The First Hybrid International Educational Comprehensive Cleft Care Workshop.

OBJECTIVE: Describe the first hybrid global simulation-based comprehensive cleft care workshop, evaluate impact on participants, and compare experiences based on in-person versus virtual attendance. DESIGN: Cross-sectional survey-based evaluation. SETTING: International comprehensive cleft care workshop. PARTICIPANTS: Total of 489 participants. INTERVENTIONS: Three-day simulation-based hybrid comprehensive cleft care workshop. MAIN OUTCOME MEASURES: Participant demographic data, perceived barriers and interventions needed for global comprehensive cleft care delivery, participant workshop satisfaction, and perceived short-term impact on practice stratified by in-person versus virtual attendance. RESULTS: The workshop included 489 participants from 5 continents. The response rate was 39.9%. Participants perceived financial factors (30.3%) the most significant barrier and improvement in training (39.8%) as the most important intervention to overcome barriers facing cleft care delivery in low to middle-income countries. All participants reported a high level of satisfaction with the workshop and a strong positive perceived short-term impact on their practice. Importantly, while this was true for both in-person and virtual attendees, in-person attendees reported a significantly higher satisfaction with the workshop (28.63 ± 3.08 vs 27.63 ± 3.93; P = .04) and perceived impact on their clinical practice (22.37 ± 3.42 vs 21.02 ± 3.45 P = .01). CONCLUSION: Hybrid simulation-based educational comprehensive cleft care workshops are overall well received by participants and have a positive perceived impact on their clinical practices. In-person attendance is associated with significantly higher satisfaction and perceived impact on practice. Considering that financial and health constraints may limit live meeting attendance, future efforts will focus on making in-person and virtual attendance more comparable.

Three-Dimensional High-Throughput Cell Encapsulation Platform to Study Changes in Cell-Matrix Interactions.

In their native extracellular microenvironment, cells respond to a complex array of biochemical and mechanical cues that can vary in both time and space. High-throughput methods that allow characterization of cell-laden matrices are valuable tools to screen through many combinations of variables, ultimately helping to evolve and test hypotheses related to cell-ECM signaling. Here, we developed a platform for high-throughput encapsulation of cells in peptide-functionalized poly(ethylene glycol) hydrogels. Hydrogels were synthesized using a thiol-ene, photoclick reaction, which allowed the cell matrix environment to be modified in real time. Matrix signals were dynamically altered by in situ tethering of RGDS (0-1.5 mM), a fibronectin-derived adhesive peptide that induced more elongation than RLD or IKVAV, and/or by increasing the matrix modulus (1 to 6 kPa). This method was demonstrated with aortic valvular interstitial cells (VICs), a population of cells responsible for the pathological fibrosis and matrix remodeling that leads to aortic stenosis. VIC response to cell-matrix interactions was characterized by quantifying cell morphology and the fraction of cells exhibiting α-smooth muscle actin (αSMA) stress fibers, a hallmark of the myofibroblast phenotype. VICs elongated in response to RGDS addition, with a dramatic change in morphology within 24 h. Myofibroblast activation was also dependent on RGDS addition, with VICs exhibiting high activation (16-24%) in 1 kPa gels with RGDS. Response to RGDS was path-dependent, with the amount of time exposed to the adhesive ligand important in determining VIC morphology and activation. Although VIC aspect ratios were dependent on the amount of time spent in a stiff vs soft gel, low levels of VIC activation (≤4%) were observed in any gels cultured in higher modulus (6 kPa vs 1 kPa) microenvironments.

Microarray analyses to quantify advantages of 2D and 3D hydrogel culture systems in maintaining the native valvular interstitial cell phenotype.

Valvular interstitial cells (VICs) actively maintain and repair heart valve tissue; however, persistent activation of VICs to a myofibroblast phenotype can lead to aortic stenosis. To better understand and quantify how microenvironmental cues influence VIC phenotype and myofibroblast activation, we compared expression profiles of VICs cultured on poly(ethylene glycol) (PEG) gels to those cultured on tissue culture polystyrene (TCPS), as well as fresh isolates. In general, VICs cultured in hydrogel matrices had lower levels of activation (<10%), similar to levels seen in healthy valve tissue, while VICs cultured on TCPS were ∼75% activated myofibroblasts. VICs cultured on TCPS also exhibited a higher magnitude of perturbations in gene expression than soft hydrogel cultures when compared to the native phenotype. Using peptide-modified PEG gels, VICs were seeded on (2D), as well as encapsulated in (3D), matrices of the same composition and modulus. Despite similar levels of activation, VICs cultured in 2D had distinct variations in transcriptional profiles compared to those in 3D hydrogels. Genes related to cell structure and motility were particularly affected by the dimensionality of the culture platform, with higher expression levels in 2D than in 3D. These results indicate that dimensionality may play a significant role in dictating cell phenotype (e.g., through differences in polarity, diffusion of soluble signals), and emphasize the importance of using multiple metrics when characterizing cell phenotype.

