The PARP inhibitor Rucaparib synergizes with radiation to attenuate atypical teratoid rhabdoid tumor growth.

Background: Atypical teratoid rhabdoid tumors (ATRT) are highly aggressive pediatric brain tumors. The available treatments rely on toxic chemotherapy and radiotherapy, which themselves can cause poor outcomes in young patients. Poly (ADP-ribose) polymerases (PARP), multifunctional enzymes which play an important role in DNA damage repair and genome stability have emerged as a new target in cancer therapy. An FDA-approved drug screen revealed that Rucaparib, a PARP inhibitor, is important for ATRT cell growth. This study aims to investigate the effect of Rucaparib treatment in ATRT. Methods: This study utilized cell viability, colony formation, flow cytometry, western blot, immunofluorescence, and immunohistochemistry assays to investigate Rucaparib's effectiveness in BT16 and MAF737 ATRT cell lines. In vivo, intracranial orthotopic xenograft model of ATRT was used. BT16 cell line was transduced with a luciferase-expressing vector and injected into the cerebellum of athymic nude mice. Animals were treated with Rucaparib by oral gavaging and irradiated with 2 Gy of radiation for 3 consecutive days. Tumor growth was monitored using In Vivo Imaging System. Results: Rucaparib treatment decreased ATRT cell growth, inhibited clonogenic potential of ATRT cells, induced cell cycle arrest and apoptosis, and led to DNA damage accumulation as shown by increased expression of γH2AX. In vivo, Rucaparib treatment decreased tumor growth, sensitized ATRT cells to radiation and significantly increased mice survival. Conclusion: We demonstrated that Rucaparib has potential to be a new therapeutic strategy for ATRT as seen by its ability to decrease ATRT tumor growth both in vitro and in vivo.

Venetoclax Cooperates with Ionizing Radiation to Attenuate Diffuse Midline Glioma Tumor Growth.

PURPOSE: Tumor relapse after radiotherapy is a major hurdle in treating pediatric H3K27M-mutant diffuse midline gliomas (DMG). Radiotherapy-induced stress increases association of BCL2 family of proteins with BH3 pro-apoptotic activators preventing apoptosis. We hypothesized that inhibition of radiotherapy-induced BCL2 with a clinically relevant inhibitor, venetoclax, will block BCL2 activity leading to increased apoptosis. BCL2 has never been implicated in DMG as a radiotherapy-induced resistant mechanism. EXPERIMENTAL DESIGN: We performed an integrated genomic analysis to determine genes responsible for radioresistance and a targeted drug screen to identify drugs that synergize with radiation in DMG. Effect of venetoclax on radiation-naïve and 6 Gy radiation on cells was evaluated by studying cell death, changes in BCL2 phosphorylation, reactive oxygen species (ROS), and apoptosis, as well as BCL2 association with BH3 apoptosis initiators. The efficacy of combining venetoclax with radiation was evaluated in vivo using orthotopic xenograft models. RESULTS: BCL2 was identified as a key regulator of tumor growth after radiation in DMGs. Radiation sensitizes DMGs to venetoclax treatment independent of p53 status. Venetoclax as a monotherapy was not cytotoxic to DMG cells. Postradiation venetoclax treatment significantly increased cell death, reduced BCL2-BIM association, and augmented mitochondrial ROS leading to increased apoptosis. Combining venetoclax with radiotherapy significantly enhanced the survival of mice with DMG tumors. CONCLUSIONS: This study shows that venetoclax impedes the antiapoptotic function of radiation-induced BCL2 in DMG, leading to increased apoptosis. Results from these preclinical studies demonstrate the potential use of the BCL2 inhibitor venetoclax combined with radiotherapy for pediatric DMG.

Targeting the TP53/MDM2 axis enhances radiation sensitivity in atypical teratoid rhabdoid tumors.

