Endotoxin contamination alters macrophage-cancer cell interaction and therapeutic efficacy in pre-clinical 3D in vitro models.

The rapid developments in biofabrication, in particular 3D bioprinting, in the recent years have facilitated the need for novel biomaterials that aim to replicate the target tissue in great detail. The presence of endotoxins in these biomaterials is often an overlooked problem. In pre-clinical 3D in vitro models, endotoxins can have significant influence on cell behavior and credibility of the model. In this study we demonstrate the effects of high levels of endotoxins in commercially-available gelatin on the macrophage-cancer cell crosstalk in a 3D bioprinted co-culture model. First, it is demonstrated that, while presenting the same mechanical and structural stimuli, high levels of endotoxin can have significant influence on the metabolic activity of macrophages and cancer cells. Furthermore, this study shows that high endotoxin contamination causes a strong inflammatory reaction in macrophages and significantly inhibits the effects of a paracrine macrophage-cancer cell co-culture. At last, it is demonstrated that the differences in endotoxin levels can drastically alter the efficacy of novel macrophage modulating immunotherapies, AS1517499 and 3-methyladenine. Altogether, this study shows that endotoxin contamination in biomaterials can significantly alter intra- and intercellular communication and thereby drug efficacy, which might lead to misinterpretation of the potency and safety of the tested compounds.

Microarchitectural mimicking of stroma-induced vasculature compression in pancreatic tumors using a 3D engineered model.

Fibrotic tumors, such as pancreatic ductal adenocarcinoma (PDAC), are characterized for high desmoplastic reaction, which results in high intra-tumoral solid stress leading to the compression of blood vessels. These microarchitectural alterations cause loss of blood flow and poor intra-tumoral delivery of therapeutics. Currently, there is a lack of relevant in vitro models capable of replicating these mechanical characteristics and to test anti-desmoplastic compounds. Here, a multi-layered vascularized 3D PDAC model consisting of primary human pancreatic stellate cells (PSCs) embedded in collagen/fibrinogen (Col/Fib), mimicking tumor tissue within adjunct healthy tissue, is presented to study the fibrosis-induced compression of vasculature in PDAC. It is demonstrated how the mechanical and biological stimulation induce PSC activation, extracellular matrix production and eventually vessel compression. The clinical relevance is confirmed by correlating with patient transcriptomic data. Furthermore, the effects of gradual vessel compression on the fluid dynamics occurring within the channel is evaluated in silico. Finally, it is demonstrated how cancer-associated fibroblast (CAF)-modulatory therapeutics can inhibit the cell-mediated compression of blood vessels in PDAC in vitro, in silico and in vivo. It is envisioned that this 3D model is used to improve the understanding of mechanical characteristics in tumors and for evaluating novel anti-desmoplastic therapeutics.

Effect of oversized partial postdilatation on coatings of contemporary durable polymer-based drug-eluting stents: a scanning electron microscopy study.

BACKGROUND: Oversized postdilation of drug-eluting stents (DES) is often performed to avoid stent malapposition. In stents implanted in long lesion or major bifurcations, extremely oversized partial postdilation may be required, which exposes DES coating to extreme forces. This study aims to assess shape and incidence of coating irregularities on durable polymer-based DES following extremely oversized partial postdilatation. METHODS: Fifteen DES samples (3 3.5 mm stents of Cypher Select plus [Cordis Europa, Roden, the Netherlands], Taxus Liberté[Boston Scientific Corp., Natick, MA, USA], Endeavor Sprint [Medtronic Vascular, Santa Rosa, CA, USA], Endeavor Resolute [Medtronic Vascular, Santa Rosa, CA, USA], and Xience V [Abbott Vascular, Santa Clara, CA, USA]) were deployed in sterile water (37 °C) at 14 atm, followed by a proximal postdilation with noncompliant 5.0-mm balloons at 18 atm. Stents were then examined with scanning electron microscopy. RESULTS: Thorough examination of a total of 660 scanning electron microscopic images demonstrated that shape and incidence of coating irregularities in the postdilated and/or transitional DES regions differed only mildly from the nonpostdilated regions. Cypher Select plus showed more peeling without bare metal aspect in the postdilated and transitional regions, and cracks were wider (P < 0.001) in the postdilated and transitional regions; in Taxus Liberté one additional irregularity (torn webbing) and more wrinkles were observed (P < 0.05 for both); in Endeavor Resolute wider cracks were found in the extremely postdilated region only (P < 0.001). Endeavor Sprint and Xience V showed no differences in shape or incidence of coating irregularities between oversized and nonoversized stent regions. CONCLUSIONS: Bench side assessment of five contemporary durable polymer-based DES with scanning electron microscopy suggests that even very aggressive stent postdilatation results in no more than mild differences in coating irregularities between postdilated and nonpostdilated stent regions.

Micro-computed tomographic assessment following extremely oversized partial postdilatation of drug-eluting stents.

AIMS: To assess the spatial geometry of drug-eluting stents (DES) following extremely oversized proximal postdilatation. Interventions of distal left main (LM) disease generally require stenting across the LM bifurcation with inherent vessel tapering along this segment and a high likelihood of stent malapposition, which can be avoided by such postdilations. METHODS AND RESULTS: Sixteen DES (four 3.5 mm-samples of Cypher Select Plus, Taxus Liberté, Endeavor Resolute, Xience V) were deployed in water; 12 samples were then proximally postdilated with noncompliant 5.0 mm balloons at 18 atm. All samples were examined by micro-computed tomography. Taxus Liberté, Endeavor Resolute, and Xience V, showed increased cell areas in the transitional region (just distal to postdilated region), while Cypher Select showed its largest cells inside the postdilated region. Overall, the largest maximum cell area was observed in Endeavor Resolute, while Cypher Select showed the smallest (p<0.001, for both). In addition, the size of the very proximal postdilated cells was relatively small in most DES except Xience V. CONCLUSIONS: Extremely oversized partial stent postdilatation demonstrated significant between-DES differences in final spatial stent configuration and maximum cell size. These data could be of practical interest with regard to coronary interventions in LM stems with stenting across the LM bifurcation.

Coating irregularities of durable polymer-based drug-eluting stents as assessed by scanning electron microscopy.

AIMS: To classify and quantify post-expansion irregularities in durable polymer-based coatings of drug-eluting stents (DES). METHODS AND RESULTS: Taxus Liberté, Endeavor Sprint, Endeavor Resolute and Xience V DES (three samples of each) were explored by light microscopy and scanning electron microscopy (SEM) following expansion at 14 atm in water. Incidence and size of irregularities were measured during thorough quantitative examinations of a 360 SEM images. DES types examined showed a significant difference in the incidence of irregularities (p<0.0001; 6.6+/-4.2/image at 60-fold magnification) with typical patterns specific for each DES. All types showed areas with bare metal-aspects, but incidence, shape, and size differed largely: Sprint showed the largest areas. Cracks were only found in Sprint and Resolute, while wrinkles were seen exclusively in Taxus Liberté and Xience V (p<0.0001). The coating of each DES type showed some inhomogeneity of distribution, but the incidence differed (p<0.0001) and was least in Taxus Liberté, which, on the other hand, was the only DES that showed webbing with large bare-metal exposure. CONCLUSIONS: The incidence and size of various coating irregularities on different types of DES varied widely. These data may be considered in ongoing discussions on the differences between DES and may serve as reference to compare novel DES.