Synthetic Self-assembling Peptide Research Articles

Development of 3D melanoma cultures on a hyaluronic acid-based scaffold with synthetic self-assembling peptides: Electroporation enhancement

Electrochemotherapy (ECT) with bleomycin is an effective antitumor treatment. Still, researchers are investigating new drugs and electroporation conditions to improve its efficacy. To this aim, in vivo assays are accurate but expensive and ethically questionable. Conversely, in vitro assays, although cheaper and straightforward, do not reflect the architecture of the biological tissue because they lack a tridimensional (3D) structure (as in the case of two-dimensional [2D] in vitro assays) or do not include all the extracellular matrix components (as in the case of 3D in vitro scaffolds). To address this issue, 3D in vitro models have been proposed, including spheroids and hydrogel-based cultures, which require a suitable low-conductive medium to allow cell membrane electroporation.In this study, a synthetic scaffold based on hyaluronic acid (HA) and self-assembling peptides (SAPs; EAbuK), condensed with a Laminin-derived adhesive sequence (IKVAV), is proposed as a reliable alternative. We compare SKMEL28 cells cultured in the HA-EAbuK-IKVAV scaffold to the control (HA only scaffold). Three days after seeding, the culture on the HA-EAbuK-IKVAV scaffold showed collagen production. SKMEL28 cells cultured on the HA-EAbuK-IKVAV scaffold started to be electroporated at 400 V/cm, whereas, at the same electric field intensity, those cultured on HA were not. As a reference, 2D experiments showed that electroporation of SKMEL28 cells starts at 600 V/cm using an electroporation buffer and at 800 V/cm in a culture medium, but with very low efficiency (<50 % of cells electroporated). 3D cultures on HA-EAbuK-IKVAV allowed the simulation of a more reliable microenvironment and may represent a valuable tool for studying electroporation conditions. Using Finite Element Analysis (FEA) to compute the transmembrane potential, we detected the influence of inhomogeneity of the extracellular matrix on electroporation effect. Our 3D cell culture electroporation simulations showed that the transmembrane potential increased when collagen surrounded the cells. Of note, in the collagen-enriched HA-EAbuK-IKVAV scaffold, EP was already improved at lower electric field intensities. This study shows the influence of the extracellular matrix on electric conductivity and electric field distribution on cell membrane electroporation and supports the adoption of more reliable 3D scaffolds in experimental electroporation studies.

Evaluation of a synthetic peptide-based bioink (PeptiInk Alpha 1) for in vitro 3D bioprinting of cartilage tissue models

Cartilage pathology in human disease is poorly understood and requires further research. Various attempts have been made to study cartilage pathologies using in vitro human cartilage models as an alternative for preclinical research. Three-dimensional (3D) bioprinting is a technique that has been used to 3D-bioprint cartilage tissue models in vitro using animal-derived materials such as gelatine or hyaluronan, which present challenges in terms of scalability, reproducibility, and ethical concerns. We present an assessment of synthetic self-assembling peptides as bioinks for bioprinted human in vitro cartilage models. Primary human chondrocytes were mixed with PeptiInk Alpha 1, 3D-bioprinted and cultured for 14 days, and compared with 3D chondrocyte pellet controls. Cell viability was assessed through LIVE/DEAD assays and DNA quantification. High cell viability was observed in the PeptiInk culture, while a fast decrease in DNA levels was observed in the 3D pellet control. Histological evaluation using hematoxylin and eosin staining and immunofluorescence labeling for SOX-9, collagen type II, and aggrecan showed a homogeneous cell distribution in the 3D-bioprinted PeptiInks as well as high expression of chondrogenic markers in both control and PeptiInk cultures. mRNA expression levels assessed by - qRT-PCR (quantitative real time-polymerase chain reaction) confirmed chondrogenic cell behavior. These data showed promise in the potential use of PeptiInk Alpha 1 as a bioprintable manufacturing material for human cartilage in vitro models.

Efficacy of a novel self-assembling peptide gel for hemostasis in refractory neoplastic bleeding.

Video 1Outline of how self-assembling peptide gel can be used exvivo.