Atypical teratoid rhabdoid tumor (ATRT) is a highly aggressive pediatric brain tumor. Despite radiation, aggressive chemotherapy and autologous stem cell rescue, children usually have a poor survival time. In the present study, the role of TP53/MDM2 interaction in ATRT was investigated. A functional genomic screen identified the TP53/MDM2 axis as a therapeutic target in the central nervous system (CNS) ATRT. Gene expression analysis revealed that all ATRT sub­groups expressed high levels of MDM2, which is a negative regulator of TP53. Using cell viability, colony formation and methylcellulose assays it was found that genetic MDM2 inhibition with short hairpin RNA or chemical MDM2 inhibition with small molecule inhibitors, Nutlin3 and idasanutlin (RG7388) decreased the growth of ATRT cell lines. Furthermore, idasanutlin significantly decreased the growth of intracranial orthotopic ATRT brain tumors, as evaluated using T2 MRI, and prolonged survival time relative to control animals. MRI of intracranial tumors showed that diffusion coefficient, an effective marker for successful treatment, significantly increased with idasanutlin treatment showing tumor necrosis/apoptosis. Immunohistochemistry revealed an increased number of caspase­3­positive cells in the idasanutlin treatment group, confirming the induction of apoptosis in vivo. Using flow cytometry and western blot analysis we show that inhibition of MDM2 enhanced radiation sensitivity in vitro by potentiating DNA damage via the induction of the TP53/Bax/Puma proapoptotic axis. Furthermore, DNA damage was associated with increased mitochondrial reactive oxygen species accumulation. The present study demonstrated that MDM2 expression level was increased in ATRT patient samples and MDM2 inhibition suppressed ATRT cell growth in vitro, and leads to apoptosis in vivo. MDM2 inhibition potentiates DNA damage and sensitizes ATRT cells to radiation. These findings highlight the TP53/MDM2 axis as a rational therapeutic target in CNS ATRT.

A novel PLK1 inhibitor onvansertib effectively sensitizes MYC-driven medulloblastoma to radiotherapy.

BACKGROUND: Group 3 medulloblastoma (MB) is often accompanied by MYC amplification. PLK1 is an oncogenic kinase that controls cell cycle and proliferation and has been preclinically validated as a cancer therapeutic target. Onvansertib (PCM-075) is a novel, orally available PLK1 inhibitor, which shows tumor growth inhibition in various types of cancer. We aim to explore the effect of onvansertib on MYC-driven medulloblastoma as a monotherapy or in combination with radiation. METHODS: Crisper-Cas9 screen was used to discover essential genes for MB tumor growth. Microarray and immunohistochemistry on pediatric patient samples were performed to examine the expression of PLK1. The effect of onvansertib in vitro was measure by cell viability, colony-forming assays, extreme limiting dilution assay, and RNA-Seq. ALDH activity, cell-cycle distribution, and apoptosis were analyzed by flow cytometry. DNA damage was assessed by immunofluorescence staining. Medulloblastoma xenografts were generated to explore the monotherapy or radio-sensitizing effect. RESULTS: PLK1 is overexpressed in Group 3 MB. The IC50 concentrations of onvansertib in Group 3 MB cell lines were in a low nanomolar range. Onvansertib reduced colony formation, cell proliferation, stem cell renewal and induced G2/M arrest in vitro. Moreover, onvansertib in combination with radiation increased DNA damage and apoptosis compared with radiation treatment alone. The combination radiotherapy resulted in marked tumor regression in xenografts. CONCLUSIONS: These findings demonstrate the efficacy of a novel PLK1 inhibitor onvansertib in vitro and in xenografts of Group 3 MB, which suggests onvansertib is an effective strategy as monotherapy or in combination with radiotherapy in MB.

Senescence Induced by BMI1 Inhibition Is a Therapeutic Vulnerability in H3K27M-Mutant DIPG.