Transcriptional profiles of valvular interstitial cells cultured on tissue culture polystyrene, on 2D hydrogels, or within 3D hydrogels.

Valvular interstitial cells (VICs) actively maintain and repair heart valve tissue; however, persistent activation of VICs to a myofibroblast phenotype can lead to aortic stenosis (Chen and Simmons, 2011) [1]. To better understand and quantify how microenvironmental cues influence VIC phenotype, we compared expression profiles of VICs cultured on/in poly(ethylene glycol) (PEG) gels to those cultured on tissue culture polystyrene (TCPS), as well as fresh isolates. Here, we present both the raw and processed microarray data from these culture conditions. Interpretation of this data can be found in a research article entitled "Microarray analyses to quantify advantages of 2D and 3D hydrogel culture systems in maintaining the native valvular interstitial cell phenotype" (Mabry et al., 2015) [2].

Dynamic stiffening of poly(ethylene glycol)-based hydrogels to direct valvular interstitial cell phenotype in a three-dimensional environment.

Valvular interstitial cells (VICs) are active regulators of valve homeostasis and disease, responsible for secreting and remodeling the valve tissue matrix. As a result of VIC activity, the valve modulus can substantially change during development, injury and repair, and disease progression. While two-dimensional biomaterial substrates have been used to study mechanosensing and its influence on VIC phenotype, less is known about how these cells respond to matrix modulus in a three-dimensional environment. Here, we synthesized MMP-degradable poly(ethylene glycol) (PEG) hydrogels with elastic moduli ranging from 0.24 kPa to 12 kPa and observed that cell morphology was constrained in stiffer gels. To vary gel stiffness without substantially changing cell morphology, cell-laden hydrogels were cultured in the 0.24 kPa gels for 3 days to allow VIC spreading, and then stiffened in situ via a second, photoinitiated thiol-ene polymerization such that the gel modulus increased from 0.24 kPa to 1.2 kPa or 13 kPa. VICs encapsulated within soft gels exhibited αSMA stress fibers (∼ 40%), a hallmark of the myofibroblast phenotype. Interestingly, in stiffened gels, VICs became deactivated to a quiescent fibroblast phenotype, suggesting that matrix stiffness directs VIC phenotype independent of morphology, but in a manner that depends on the dimensionality of the culture platform. Collectively, these studies present a versatile method for dynamic stiffening of hydrogels and demonstrate the significant effects of matrix modulus on VIC myofibroblast properties in three-dimensional environments.

Photoresponsive elastic properties of azobenzene-containing poly(ethylene-glycol)-based hydrogels.

The elastic modulus of the extracellular matrix is a dynamic property that changes during various biological processes, such as disease progression or wound healing. Most cell culture platforms, however, have traditionally exhibited static properties, making it necessary to replate cells to study the effects of different elastic moduli on cell phenotype. Recently, much progress has been made in the development of substrates with mechanisms for either increasing or decreasing stiffness in situ, but there are fewer examples of substrates that can both stiffen and soften, which may be important for simulating the effects of repeated ECM injury and resolution. In the work presented here, poly(ethylene glycol)-based hydrogels reversibly stiffen and soften with multiple light stimuli via photoisomerization of an azobenzene-containing cross-linker. Upon irradiation with cytocompatible doses of 365 nm light (10 mW/cm(2), 5 min), isomerization to the azobenzene cis configuration leads to a softening of the hydrogel up to 100-200 Pa (shear storage modulus, G'). This change in gel properties is maintained over a time scale of several hours due to the long half-life of the cis isomer. The initial modulus of the gel can be recovered upon irradiation with similar doses of visible light. With applications in mechanobiology in mind, cytocompatibility with a mechanoresponsive primary cell type is demonstrated. Porcine aortic valvular interstitial cells were encapsulated in the developed hydrogels and shown to exhibit high levels of survival, as well as a spread morphology. The developed hydrogels enable a route to the noninvasive control of substrate modulus independent of changes in the chemical composition or network connectivity, allowing for investigations of the effect of dynamic matrix stiffness on adhered cell behavior.