Application of a 3D hydrogel-based model to replace use of animals for passaging patient-derived xenografts

PurposeThis 3D in vitro cancer model for propagation of patient-derived cells, using a synthetic self-assembling peptide gel, allows the formation of a fully characterised, tailorable tumour microenvironment. Unlike many existing 3D cancer models, the peptide gel is inert, apart from molecules and motifs deliberately added or produced by cells within the model.MethodsBreast cancer patient-derived xenografts (PDXs) were disaggregated and embedded in a peptide hydrogel. Growth was monitored by microscopic examination and at intervals, cells were extracted from the gels and passaged on into fresh gels. Passaged cells were assessed by qPCR and immunostaining techniques for the retention of characteristic markers.ResultsBreast cancer PDXs were shown to be capable of expansion over four or more passages in the peptide gel. Contaminating mouse cells were found to be rapidly removed by successive passages. The resulting human cells were shown to be compatible with a range of common assays useful for assessing survival, growth and maintenance of heterogeneity.ConclusionsBased on these findings, the hydrogel has the potential to provide an effective and practical breast cancer model for the passage of PDXs which will have the added benefits of being relatively cheap, fully-defined and free from the use of animals or animal products. Encapsulated cells will require further validation to confirm the maintenance of cell heterogeneity, genotypes and phenotypes across passage, but with further development, including the addition of bespoke cell and matrix components of the tumour microenvironment, there is clear potential to model other cancer types.

Efficacy of self-assembling peptide in mitigating delayed bleeding after advanced endoscopic resection of gastrointestinal lesions: A meta-analysis.

Background and study aims Advanced endoscopic resection techniques carry a risk of delayed bleeding (DB). A novel fully synthetic self-assembling peptide (SAP) has shown promising results in mitigating this risk. In this meta-analysis, we evaluated all available data and analyzed the effectiveness of SAP in reducing DB after advanced endoscopic resection of gastrointestinal luminal lesions. Patients and methods Electronic databases (PubMed, Embase, and Cochrane Library) from January 2010 through October 2022 were searched for publications addressing the use of SAP solution in patients undergoing advanced endoscopic resection of gastrointestinal lesions. Pooled proportions were calculated using fixed (inverse variance) and random-effects (DerSimonian-Laird) models. Results The initial search identified 277 studies, of which 63 relevant articles were reviewed. The final analysis included data from six studies comprising 307 patients that met inclusion criteria. The pooled rate of DB was 5.73 % (95 % confidence interval [CI] = 3.42-8.59). Mean patient age was 69.40 years ± 1.82. The weighted mean size of resected lesions was 36.20 mm (95 % CI = 33.37-39.02). Endoscopic submucosal dissection was used in 72.69 % (95 % CI = 67.62-77.48), while endoscopic mucosal resection was used in 26.42 % (95 % CI = 21.69-31.44) of the procedures. Among the 307 patients, 36 % were on antithrombotic medications. No adverse events (AEs) were attributable to using SAP, with a pooled rate of 0.00 % (95 % CI = 0.00-1.49). Conclusions SAP solution appears promising in reducing post-procedural DB after advanced endoscopic resection of high-risk gastrointestinal lesions with no reported AEs.

Growth and differentiation of human induced pluripotent stem cell (hiPSC)-derived kidney organoids using fully synthetic peptide hydrogels

Human induced pluripotent stem cell (hiPSC)-derived kidney organoids have prospective applications ranging from basic disease modelling to personalised medicine. However, there remains a necessity to refine the biophysical and biochemical parameters that govern kidney organoid formation. Differentiation within fully-controllable and physiologically relevant 3D growth environments will be critical to improving organoid reproducibility and maturation. Here, we matured hiPSC-derived kidney organoids within fully synthetic self-assembling peptide hydrogels (SAPHs) of variable stiffness (storage modulus, G′). The resulting organoids contained complex structures comparable to those differentiated within the animal-derived matrix, Matrigel. Single-cell RNA sequencing (scRNA-seq) was then used to compare organoids matured within SAPHs to those grown within Matrigel or at the air-liquid interface. A total of 13,179 cells were analysed, revealing 14 distinct clusters. Organoid compositional analysis revealed a larger proportion of nephron cell types within Transwell-derived organoids, while SAPH-derived organoids were enriched for stromal-associated cell populations. Notably, differentiation within a higher G’ SAPH generated podocytes with more mature gene expression profiles. Additionally, maturation within a 3D microenvironment significantly reduced the derivation of off-target cell types, which are a known limitation of current kidney organoid protocols. This work demonstrates the utility of synthetic peptide-based hydrogels with a defined stiffness, as a minimally complex microenvironment for the selected differentiation of kidney organoids.