Diffuse intrinsic pontine glioma (DIPG) is an incurable brain tumor of childhood characterized by histone mutations at lysine 27, which results in epigenomic dysregulation. There has been a failure to develop effective treatment for this tumor. Using a combined RNAi and chemical screen targeting epigenomic regulators, we identify the polycomb repressive complex 1 (PRC1) component BMI1 as a critical factor for DIPG tumor maintenance in vivo. BMI1 chromatin occupancy is enriched at genes associated with differentiation and tumor suppressors in DIPG cells. Inhibition of BMI1 decreases cell self-renewal and attenuates tumor growth due to induction of senescence. Prolonged BMI1 inhibition induces a senescence-associated secretory phenotype, which promotes tumor recurrence. Clearance of senescent cells using BH3 protein mimetics co-operates with BMI1 inhibition to enhance tumor cell killing in vivo.

Single-Cell RNA Sequencing of Childhood Ependymoma Reveals Neoplastic Cell Subpopulations That Impact Molecular Classification and Etiology.

Ependymoma (EPN) is a brain tumor commonly presenting in childhood that remains fatal in most children. Intra-tumoral cellular heterogeneity in bulk-tumor samples significantly confounds our understanding of EPN biology, impeding development of effective therapy. We, therefore, use single-cell RNA sequencing, histology, and deconvolution to catalog cellular heterogeneity of the major childhood EPN subgroups. Analysis of PFA subgroup EPN reveals evidence of an undifferentiated progenitor subpopulation that either differentiates into subpopulations with ependymal cell characteristics or transitions into a mesenchymal subpopulation. Histological analysis reveals that progenitor and mesenchymal subpopulations co-localize in peri-necrotic zones. In conflict with current classification paradigms, relative PFA subpopulation proportions are shown to determine bulk-tumor-assigned subgroups. We provide an interactive online resource that facilitates exploration of the EPN single-cell dataset. This atlas of EPN cellular heterogeneity increases understanding of EPN biology.

Super Elongation Complex as a Targetable Dependency in Diffuse Midline Glioma.

Histone 3 gene mutations are the eponymous drivers in diffuse midline gliomas (DMGs), aggressive pediatric brain cancers for which no curative therapy currently exists. These recurrent oncohistones induce a global loss of repressive H3K27me3 residues and broad epigenetic dysregulation. In order to identify therapeutically targetable dependencies within this disease context, we performed an RNAi screen targeting epigenetic/chromatin-associated genes in patient-derived DMG cultures. This identified AFF4, the scaffold protein of the super elongation complex (SEC), as a molecular dependency in DMG. Interrogation of SEC function demonstrates a key role for maintaining clonogenic potential while promoting self-renewal of tumor stem cells. Small-molecule inhibition of SEC using clinically relevant CDK9 inhibitors restores regulatory RNA polymerase II pausing, promotes cellular differentiation, and leads to potent anti-tumor effect both in vitro and in patient-derived xenograft models. These studies present a rationale for further exploration of SEC inhibition as a promising therapeutic approach to this intractable disease.

Performance of marrow stromal cell-seeded small-caliber multilayered vascular graft in a senescent sheep model.

Failure of small-caliber grafts, used as bypass or reconstructive grafts in cardiovascular treatments, is often caused by thrombosis and stenosis. We have developed a multilayered, compliant graft with an electrospun heparin-encapsulated core and collagen-chitosan shell. Herein, the performances of acellular and cell-seeded grafts were evaluated in adult sheep for preclinical assessment. Allogeneic ovine marrow stroma cells (MSCs) were uniformly attached to the lumen of cell-seeded grafts. Interposition grafts were used for carotid arteries. Four grafts were tested for each type. Upon implantation, all grafts successfully restored perfusion and rhythmically deformed under pulsatile arterial flow. Weekly ultrasonography and Doppler revealed that all grafts remained patent for perfusion during the course of one-month study. No formation of blood clots or other complications were found. The diameter of graft lumen did not vary significantly over the time or with the graft type, while narrowing at anastomosis and significant thickening of graft wall were found in both types of grafts. More significant neotissue formation was found at anastomotic sections of acellular controls compared to cell-seeded grafts. Results from histological and immunofluorescent analyses revealed moderate intimal hyperplasia (IH) at anastomosis. When compared to cell-seeded grafts, acellular controls presented thicker IH composed of α-smooth muscle actin positive cells and ground substances, which correlated with reduced and more disturbing flow. IH was thickest at anastomosis and tapered off to a minimum in the mid-section. Few PECAM-positive cells appeared on cell-seeded grafts but not acellular controls. Additionally, lesser graft thickening was found in cell-seed grafts, which might be associated with the function of stromal cells in altering the fibrotic process during tissue repair. Results suggest that MSCs held the potential to reduce hyperplasia and improve healing in an aged, large animal model for vascular grafting.

