Cancer Stem Cell Microenvironment Models with Biomaterial Scaffolds In Vitro

Defined by its potential for self-renewal, differentiation and tumorigenicity, cancer stem cells (CSCs) are considered responsible for drug resistance and relapse. To understand the behavior of CSC, the effects of the microenvironment in each tissue are a matter of great concerns for scientists in cancer biology. However, there are many complicated obstacles in the mimicking the microenvironment of CSCs even with current advanced technology. In this context, novel biomaterials have widely been assessed as in vitro platforms for their ability to mimic cancer microenvironment. These efforts should be successful to identify and characterize various CSCs specific in each type of cancer. Therefore, extracellular matrix scaffolds made of biomaterial will modulate the interactions and facilitate the investigation of CSC associated with biological phenomena simplifying the complexity of the microenvironment. In this review, we summarize latest advances in biomaterial scaffolds, which are exploited to mimic CSC microenvironment, and their chemical and biological requirements with discussion. The discussion includes the possible effects on both cells in tumors and microenvironment to propose what the critical factors are in controlling the CSC microenvironment focusing the future investigation. Our insights on their availability in drug screening will also follow the discussion

NOR-3, a donor of nitric oxide, increases intracellular Zn²⁺ concentration and decreases cellular thiol content: A model experiment using rat thymocytes, FluoZin-3, and 5-chloromethylfluorescein

Our previous study showed that nitroprusside, a donor of nitric oxide (NO), increased intracellular Zn2+ concentration without affecting cellular content of glutathione (GSH) although it has been proposed that the cytotoxicity of NO is resulted from its interaction with glutathione and zinc. Nitroprusside releases not only NO but also cyanides (Fe(II)CN and Fe(III)CN), CN-, Fe2+, and Fe3+. Therefore, such decomposition products may mask NO-induced action on cellular GSH content. In this study, we used NOR-3 as a donor of NO to reveal the effects of NO on intracellular Zn2+ concentration and cellular GSH content in a cytometric manner with fluorescent probes, FluoZin-3-AM and 5-chloromethylfluorescein diacetate. NOR-3 at 1-3 mM significantly increased intracellular Zn2+ concentration and decreased cellular GSH content. After the removal of extracellular Zn2+ by diethylenetriamine-N,N,N',N",N"-pentaacetic acid (DTPA, a chelator for Zn2+), the increase in intracellular Zn2+ concentration by NOR-3 was still observed although DTPA significantly attenuated the increase in intracellular Zn2+ concentration by NOR-3. Results suggest an involvement of both intracellular Zn2+ release and increase in membrane Zn2+ permeability. It is likely that NO induces oxidative stress, leading to an increase in intracellular Zn2+ concentration

がん遺伝子検査法としてのFRET-PHFA法の開発及び全自動遺伝子解析に向けた基礎評価

主1工学_物質創造工

High-throughput drug screening models of mature adipose tissues which replicate the physiology of patients’ Body Mass Index (BMI)

Obesity is a complex and incompletely understood disease, but current drug screening strategies mostly rely on immature in vitro adipose models which cannot recapitulate it properly. To address this issue, we developed a statistically validated high-throughput screening model by seeding human mature adipocytes from patients, encapsulated in physiological collagen microfibers. These drop tissues ensured the maintenance of adipocyte viability and functionality for controlling glucose and fatty acids uptake, as well as glycerol release. As such, patients’ BMI and insulin sensitivity displayed a strong inverse correlation: the healthy adipocytes were associated with the highest insulin-induced glucose uptake, while insulin resistance was confirmed in the underweight and severely obese adipocytes. Insulin sensitivity recovery was possible with two type 2 diabetes treatments, rosiglitazone and melatonin. Finally, the addition of blood vasculature to the model seemed to more accurately recapitulate the in vivo physiology, with particular respect to leptin secretion metabolism

がん遺伝子検査法としてのFRET-PHFA法の開発及び全自動遺伝子解析に向けた基礎評価

主1工学_物質創造工

Collagen Microfibers Induce Blood Capillary Orientation and Open Vascular Lumen

Achieving vascularization of engineered tissues or structures is a major challenge in the field of tissue engineering. Hitherto, studies on vascularization have demonstrated limited control of vascular network geometry, such as vasculature direction and network density. An open vascular lumen is crucial to ensure that cells survive and that metabolic activity is fully functional in large‐sized tissues. Herein, a method based on high water‐dispersible collagen microfibers (CMF) to fabricate capillary orientation‐controllable 3D tissue with an open vascular lumen using a dispensing machine is reported. A twenty micrometers‐long CMF (CMF‐20) with high dispersion property are shown to be more effective for dispensing a homogenous tissue and inducing formation of an interconnected capillary network than two hundred micrometers‐long CMF (CMF‐200). One of the advantages is the prevention of shrinkage on the z‐axis of hydrogel‐based tissue which acts as a microscaffold. The gaps between the fibers can support endothelial cell migration and maturation, thus forming a larger vascular lumen compared to CMF‐free controls. Besides, shear forces produced by the dispensing process cause the collagen microfibers to align, and these microfibers guide cell alignment by integrin‐induced adhesion. The findings based on CMF to allow blood capillary alignment and vascular lumen stabilization will be an important technology in tissue engineering