Increased Stiffness Downregulates Focal Adhesion Kinase Expression in Pancreatic Cancer Cells Cultured in 3D Self-Assembling Peptide Scaffolds.

The focal adhesion kinase (FAK) is a non-receptor tyrosine kinase that participates in integrin-mediated signal transduction and contributes to different biological processes, such as cell migration, survival, proliferation and angiogenesis. Moreover, FAK can be activated by autophosphorylation at position Y397 and trigger different signaling pathways in response to increased extracellular matrix stiffness. In addition, FAK is overexpressed and/or hyperactivated in many epithelial cancers, and its expression correlates with tumor malignancy and invasion potential. One of the characteristics of solid tumors is an over deposition of ECM components, which generates a stiff microenvironment that promotes, among other features, sustained cell proliferation and survival. Researchers are, therefore, increasingly developing cell culture models to mimic the increased stiffness associated with these kinds of tumors. In the present work, we have developed a new 3D in vitro model to study the effect of matrix stiffness in pancreatic ductal adenocarcinoma (PDAC) cells as this kind of tumor is characterized by a desmoplastic stroma and an increased stiffness compared to its normal counterpart. For that, we have used a synthetic self-assembling peptide nanofiber matrix, RAD16-I, which does not suffer a significant degradation in vitro, thus allowing to maintain the same local stiffness along culture time. We show that increased matrix stiffness in synthetic 3D RAD16-I gels, but not in collagen type I scaffolds, promotes FAK downregulation at a protein level in all the cell lines analyzed. Moreover, even though it has classically been described that stiff 3D matrices promote an increase in pFAKY397/FAK proteins, we found that this ratio in soft and stiff RAD16-I gels is cell-type-dependent. This study highlights how cell response to increased matrix stiffness greatly depends on the nature of the matrix used for 3D culture.

Growth and differentiation of human induced pluripotent stem cell (hiPSC) derived kidney organoids using fully synthetic peptide hydrogels

Human induced pluripotent stem cell (hiPSC)‐derived kidney organoids have prospective applications ranging from basic disease modelling to personalised medicine. However, there remains a necessity to refine the biophysical and biochemical parameters that govern kidney organoid formation. Differentiation within fully‐controllable and physiologically relevant 3D growth environments will be critical to improving organoid reproducibility and maturation. Here, we matured hiPSC‐derived kidney organoids within fully synthetic self‐assembling peptide hydrogels (SAPHs) of variable stiffness (storage modulus, G′). The resulting organoids contained complex structures comparable to those differentiated within the animal‐derived matrix, Matrigel. Single‐cell RNA sequencing (scRNA‐seq) was then used to compare organoids matured within SAPHs to those grown within Matrigel or at the air‐liquid interface. A total of 13,179 cells were analysed, revealing 14 distinct clusters. Organoid compositional analysis revealed a larger proportion of nephron cell types within Transwell‐derived organoids, while SAPH derived organoids were enriched for stromal‐associated cell populations. Notably, differentiation within a higher G’ SAPH generated podocytes with more mature gene expression profiles. Additionally, maturation within a 3D microenvironment significantly reduced the derivation of offtarget cell types, which are a known limitation of current kidney organoid protocols. This work demonstrates the utility of synthetic peptide‐based hydrogels with a defined stiffness, as a minimally complex microenvironment for the selected differentiation of kidney organoids.