Development of an electrospun biomimetic polyurea scaffold suitable for vascular grafting.

The optimization of biomechanical and biochemical properties of a vascular graft to render properties relevant to physiological environments is a major challenge today. These critical properties of a vascular graft not only regulate its stability and integrity, but also control invasion of cells for scaffold remodeling permitting its integration with native tissue. In this work, we have synthesized a biomimetic scaffold by electrospinning a blend of a polyurea, poly(serinol hexamethylene urea) (PSHU), and, a polyester, poly-ε-caprolactone (PCL). Mechanical properties of the scaffold were varied by varying polymer blending ratio and electrospinning flow rate. Mechanical characterization revealed that scaffolds with lower PSHU content relative to PCL content resulted in elasticity close to native mammalian arteries. We also found that increasing electrospinning flow rates also increased the elasticity of the matrix. Optimization of elasticity generated scaffolds that enabled vascular smooth muscle cells (SMCs) to adhere, grow and maintain a SMC phenotype. The 30/70 scaffold also underwent slower degradation than scaffolds with higher PSHU content, thereby, providing the best option for in vivo remodeling. Further, Gly-Arg-Gly-Asp-Ser (RGD) covalently conjugated to the polyurea backbone in 30/70 scaffold resulted in significantly increased clotting times. Reducing surface thrombogenicity by the conjugation of RGD is critical to avoiding intimal hyperplasia. Hence, biomechanical and biochemical properties of a vascular graft can be balanced by optimizing synthesis parameters and constituent components. For these reasons, the optimized RGD-conjugated 30/70 scaffold electrospun at 2.5 or 5 mL/h has great potential as a suitable material for vascular grafting applications. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 278-290, 2018.

Novice Meditators of an Easily Learnable Audible Mantram Sound Self-Induce an Increase in Vagal Tone During Short-term Practice: A Preliminary Study.

INTRODUCTION: The autonomic nervous system (ANS) plays a vital role in regulation of the physiological processes during normal and pathological conditions. Heart rate variability (HRV) is regarded as a major indicator of the self-regulatory strength and balance of parasympathetic nervous system (PNS) and sympathetic nerve system (SNS) impulses, as interpreted by the intrinsic nervous system of the heart. The present study focuses on the evaluation of the effects of audible and mindful practice of chanting meditation on HRV and on the cognitive disposition. METHODOLOGY: The subjects were randomly assigned to 2 different sounds based on monotone repetition. One was the Hare Krishna Mantram (HKM-Sanskrit experimental sound) and the other was a concocted sound (Sanskrit placebo). Changes in vagal tone were measured with respect to both time domain and frequency domains. Five-min baseline and postmeditation measurements were obtained on different days over the next 6 wk. The subjects who chanted the placebo/sham sound switched to the experimental sound at the 4-wk mark for the next 2 wk. All subjects completed an experience survey. RESULTS: Paired t test results for all HRV parameters achieved statistical significance in the test group. Statistical significance in all the aforementioned measures of HRV was also observed on switching the control group placebo sound chanting to the experimental sound. HRV wave forms showed relaxation, a pattern and experience survey suggests an increase in attributes associated with higher self-regulation. CONCLUSION: This study suggests statistically significant efficacy data and that a larger randomized study is feasible to test the potential of the audible repetition of the HKM in clinical settings. It may therefore enable beneficial lifestyle for health creation and thus play a role in the prevention of chronic diseases. Further, large scale studies are required for a better perspective on the effect of mantram repetition on the HRV.

