Cell sheet engineering enables scaffold-free tissue reconstruction, while preserving native cell–cell junctions and extracellular matrix architecture. However, the clinical translation of this technology has been hindered by high production costs, limited open-source protocols, regulatory challenges and functional limitations, in particular with regards to mechanical stability. In this Review, we first describe how cell sheets are fabricated using stimuli-responsive substrates and examine their applications in cardiac, hepatic, uterine, tendon, ligament and paediatric contexts. We then discuss key mechanical considerations in cell sheet engineering, including strategies to modulate mechanical properties through sheet stacking, anisotropy control through micropatterning, and cyclic mechanical conditioning. By addressing technical innovations and remaining challenges, we provide a framework for expanding the utility of cell sheet engineering to mechanically demanding tissue environments. Cell sheet engineering offers a scaffold-free approach to tissue reconstruction. This Review outlines fabrication methods, key mechanical design strategies and innovations aimed at enabling the use of cell sheets in mechanically demanding tissues.

Maternal and fetal health during pregnancy is closely tied to contractile modulation of the uterine myometrium, as dysfunctional contractions underlie pregnancy complications such as preterm labor and postpartum hemorrhaging. Structural abnormalities increase the risk for pregnancy complications, yet the complex interplay between biochemical and biomechanical cues in the myometrial microenvironment is not well defined. Significant gaps in our understanding of myometrial physiology and the lack of physiologically relevant research tools to enable systematic investigation have resulted in a largely ineffective and nonspecific therapeutic landscape for myometrial contractile modulation during pregnancy. We address this unmet need by developing a widely accessible approach for in vitro hydrogel platform fabrication to advance the study of agonist-mediated calcium responses in myometrial cells. We directly fabricated polyacrylamide hydrogels in polypropylene multiwell plates and Petri dishes and validated their mechanical properties and use as a mechanically tunable cell culture substrate. Using "physiological" matrix conditions during pregnancy, a fluorescent calcium mobilization assay was implemented in a plate-reader-based workflow to determine the dose sensitivity of myometrial cells to the endogenous agonist oxytocin. Using hydrogels with elastic moduli spanning physiological, pathological, and supraphysiological conditions in the myometrium, we assessed stiffness-mediated effects in myometrial cell behavior, including agonist-mediated calcium responses. We observed the mechanosensitive modulation of calcium response curve amplitude in oxytocin-stimulated myometrial cells. Myometrial cell behavior on mechanically tunable substrates was compared against widely used tissue culture plastic, and the observed changes to myometrial cell morphology and calcium responses highlight the significant influence of supraphysiological substrates. To explore complex pathological microenvironments, myometrial cells were exposed to the proinflammatory trigger lipopolysaccharide (LPS) before agonist stimulation. In our studies, LPS exposure altered calcium responses during oxytocin stimulation in cells cultured on supraphysiological substrates. Altogether, we present an easily adoptable in vitro hydrogel platform with scalable fabrication and versatile application use, including new directions in myometrial mechanobiology.Impact StatementWe investigated mechanosensitive modulation of agonist-mediated calcium responses in myometrial cells during pregnancy using a high-throughput in vitro polyacrylamide hydrogel platform. Polypropylene labware enabled scalable fabrication of an in vitro hydrogel platform without being costly or labor-intensive. Our studies found that oxytocin-stimulated calcium responses in myometrial cells are sensitive to nonphysiological matrix conditions. This work highlights the importance of using physiologically relevant engineered microenvironments and represents new directions for understanding myometrial mechanobiology. Our platform can help accelerate the identification of agonists and antagonists to counteract dysfunctional myometrial contractions and can be used more broadly in other tissues that exhibit mechanosensitive pathologies.