Comparing natural hydrogels to self-assembling peptides in spinal cord injury treatment: a systematic review

Background: In many cases, central nervous system (CNS) injury is unchanging due to the absence of neuronal regeneration and repair capabilities. In recent years, regenerative medicine, and especially hydrogels, has reached a significant amount of attention for their promising results for the treatment of spinal cord injury (SCI) currently considered permanent. Hydrogels are categorized based on their foundation: synthetic, natural, and combination. The objective of this study was to compare the properties and efficacy of commonly used hydrogels, like collagen, and other natural peptides with synthetic self-assembling peptide hydrogels in the treatment of SCI. Methods: Articles were searched in PubMed, Scopus, Web of Science, and Embase. All studies from 1985 until January 2020 were included in the primary search. Eligible articles were included based on the following criteria: administering hydrogels (both natural and synthetic) for SCI treatment, solely focusing on spinal cord injury treatment, and published in a peer-reviewed journal. Data on axonal regeneration, revascularization, elasticity, drug delivery efficacy, and porosity were extracted. Results: A total of 24 articles were included for full-text review and data extraction. There was only one experimental study comparing collagen I (natural hydrogel) and polyethylene glycol (PEG) in an in vitro setting. The included study suggested the behavior of cells with PEG is more expectable in the injury site, which makes it a more reliable scaffold for neurites. Conclusions: There is limited research comparing and evaluating both types of natural and self-assembling peptides (SAPs) in the same animal or in vitro study, despite its importance. Although we assume that the remodeling of natural scaffolds may lead to a stable hydrogel, there was not a definitive conclusion that synthetic hydrogels are more beneficial than natural hydrogels in neuronal regeneration.

Enhancing Peptide Biomaterials for Biofabrication.

Biofabrication using well-matched cell/materials systems provides unprecedented opportunities for dealing with human health issues where disease or injury overtake the body’s native regenerative abilities. Such opportunities can be enhanced through the development of biomaterials with cues that appropriately influence embedded cells into forming functional tissues and organs. In this context, biomaterials’ reliance on rigid biofabrication techniques needs to support the incorporation of a hierarchical mimicry of local and bulk biological cues that mimic the key functional components of native extracellular matrix. Advances in synthetic self-assembling peptide biomaterials promise to produce reproducible mimics of tissue-specific structures and may go some way in overcoming batch inconsistency issues of naturally sourced materials. Recent work in this area has demonstrated biofabrication with self-assembling peptide biomaterials with unique biofabrication technologies to support structural fidelity upon 3D patterning. The use of synthetic self-assembling peptide biomaterials is a growing field that has demonstrated applicability in dermal, intestinal, muscle, cancer and stem cell tissue engineering.

Tuneable Hybrid Hydrogels via Complementary Self-Assembly of a Bioactive Peptide with a Robust Polysaccharide

Synthetic materials designed for improved biomimicry of the extracellular matrix must contain fibrous, bioactive, and mechanical cues. Self-assembly of low molecular weight gelator (LMWG) peptides Fmoc-DIKVAV (Fmoc-aspartic acid-isoleucine-lysine-valine-alanine-valine) and Fmoc-FRGDF (Fmoc-phenylalanine-arginine-glycine-aspartic acid-phenylalanine) creates fibrous and bioactive hydrogels. Polysaccharides such as agarose are biocompatible, degradable, and non-toxic. Agarose and these Fmoc-peptides have both demonstrated efficacy in vitro and in vivo. These materials have complementary properties; agarose has known mechanics in the physiological range but is inert and would benefit from bioactive and topographical cues found in the fibrous, protein-rich extracellular matrix. Fmoc-DIKVAV and Fmoc-FRGDF are synthetic self-assembling peptides that present bioactive cues "IKVAV" and "RGD" designed from the ECM proteins laminin and fibronectin. The work presented here demonstrates that the addition of agarose to Fmoc-DIKVAV and Fmoc-FRGDF results in physical characteristics that are dependent on agarose concentration. The networks are peptide-dominated at low agarose concentrations, and agarose-dominated at high agarose concentrations, resulting in distinct changes in structural morphology. Interestingly, at mid-range agarose concentration, a hybrid network is formed with structural similarities to both peptide and agarose systems, demonstrating reinforced mechanical properties. Bioactive-LMWG polysaccharide hydrogels demonstrate controllable microenvironmental properties, providing the ability for tissue-specific biomaterial design for tissue engineering and 3D cell culture.

