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Covalent adaptable networks (CANs) are polymeric networks with cross-links that can break and reform in response to external stimuli, including pH, shear, and temperature, making them potential materials for use as injectable cell delivery vehicles. In the native niche, cells rearrange the extracellular matrix (ECM) to undergo basic functions including migration, spreading, and proliferation. Bond rearrangement enables these hydrogels to mimic viscoelastic properties of the native ECM which promote migration and delivery from the material to the native tissue. In this work, we characterize thioester CANs to inform their design as effective cell delivery vehicles. Using bulk rheology, we characterize the rearrangement of these networks when they are subjected to strain, which mimics the strain applied by a syringe, and using multiple particle tracking microrheology (MPT) we measure cell-mediated remodeling of the pericellular region. Thioester networks are formed by photopolymerizing 8-arm poly(ethylene glycol) (PEG)-thiol and PEG-thioester norbornene. Bulk rheology measures scaffold properties during low and high strain and demonstrates that thioester scaffolds can recover rheological properties after high strain is applied. We then 3D encapsulated human mesenchymal stem cells (hMSCs) in thioester scaffolds. Using MPT, we characterize degradation in the pericellular region. Encapsulated hMSCs degrade these scaffolds within ≈4 days post-encapsulation. We hypothesize that this degradation is mainly due to cytoskeletal tension that cells apply to the matrix, causing adaptable thioester bonds to rearrange, leading to degradation. To verify this, we inhibited cytoskeletal tension using blebbistatin, a myosin-II inhibitor. Blebbistatin-treated cells can degrade these networks only by secreting enzymes including esterases. Esterases hydrolyze thioester bonds, which generate free thiols, leading to bond exchange. Around treated cells, we measure a decrease in the extent of pericellular degradation. We also compare cell area, eccentricity, and speed of untreated and treated cells. Inhibiting cytoskeletal tension results in significantly smaller cell area, more rounded cells, and lower cell speeds when compared to untreated cells. Overall, this work shows that cytoskeletal tension plays a major role in hMSC-mediated degradation of thioester networks. Cytoskeletal tension is also important for the spreading and motility of hMSCs in these networks. This work informs the design of thioester scaffolds for tissue regeneration and cell delivery.
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BACKGROUND: There has been a proliferation of physicians of different levels of experience and training offering nonsurgical cosmetic procedures. Rising demand, compounded by increasing utilization of new and existing technologies by numerous physician specialties, compels discussion of adequate standardized training and patient safety. METHODS: A retrospective chart review of patients who presented to our single site dermatology clinic for managment of complications following chemical peel, laser or energy-based device treatments performed by core cosmetic physicians between the years of 2013 and 2024 was conducted. Core cosmetic physicians included plastic surgery, facial surgery/otolaryngology, oculoplastic surgery, and dermatology. Charts were reviewed for documentation of the type of complication, procedure causing the complication, and physician credentials, and referral source. RESULTS: Twenty-five patients were identified as having complications from chemical peeling, laser treatment or energy-based devices. Devices implicated included CO2 laser (fractional or fully ablative), chemical peels, 1064 nm long-pulsed Nd:YAG laser, 1320 nm Nd:YAG laser, intense pulsed light, 595 nm pulsed dye laser, Q-switched Nd:YAG laser, radiofrequency with and without microneedling, and 1550 nm erbium-doped fiber laser. Complications included hypertrophic scarring, atrophic scarring, post-inflammatory erythema, post-inflammatory hyperpigmentation, and post-inflammatory hypopigmentation. CONCLUSIONS: Even in experienced hands, complications can arise. It is imperative that all physicians offering cosmetic treatments are equipped to recognize clinical endpoints, identify and manage complications, or make a timely referral to decrease the risk of a permanent and potentially devastating esthetic outcome for patients.