Comparison of magnetic resonance elastography and diffusion-weighted imaging for differentiating benign and malignant liver lesions.

OBJECTIVES: Comparison of magnetic resonance elastography (MRE) and diffusion-weighted imaging (DWI) for differentiating malignant and benign focal liver lesions (FLLs). METHODS: Seventy-nine subjects with 124 FLLs (44 benign and 80 malignant) underwent both MRE and DWI. MRE was performed with a modified gradient-echo sequence and DWI with a free breathing technique (b = 0.500). Apparent diffusion coefficient (ADC) maps and stiffness maps were generated. FLL mean stiffness and ADC values were obtained by placing regions of interest over the FLLs on stiffness and ADC maps. The accuracy of MRE and DWI for differentiation of benign and malignant FLL was compared using receiver operating curve (ROC) analysis. RESULTS: There was a significant negative correlation between stiffness and ADC (r = -0.54, p < 0.0001) of FLLs. Malignant FLLs had significantly higher mean stiffness (7.9kPa vs. 3.1kPa, p < 0.001) and lower mean ADC (129 vs. 200 × 10(-3)mm(2)/s, p < 0.001) than benign FLLs. The sensitivity/specificity/positive predictive value/negative predictive value for differentiating malignant from benign FLLs with MRE (cut-off, >4.54kPa) and DWI (cut-off, <151 × 10(-3)mm(2)/s) were 96.3/95.5/97.5/93.3% (p < 0.001) and 85/81.8/88.3/75% (p < 0.001), respectively. ROC analysis showed significantly higher accuracy for MRE than DWI (0.986 vs. 0.82, p = 0.0016). CONCLUSION: MRE is significantly more accurate than DWI for differentiating benign and malignant FLLs. KEY POINTS: • MRE is superior to DWI for differentiating benign and malignant focal liver lesions. • Benign lesions with large fibrous components may have higher stiffness with MRE. • Cholangiocarcinomas tend to have higher stiffness than hepatocellular carcinomas. • Hepatocellular adenomas tend to have lower stiffness than focal nodular hyperplasia. • MRE is superior to conventional MRI in differentiating benign and malignant liver lesions.

A fibrin/hyaluronic acid hydrogel for the delivery of mesenchymal stem cells and potential for articular cartilage repair.

BACKGROUND: Osteoarthritis (OA) is a degenerative joint disease affecting approximately 27 million Americans, and even more worldwide. OA is characterized by degeneration of subchondral bone and articular cartilage. In this study, a chondrogenic fibrin/hyaluronic acid (HA)-based hydrogel seeded with bone marrow-derived mesenchymal stem cells (BMSCs) was investigated as a method of regenerating these tissues for OA therapy. This chondrogenic hydrogel system can be delivered in a minimally invasive manner through a small gauge needle, forming a three-dimensional (3D) network structure in situ. However, an ongoing problem with fibrin/HA-based biomaterials is poor mechanical strength. This was addressed by modifying HA with methacrylic anhydride (MA) (HA-MA), which reinforces the fibrin gel, thereby improving mechanical properties. In this study, a range of fibrinogen (the fibrin precursor) and HA-MA concentrations were explored to determine optimal conditions for increased mechanical strength, BMSC proliferation, and chondrogenesis potential in vitro. RESULTS: Increased mechanical strength was achieved by HA-MA reinforcement within fibrin hydrogels, and was directly correlated with increasing HA-MA concentration. Live/dead staining and metabolic assays confirmed that the crosslinked fibrin/HA-MA hydrogels provided a suitable 3D environment for BMSC proliferation. Quantitative polymerase chain reaction (qPCR) of BMSCs incubated in the fibrin/HA-MA hydrogel confirmed decreased expression of collagen type 1 alpha 1 mRNA with an increase in Sox9 mRNA expression especially in the presence of a platelet lysate, suggesting early chondrogenesis. CONCLUSION: Fibrin/HA-MA hydrogel may be a suitable delivery method for BMSCs, inducing BMSC differentiation into chondrocytes and potentially aiding in articular cartilage repair for OA therapy.