Three-dimensional printing of extracellular matrix (ECM)-mimicking scaffolds: A critical review of the current ECM materials.

The loss of tissues and organs through injury and disease has stimulated the development of therapeutics that have the potential to regenerate and replace the affected tissue. Such therapeutics have the benefit of reducing the reliance and demand for life-saving organ transplants. Of the several regenerative strategies, 3D printing has emerged as the forerunner in regenerative attempts due to the fact that biologically and anatomically correct 3D structures can be fabricated according to the specified need. Despite the progress in this field, improvement is still limited by the difficulty in fabricating scaffolds that adequately mimic the native cellular microenvironment. In response, despite the complexities of the native extracellular matrix (ECM), the inclusion of ECM components into bioinks has emerged as a cutting-edge research area in terms of providing possible ECM-mimicking abilities of the 3D printed constructs. Furthermore, the development of ECM-mimicking scaffolds can potentially assist in improving personalized patient treatments. This review provides a critical analysis of selected naturally occurring ECM components as well as synthetic self-assembling peptides in their ability to provide the required ECM microenvironment for tissue regeneration. The success and possible short comings of each material, as well as the specific characteristics of each bioink, are evaluated.

Extracellular Matrix/Amorphous Magnesium Phosphate Bioink for 3D Bioprinting of Craniomaxillofacial Bone Tissue.

Bioprinting, a promising field in regenerative medicine, holds great potential to create three-dimensional, defect-specific vascularized bones with tremendous opportunities to address unmet craniomaxillofacial reconstructive challenges. A cytocompatible bioink is a critical prerequisite to successfully regenerate functional bone tissue. Synthetic self-assembling peptides have a nanofibrous structure resembling the native extracellular matrix (ECM), making them an excellent bioink component. Amorphous magnesium phosphates (AMPs) have shown greater levels of resorption while maintaining high biocompatibility, osteoinductivity, and low inflammatory response, as compared to their calcium phosphate counterparts. Here, we have established a novel bioink formulation (ECM/AMP) that combines an ECM-based hydrogel containing 2% octapeptide FEFEFKFK and 98% water with AMP particles to realize high cell function with desirable bioprintability. We analyzed the osteogenic differentiation of dental pulp stem cells (DPSCs) encapsulated in the bioink, as well as in vivo bone regeneration, to define the potential of the formulated bioink as a growth factor-free bone-forming strategy. Cell-laden AMP-modified bioprinted constructs showed an improved cell morphology but similar cell viability (∼90%) compared to their AMP-free counterpart. In functional assays, the cell-laden bioprinted constructs modified with AMP exhibited a high level of mineralization and osteogenic gene expression without the use of growth factors, thus suggesting that the presence of AMP-triggered DPSCs' osteogenic differentiation. Cell-free ECM-based bioprinted constructs were implanted in vivo. In comparison with the ECM group, bone volume per total volume for ECM/1.0AMP was approximately 1.7- and 1.4-fold higher at 4 and 8 weeks, respectively. Further, a significant increase in the bone density was observed in ECM/1.0AMP from 4 to 8 weeks. These results demonstrate that the presence of AMP in the bioink significantly increased bone formation, thus showing promise for in situ bioprinting strategies. We foresee significant potential in translating this innovative bioink toward the regeneration of patient-specific bone tissue for regenerative dentistry.

Practical Considerations in the Design and Use of Immunologically Active Fibrillar Peptide Assemblies.

The design, formulation, and immunological evaluation of self-assembling peptide materials is relatively straightforward. Indeed, one of the advantages of synthetic self-assembling peptides is that one can progress from initial concept to in vivo testing in a matter of days. However, because these materials are supramolecular, working with them is not without some practical challenges, and subtle changes in design, synthesis, handling, and formulation can affect the materials' immunogenicity. This chapter is intended to communicate some of these practical aspects of working with these materials that are not always enumerated in conventional research papers. Epitope considerations, peptide synthesis, purification, storage, nanofiber formation, quality control, immunological evaluation, and the overall phenotypic characteristics of the immune responses to be expected from these materials are discussed.

