Title Značaj izražaja CRKL, AIFM3 i UBASH3A tijekom razvoja humanog bubrega Title (english) Expession of CRKL, AIFM3 and UBASH3A genes during human kidney development Author Mirela Lozić Mentor Katarina Vukojević (mentor)Committee member Merica Glavina Durdov (predsjednik povjerenstva)Committee member Tatijana Zemunik (član povjerenstva)Committee member Ivica Grković (član povjerenstva)Granter University of Split Defense date and country 2022, Croatia Scientific / art field, BIOMEDICINE AND HEALTHCARE Universal decimal classificationUDC ) 61 - Medical sciences Abstract CAKUT je skraćenica za kongenitalne anomalije bubrega i urološkog trakta. U ovoj retrospektivnoj studiji istražen je mRNA transkript i prostorno-vremenski obrazac izražaja proteinskog produkta tri CAKUT kandidat gena, CRKL, UBASH3A i AIFM3 na uzorcima humanog bubrega u razvoju i uspoređeni s obrascem izražaja u CAKUT-u i podocitopatijama. Izražaj njihovih proteina analiziran je imunofluorescencijom na 14 bubrega u razvoju, 28 CAKUT-a i 10 podocitopatija, a transkripti mRNA RT-qPCR-a na uzorcima humanog bubrega u razvoju.
Izražaj AIFM3 bio je pretežito u tubulima, a manje u glomerulima i maksimalan u neonatalnom razdoblju (Ph4 1M), moguće zbog uloge AIFM3 u energetskom metabolizmu. Kolokalazacija AIFM3 i AIF je bila općenito slaba, najčešće nađena u Ph4 1M. Izražaj UBASH3A nađen je u svim razvojnim fazama u strukturama nefrona i stanicama tubula, a najveći u prenatalnoj Ph4 kad fetus najbrže raste. U nezrelim glomerulima UBASH3A je nađen u podocitima. CRKL je bio izražen rijetko, ali intenzivno u pojedinačnim stanicama, najviše u Ph3 kad se formira većina nefrona.
Istraživanjem prostornog izražaja AIFM3 i CRKL u bubrezima i mokraćovodima 28 ispitanika s CAKUT fenotipom, metodom imunofluroscencije nađen je pojačani izražaj AIFM3 u epitelu s prekomjernom proliferacijom i apoptozom. CRKL je imao isti točkasti obrazac izražaja u bubregu zahvaćenom CAKUT-om i kontroli pa se može pretpostaviti da normalno obavlja svoju ulogu u ispitanika s CAKUT-om. UBASH3A je bio izražen u ispitanika s DS i CAKUT fenotipom, ali različito od kontrole u intenzitetu izražaja, raspodjeli i količini. Mogući razlozi su veliki broj abnormalnih produkata gena i/ili apoptotski procesi zbog uloge UBASH3A u pojačavanju apoptoze ovisne o AIF-u. Za razliku od kontrole, u podocitopatijama je utvrđen negativan izražaj UBASH3A.
Dinamika i obrazac izražaja CRKL, UBASH3A i AIFM3 u različitim strukturama tijekom razvoja humanog bubrega razlikuju se od obrasca u bolesti i ukazuju na ulogu CRKL, UBASH3A i AIFM3 gena u nastanku CAKUT-a.
Abstract (english) CAKUT is an acronym for congenital anomalies of the kidney and urinary tract. The aim of this retrospective study was to analyze the mRNA transcript and the spatio-temporal pattern of expression of the protein products of the CRKL, UBASH3A and AIFM3 genes in developing human kidneys as candidate genes for CAKUT and to compare the normal pattern of their protein products with those in CAKUT and podocytopathies.
Immunofluorescent protein levels were analyzed in kidney tissue in 14 developing kidneys, 28 CAKUT and 10 podocytopathies, and mRNA transcripts by RT-qPCR in 14 developing kidneys. The results were analyzed using the GraphPad statistical program and p<0.05 was selected as the significant difference.
AIFM3 was found predominantly in tubules and less in glomeruli and was maximally expressed in the neonatal period (Ph4 1M), probably due to the role of AIFM3 in energy metabolism. Co-localization of AIFM3 and AIF was generally weak, most commonly found in Ph4 1M. UBASH3A expression was found in all stages and structures of the nephron, the highest in prenatal Ph4 when growth is fastest. In glomeruli, UBASH3A was found in podocytes. CRKL is rarely but intensely expressed in certain cells, mainly in Ph3, when most nephrons are formed.
In 28 patients with the CAKUT phenotype, increased expression of AIFM3 was found in the epithelium with excessive proliferation and apoptosis. CRKL had the same punctate nuclear pattern as in the control, so it can be assumed that it performs its role normally in patients with CAKUT. UBASH3A was expressed in a patient with DS and CAKUT phenotypes, but differently from the control regarding expression intensity, distribution and quantity. Possible reasons are a large number of abnormal gene products and/or apoptotic processes due to the role of UBASH3A in enhancing AIF-dependent apoptosis. In contrast to the control, a negative expression of UBASH3A was found in podocytopathies.
The dynamics of expression of CRKL, UBASH3A and AIFM3 in different structures in the development of the human kidney and the difference from the pathological pattern indicate their role in the development of CAKUT.
Keywords
CRKL
UBASH3A
AIFM3
bubreg
razvoj
Keywords (english)
CRKL
UBASH3A
AIFM3
kidney
development
Language croatian URN:NBN urn:nbn:hr:171:380248 Study programme Title: Biology of Neoplasms Type of resource Text File origin Born digital Access conditions Open access Terms of use Created on 2022-09-29 11:33:35
