ABSTRACT
Pancreatic ß cells are responsible for insulin secretion and are important for glucose regulation in a healthy body and diabetic disease patient without prelabeling of islets. While the conventional biomarkers for diabetes have been glucose and insulin concentrations in the blood, the direct determination of the pancreatic ß cell mass would provide critical information for the disease status and progression. By combining fluorination and diversity-oriented fluorescence library strategy, we have developed a multimodal pancreatic ß cell probe PiF for both fluorescence and for PET (positron emission tomography). By simple tail vein injection, PiF stains pancreatic ß cells specifically and allows intraoperative fluorescent imaging of pancreatic islets. PiF-injected pancreatic tissue even facilitated an antibody-free islet analysis within 2 h, dramatically accelerating the day-long histological procedure without any fixing and dehydration step. Not only islets in the pancreas but also the low background of PiF in the liver allowed us to monitor the intraportal transplanted islets, which is the first in vivo visualization of transplanted human islets without a prelabeling of the islets. Finally, we could replace the built-in fluorine atom in PiF with radioactive 18F and successfully demonstrate in situ PET imaging for pancreatic islets.
Subject(s)
Fluorescent Dyes/chemistry , Insulin-Secreting Cells/cytology , Xanthenes/chemistry , Animals , Diabetes Mellitus, Experimental/pathology , Fluorescence , Fluorescent Dyes/chemical synthesis , Fluorescent Dyes/pharmacokinetics , Fluorescent Dyes/toxicity , Humans , Insulin-Secreting Cells/transplantation , Islets of Langerhans Transplantation , Liver/cytology , Mice, Inbred C57BL , Mice, Inbred ICR , Positron-Emission Tomography , Rats , Xanthenes/chemical synthesis , Xanthenes/pharmacokinetics , Xanthenes/toxicityABSTRACT
Selection of a specific neural stem/progenitor cells (NSPCs) has attracted broad attention in regenerative medicine for neurological disorders. Here, we report a fluorescent probe, CDg13, and its application for isolating strong neurogenic NSPCs. In comparison to the NSPCs isolated by other biomarkers, CDg13-stained NSPCs showed higher capability to differentiate into neurons. Target identification revealed that the fluorescence intensity of the probe within cells is inversely proportional to the expression levels of mouse and human Abcg2 transporters. These findings suggest that low Abcg2 expression is a biomarker for neurogenic NSPCs in mouse brain. Furthermore, CDg13 can be used to isolate Abcg2low cells from heterogeneous cell populations.
Subject(s)
Benzamides/chemistry , Fluorescent Dyes/chemistry , Neural Stem Cells/metabolism , Neurons/metabolism , Xanthenes/chemistry , ATP Binding Cassette Transporter, Subfamily G, Member 2/antagonists & inhibitors , ATP Binding Cassette Transporter, Subfamily G, Member 2/genetics , ATP Binding Cassette Transporter, Subfamily G, Member 2/metabolism , Animals , Benzamides/metabolism , Biomarkers/metabolism , Brain/cytology , Brain/metabolism , Cell Differentiation , Cells, Cultured , Embryo, Mammalian/cytology , Humans , Mice , Microscopy, Fluorescence , Neoplasm Proteins/antagonists & inhibitors , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Neural Stem Cells/chemistry , Neural Stem Cells/cytology , Neurons/cytology , Propionates/chemistry , Propionates/metabolism , Quinolines/chemistry , Quinolines/metabolism , RNA Interference , RNA, Small Interfering/metabolism , Xanthenes/metabolismABSTRACT
Two-photon (TP) microscopy has an advantage for live tissue imaging which allows a deeper tissue penetration up to 1 mm comparing to one-photon (OP) microscopy. While there are several OP fluorescence probes in use for pancreatic islet imaging, TP imaging of selective cells in live islet still remains a challenge. Herein, we report the discovery of first TP live pancreatic islet imaging probe; TP-α (Two Photon-alpha) which can selectively stain glucagon secreting alpha cells. Through fluorescent image based screening using three pancreatic cell lines, we discovered TP-α from a TP fluorescent dye library TPG (TP-Green). In vitro fluorescence test showed that TP-α have direct interaction and appear glucagon with a significant fluorescence increase, but not with insulin or other hormones/analytes. Finally, TP-α was successfully applied for 3D imaging of live islets by staining alpha cell directly. The newly developed TP-α can be a practical tool to evaluate and identify live alpha cells in terms of localization, distribution and availability in the intact islets.