strong need for mapping methods, particularly integrative ones. Here, we report the identification of an energetic epitope by determining the interfacial hot-spot that dominates the binding affinity for an anti-interleukin-23 (anti-IL-23) antibody by using the complementary approaches of hydrogen/deuterium exchange mass spectrometry (HDX-MS), fast photochemical oxidation of proteins (FPOP), alanine shave mutagenesis, and binding analytics. Five peptide regions on IL-23 with reduced backbone amide solvent accessibility upon antibody binding were identified by HDX-MS, and five different peptides over the same three regions were identified by FPOP. In addition, FPOP analysis at the residue level reveals potentially key interacting residues. Mutants with 3-5 residues changed to alanine have no measurable differences from wild-type IL-23 except for binding of and signaling blockade by the 7B7 anti-IL-23 antibody. The M5 IL-23 mutant differs from wild-type by five alanine substitutions and represents the dominant energetic epitope of 7B7. M5 shows a dramatic decrease in binding to BMS-986010 (which contains the 7B7 Fab, where Fab is fragment antigen-binding region of an antibody), yet it maintains functional activity, binding to p40 and p19 specific reagents, and maintains biophysical properties similar to wild-type IL-23 (monomeric state, thermal stability, and secondary structural features).

2016 AAHA/IAAHPC End-of-Life Care Guidelines.

End-of-life (EOL) care and decisionmaking embody the critical final stage in a pet's life and are as important and meaningful as the sum of the clinical care provided for all prior life stages. EOL care should focus on maximizing patient comfort and minimizing suffering while providing a collaborative and supportive partnership with the caregiver client. Timely, empathetic, and nonjudgmental communication is the hallmark of effective client support. Veterinarians should not allow an EOL patient to succumb to a natural death without considering the option of euthanasia and ensuring that other measures to alleviate discomfort and distress are in place. Animal hospice care addresses the patient's unique emotional and social needs as well as the physical needs traditionally treated in clinical practice. An EOL treatment plan should consist of client education; evaluating the caregiver's needs and goals for the pet; and a collaborative, personalized, written treatment plan involving the clinical staff and client. Primary care practices should have a dedicated team to implement palliative and hospice care for EOL patients. How the healthcare team responds to a client's grief after the loss of a pet can be a key factor in the client's continued loyalty to the practice. Referral to professional grief-support counseling can be a helpful option in this regard.

Quantification of in vivo site-specific Asp isomerization and Asn deamidation of mAbs in animal serum using IP-LC-MS.

BACKGROUND: Isomerization of aspartic acid and deamidation of asparagine are two common amino acid modifications that are of particular concern if located within the complementarity-determining region of therapeutic antibodies. Questions arise as to the extent of modification occurring in circulation due to potential exposure of the therapeutic antibody to different pH regimes. RESULTS: To enable evaluation of site-specific isomerization and deamidation of human mAbs in vivo, immunoprecipitation (IP) has been combined with LC-MS providing selective enrichment, separation and detection of naive and modified forms of tryptic peptides comprising complementarity-determining region sequences. CONCLUSION: IP-LC-MS can be applied to simultaneously quantify in vivo drug concentrations and measure the extent of isomerization or deamidation in PK studies conducted during the drug discovery stage.

Efficacy of a single dose of trazodone hydrochloride given to cats prior to veterinary visits to reduce signs of transport- and examination-related anxiety.

OBJECTIVE To evaluate the efficacy of a single dose of trazodone for reducing anxiety in cats during transport to a veterinary hospital and facilitating handling during veterinary examination. DESIGN Double-blind, placebo-controlled, randomized crossover study. ANIMALS 10 healthy client-owned cats (2 to 12 years of age) with a history of anxiety during transport or veterinary examination. PROCEDURES Each cat was randomly assigned to first receive trazodone hydrochloride (50 mg) or a placebo PO. The assigned treatment was administered, and each cat was placed in a carrier and transported by car to a veterinary clinic, where it received a structured veterinary examination. Owners scored their cat's signs of anxiety before, during, and after transport and examination. The veterinarian also assessed signs of anxiety during examination. After a 1- to 3-week washout period, each cat received the opposite treatment and the protocol was repeated. RESULTS Compared with placebo, trazodone resulted in a significant improvement in the cats' signs of anxiety during transport. Veterinarian and owner scores for ease of handling during veterinary examination also improved with trazodone versus the placebo. No significant differences were identified between treatments in heart rate or other physiologic variables. The most common adverse event related to trazodone administration was signs of sleepiness. CONCLUSIONS AND CLINICAL RELEVANCE Oral administration of a single dose of trazodone to cats prior to a veterinary visit resulted in fewer signs of transport- and examination-related anxiety than did a placebo and was generally well tolerated by most cats. Use of trazodone in this manner may promote veterinary visits and, consequently, enhance cat welfare.

Training veterinary students in shelter medicine: a service-learning community-classroom technique.

