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Oulun yliopiston väitöskirjat

NOVEL CULTURE AND ORGANOID TECHNOLOGIES TO STUDY MAMMALIAN KIDNEY DEVELOPMENT, ACTA UNIVERSITATIS OULUENSIS D Medica 1454

ISBN-10:	978-952-62-1824-3
Kieli:	englanti
Kustantaja:	Oulun yliopisto
Oppiaine:	Lääketiede
Painos:	Osajulkaisuväitöskirjan yhteenveto-osa
Painosvuosi:	2018
Sijainti:	Print Tietotalo
Sivumäärä:	76
Tekijät:	SAARELA ULLA

15.00 €

Kidney diseases affect an increasing number of people worldwide, and there is a growing demand to develop new treatments and increase the number of transplantable organs. New treatments can be designed when new knowledge is gained by studying the details of kidney development. The ex vivo culture techniques have been used for over a century to study the development of kidneys, but they are not optimal for long-term imaging and following the nephrogenesis process over time. Kidney organoids, which are cellular aggregates resembling the in vivo kidney, together with intact embryonic kidneys, present a platform for these studies. However, there are limitations when working with primary embryonic kidney cells. Primary embryonic metanephric mesenchymal cells are usually low in number and lose the ability to undergo nephrogenesis rapidly. New ways to culture, biobank, and transfect cells can offer ways for functional testing of the effects of different genes on the nephrogenesis. This study presents new tools for studying nephrogenesis. Time-lapse imaging of organ development may be enhanced by using a Fixed Z-direction (FiZD) culture system where the kidney explant is grown in a restricted 70μm space. The technique enables the segmentation of the individual cells in a two-dimensional image and a dynamic analysis of the time-lapse data. This study also presents a technique of dissociation and reaggregation of the uninduced kidney metanephric mesenchyme (MM). With this novel method of culturing the dissociated MM cells in a growth factor medium for 24 hours, the cells can keep their competence for nephrogenesis. This technique allows the genetic manipulation of the MM cells before the induction to form nephrons, allowing functional testing of genes in the metanephric mesenchyme. This study further presents different techniques for gene editing of MM cells and introduces biobanking of primary kidney cells. It is shown here that the MM and ureteric bud (UB) cells have the capability to remember their fates and build nephron-like structures or continue branching after the cryopreservation in the liquid nitrogen. The methods introduced here provide new ways to create kidney organoids, manipulate their genome, and biobank the primary embryonic kidney cells. The developed FiZD culture system enhances the imaging of kidney development compared to the previously used culture methods. Using this method, the morphogenesis of the developing kidney can be followed more precisely, even in a single cell level. This culture method may also be used to culturing other organs, such as ovary, and may help provide insights into the development of other tissues as well.

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