Clinical outcomes of the Furlow Z-plasty for primary cleft palate repair.

OBJECTIVE: To review the clinical outcomes following the Furlow Z-plasty for primary cleft palate repair. The primary objective was to determine if the presence of an associated sequence or syndrome (i.e., Pierre Robin sequence), age at palate repair, cleft type, or surgeon experience influenced speech outcomes after a Furlow Z-plasty. DESIGN: The outcomes of 140 patients who underwent palate repair were analyzed retrospectively. Speech evaluations were performed to score the severity of hypernasality, nasal escape, articulation errors, and velopharyngeal insufficiency. RESULTS: The mean age at latest evaluation was 4 years 9 months (age range 2 years old to 12 years old and 4 months). Of the 140 patients, 83% had no evidence of hypernasality, 91% had no presence of nasal escape, and 69% had no articulation errors. Overall, 84% of patients had no evidence of velopharyngeal insufficiency. Secondary posterior pharyngeal flap to correct velopharyngeal insufficiency was required in only 2.1% of patients. The formation of an oronasal fistula occurred in only 3.6% of patients. Nonsyndromic patients with Pierre Robin sequence and syndromic patients did just as well as purely nonsyndromic patients in terms of velopharyngeal insufficiency, hypernasality, and nasal escape. Syndromic patients were more likely to make mild-to-moderate articulation errors. In addition, age at palate repair, cleft type, and surgeon experience had no statistically significant effect on speech results. CONCLUSIONS: The Furlow Z-plasty yielded excellent speech results in our patient population with minimal and acceptable rates of fistula formation, velopharyngeal insufficiency, and the need for additional corrective surgery.

Midmaxillary internal distraction osteogenesis: ideal surgery for the mature cleft patient.

BACKGROUND: Distraction osteogenesis is a recent technique often used for maxillary advancement to correct skeletofacial deformities in cleft patients. Conventional orthognathics falls short of achieving adequate stable results. Osteodistraction with Le Fort I osteotomy may create velopharyngeal incompetence. The authors propose a new technique for distracting only the anterior maxilla to improve aesthetics and occlusion while preserving existing speech patterns, using skeletally fixated intraoral devices. METHODS: Seven patients with cleft lip and palate aged 15 years 11 months to 26 years 5 months were selected. All osteotomies included horizontal anterior maxillary osteotomies created to the first molar and vertical osteotomies created between the second premolar and the first molars. Patients were evaluated preoperatively and postoperatively by a speech pathologist by means of fiberoptic video nasoendoscopy. RESULTS: Absolute bony anteroposterior movement ranged from 8 to 18.0 mm (average, 11.29 mm). Changes in skeletal facial profile (N- A-Pg) demonstrated approximately 18.61 degrees of correction in the conversion toward convex profiles. Patients were advanced to stable class I or mild class II relationships, and open bites were closed without development of hypernasal speech. CONCLUSIONS: This novel technique permits significant anterior movements, allowing dramatic improvements in functional and facial aesthetic outcomes. Furthermore, intraoral appliances allowed greater acceptance in this age group. Disadvantages include complicated orthodontic setup and surgical procedure, as well as the cost of occasionally necessary dental implants. No postoperative relapse was seen at an average follow-up of 33 months. This approach renders comfortable yet effective patient care, yielding optimum results while circumventing shortcomings of conventional techniques.

Impact of cleft lip and/or palate on nutritional health and oral-motor development.

Infants born with a cleft may require modifications in feeding practices prior to surgical closure of the defect; however, few changes in dietary recommendations are necessary. Often, the delivery method of breast milk or formula can be altered in order to require less effort by the infant and decrease caloric output, thereby increasing the calories ingested to facilitate weight gain and growth. This adaptation may not be necessary when the cleft does not include the palate but can be implemented fairly easily when the baby appears to have difficulty obtaining adequate nutritional intake. Before and after any surgical intervention, the goal remains to continue to supply the infant with sufficient caloric intake to heal and to continue to grow. Although some surgeons may demand modifications in how the baby is fed postoperatively, many advocate cautious reinstitution of normal feeding practices. Early referral for dental care should be encouraged in children born with clefts because these children (even the very young) demonstrate higher dental needs. Education provided to parents regarding causes of and methods to reduce dental disease could help decrease its incidence and help these children require less invasive and difficult rehabilitation therapy.