Designer bFGF-incorporated d-form self-assembly peptide nanofiber scaffolds to promote bone repair

d-Form and l-form peptide nanofiber scaffolds can spontaneously form stable β-sheet secondary structures and nanofiber hydrogel scaffolds, and hold some promise in hemostasis and wound healing. We report here on the synthetic self-assembling peptide d-RADA16 and l-RADA16 are both found to produce stable β-sheet secondary structure and nanofiber hydrogel scaffolds based on circular dichroism (CD) spectroscopy, transmission electron microscopy (TEM) and rheology analysis etc. d-RADA16 hydrogel and l-RADA16 hydrogel can enhance obvious bone repair in femoral condyle defects of the Sprague-Dawley (SD) rat model compared to PBS treatment. Based on micro-computed tomography (CT), it was revealed that d-RADA16 hydrogel and l-RADA16 hydrogel were capable to obtain the extensive bone healing. Histological evaluation also found that these two hydrogels facilitate the presence of more mature bone tissue within the femoral condyle defects. Additionally, d-RADA16 hydrogel showed some potential in storing and releasing basic-fibroblast growth factor (bFGF) which was able to further promote bone regeneration based on micro-CT analysis. These results indicate that d-form peptide nanofiber hydrogel have some special capacity for bone repair.

Bioprinting synthetic self-assembling peptide hydrogels for biomedical applications

Three-dimensional (3D) bioprinting is a disruptive technology for creating organotypic constructs for high-throughput screening and regenerative medicine. One major challenge is the lack of suitable bioinks. Short synthetic self-assembling peptides are ideal candidates. Several classes of peptides self-assemble into nanofibrous hydrogels resembling the native extracellular matrix. This is a conducive microenvironment for maintaining cell survival and physiological function. Many peptides also demonstrate stimuli-responsive gelation and tuneable mechanical properties, which facilitates extrusion before dispensing and maintains the shape fidelity of the printed construct in aqueous media. The inherent biocompatibility and biodegradability bodes well for in vivo applications as implantable tissues and drug delivery matrices, while their short length and ease of functionalization facilitates synthesis and customization. By applying self-assembling peptide inks to bioprinting, the dynamic complexity of biological tissue can be recreated, thereby advancing current biomedical applications of peptide hydrogel scaffolds.

Synergetic Use of Neural Precursor Cells and Self-assembling Peptides in Experimental Cervical Spinal Cord Injury

Spinal cord injuries (SCI) cause serious neurological impairment and psychological, economic, and social consequences for patients and their families. Clinically, more than 50% of SCI affect the cervical spine. As a consequence of the primary injury, a cascade of secondary mechanisms including inflammation, apoptosis, and demyelination occur finally leading to tissue scarring and development of intramedullary cavities. Both represent physical and chemical barriers to cell transplantation, integration, and regeneration. Therefore, shaping the inhibitory environment and bridging cavities to create a supportive milieu for cell transplantation and regeneration is a promising therapeutic target. Here, a contusion/compression model of cervical SCI using an aneurysm clip is described. This model is more clinically relevant than other experimental models, since complete transection or ruptures of the cord are rare. Also in comparison to the weight drop model, which in particular damage the dorsum columns, circumferential compression of the spinal cord appears advantageous. Clip closing force and duration can be adjusted to achieve different injury severity. A ring spring facilitates precise calibration and constancy of clip force. Under physiological conditions, synthetic self-assembling peptides (SAP) self-assemble into nanofibers and thus, are appealing for application in SCI. They can be injected directly into the lesion minimizing damage to the cord. SAPs are biocompatible structures erecting scaffolds to bridge intramedullary cavities and thus, equip the damaged cord for regenerative treatments. K2(QL)6K2 (QL6) is a novel SAP introduced by Dong et al. In comparison to other peptides, QL6 self-assembles into β-sheets at neutral pH. 14 days after SCI, after the acute stage, SAPs are injected into the center of the lesion and neural precursor cells (NPC) are injected into adjacent dorsal columns. In order to support cell survival, transplantation is combined with continuous subdural administration of growth factors by osmotic micro pumps for 7 days.