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Abrasão Química , Técnicas Cosméticas , Humanos , Estudos Retrospectivos , Abrasão Química/efeitos adversos , Técnicas Cosméticas/instrumentação , Técnicas Cosméticas/efeitos adversos , Feminino , Masculino , Terapia a Laser/efeitos adversos , Terapia a Laser/instrumentação , Pessoa de Meia-Idade , Adulto , Complicações Pós-Operatórias/etiologia , Lasers de Estado Sólido/uso terapêuticoRESUMO
Polymer-peptide hydrogels are being designed as implantable materials that deliver human mesenchymal stem cells (hMSCs) to treat wounds. Most wounds can progress through the healing process without intervention. During the normal healing process, cytokines are released from the wound to create a concentration gradient, which causes directed cell migration from the native niche to the wound site. Our work takes inspiration from this process and uniformly tethers cytokines into the scaffold to measure changes in cell-mediated degradation and motility. This is the first step in designing cytokine concentration gradients into the material to direct cell migration. We measure changes in rheological properties, encapsulated cell-mediated pericellular degradation and migration in a hydrogel scaffold with covalently tethered cytokines, either tumor necrosis factor-α (TNF-α) or transforming growth factor-ß (TGF-ß). TNF-α is expressed in early stages of wound healing causing an inflammatory response. TGF-ß is released in later stages of wound healing causing an anti-inflammatory response in the surrounding tissue. Both cytokines cause directed cell migration. We measure no statistically significant difference in modulus or the critical relaxation exponent when tethering either cytokine in the polymeric network without encapsulated hMSCs. This indicates that the scaffold structure and rheology is unchanged by the addition of tethered cytokines. Increases in hMSC motility, morphology and cell-mediated degradation are measured using a combination of multiple particle tracking microrheology (MPT) and live-cell imaging in hydrogels with tethered cytokines. We measure that tethering TNF-α into the hydrogel increases cellular remodeling on earlier days postencapsulation and tethering TGF-ß into the scaffold increases cellular remodeling on later days. We measure tethering either TGF-ß or TNF-α enhances cell stretching and, subsequently, migration. This work provides rheological characterization that can be used to design new materials that present chemical cues in the pericellular region to direct cell migration.
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Movimento Celular , Hidrogéis , Células-Tronco Mesenquimais , Reologia , Fator de Necrose Tumoral alfa , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Hidrogéis/química , Fator de Necrose Tumoral alfa/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta/química , Citocinas/metabolismo , Cicatrização , Células CultivadasRESUMO
Background: Gestational diabetes mellitus (GDM) complicates â¼10% of pregnancies, with the highest rates among Asian women. Evidence suggests that GDM is associated with an increased risk for future chronic health conditions, yet data for Asian women are sparse. We explored the association between prior GDM and metabolic dysfunction with nationally representative data to obtain Asian-specific estimates. Methods: For this cross-sectional study, data were drawn from the National Health and Nutrition Examination Survey for 7195 women with a prior pregnancy. GDM (yes/no) was defined using the question "During pregnancy, were you ever told by a doctor or other health professional that you had diabetes, sugar diabetes, or gestational diabetes?." Current metabolic dysfunction (yes/no) was based on having at least one of four indicators: systolic blood pressure (SBP, ≥130 mmHg), waist circumference (≥88 cm), high-density lipoprotein (HDL) cholesterol (<50 mg/dL), and glycosylated hemoglobin (HbA1c) (≥6.5%). Logistic regression estimated odds ratios (ORs) and 95% confidence intervals (CIs) for the association between prior GDM and metabolic outcomes, overall and by race. Models included sampling weights and demographic and behavioral factors. Results: Overall, women with prior GDM had 46% greater odds of high waist circumference (OR: 1.5; 95% CI: 1.1-2.0) and 200% greater odds (OR: 3.0; 95% CI: 2.1-4.2) of high HbA1c. Prior GDM was not associated with high blood pressure or low HDL cholesterol. In race-specific analyses, prior GDM was associated with increased risk of elevated HbA1c among Asian (OR: 6.6; 95% CI: 2.5-17.2), Mexican American (OR: 3.0; 95% CI: 1.5-5.8), Black (OR: 3.0; 95% CI: 1.7-5.5), and White (OR: 2.6; 95% CI: 1.5-4.6) women. Prior GDM was associated with elevated SBP among Mexican American women and low HDL among Black women. Discussion: Prior GDM is associated with elevated HbA1c among all women, yet is a stronger predictor of elevated HbA1c among Asian women than other women. Race-specific associations between prior GDM and metabolic dysfunction were observed among Mexican American and Black women. Further research is warranted to understand the observed race/ethnic-specific associations.
