Adhesion Characteristics of Human Pancreatic Islets, Duct Epithelial Cells, and Acinar Cells to a Polymer Scaffold

Nakashima, Yoshiki; Iguchi, Hiroki; Takakura, Kenta; Nakamura, Yuta; Izumi, Kenji; Koba, Naoya; Haneda, Satoshi; Tsukahara, Masayoshi

doi:10.1177/09636897221120500

Cited by 3 publications

(1 citation statement)

References 68 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Real-time PCR was performed using a StepOnePlus system (Life Technologies, Carlsbad, CA, USA). Luna Universal qPCR Master Mix (New England Biolabs Inc., Ipswich, MA, USA) was used according to the manufacturer's instructions 74,75 . The PCR protocol was as follows: (1) initial denaturation at 95°C for 10 min; For the analysis of real-time PCR data using a TaqMan™ Array96-Well FAST Plate, 18S, GAPDH, HPRT1, and GUSB were used as housekeeping genes.…”

Section: Cell Differentiation Assaysmentioning

confidence: 99%

Human induced pluripotent stem cell culture on a non-coated dish using medium supplemented with milk fat globule epidermal growth factor 8

Nakashima

Tsukahara

2023

Preprint

Self Cite

View full text Add to dashboard Cite

We propose a method of conditioning medium for non-coated dishes of human induced pluripotent stem cells (hiPSCs) by adding milk fat globule epidermal growth factor 8 (MFG-E8) (1-2 μg/mL) and 5%-10% gelatin solution (1.0 mg/mL) to medium (MFGE8-G). hiPSCs cultured in MFGE8-G StemFit Ak03 and CTS Essential 8 Medium showed cell adhesion. In a multiomics analysis, hiPSCs cultured in MFGE8-G showed an elevated expression of integrins, especially integrin α5β1. The expression of polo-like kinase 1 (PLK1), a cell cycle-regulating serine/threonine kinase, and mitogen-activated protein kinase kinase was increased in hiPSCs cultured on iMatrix-511. An mRNA expression analysis of histone methyltransferases and demethyltransferases revealed that hiPSCs cultured in MFGE8-G had an increased expression of SET And MYND Domain Containing 2, a methyltransferase for H3H4 and H3K36, and EZH2, a methyltransferase for H3K27. Conversely, the expression of H3K9, H3K20, and H3K36 demethylases was generally higher in hiPSCs cultured on iMatrix-511. A master regulation analysis revealed that integrin α5β1, fibroblast growth factor receptor, and protein phosphatase 1 were activated in hiPSCs cultured in MFGE8-G. The hiPSCs cultured on iMatrix-511 showed upregulated Mitogen-activated protein kinase 2 and the DNA checkpoint protein Cdc25. hiPSCs cultured on MFGE8-G demonstrated integrin-mediated cell adhesion of MFG-E8 with gelatin.

show abstract

Section: Cell Differentiation Assaysmentioning

confidence: 99%

Human induced pluripotent stem cell culture on a non-coated dish using medium supplemented with milk fat globule epidermal growth factor 8

Nakashima

Tsukahara

2023

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

ECM Proteins Nidogen‐1 and Decorin Restore Functionality of Human Islets of Langerhans upon Hypoxic Conditions

Jeyagaran,

Urbanczyk,

Carvajal‐Berrio

et al. 2024

Adv Healthcare Materials

View full text Add to dashboard Cite

Transplantation of donor islets of Langerhans is a potential therapeutic approach for patients with diabetes mellitus; however, its success is limited by islet death and dysfunction during the initial hypoxic conditions at the transplantation site. This highlights the need to support the donor islets in the days post‐transplantation until the site is vascularized. It was previously demonstrated that the extracellular matrix (ECM) proteins nidogen‐1 (NID1) and decorin (DCN) improve the functionality and survival of the β‐cell line, EndoC‐βH3, and the viability of human islets post‐isolation. To advance the use of these ECM proteins toward a clinical application and elucidate the mechanisms of action in primary islets, the study assesses the effects of ECM proteins NID1 and DCN on isolated human donor islets cultured in normoxic and hypoxic conditions. NID1‐ and DCN‐treatment restore β‐cell functionality of human donor islets in a hypoxic environment through upregulation of genes involved in glycolytic pathways and reducing DNA fragmentation in hypoxic conditions comparable to normoxic control islets. The results demonstrate that the utilization of NID1 or DCN with islets of Langerhans may have the potential to overcome the hypoxia‐induced cell death observed post‐transplantation and improve transplant outcomes.

show abstract

3D evaluation of the extracellular matrix of hypoxic pancreatic islets using light sheet fluorescence microscopy

Ramirez,

Bastien,

Chae

et al. 2024

Islets

View full text Add to dashboard Cite

Pancreatic islet transplantation is a promising treatment for type 1 diabetes, but the survival and function of transplanted islets are hindered by the loss of extracellular matrix (ECM) during islet isolation and by low oxygenation upon implantation. This study aimed to evaluate the impact of hypoxia on ECM using a cutting-edge imaging approach based on tissue clearing and 3D microscopy. Human and rat islets were cultured under normoxic (O 2 21%) or hypoxic (O 2 1%) conditions. Immunofluorescence staining targeting insulin, glucagon, CA9 (a hypoxia marker), ECM proteins (collagen 4, fibronectin, laminin), and E-cadherin (intercellular adhesion protein) was performed on fixed whole islets. The cleared islets were imaged using Light Sheet Fluorescence Microscopy (LSFM) and digitally analyzed. The volumetric analysis of target proteins did not show significant differences in abundance between the experimental groups. However, 3D projections revealed distinct morphological features that differentiated normoxic and hypoxic islets. Under normoxic conditions, ECM could be found throughout the islets. Hypoxic islets exhibited areas of scattered nuclei and central clusters of ECM proteins, indicating central necrosis. E-cadherin was absent in these areas. Our results, demonstrating a diminution of islets’ functional mass in hypoxia, align with the functional decline observed in transplanted islets experiencing low oxygenation after grafting. This study provides a methodology combining tissue clearing, multiplex immunofluorescence, Light Sheet Fluorescence Microscopy, and digital image analysis to investigate pancreatic islet morphology. This 3D approach allowed us to highlight ECM organizational changes during hypoxia from a morphological perspective.

show abstract

Adhesion Characteristics of Human Pancreatic Islets, Duct Epithelial Cells, and Acinar Cells to a Polymer Scaffold

Cited by 3 publications

References 68 publications

Human induced pluripotent stem cell culture on a non-coated dish using medium supplemented with milk fat globule epidermal growth factor 8

Human induced pluripotent stem cell culture on a non-coated dish using medium supplemented with milk fat globule epidermal growth factor 8

ECM Proteins Nidogen‐1 and Decorin Restore Functionality of Human Islets of Langerhans upon Hypoxic Conditions

3D evaluation of the extracellular matrix of hypoxic pancreatic islets using light sheet fluorescence microscopy

Contact Info

Product

Resources

About