The "Voices" under this Perspective underline the importance of interdisciplinary collaboration and partnerships across several disciplines, such as medical science and technology, medicine, bioengineering, and computational approaches, in bridging the gap between research, manufacturing, and clinical applications. Effective communication is key to bridging team gaps, enhancing trust, and resolving conflicts, thereby fostering teamwork and individual growth toward shared goals. Drawing from the success of the COVID-19 vaccine development, we advocate the application of similar collaborative models in other complex health areas such as nanomedicine and biomedical engineering. The role of digital technology and big data in healthcare innovation is highlighted along with the necessity for specialized education in collaborative practices. This approach is decisive in advancing healthcare solutions, leading to improved treatment and patient outcomes.

Fibrin is a major component in early adhesion formation and, consequently, is an attractive molecular target for imaging agents. While previous studies have indicated that the peptide CREKA (Cys-Arg-Glu-Lys-Ala) can bind to fibrin and direct an attached imaging agent to the desired tissue, the molecular mechanism by which CREKA targets fibrin remains unknown. This study aims to determine the binding mode of peptides capable of interacting with fibrin for tissue targeting. Existing fibrin and fibrinogen structures were analyzed, and computational mapping was performed to identify binding hot spots on the surface of the fibrin protein. Models of the binding mode of the potential fibrin targeting peptide, CREKA and its analogs, were developed. The models indicate that the peptides bind in the "A" and "B" holes of fibrin, and therefore, peptide binding would be expected to limit fibrin polymerization. Dynamic light scattering shows that the effects of the peptides on fibrin polymerization are consistent with the modeling. Our findings provide detailed molecular models of peptide-fibrin binders as starting points for the development of optimized peptides for tissue targeting. This study lays the groundwork for the development of targeted contrast agents for the detection of forming adhesions and could be applied more broadly in scenarios such as thrombosis and stroke, where targeting fibrin is also desired.

The nanomedicine field continues to gain momentum, with several groundbreaking clinical trials underway. However, despite the promise of advanced antifouling nanoparticles incorporating poly(ethylene glycol)—a key component in the development of COVID-19 vaccines—the clinical translation of nanomedicine remains limited. This is primarily due to the relatively low delivery efficacy, with passive targeting relying on the enhanced permeability and retention effect, and active targeting leading to only modest improvements in target tissue accumulation. Improving the targeting, biocompatibility, and functionality of nanoparticles has the potential to create more effective, personalized, and minimally invasive therapies. This review aims to highlight the rise of a previously unidentified order of immune-minded nanomaterials and explores how mechanobiological principles and biomechanical nanotools are revolutionizing our understanding of nano-bio interactions in relation to disease. By considering mechanical properties such as stiffness, surface topology, and behavior under physiological flow conditions, researchers can better engineer nanoparticles for improved therapeutic outcomes.

The automated soft matter indenter (ASMI) is a platform for rapidly performing mechanical characterization of samples with elastic moduli in the range 7 kPa to 67 MPa with a sample acquisition time between 1 and 10 min. It is a low-cost system based upon open-source software, a modified mill, and an educational force sensor with a total bill of materials <$500. This system tests batches of up to 96 samples based on a standard well-plate sample holder without requiring any human intervention. Using the ASMI, users can obtain mechanical data in a programmable manner that enables high-throughput workflows, precisely testing time-dependent phenomena, and integration with other processing steps for closed-loop optimization.

To optimize microbubble formulations for clinical applications, the size distribution, concentration, and acoustic intensity must be rapidly measurable to allow for the successful iteration of microbubble design. In this paper, a comprehensive method was developed to compare microbubble formulations with different lipid shell compositions using optical and acoustic methods of measurement to collect the size distribution, concentration, and mean scattering intensity. An open-source ImageJ macro code was modified for the selective counting and sizing of brightfield microbubble images. A high-throughput agarose phantom was designed to collect multiple scattering reflections of microbubble samples to estimate the echogenicity of each microbubble solution. The information contained in the size distribution and concentration, combined with the instantaneous scattering power, can identify modifications needed for prototyping specific microbubble formulations.