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Diabetes Gestacional , Inquéritos Nutricionais , Humanos , Feminino , Diabetes Gestacional/epidemiologia , Diabetes Gestacional/sangue , Diabetes Gestacional/etnologia , Gravidez , Adulto , Estudos Transversais , Estados Unidos/epidemiologia , Adulto Jovem , Hemoglobinas Glicadas/metabolismo , Hemoglobinas Glicadas/análise , Fatores de Risco , Doenças Metabólicas/epidemiologia , Doenças Metabólicas/diagnóstico , Pressão Sanguínea , Circunferência da Cintura , Pessoa de Meia-Idade , HDL-Colesterol/sangueRESUMO
Covalent adaptable networks are designed for applications including cell and drug delivery and tissue regeneration. These applications require network degradation at physiological conditions and on a physiological timescale with microstructures that can: (1) support, protect and deliver encapsulated cells or molecules and (2) provide structure to surrounding tissue. Due to this, the evolving microstructure and rheological properties during scaffold degradation must be characterized. In this work, we characterize degradation of covalent adaptable poly(ethylene glycol) (PEG)-thioester networks with different amounts of excess thiol. Networks are formed between PEG-thiol and PEG-thioester norbornene using photopolymerization. These networks are adaptable because of a thioester exchange reaction that takes place in the presence of excess thiol. We measure degradation of PEG-thioester networks with L-cysteine using multiple particle tracking microrheology (MPT). MPT measures the Brownian motion of fluorescent probe particles embedded in a material and relates this motion to rheological properties. Using time-cure superposition (TCS), we characterize the microstructure of these networks at the gel-sol phase transition by calculating the critical relaxation exponent, n, for each network with different amounts of excess thiol. Based on the measured n values, networks formed with 0% and 50% excess thiol are tightly cross-linked and elastic in nature. While networks formed with 100% excess are similar to ideal, percolated networks, which have equal viscous and elastic components. MPT measurements during degradation of these networks also measure a non-monotonic increase in probe motility. We hypothesize that this is network rearrangement near the phase transition. We then measure macroscopic material properties including the equilibrium modulus and stress relaxation. We measure a trend in bulk network properties that agrees with the values of n. Elastic modulus and stress relaxation measurements show that networks with 50% excess thiol are more elastic compared to the other two networks. As the amount of excess thiol is increased from 0% to 50%, the networks become more elastic. Further increasing excess thiol to 100% reduces the elastically effective cross-links. We hypothesize that these properties are due to network non-idealities, resulting in networks with 50% excess thiol that are more elastic. This work characterizes dynamic rheological properties during degradation, which mimics processes that could occur during implantation. This work provides information that can be used in the future design of implantable materials enabling both the rheological properties and timescale of degradation to be specified.
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Rheological modifiers are used to tune rheology or induce phase transitions of products. Microfibrillated cellulose (MFC), a renewable material, has the potential to be used for rheological modification. However, the lack of studies on the evolution in rheological properties and structure during its phase transitions has prevented MFC from being added to consumer, fabric, and home care products. In this work, we characterize surface-oxidized MFC (OMFC), a negatively charged colloidal rod suspension. We measure the rheological properties and structure of OMFC during sol-gel phase transitions induced by either anionic or cationic surfactant using multiple particle tracking microrheology (MPT). MPT tracks the Brownian motion of fluorescent probe particles embedded in a sample, which is related to the sample's rheological properties. Using MPT, we measure that OMFC gelation evolution is dependent on the charge of the surfactant that induces the phase transition. OMFC gelation is gradual in anionic surfactant. In cationic surfactant, gelation is rapid followed by length scale-dependent colloidal fiber rearrangement. Initial OMFC concentration is directly related to how tightly associated the network is at the phase transition, with an increase in concentration resulting in a more tightly associated network with smaller pores. Bulk rheology measures that OMFC forms a stiffer structure but yields at lower strains in cationic surfactant than in anionic surfactant. This study characterizes the role of surfactant in inducing phase transitions, which can be used as a guide for designing future products.
