Impact of Undergraduate Research Training Programs: An Illustrative Example of Finding a Comparison Group and Evaluating Academic and Graduate School Outcomes.

In this study, researchers at a large, urban, comprehensive minority-serving institution used propensity score matching to identify a unique comparison group to study academic and graduate school outcomes in students served by the National Institutes of Health-funded Building Infrastructure Leading to Diversity (BUILD) Initiative. Acknowledging that students' self-selection biases may confound findings, the use of propensity methods to match students served with those who were not (but were otherwise eligible) provides a valuable tool for evaluators and practitioners to combat this challenge and better evaluate their effectiveness and impact on students' success. This study's findings indicate that BUILD participants had higher academic and graduate school success with regard to cumulative GPA, units attempted and completed, graduation status, and application and admission to graduate programs.

Evaluation of a bioengineered construct for tissue engineering applications.

Effective biomaterial options for tissue repair and regeneration are limited. Current biologic meshes are derived from different tissue sources and are generally sold as decellularized tissues. This work evaluated two collagen based bioengineered constructs and a commercial product in a model of abdominal full thickness defect repair. To prepare the bioengineered construct, collagen type 1 from porcine skin was isolated using an acid solubilization method. After purification, the collagen was formed into collagen sheets that were physically bonded to form a mechanically robust construct that was subsequently laser micropatterned with pores as a means to promote tissue integration (collagen only construct). A second engineered construct consisted of the aforementioned collagen construct embedded in an RGD-functionalized alginate gel that serves as a bioactive interface (collagen-alginate construct). The commercial product is a biologic mesh derived from bovine pericardium (Veritas® ). We observed enhanced vascularization in the midportion of the engineered collagen-alginate construct 2 weeks after implantation. Overall, the performance of the bioengineered constructs was similar to that of the commercial product with comparable integration strength at 8 weeks. Bioengineered constructs derived from monomeric collagen demonstrate promise for a variety of load bearing applications in tissue engineering. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2345-2354, 2018.

An immobilized liquid interface prevents device associated bacterial infection in vivo.

Virtually all biomaterials are susceptible to biofilm formation and, as a consequence, device-associated infection. The concept of an immobilized liquid surface, termed slippery liquid-infused porous surfaces (SLIPS), represents a new framework for creating a stable, dynamic, omniphobic surface that displays ultralow adhesion and limits bacterial biofilm formation. A widely used biomaterial in clinical care, expanded polytetrafluoroethylene (ePTFE), infused with various perfluorocarbon liquids generated SLIPS surfaces that exhibited a 99% reduction in S. aureus adhesion with preservation of macrophage viability, phagocytosis, and bactericidal function. Notably, SLIPS modification of ePTFE prevents device infection after S. aureus challenge in vivo, while eliciting a significantly attenuated innate immune response. SLIPS-modified implants also decrease macrophage inflammatory cytokine expression in vitro, which likely contributed to the presence of a thinner fibrous capsule in the absence of bacterial challenge. SLIPS is an easily implementable technology that provides a promising approach to substantially reduce the risk of device infection and associated patient morbidity, as well as health care costs.

Engineered composite fascia for stem cell therapy in tissue repair applications.

A critical challenge in tissue regeneration is to develop constructs that effectively integrate with the host tissue. Here, we describe a composite, laser micromachined, collagen-alginate construct containing human mesenchymal stem cells (hMSCs) for tissue repair applications. Collagen type I was fashioned into laminated collagen sheets to form a mechanically robust fascia that was subsequently laser micropatterned with pores of defined dimension and spatial distribution as a means to modulate mechanical behavior and promote tissue integration. Significantly, laser micromachined patterned constructs displayed both substantially greater compliance and suture retention strength than non-patterned constructs. hMSCs were loaded in an RGD-functionalized alginate gel modified to degrade in vivo. Over a 7 day observation period in vitro, high cell viability was observed with constant levels of VEGF, PDGF-ß and MCP-1 protein expression. In a full thickness abdominal wall defect model, the composite construct prevented hernia recurrence in Wistar rats over an 8-week period with de novo tissue and vascular network formation and the absence of adhesions to underlying abdominal viscera. As compared to acellular constructs, constructs containing hMSCs displayed greater integration strength (cell seeded: 0.92 ± 0.19 N/mm vs. acellular: 0.59 ± 0.25 N/mm, p=0.01), increased vascularization (cell seeded: 2.7-2.1/hpf vs. acellular: 1.7-2.1/hpf, p<0.03), and increased infiltration of macrophages (cell seeded: 2021-3630 µm(2)/hpf vs. acellular: 1570-2530 µm(2)/hpf, p<0.05). A decrease in the ratio of M1 macrophages to total macrophages was also observed in hMSC-populated samples. Laser micromachined collagen-alginate composites containing hMSCs can be used to bridge soft tissue defects with the capacity for enhanced tissue repair and integration. STATEMENT OF SIGNIFICANCE: Effective restoration of large soft tissue defects caused by trauma or treatment complications represents a critical challenge in the clinic. In this study, a novel composite construct was engineered and evaluated for stem cell delivery and tissue repair. Laser micromachining was used to fabricate patterned, microporous constructs designed with pores of defined size and distribution as a means to tune mechanical responses, accommodate and protect incorporated cells, and enhance tissue integration. The construct was embedded within an engineered alginate gel containing hMSCs. Upon repair of a full thickness abdominal wall defect in a rat model, the composite construct modulated host innate immunity towards a reparative phenotypic response, promoted neovascularization and associated matrix production, and increased the strength of tissue integration.