AC5 Surgical Hemostat™ as an effective hemostatic agent in an anticoagulated rat liver punch biopsy model

Intra-operative and postoperative bleeding is a major concern in surgical procedures for patients taking anticoagulant medications, or where anticoagulants are used to prevent potential life-threatening embolic complications. Heparin is the anticoagulant used most frequently and has an immediate effect on blood clotting, lasting 4 to 6h. Although synthetic self-assembling peptides have been shown to achieve rapid hemostasis in small animals, none have adequately addressed the potential for hemostasis in the presence of anticoagulant therapy in-vivo. Our goal was to investigate the hemostatic activity of a known synthetic self-assembling peptide in animals treated and untreated with heparin anticoagulation therapy. Using a rat liver puncture model, animals were treated with known synthetic peptide AC5 Surgical Hemostatic Device™, or saline controls. Time-to-hemostasis and coagulation times were recorded in both heparinized and non-heparinized animals. Here we show that AC5™ was able to achieve rapid hemostasis equivalently in both heparinized and non-heparinized animals. From the Clinical EditorIntra-operative and postoperative bleeding is a major concern in surgical procedures for patients taking anticoagulant medications. In this work the effective hemostasis was demonstrated both in heparinized and non-heparinized animals using self-assembling peptides.

Toward a 3D Cellular Model for Studying In Vitro the Outcome of Photodynamic Treatments: Accounting for the Effects of Tissue Complexity

Clinical therapies have traditionally been developed using two-dimensional (2D) cell culture systems, which fail to accurately capture tissue complexity. Therefore, three-dimensional (3D) cell cultures are more attractive platforms to integrate multiple cues that arise from the extracellular matrix and cells, closer to an in vivo scenario. Here we report the development of a 3D cellular model for the in vitro assessment of the outcome of oxygen- and drug-dependent therapies, exemplified by photodynamic therapy (PDT). Using a synthetic self-assembling peptide as a cellular scaffold (RAD16-I), we were able to recreate the in vivo limitation of oxygen and drug diffusion and its biological effect, which is the development of cellular resistance to therapy. For the first time, the production and decay of the cytotoxic species singlet oxygen could be observed in a 3D cell culture. Results revealed that the intrinsic mechanism of action is maintained in both systems and, hence, the dynamic mass transfer effects accounted for the major differences in efficacy between the 2D and 3D models. We propose that this methodological approach will help to improve the efficacy of future oxygen- and drug-dependent therapies such as PDT.

Effects of Self-Assembling Peptide Hydrogel Scaffold on Bone Regeneration with Recombinant Human Bone Morphogenetic Protein-2

The objective of this pilot study was to histologically evaluate bone regeneration using a self-assembling peptide hydrogel scaffold with recombinant human bone morphogenetic protein-2 (rhBMP-2) in a rabbit calvaria model. Five adult New Zealand White rabbits were used for the study. Each received four titanium cylinders, which were placed into perforated slits made in the outer cortical bone of the calvaria. The cylinders were filled with the following test materials: (1) unfilled control; (2) rhBMP-2; (3) PuraMatrix (PM), a synthetic self-assembling peptide (RADA16-I) consisting of a 16-amino acid sequence and with a three-dimensional structure; and (4) PM/rhBMP-2. Each cylinder was covered with a titanium lid. After 8 weeks, the animals were sacrificed, and ground sections were obtained for histomorphometric analysis. Histomorphometric analysis showed that regenerated tissue in the cylinder with PM/rhBMP-2 was significantly increased compared to the empty control. The mean area values of regenerated tissue in the cylinders were 35.80% ± 10.35% (control), 47.94% ± 5.65% (rhBMP-2), 48.94% ± 11.33% (PM), and 58.06% ± 14.84% (PM/rhBMP-2). The mean area values of newly formed bone in the cylinders were 9.39% ± 4.34% (control), 14.03% ± 2.25% (rhBMP-2), 13.99% ± 2.15% (PM), and 16.61% ± 3.79% (PM/rhBMP-2). Neither rhBMP-2 nor PM alone significantly enhanced bone regeneration compared to the empty control cylinder. PM with rhBMP-2 significantly enhanced bone regeneration on the bone augmentation model in a rabbit. PM promises to be a useful alternative synthetic material as a carrier for rhBMP-2 for bone regeneration.