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OBJECTIVE: Evaluate whether general practitioners' formal small animal (canine and feline) nutrition instruction in veterinary school and the amount and type of continuing education engagement affect perceived self-reported confidence and frequency in discussing nutrition with clients. SAMPLE: 403 small animal veterinarians who responded to an online survey distributed through the American Animal Hospital Association. PROCEDURES: Veterinarians were surveyed regarding perceived amount of formal instruction received in veterinary school, interest, time committed to self-education, and confidence in both self and staff knowledge in small animal nutrition. RESULTS: Of those veterinarians who responded to the survey, 57.1% (201/352) reported they received "none" or "very little" formal instruction in small animal nutrition, while 151 of 352 answered "some" or "a significant amount." Veterinarians with more formal instruction and veterinarians who reported spending more time in self-education had increased confidence in their own nutritional knowledge (P < .01) and that of their staff (P < .01). CLINICAL RELEVANCE: Veterinarians with self-reported significant formal instruction and veterinarians with higher continuing education engagement were more confident in their knowledge and their staff's knowledge regarding therapeutic and nontherapeutic small animal nutrition. Therefore, it is important for the profession to address veterinary nutrition education gaps in order to increase the veterinary healthcare team's engagement in nutritional discussions with their clients for both healthy and sick pets.
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Doenças do Gato , Doenças do Cão , Clínicos Gerais , Médicos Veterinários , Animais , Gatos , Cães , Humanos , Inquéritos e Questionários , Educação ContinuadaRESUMO
Injectable poly(ethylene glycol) (PEG)-based hydrogels were reversibly cross-linked through thia-conjugate addition bonds and demonstrated to shear thicken at low shear rates. Cross-linking bond exchange kinetics and dilute polymer concentrations were leveraged to tune hydrogel plateau moduli (from 60 to 650 Pa) and relaxation times (from 2 to 8 s). Under continuous flow shear rheometry, these properties affected the onset of shear thickening and the degree of shear thickening achieved before a flow instability occurred. The changes in viscosity were reversible whether the shear rate increased or decreased, suggesting that chain stretching drives this behavior. Given the relevance of dynamic PEG hydrogels under shear to biomedical applications, their injectability was investigated. Injection forces were found to increase with higher polymer concentrations and slower bond exchange kinetics. Altogether, these results characterize the nonlinear rheology of dilute, dynamic covalent tetra-PEG hydrogels and offer insight into the mechanism driving their shear thickening behavior.
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OBJECTIVE: To evaluate the voluntary acceptance of 10 commercially available compounding flavors in cats. ANIMALS: 46 healthy cats between 1 and 12 years of age. PROCEDURES: Each cat underwent a 14-day study period consisting of a 4-day acclimation period followed by a 10-day trial period in which each cat was randomly offered 10 different compounding flavors. Owners completed a presurvey along with a daily observation logbook. Kits, including residual amounts of flavors, were returned and weighed to determine residual weight and calculate the amount ingested. RESULTS: Overall, cats did not voluntarily accept most of the compounding flavors; 58.8% (124/211) and 84.5% (267/311) of offered samples of oil-based and water-based compounding flavors, respectively, were rejected or minimally accepted. Cats were significantly (P < .001) more likely to accept oil-based flavors, compared to water-based flavors. The sweet water-based flavors were least accepted, compared to water-based control and water-based savory flavors (P = .040 and P < .001, respectively). Owner-perceived acceptance was moderately correlated with residual flavor weights (Kendall tau [τ] = -0.466; P < .001). Owners were not able to accurately predict which flavors their cats would accept. CLINICAL RELEVANCE: Cats should be offered oil-based compounding flavorings when available, whereas water-based sweet flavorings should be avoided. Owner perception of acceptance is a valid metric to assess flavor acceptance, which can be used in future studies evaluating flavor acceptance. Owners may not accurately predict their cats' flavor preferences, limiting their ability to guide optimal flavor selection.
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Paladar , Água , Gatos , AnimaisRESUMO
SUMMARY: An aesthetically pleasing umbilicus is a critical component to the overall cosmesis and resultant patient satisfaction after deep inferior epigastric artery perforator (DIEP) flap breast reconstruction. Because of variables in body habitus, comorbidities, and technical aspects of the procedure, patients undergoing DIEP flap breast reconstruction are at a higher risk of umbilical complications and poor aesthetic appearance of the neoumbilicus compared with those undergoing cosmetic abdominoplasty. To minimize these potential problems and maximize the overall aesthetic appearance of the abdomen, the authors propose an algorithmic approach to umbilical inset after DIEP flap harvest that takes into account several critical factors: the thickness of the subcutaneous tissue of the abdominal flap, the length of the umbilical stalk, and the depth of the umbilical bowl. This simple algorithmic approach is a useful tool that will assist surgeons in minimizing umbilical complications and delivering a superior cosmetic appearance to the abdominal donor site in DIEP flap reconstruction.
