Light Microscopy core for NIDDK

National Institute Of Diabetes And Digestive And Kidney Diseases

Investigators

Abstract

The mission of the National Institutes of Health (NIH) is to seek fundamental knowledge about the nature and behavior of living systems and to apply that knowledge to enhance health, lengthen life, and reduce illness and disability. As part of the NIH, the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) Intramural Research Program (IRP) conducts biomedical research and training related to diabetes mellitus; endocrine, bone, and metabolic diseases; digestive diseases, including liver diseases and nutritional disorders; and kidney, urologic, and hematologic diseases. Intramural research is conducted in the Institute's laboratories and clinical facilities in Bethesda, Maryland and in Phoenix, Arizona. There are currently 17 Laboratories and Branches (i.e. 17 departments with different focus areas) in the NIDDK IRP. Most if not all these areas of research, either occasionally or on an ongoing basis, require light microscopy (LM) methods to qualitatively complement other assays, or as the basis of a quantitative assay. The There are three main categories of LM. Conventional widefield, transmitted light techniques, such as darkfield or differential interference contrast (DIC), are still useful because of their innate ability to show cellular morphology. Second, an old standby, immunohistochemistry (IHC)-- using absorptive probes to colorize tissue in transmitted light. This technique has been used for decades (e.g. in pathology), but recent advances with new probes and color deconvolution have allowed the field to grow, although requiring more finesse when being quantitated. The third and most common category of LM is characterized by the use of fluorescent probes targeting biomolecules (specific proteins, DNA, etc) of interest, making them glow with different colors on a black background. Such fluorescent dyes can even be used to monitor subcellular events in real-time. There are many sub-categories of fluorescence microscopy, e.g. confocal, super-resolution, and TIRF (Total Internal Reflectance-induced Fluorescence). Each of these involve specialized technique in sample prep, image acquisition, and/or image analysis. Some NIDDK labs might be comfortable without advanced help on one of more of the techniques described above, and might also be able to set aside a budget for equipment they would use regularly; but to fill any gaps, the NIDDK Advanced Light Microscopy & Image Analysis Core (ALMIAC) remains a reliable resource for cutting-edge light microscopy equipment, software, and expertise. For the FY2021 budget year (October 2020 - September 2021), there were >40 researchers using ALMIAC resources, representing >20 Sections (individual labs), from at least 9 (out of 17) Laboratories and Branches. In other words, from a lab/branch perspective, >50% of NIDDK used ALMIAC resources. Out of the 40+ individual researchers helped, there were at least 9 NIDDK publications during FY 2021 that used images acquired and/or analyzed with the help of ALMIAC resources: .....Leah F. Rosin, Jose Gil Jr., Ines A. Drinnenberg, Elissa P. Lei. Oligopaint DNA FISH reveals telomere-based meiotic pairing dynamics in the silkworm, Bombyx mori. PLoS Genet. 2021 Jul 28;17(7):e1009700. https://doi.org/10.1371/journal.pgen.1009700. PMID: 34319984 .....Indira Bag, Shue Chen, Leah F. Rosin, Yang Chen, Chen-Yu Liu, Guo-Yun Yu & Elissa P. Lei. M1BP cooperates with CP190 to activate transcription at TAD borders and promote chromatin insulator activity. Nat Commun. 2021 Jul 7;12(1):4170. https://doi.org/10.1038/s41467-021-24407-y. PMID: 34234130 .....Sarju J. Patel, Olga Protchenko, Minoo Shakoury-Elizeh, Ethan Baratz, Shyamalagauri Jadhav, and Caroline C. Philpott. The iron chaperone and nucleic acid-binding activities of poly(rC)-binding protein 1 are separable and independently essential. Proc Natl Acad Sci U S A. 2021 Jun 22;118(25):e2104666118. https://doi.org/10.1073/pnas.2104666118. PMID: 34161287 .....Shanu Jain, Sai P Pydi, Young-Hwan Jung, Mirko Scortichini, Efrat L Kesner, Tadeusz P Karcz, Donald N Cook, Oksana Gavrilova, Jrgen Wess, Kenneth A Jacobson. Adipocyte P2Y14 receptors play a key role in regulating whole-body glucose and lipid homeostasis. JCI Insight. 2021 May 24;6(10):e146577. PMID: 34027896 .....Temesgen E. Andargie, Naoko Tsuji, Fayaz Seifuddin, Moon Kyoo Jang, Peter S.T. Yuen, Hyesik Kong, Ilker Tunc, Komudi Singh, Ananth Charya, Kenneth Wilkins, Steven Nathan, Andrea Cox, Mehdi Pirooznia, Robert A. Star, and Sean Agbor-Enoh. Cell-free DNA maps COVID-19 tissue injury and risk of death and can cause tissue injury. JCI Insight. 2021 Apr 8;6(7):e147610. https://doi.org/10.1172/jci.insight.147610. PMID: 33651717 .....Peng Wang & Yihong Ye. Filamentous recombinant human Tau activates primary astrocytes via an integrin receptor complex. Nat Commun. 2021 Jan 4;12(1):95. https://doi.org/10.1038/s41467-020-20322-w. PMID: 33398028 .....Shanu Jain, Sai P Pydi, Kiran S Toti, Bernard Robaye, Marco Idzko, Oksana Gavrilova , Jrgen Wess, Kenneth A Jacobson. Lack of adipocyte purinergic P2Y 6 receptor greatly improves whole body glucose homeostasis. Proc Natl Acad Sci U S A. 2020 Dec 1;117(48):30763-30774. PMID: 33199639 .....Qi Zhang, Catherine Zhengzheng Chen, Manju Swaroop, Miao Xu, Lihui Wang, Juhyung Lee, Amy Qiu Wang, Manisha Pradhan, Natalie Hagen, Lu Chen, Min Shen, Zhiji Luo, Xin Xu, Yue Xu, Wenwei Huang, Wei Zheng & Yihong Ye. Heparan sulfate assists SARS-CoV-2 in cell entry and can be targeted by approved drugs in vitro. Cell Discov. 2020 Nov 4;6(1):80. https://doi.org/10.1038/s41421-020-00222-5. PMID: 33298900 In addition to the specific research projects mentioned above, several areas of improvement in capability and workflow include: 1) Installation and deployment of new spinning disk confocal microscope with super-resolution capability, the Nikon CSU-W1 SoRa. 2) With the help of NIDDK CTB (Computer Technology Branch), continued work on the beta version of the NIDDK ALMIAC website. The website should be ready for full deployment (internally to NIDDK) later this calendar year. 3) Upgrades of 4 microscope computers and one workstation computer, to accommodate Windows 10 compatibility and bring hardware to current standards. 4) With the help of SIP students, additional written protocols for microscope use and image processing, including the use of denoising algorithms and stitching images using the freely available ImageJ/Fiji software.