Localized delivery of mechano-growth factor E-domain peptide via polymeric microstructures improves cardiac function following myocardial infarction.

The Insulin like growth factor-I isoform mechano-growth factor (MGF), is expressed in the heart following myocardial infarction and encodes a unique E-domain region. To examine E-domain function, we delivered a synthetic peptide corresponding to the unique E-domain region of the human MGF (IGF-1Ec) via peptide eluting polymeric microstructures to the heart. The microstructures were made of poly (ethylene glycol) dimethacrylate hydrogel and bioengineered to be the same size as an adult cardiac myocyte (100 × 15 × 15 µm) and with a stiffness of 20 kPa. Peptide eluting microrods and empty microrods were delivered via intramuscular injection following coronary artery ligation in mice. To examine the physiologic consequences, we assessed the impact of peptide delivery on cardiac function and cardiovascular hemodynamics using pressure-volume loops and gene expression by quantitative RT-PCR. A significant decline in both systolic and diastolic function accompanied by pathologic hypertrophy occurred by 2 weeks which decompensated further by 10 weeks post-infarct in the untreated groups. Delivery of the E-domain peptide eluting microrods decreased mortality, ameliorated the decline in hemodynamics, and delayed decompensation. This was associated with the inhibition of pathologic hypertrophy despite increasing vascular impedance. Delivery of the empty microrods had limited effects on hemodynamics and while pathologic hypertrophy persisted there was a decrease in ventricular stiffness. Our data show that cardiac restricted administration of the MGF E-domain peptide using polymeric microstructures may be used to prevent adverse remodeling of the heart and improve function following myocardial infarction.

Discrete microstructural cues for the attenuation of fibrosis following myocardial infarction.

Chronic fibrosis caused by acute myocardial infarction (MI) leads to increased morbidity and mortality due to cardiac dysfunction. We have developed a therapeutic materials strategy that aims to mitigate myocardial fibrosis by utilizing injectable polymeric microstructures to mechanically alter the microenvironment. Polymeric microstructures were fabricated using photolithographic techniques and studied in a three-dimensional culture model of the fibrotic environment and by direct injection into the infarct zone of adult rats. Here, we show dose-dependent down-regulation of expression of genes associated with the mechanical fibrotic response in the presence of microstructures. Injection of this microstructured material into the infarct zone decreased levels of collagen and TGF-ß, increased elastin deposition and vascularization in the infarcted region, and improved functional outcomes after six weeks. Our results demonstrate the efficacy of these discrete anti-fibrotic microstructures and suggest a potential therapeutic materials approach for combatting pathologic fibrosis.

Sustained delivery of MGF peptide from microrods attracts stem cells and reduces apoptosis of myocytes.

Local release of drugs may have many advantages for tissue repair but also presents major challenges. Bioengineering approaches allow microstructures to be fabricated that contain bioactive peptides for sustained local delivery. Heart tissue damage is associated with local increases in mechano growth factor (MGF), a member of the IGF-1 family. The E domain of MGF peptide is anti-apoptotic and a stem cell homing factor. The objectives of this study were to fabricate a microrod delivery device of poly (ethylene glycol) dimethacrylate (PEGDMA) hydrogel loaded with MGF peptide and to determine the elution profile and bioactivity of MGF. The injectable microrods are 30 kPa stiffness and 15 µm widths by 100 µm lengths, chosen to match heart stiffness and myocyte size. Successful encapsulation of native MGF peptide within microrods was achieved with delivery of MGF for 2 weeks, as measured by HPLC. Migration of human mesenchymal stem cells (hMSCs) increased with MGF microrod treatment (1.72 ± 0.23, p < 0.05). Inhibition of the apoptotic pathway in neonatal rat ventricular myocytes was induced by 8 h of hypoxia (1 % O2). Protection from apoptosis by MGF microrod treatment was shown by the TUNEL assay and increased Bcl-2 expression (2 ± 0.19, p < 0.05). Microrods without MGF regulated the cytoskeleton, adhesion, and proliferation of hMSCs, and MGF had no effect on these properties. Therefore, the combination microdevice provided both the mechanical cues and 2-week MGF bioactivity to reduce apoptosis and recruit stem cells, suggesting potential use of MGF microrods for cardiac regeneration therapy in vivo.