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Abdominoplastia , Mamoplastia , Retalho Perfurante , Humanos , Umbigo/cirurgia , Retalho Perfurante/irrigação sanguínea , Estudos Retrospectivos , Mamoplastia/métodos , Artérias Epigástricas/cirurgiaRESUMO
Human mesenchymal stem cells (hMSCs) are instrumental in the wound healing process. They migrate to wounds from their native niche in response to chemical signals released during the inflammatory phase of healing. At the wound, hMSCs downregulate inflammation and regulate tissue regeneration. Delivering additional hMSCs to wounds using cell-laden implantable hydrogels has the potential to improve healing outcomes and restart healing in chronic wounds. For these materials to be effective, cells must migrate from the scaffold into the native tissue. This requires cells to traverse a step-change in material properties at the implant-tissue interface. Migration of cells in material with highly varying properties is not well characterized. We measure 3D encapsulated hMSC migration and remodeling in a well-characterized hydrogel with a step-change in stiffness. This cell-degradable hydrogel is composed of 4-arm poly(ethylene glycol)-norbornene cross-linked with an enzymatically-degradable peptide. The scaffold is made with two halves of different stiffnesses separated by an interface where stiffness changes rapidly. We characterize changes in structure and rheology of the pericellular region using multiple particle tracking microrheology (MPT). MPT measures Brownian motion of embedded particles and relates it to material rheology. We measure more remodeling in the soft region of the hydrogel than the stiff region on day 1 post-encapsulation and similar remodeling everywhere on day 6. In the interface region, we measure hMSC-mediated remodeling along the interface and migration towards the stiff side of the scaffold. These results can improve materials designed for cell delivery from implants to a wound to enhance healing.
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Hidrogéis , Células-Tronco Mesenquimais , Módulo de Elasticidade , Humanos , Hidrogéis/química , Polietilenoglicóis/química , CicatrizaçãoRESUMO
OBJECTIVE: To determine what perceived factors prevent small animal general practitioners from discussing pet nutrition with clients during healthy and sick pet appointments. SAMPLE: 403 veterinarians in small animal general practice. PROCEDURES: An online survey was used to gather veterinarians' opinions on perceived barriers, knowledge levels, and confidence regarding pet nutrition discussions. RESULTS: Reported barriers to discussing nutrition during healthy pet appointments included client resistance to changing brand (149/359), time constraints (146/359), misinformation online (138/359), and difficulty keeping up with products (132/359). Reported barriers to discussing nutrition during sick pet appointments included client cost concerns (101/349), pet not accepting new food (99/349), and time constraints (83/349). Veterinarians reported discussing nutrition less during healthy pet appointments, compared to sick pet appointments, and were significantly less confident with their knowledge regarding nontherapeutic food, compared to therapeutic food. Veterinarians also reported that they perceived conversations about therapeutic foods to be more positive than conversations about nontherapeutic foods, and veterinarians with more years in practice more commonly reported that there was nothing that would dissuade them from discussing nutrition. Veterinarians who reported barriers to discussing nutrition described a need for resources and reliable information for health-care teams and clients. CLINICAL RELEVANCE: Results demonstrated a substantial gap between veterinarians' assertion that nutrition conversations are indicated and the frequency with which they discuss nutrition during appointments. Veterinarians reported that they felt their nutrition conversations were frequently positive; therefore, it is important to overcome barriers to engage with clients about pet nutrition.
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Clínicos Gerais , Médicos Veterinários , Animais , Humanos , Comunicação , Inquéritos e Questionários , Equipe de Assistência ao PacienteRESUMO
BACKGROUND: Radiofrequency treatment is a relatively new and increasingly popular option for patients desiring skin tightening and an improvement in facial or body contour without undergoing an excisional surgical procedure. A systematic review of the literature was performed to investigate the safety and efficacy of monopolar and bipolar radiofrequency devices for facial and body rejuvenation. METHODS: A computerized search of the MEDLINE database was performed for clinical studies investigating the use of monopolar and bipolar radiofrequency devices in facial and body rejuvenation. Data on the type of device, treated areas, number of patients, number of treatments, follow-up, complications, and outcomes were collected. RESULTS: The systematic review was performed in September of 2020. A total of 207 articles examined the use of radiofrequency technology for cosmetic purposes, and 23 articles remained after all inclusion and exclusion criteria were considered. Nine articles evaluated monopolar devices and five articles evaluated bipolar devices for treatment of the face. Three articles evaluated monopolar devices and six articles evaluated bipolar devices for treatment of various body areas. CONCLUSION: There is clinical evidence that monopolar and bipolar radiofrequency devices produce measurable improvement in skin laxity of the face and body with an acceptable complication profile. The majority of reported complications are minor and transient in nature; major complication rates are higher with the use of monopolar devices than with the use of bipolar devices.