Background: Cardiac allograft vasculopathy (CAV) is a rapidly progressive form of coronary atherosclerosis limiting long-term survival after heart transplantation. Objectives: We evaluated the diagnostic and prognostic yield of quantitative stress cardiovascular magnetic resonance (CMR) perfusion for CAV detection in heart transplant recipients. Methods: Patients who received orthotopic heart transplants and underwent stress CMR for CAV assessment were included in the study and followed up for almost 2 years (median 1.8; IQR 0.9,2.7). The diagnostic accuracy of qualitative and quantitative stress CMR was assessed by calculating sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV), using invasive or CT coronary angiography as the reference for CAV detection. The area under the curve (AUC) was compared for qualitative and quantitative stress CMR. Adjusted hazard ratios for major adverse cardiac events (MACE), including death and unplanned cardiac hospitalizations were derived in all patients. The global myocardial perfusion reserve index (MPRi) was obtained by normalization to the rate-pressure product. Results: In a cohort of 60 patients, n=18 (30%) had significant CAV (grade 2 or 3), and n=11 (18.3%) experienced MACE. At the Youden index threshold of 2.1, the myocardial perfusion reserve index (MPRi) demonstrated a sensitivity of 85.7%, a specificity of 70.3%, a PPV of 52.2%, and an NPV of 92.9%. The MPRi was significantly more accurate than visual assessment (p < 0.001) in identifying underlying CAV (Figure 1) and it was an independent predictor of MACE (HR:0.26;95%CI:0.07,0.93; log-rank p=0.022; Figure 2), while the visual presence of inducible myocardial perfusion defect did not (HR:2.23;95%CI:0.57,8.66; p=0.2). Conclusions: In patients with previous heart transplantation, quantitative stress CMR perfusion has incremental diagnostic and prognostic value over qualitative stress CMR for the non-invasive detection of CAV.

In the United States, 1 in 10 infants are born preterm. The majority of neonatal deaths and nearly a third of infant deaths are linked to preterm birth. Preterm birth is initiated when the quiescent state of the uterus ends prematurely, leading to contractions and parturition beginning as early as 32 weeks, though the origins are not well understood. To enable research and discovery of therapeutics with potential to better address preterm birth, the capability to study isolated cell processes of pregnant uterine tissue in vitro is needed. Our development of an in vitro model of the myometrium utilizing human uterine smooth muscle cells (uSMCs) responsible for contractions provides a methodology to examine cellular mechanisms of late-stage pregnancy potentially involved in preterm birth. We discuss culture of uSMCs on a flexible polydimethylsiloxane (PDMS) substrate functionalized with cationic poly-l-lysine (PLL), followed by extracellular matrix (ECM) protein coating. Previous work exploring uSMC behavior on PDMS substrates have utilized collagen-I coatings, however, we demonstrated the first exploration of human uSMC response to strain on fibronectin-coated flexible membranes, importantly reflecting the significant increase of fibronectin content found in the myometrial ECM during late-stage pregnancy. Using the model we developed, we conducted proof-of-concept studies to investigate the impact of substrate strain on uSMC cell morphology and gene expression. It was found that PLL and varied ECM protein coatings (collagen I, collagen III, and fibronectin) altered cell nuclei morphology and density on PDMS substrates. Additionally, varied strain rates applied to uSMC substrates significantly impacted uSMC gene expression of IL-6, a cytokine associated with instances of preterm labor. These results suggest that both surface and mechanical properties of in vitro systems impact primary human uSMC phenotype and offer uSMC culture methodologies that can be utilized to further the understanding of cellular pathways involved in the uterus under mechanical load.

Recurrent myocardial ischemia can lead to left ventricular (LV) dysfunction in patients with coronary artery disease (CAD). In this observational cohort study, we assessed for chronic metabolomic and transcriptomic adaptations within LV myocardium of patients undergoing coronary artery bypass grafting. During surgery, paired transmural LV biopsies were acquired on the beating heart from regions with and without evidence of inducible ischemia on preoperative stress perfusion cardiovascular magnetic resonance. From 33 patients, 63 biopsies were acquired, compared to analysis of LV samples from 11 donor hearts. The global myocardial adenosine triphosphate (ATP):adenosine diphosphate (ADP) ratio was reduced in patients with CAD as compared to donor LV tissue, with increased expression of oxidative phosphorylation (OXPHOS) genes encoding the electron transport chain complexes across multiple cell types. Paired analyses of biopsies obtained from LV segments with or without inducible ischemia revealed no significant difference in the ATP:ADP ratio, broader metabolic profile or expression of ventricular cardiomyocyte genes implicated in OXPHOS. Differential metabolite analysis suggested dysregulation of several intermediates in patients with reduced LV ejection fraction, including succinate. Overall, our results suggest that viable myocardium in patients with stable CAD has global alterations in bioenergetic and transcriptional profile without large regional differences between areas with or without inducible ischemia.