Vascular stiffening in pulmonary hypertension: cause or consequence? (2013 Grover Conference series).

Recent studies have indicated that systemic arterial stiffening is a precursor to hypertension and that hypertension, in turn, can perpetuate arterial stiffening. Pulmonary artery (PA) stiffening is also well documented to occur in pulmonary hypertension (PH), and there is evidence that pulmonary vascular stiffness (PVS) may be a better predictor of outcome than pulmonary vascular resistance (PVR). We have hypothesized that the decreased flow-damping function of elastic PAs in PH likely initiates and/or perpetuates dysfunction of pulmonary microvasculature. Recent studies have shown that large-vessel stiffening increases flow pulsatility in the distal pulmonary vasculature, leading to endothelial dysfunction within a proinflammatory, vasoconstricting, and profibrogenic environment. The intricate role of stiffening-stimulated high pulsatile flow in endothelial cell dysfunction includes stepwise molecular events underlying PA hypertrophy, inflammation, endothelial-mesenchymal transition, and fibrosis. In addition to contributing to microenvironmental alterations of the distal vasculature, disordered proximal-distal PA coupling likely also plays a role in increasing ventricular afterload, ultimately causing right ventricle (RV) dysfunction and death. Current therapeutic treatments do not provide a realistic approach to destiffening arteries and, thus, to potentially abrogating the effects of high pulsatile flow on the distal pulmonary vasculature or the increased work imposed by stiffening on the RV. Scrutinizing the effect of PA stiffening on high pulsatile flow-induced cellular and molecular changes, and vice versa, might lead to important new therapeutic options that abrogate PA remodeling and PH development. With a clear understanding that PA stiffening may contribute to the progression of PH to an irreversible state by contributing to chronic microvascular damage in lungs, future studies should be aimed first at defining the underlying mechanisms leading to PA stiffening and then at improved treatment approaches based on these findings.

Erratum to: A fibrin/hyaluronic acid hydrogel for the delivery of mesenchymal stem cells and potential for articular cartilage repair.

[This corrects the article DOI: 10.1186/1754-1611-8-10.].

DIA2: Web-based Cyberinfrastructure for Visual Analysis of Funding Portfolios.

We present a design study of the Deep Insights Anywhere, Anytime (DIA2) platform, a web-based visual analytics system that allows program managers and academic staff at the U.S. National Science Foundation to search, view, and analyze their research funding portfolio. The goal of this system is to facilitate users' understanding of both past and currently active research awards in order to make more informed decisions of their future funding. This user group is characterized by high domain expertise yet not necessarily high literacy in visualization and visual analytics-they are essentially casual experts-and thus require careful visual and information design, including adhering to user experience standards, providing a self-instructive interface, and progressively refining visualizations to minimize complexity. We discuss the challenges of designing a system for casual experts and highlight how we addressed this issue by modeling the organizational structure and workflows of the NSF within our system. We discuss each stage of the design process, starting with formative interviews, prototypes, and finally live deployments and evaluation with stakeholders.

Mesoporous silica nanoparticles as a breast-cancer targeting ultrasound contrast agent.