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Técnicas Cosméticas , Envelhecimento da Pele , Humanos , Rejuvenescimento , PeleRESUMO
Rheological modifiers tune product rheology with a small amount of material. To effectively use rheological modifiers, characterizing the rheology of the system at different compositions is crucial. Two colloidal rod system, hydrogenated castor oil and polyamide, are characterized in a formulation that includes a surfactant (linear alkylbenzene sulfonate) and a depletant (polyethylene oxide). We characterize both rod systems using multiple particle tracking microrheology (MPT) and bulk rheology and build phase diagrams over a large component composition space. In MPT, fluorescent particles are embedded in the sample and their Brownian motion is measured and related to rheological properties. From MPT, we determine that in both systems: (1) microstructure is not changed with increasing colloid concentration, (2) materials undergo a sol-gel transition as depletant concentration increases and (3) the microstructure changes but does not undergo a phase transition as surfactant concentration increases in the absence of depletant. When comparing MPT and bulk rheology results different trends are measured. Using bulk rheology we observe: (1) elasticity of both systems increase as colloid concentration increases and (2) the storage modulus does not change when PEO or LAS concentration is increased. The differences measured with MPT and bulk rheology are likely due to differences in sensitivity and measurement method. This work shows the utility of using both techniques together to fully characterize rheological properties over a large composition space. These gelation phase diagrams will provide a guide to determine the composition needed for desired rheological properties and eliminate trial-and-error experiments during product formulation.
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This pilot survey study describes student expectations and experiences at WisCARES, a low-cost veterinary medical teaching clinic where students from multiple disciplines collaborate. We hypothesized that prior to the workday, students would describe different expectations of working in an interdisciplinary access to care clinic than what they ultimately experienced. We surveyed 62 students from the School of Veterinary Medicine (46) and pharmacy (16) who spent a clinic day at WisCARES. Before introductory rounds, students completed a short survey consisting of four open-ended questions about their learning expectations; at the end of the day, they reviewed their initial responses and added what they actually learned. Qualitative information was categorized and analyzed using descriptive statistics. Thirteen major themes emerged: diversity, confidence, communication, case lead/case management, financial experience, helping people, teamwork, technical skills, inter-professional experience, mentoring, non-specific positive regard, appreciation for resources, and rounds. Students reported improved confidence in managing and leading cases with specific positive outcomes in communicating with clients, particularly regarding leading financial conversations. Developing greater insight into diversity was a common theme expressed in students' expectations but was less frequently noted as an end-of-day outcome. Veterinary students less frequently described the value of the inter-professional environment and collaboration, but this was a major theme noted among pharmacy students. Student feedback was positive overall. The current study is useful in identifying areas for improving collaborative instruction and access to care professional student learning opportunities.
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Educação em Veterinária , Estudantes de Farmácia , Animais , Acessibilidade aos Serviços de Saúde , Hospitais Veterinários , Humanos , MotivaçãoRESUMO
Implantable hydrogels are designed to treat wounds by providing structure and delivering additional cells to damaged tissue. These materials must consider how aspects of the native wound, including environmental chemical cues, affect and instruct delivered cells. One cell type researchers are interested in delivering are human mesenchymal stem cells (hMSCs) due to their importance in healing. Wound healing involves recruiting and coordinating a variety of cells to resolve a wound. hMSCs coordinate the cellular response and are signaled to the wound by cytokines, including transforming growth factor-ß (TGF-ß) and tumor necrosis factor-α (TNF-α), present in vivo. These cytokines change hMSC secretions, regulating material remodeling. TGF-ß, present from inflammation through remodeling, directs hMSCs to reorganize collagen, increasing extracellular matrix (ECM) structure. TNF-α, present primarily during inflammation, cues hMSCs to clear debris and degrade ECM. Because cytokines change how hMSCs degrade their microenvironment and are naturally present in the wound, they also affect how hMSCs migrate out of the scaffold to conduct healing. Therefore, the effects of cytokines on hMSC remodeling are important when designing materials for cell delivery. In this work, we encapsulate hMSCs in a polymer-peptide hydrogel and incubate the scaffolds in media with TGF-ß or TNF-α at concentrations similar to those in wounds. Multiple particle tracking microrheology (MPT) measures hMSC-mediated scaffold degradation in response to these cytokines, which mimics aspects of the in vivo microenvironment post-implantation. MPT uses video microscopy to measure Brownian motion of particles in a material, quantifying structure and rheology. Using MPT, we measure increased hMSC-mediated remodeling when cells are exposed to TNF-α and decreased remodeling after exposure to TGF-ß when compared to untreated hMSCs. This agrees with previous studies that measure: (1) TNF-α encourages matrix reorganization and (2) TGF-ß signals the formation of new matrix. These results enable material design that anticipates changes in remodeling after implantation, improving control over hMSC delivery and healing.