Background: Stress perfusion cardiac magnetic resonance (CMR) imaging is recommended in international guidelines in class I in patients with known or suspected CAD. Patients undergoing cardiac transplantation (OHT) are at risk of developing cardiac allograft vasculopathy (CAV). Adenosine is relatively contraindicated after transplantation because of a presumed risk of atrioventricular block in denervated hearts. We sought to evaluate the safety and feasibility of adenosine stress CMR in patients with OHT and suspected CAV. Methods: In an outpatient cohort study, we measured the incidence of adverse effects of adenosine stress CMR in consecutive OHT recipients undergoing quantitative stress perfusion CMR for suspected CAV. We additionally stratified patients based on presence of significant CAV (grades 2-3 versus 0-1) on either invasive or computed tomography coronary angiography to evaluate potential CMR biomarkers of significant coronary disease in this population. Results: We identified 50 adenosine stress CMR studies performed in 45 individual patients at a median of 19 years after transplantation (IQR 4.7-24.4); 9% of studies were conducted within the first year post-transplantation. No life-threatening adverse events, brief or prolonged atrioventricular block or other arrythmias occurred after adenosine infusion. Significant myocardial ischemia (≥ 1.5 segments) was reported in 17% of patients. When stratified by CAV severity, stress MBF index, but not inducible ischemia on visual inspection, was found to significantly differentiate patients with severe CAV from those without significant coronary disease (ANOVA p = 0.028). Conclusion: Adenosine stress perfusion CMR is safe and feasible in OHT recipients. Quantitative measurement of stress MBF index may represent a promising marker of coronary disease with incremental value over qualitative assessment of ischemia, and should be evaluated in future studies.

Peritoneal adhesions are bands of fibrous tissue that can bind adjacent tissue or organs together and have a high probability of occurrence after a patient undergoes deep abdominal surgery. These adhesions can cause complications, such as chronic pain, intestinal obstruction or constriction, organ displacement, and even death, yet there exists no common diagnostic for these adhesions prior to symptomatic appearance. Furthermore, the gold standard treatment is ironically surgical removal. Nascent adhesions formed in the first 48 hafter surgery are primarily composed of fibrin. In this study, lipid shelled microbubbles have been designed as a potential theranostic agent to detect and treat adhesions using a fibrin targeting peptide called CREKA (Cys-Arg-Glu-Lys-Ala). Low amplitude through-transmission experiments were conducted to characterize the mechanical properties of the microbubble lipid shell, such as the shear modulus of elasticity and shear viscosity. Passive cavitation detection (PCD) experiments were conducted to determine the inertial cavitation threshold by which the microbubbles may break up fibrin under ultrasound exposure. [Work supported by NIH SBIR and BU Mechanical Engineering Department.]

Cardiovascular disease (CVD) represents a significant threat to women's health. Heart failure (HF) is one CVD that still has an increasing incidence and about half of all cases involve women. HF is characterised by strong sex-specific features in aetiology, clinical manifestation and outcomes. Women are more likely to have hypertensive heart disease and HF with preserved ejection fraction, they experience worse quality of life but have a better overall survival rate. Women's hearts also have unique morphological characteristics that should be considered during cardiovascular assessment. It is important to understand and highlight these sex-specific features to be able to provide a tailored diagnostic approach and therapeutic management. The aim of this article is to review these aspects together with the challenges and the unique characteristics of different imaging modalities used for the diagnosis and follow-up of women with HF.