Integrin α3 blockade enhances microtopographical down-regulation of α-smooth muscle actin: role of microtopography in ECM regulation.

Development of functional engineered matrices for regenerative therapies can benefit from an understanding of how physical cues at the microscale affect cell behavior. In this work, we use microfabricated systems to study how stiffness and microscale topographical cues in the form of "micropegs" affect extracellular matrix synthesis. Previous work from our lab has shown that microtopographical cues in 2D and 3D systems decrease cellular proliferation and regulate matrix synthesis. In this work, the combined role of stiffness and topography on ECM synthesis is investigated in a 2D micropeg system. These studies show that fibroblasts cultured on polydimethylsiloxane (PDMS) substrates with micropegs have reduced expression of collagen type I (Col I) and collagen type VI (Col VI) compared to fibroblasts cultured on flat substrates. In addition, cells on micropegged substrates exhibit down-regulation of other important regulators of ECM synthesis such as α-smooth muscle actin (α-SMA), and integrin α3 (Int α3). Interestingly, this effect is dependent on the contractility and adhesion of the cells. When cultured in the presence of RhoA kinase (ROCK) and myosin light chain kinase (MLCK) inhibitors, no significant differences in the expression of collagen, α-SMA, Int α3, and TGFB1 are observed. Additionally, disruptions in cell adhesion prevent microtopographical regulation of ECM synthesis. When using an antibody to block the extracellular domain of Int α3, no differences in the expression of collagen are observed and blocking Int α3 results in enhanced down-regulation of α-SMA on the stiffer micropegged substrates. These findings demonstrate that regulation of extracellular matrix production by cells on a synthetic substrate can be guided via physical cues at the microscale, and add to the body of knowledge on the role of integrin-mediated mechanotransduction.

Microtopographical cues in 3D attenuate fibrotic phenotype and extracellular matrix deposition: implications for tissue regeneration.

Recent studies have highlighted the role of external biophysical cues on cell morphology and function. In particular, substrate geometry and rigidity in two dimensions has been shown to impact cell growth, death, differentiation, and motility. Knowledge of how these physical cues affect cell function in three dimensions is critical for successful development of novel regenerative therapies. In this work, the effect of discrete micromechanical cues in three-dimensional (3D) system on cell proliferation, gene expression, and extracellular matrix synthesis was investigated. Poly(ethylene glycol) dimethacrylate hydrogel microrods were fabricated using photolithography and suspended in gel to create a 3D culture with microscale cues of defined mechanical properties in the physiological range (2-50 kPa). These microrods significantly affected fibroblast proliferation, matrix production, and gene expression. Cultures with stiff microrods reduced fibroblast proliferation and downregulated expression of key extracellular matrix proteins involved in scar tissue formation. In addition, the contractility marker alpha smooth muscle actin and adhesion molecule integrin alpha3 were also significantly downregulated. Cultures with soft microrods had no significant difference on fibroblast proliferation and expression of Cyclin D1, alpha smooth muscle actin, and integrin alpha3 compared to cultures with no microrods. Here, we present a new platform of potentially injectable microrods with tunable elasticity; in addition, we show that cell proliferation and gene expression are influenced by discrete physical cues in 3D.

Biophysical mechanisms of single-cell interactions with microtopographical cues.

Biophysical cues encoded in the extracellular matrix (ECM) are increasingly being explored to control cell behavior in tissue engineering applications. Recently, we showed that cell adhesion to microtopographical structures ("micropegs") can suppress proliferation in a manner that may be blunted by inhibiting cellular contractility, suggesting that this effect is related to altered cell-scaffold mechanotransduction. We now directly investigate this possibility at the microscale through a combination of live-cell imaging, single-cell mechanics methods, and analysis of gene expression. Using time-lapse imaging, we show that when cells break adhesive contacts with micropegs, they form F-actin-filled tethers that extend and then rupture at a maximum, critical length that is greater than trailing-edge tethers observed on topographically flat substrates. This critical tether length depends on myosin activation, with inhibition of Rho-associated kinase abolishing topography-dependent differences in tether length. Using cellular de-adhesion and atomic force microscopy indentation measurements, we show that the micropegs enhance cell-scaffold adhesive interactions without changing whole-cell elasticity. Moreover, micropeg adhesion increases expression of specific mechanotransductive genes, including RhoA GTPase and myosin heavy chain II, and, in myoblasts, the functional marker connexin 43. Together, our data support a model in which microtopographical cues alter the local mechanical microenvironment of cells by modulating adhesion and adhesion-dependent mechanotransductive signaling.