Ultrasound (US) is used widely in the context of breast cancer. While it is advantageous for a number of reasons, it has low specificity and requires the use of a contrast agent. Its use as a standalone diagnostic and real-time imaging modality could be achieved by development of a tumor-targeted ultrasound contrast agent (UCA); functionalizing the UCA with a tumor-targeting agent would also allow the targeted administration of anti-cancer drugs at the tumor site. In this article, clinical US techniques are used to show that mesoporous silica nanoparticles (MSNs), functionalized with the monoclonal antibody Herceptin(®), can be used as an effective UCA by increasing US image contrast. Furthermore, in vitro assays show the successful localization and binding of the MSN-Herceptin conjugate to HER2+ cancer cells, resulting in tumor-specific cytotoxicity. These results demonstrate the potential of MSNs as a stable, biocompatible, and effective therapeutic and diagnostic ("theranostic") agent for US-based breast cancer imaging, diagnosis, and treatment.

Mechanical and biocompatible characterizations of a readily available multilayer vascular graft.

There is always a considerable clinical need for vascular grafts. Considering the availability, physical and mechanical properties, and regenerative potential, we have developed and characterized readily available, strong, and compliant multilayer grafts that support cell culture and ingrowth. The grafts were made from heterogeneous materials and structures, including a thin, dense, nanofibrous core composed of poly-ε-caprolactone (PCL), and a thick, porous, hydrogel sleeve composed of genipin-crosslinked collagen-chitosan (GCC). Because the difference in physicochemical properties between PCL and GCC caused layer separation, the layer adhesion was identified as a determinant to graft property and integrity under physiological conditions. Thus, strategies to modify the layer interface, including increasing porosity of the PCL surface, decreasing hydrophobicity, and increasing interlayer crosslinking, were developed. Results from microscopic images showed that increasing PCL porosity was characterized by improved layer adhesion. The resultant graft was characterized by high compliance (4.5%), and desired permeability (528 mL/cm(2)/min), burst strength (695 mmHg), and suture strength (2.38 N) for readily grafting. Results also showed that PCL mainly contributed to the graft mechanical properties, whereas GCC reduced the water permeability. In addition to their complementary contributions to physical and mechanical properties, the distinct graft layers also provided layer-specific structures for seeding and culture of vascular endothelial and smooth muscle cells in vitro. Acellular graft constructs were readily used to replace abdominal aorta of rabbits, resulting in rapid cell ingrowth and flow reperfusion. The multilayer constructs capable of sustaining physiological conditions and promoting cellular activities could serve as a platform for future development of regenerative vascular grafts.

Roles of genipin crosslinking and biomolecule conditioning in collagen-based biopolymer: Potential for vascular media regeneration.

The vascular media, a layer of the blood vessel wall containing smooth muscle cells (SMCs), are often the target functional tissue in the construction of artificial vessel. It contributes to mechanical properties and biological functions of vessels. The present study aimed to study effects of crosslinking and biomolecule conditions in the development of mechanically strong and stable, biologically functional constructs with potential for vascular media regeneration. Genipin was used to crosslink collagen-chitosan-elastin (CCE) constructs. Results revealed that mechanical strength, stiffness, and stability of CCE constructs significantly increased with genipin concentration, but crosslinking significantly inhibited SMC contraction of and invasion in gel constructs. No contraction or invasion was observed in those crosslinked with genipin at 5 mM or above. attenuated total reflectance Fourier transform infrared results showed crosslinking changed functional groups on CCE depending on genipin concentration. To enhance biological activities on crosslinked constructs, soluble molecule factors were incorporated, and their effects on SMC activities were evaluated. These conditions include heparin, platelet-derived transforming growth factor (PDGF), high-concentrated fetal bovine serum (h-FBS), a mixture of heparin and PDGF, and a mixture of h-FBS and PDGF. The h-FBS and PDGF mixture was found to stimulate a 3.2-fold increase in SMC contraction of the crosslinked gels. It was also found that PDGF and h-FBS, separately and in combination, induced SMC invasion in the crosslinked gels, while heparin attenuated PDGF-induced SMC invasion. Our study suggests that designing high-performance acellular constructs to encourage tissue regeneration should use a combination of crosslinking condition and biomolecule factor, striking a balance between mechanical properties and biological functions.