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Células-Tronco Mesenquimais , Citocinas , Matriz Extracelular , Humanos , Hidrogéis , ReologiaRESUMO
PURPOSE: Measurement of the viscosity of concentrated protein solutions is vital for the manufacture and delivery of protein therapeutics. Conventional methods for viscosity measurements require large solution volumes, creating a severe limitation during the early stage of protein development. The goal of this work is to develop a robust technique that requires minimal sample. METHODS: In this work, a droplet-based microfluidic device is developed to quantify the viscosity of protein solutions while concentrating in micrometer-scale droplets. The technique requires only microliters of sample. The corresponding viscosity is characterized by multiple particle tracking microrheology (MPT). RESULTS: We show that the viscosities quantified in the microfluidic device are consistent with macroscopic results measured by a conventional rheometer for poly(ethylene) glycol (PEG) solutions. The technique was further applied to quantify viscosities of well-studied lysozyme and bovine serum albumin (BSA) solutions. Comparison to both macroscopic measurements and models (Krieger-Dougherty model) demonstrate the validity of the approach. CONCLUSION: The droplet-based microfluidic device provides accurate quantitative values of viscosity over a range of concentrations for protein solutions with small sample volumes (~ µL) and high compositional resolution. This device will be extended to study the effect of different excipients and other additives on the viscosity of protein solutions.
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Excipientes/química , Microfluídica/instrumentação , Muramidase/química , Soroalbumina Bovina/química , Polietilenoglicóis/química , Reologia , Imagem Individual de Molécula , Soluções , ViscosidadeRESUMO
In this work, we establish a quantitative correlation between molecular release and material degradation. We characterize a radical-initiated photopolymerized hydrogel and base-initiated Michael addition-polymerized hydrogel, which form gels through distinct crosslinking reactions. Both scaffolds use the same degradable peptide crosslinker, which enables them to be degraded through the same enzymatic degradation reaction. A fluorescently labeled poly(ethylene glycol) molecule is chemically conjugated into the scaffold and is released during enzymatic degradation. Real-time changes in scaffold rheological properties during degradation are measured using bulk rheology. Molecular release is measured by quantifying the change in fluorescence in the incubation liquid and the hydrogel scaffold. A complicating factor, previously described in the literature, is that shear may cause increased crosslinking, resulting in an increase in the storage modulus after initiation of degradation, which changes release profiles by limiting the initial release of molecules. Therefore, we also test the hypothesis that shear induces additional crosslinking in degrading hydrogel scaffolds. To determine whether shear changes rheological properties during scaffold degradation, enzymatic degradation is characterized using bulk rheology as materials undergo continuous or minimal shear. To determine the effect of shear on molecular release, shear is induced by shaking the material during incubation. Release is characterized from scaffolds that are incubated with continuous or without shaking. We determine that shear does not make a difference in scaffold degradation or release regardless of the gelation reaction. Instead, we determine that the type of hydrogel crosslinking reaction greatly affects both material degradation and molecular release. A hydrogel crosslinking by base-initiated Michael addition does undergo further crosslinking at the start of degradation. We correlate release with enzymatic degradation for both scaffolds. We determine that the material storage modulus is indirectly correlated with release during degradation. These results indicate that rheological characterization is a useful tool to characterize and predict the release of molecules from degrading hydrogels.