Shelter medicine is a rapidly developing field of great importance, and shelters themselves provide abundant training opportunities for veterinary medical students. Students trained in shelter medicine have opportunities to practice zoonotic and species-specific infectious disease control, behavioral evaluation and management, primary care, animal welfare, ethics, and public policy issues. A range of sheltering systems now exists, from brick-and-mortar facilities to networks of foster homes with no centralized facility. Exposure to a single shelter setting may not allow students to understand the full range of sheltering systems that exist; a community-classroom approach introduces students to a diverse array of sheltering systems while providing practical experience. This article presents the details and results of a series of 2-week elective clinical rotations with a focus on field and service learning in animal shelters. The overall aim was to provide opportunities that familiarized students with sheltering systems and delivered primary-care training. Other priorities included increasing awareness of public health concerns and equipping students to evaluate shelters on design, operating protocols, infectious disease control, animal enrichment, and community outreach. Students were required to participate in rounds and complete a project that addressed a need recognized by them during the rotation. This article includes costs associated with the rotation, a blueprint for how the rotation was carried out at our institution, and details of shelters visited and animals treated, including a breakdown of treatments provided. Also discussed are the student projects and student feedback on this valuable clinical experience.

Generation and characterization of human anti-human IL-21 neutralizing monoclonal antibodies.

Interleukin-21 (IL-21) is a type I four-helical bundle cytokine that exerts a variety of significant effects on many hematopoietic cells, including T and B lymphocytes and natural killer cells. IL-21 is produced predominantly by CD4+ T cells and natural killer T cells and, when aberrantly overexpressed, appears to play important roles in a wide variety of autoimmune disorders. To generate potential therapeutic reagents capable of inhibiting IL-21 for clinical use, we immunized human immunoglobulin transgenic mice with IL-21 and then identified and cloned a panel of human anti-human IL-21 binding monoclonal antibodies. IL-21 neutralizing and IL-21-binding, non-neutralizing antibodies were assigned to distinct epitope "bins" based on surface plasmon resonance competition studies. The most potent neutralizing antibodies had extremely high (sub pM) affinity for IL-21 and were able to block IL-21 activity in various biological assays using either an IL-21R-transfected pre-B-cell line or primary human B cells, and their neutralizing activity was, in some cases, superior to that of a soluble form of the high affinity heterodimeric IL-21 receptor. Characterization of this panel of IL-21 antibodies provided the basis for the selection of a therapeutic candidate antibody capable of inhibiting IL-21 activity for the treatment of autoimmune and inflammatory diseases.

A dual-targeting PDGFRbeta/VEGF-A molecule assembled from stable antibody fragments demonstrates anti-angiogenic activity in vitro and in vivo.

Targeting angiogenesis is a promising approach to the treatment of solid tumors and age-related macular degeneration (AMD). Inhibition of vascularization has been validated by the successful marketing of monoclonal antibodies (mAbs) that target specific growth factors or their receptors, but there is considerable room for improvement in existing therapies. Combination of mAbs targeting both the VEGF and PDGF pathways has the potential to increase the efficacy of anti-angiogenic therapy without the accompanying toxicities of tyrosine kinase inhibitors and the inability to combine efficiently with traditional chemotherapeutics. However, development costs and regulatory issues have limited the use of combinatorial approaches for the generation of more efficacious treatments. The concept of mediating disease pathology by targeting two antigens with one therapeutic was proposed over two decades ago. While mAbs are particularly suitable candidates for a dual-targeting approach, engineering bispecificity into one molecule can be difficult due to issues with expression and stability, which play a significant role in manufacturability. Here, we address these issues upstream in the process of developing a bispecific antibody (bsAb). Single-chain antibody fragments (scFvs) targeting PDGFRbeta and VEGF-A were selected for superior stability. The scFvs were fused to both termini of human Fc to generate a bispecific, tetravalent molecule. The resulting molecule displays potent activity, binds both targets simultaneously, and is stable in serum. The assembly of a bsAb using stable monomeric units allowed development of an anti-PDGFRB/VEGF-A antibody capable of attenuating angiogenesis through two distinct pathways and represents an efficient method for rapid engineering of dual-targeting molecules.

Engineering of stable bispecific antibodies targeting IL-17A and IL-23.

Bispecific antibodies (bsAbs) present an attractive opportunity to combine the additive and potentially synergistic effects exhibited by combinations of monoclonal antibodies (mAbs). Current challenges for engineering bsAbs include retention of the binding affinity of the parent mAb or antibody fragment, the ability to bind both targets simultaneously, and matching valency with biology. Other factors to consider include structural stability and expression of the recombinant molecule, both of which may have significant impact on its development as a therapeutic. Here, we incorporate selection of stable, potent single-chain variable fragments (scFvs) early in the engineering process to assemble bsAbs for therapeutic applications targeting the cytokines IL-17A/A and IL-23. Stable scFvs directed against human cytokines IL-23p19 and IL-17A/A were isolated from a human Fab phage display library via batch conversion of panning output from Fabs to scFvs. This strategy integrated a step for shuffling V regions during the conversion and permitted the rescue of scFv molecules in both the V(H)V(L) and the V(L)V(H) orientations. Stable scFvs were identified and assembled into several bispecific formats as fusions to the Fc domain of human IgG1. The engineered bsAbs are potent neutralizers of the biological activity of both cytokines (IC(50) < 1 nM), demonstrate the ability to bind both target ligands simultaneously and display stability and productivity advantageous for successful manufacture of a therapeutic molecule. Pharmacokinetic analysis of the bsAbs in mice revealed serum half-lives similar to human mAbs. Assembly of bispecific molecules using stable antibody fragments offers an alternative to reformatting mAbs and minimizes subsequent structure-related and manufacturing concerns.