Collagen Microfibers Induce Blood Capillary Orientation and Open Vascular Lumen

Achieving vascularization of engineered tissues or structures is a major challenge in the field of tissue engineering. Hitherto, studies on vascularization have demonstrated limited control of vascular network geometry, such as vasculature direction and network density. An open vascular lumen is crucial to ensure that cells survive and that metabolic activity is fully functional in large‐sized tissues. Herein, a method based on high water‐dispersible collagen microfibers (CMF) to fabricate capillary orientation‐controllable 3D tissue with an open vascular lumen using a dispensing machine is reported. A twenty micrometers‐long CMF (CMF‐20) with high dispersion property are shown to be more effective for dispensing a homogenous tissue and inducing formation of an interconnected capillary network than two hundred micrometers‐long CMF (CMF‐200). One of the advantages is the prevention of shrinkage on the z‐axis of hydrogel‐based tissue which acts as a microscaffold. The gaps between the fibers can support endothelial cell migration and maturation, thus forming a larger vascular lumen compared to CMF‐free controls. Besides, shear forces produced by the dispensing process cause the collagen microfibers to align, and these microfibers guide cell alignment by integrin‐induced adhesion. The findings based on CMF to allow blood capillary alignment and vascular lumen stabilization will be an important technology in tissue engineering

Collagen Microfibers Induce Blood Capillary Orientation and Open Vascular Lumen

Achieving vascularization of engineered tissues or structures is a major challenge in the field of tissue engineering. Hitherto, studies on vascularization have demonstrated limited control of vascular network geometry, such as vasculature direction and network density. An open vascular lumen is crucial to ensure that cells survive and that metabolic activity is fully functional in large‐sized tissues. Herein, a method based on high water‐dispersible collagen microfibers (CMF) to fabricate capillary orientation‐controllable 3D tissue with an open vascular lumen using a dispensing machine is reported. A twenty micrometers‐long CMF (CMF‐20) with high dispersion property are shown to be more effective for dispensing a homogenous tissue and inducing formation of an interconnected capillary network than two hundred micrometers‐long CMF (CMF‐200). One of the advantages is the prevention of shrinkage on the z‐axis of hydrogel‐based tissue which acts as a microscaffold. The gaps between the fibers can support endothelial cell migration and maturation, thus forming a larger vascular lumen compared to CMF‐free controls. Besides, shear forces produced by the dispensing process cause the collagen microfibers to align, and these microfibers guide cell alignment by integrin‐induced adhesion. The findings based on CMF to allow blood capillary alignment and vascular lumen stabilization will be an important technology in tissue engineering

Bioprinted Vascularized Mature Adipose Tissue with Collagen Microfibers for Soft Tissue Regeneration

The development of soft tissue regeneration has recently gained importance due to safety concerns about artificial breast implants. Current autologous fat graft implantations can result in up to 90% of volume loss in long-term outcomes due to their limited revascularization. Adipose tissue has a highly vascularized structure which enables its proper homeostasis as well as its endocrine function. Mature adipocytes surrounded by a dense vascular network are the specific features required for efficient regeneration of the adipose tissue to perform host anastomosis after its implantation. Recently, bioprinting has been introduced as a promising solution to recreate in vitro this architecture in large-scale tissues. However, the in vitro induction of both the angiogenesis and adipogenesis differentiations from stem cells yields limited maturation states for these two pathways. To overcome these issues, we report a novel method for obtaining a fully vascularized adipose tissue reconstruction using supporting bath bioprinting. For the first time, directly isolated mature adipocytes encapsulated in a bioink containing physiological collagen microfibers (CMF) were bioprinted in a gellan gum supporting bath. These multilayered bioprinted tissues retained high viability even after 7 days of culture. Moreover, the functionality was also confirmed by the maintenance of fatty acid uptake from mature adipocytes. Therefore, this method of constructing fully functional adipose tissue regeneration holds promise for future clinical applications