<p>Leydig-ove ćelije testisa su primarno mesto sinteze muških polnih hormona. Ovi hormoni su neophodani za reproduktivno, ali i za opšte zdravlje budući da su<br />ozbiljni zdravstveni problemi često povezani sa njihovom smanjenom produkcijom. Insulin i insulinu sličan faktor rasta 1, IGF1 <em>(engl.</em> insulin like growth factor 1), i<br />signalizacija koju pokreću preko svojih receptora (INSR i IGF1R), su jedan od ključnih faktora koji regulišu specifični razvoj tkiva, pa i samih gonada. Ipak, uloga i<br />mehanizmi delovanja ovih receptora u steroidogenim tkivima nisu u potpunosti poznati. Stoga je istraživanje uokviru ove doktorske disertacije koncipirano sa ciljem da se, na modelu prepubertalnih (P21) i adultnih (P80) mužjaka miševa sa kondicionalnom delecijom<em> Insr </em>i <em>Igf1</em>r gena u steroidogenim ćelijama (Insr/Igf1r-DKO), definiše uloga INSR i IGF1R u regulisanju diferencijacije i steroidogene funkcije Leydig-ovih ćelija. Pored toga, mužjaci i ženke P21 miševa sa istom delecijom su korišćeni za praćenje ekspresije glavnih markera mitohondrijalne biogeneze i fuzije/arhitekture u Leydigovim ćelijama, ovarijumima i nadbubrežnim žlezdama. Rezultati su potvrdili da delecija Insr i Igf1r u<br />steroidogenim tkivima utiče na diferencijaciju i funkcionalne karakteristike Leydig-ovih ćelija P21 i P80 miševa, upućujući na pojavu tzv. „feminizacije“. Broj<br />Leydig-ovih ćelija izolovanih iz P21 i P80 Insr/Igf1rDKO miševa bio je smanjen, a morfologija i ultrastruktura ovih ćelija izmenjene kod P21 Insr/Igf1rDKO miševa. Steroidogeni kapacitet i aktivnost, kao i ekspresija glavnih elemenata steroidogene mašinerije <em>(Lhcgr, Star, Cyp11a1, Cyp17a1, Hsd3b1 i 6, Hsd17b3,</em><br /><em>Sf</em>1) bili su smanjeni u Leydig-ovim ćelijama P21 i P80 <em>Insr/Igf1</em>r-DKO miševa, dok je ekspresija transkripcionih represora steroidogeneze (Arr19 i Dax1) bila povećana specifično u istim ćelijama, ali ne i u ostatku testisa.<br />Transkripcioni profil markera muškog pola (<em>Sry, Sox9, Amh</em>) bio je izmenjen u Leydig-ovim ćelijama P21 i P80 <em>Insr/Igf1r</em>-DKO miševa. Transkripcija markera ženskog pola (<em>Rspo1, Wnt4</em>) u testisima, kao i ekspresija Cyp19a1 i produkcija estradiola (E2) u Leydig-ovim ćelijama, P21 i P80 <em> Insr/Igf1r</em>-DKO miševa bile su povećane. Transkripcija markera mitohondrijalne biogenze (<em>Ppargc1a, Tfam</em>, <em>Mtnd1</em>) bila je smanjena u Leydigovim ćelijama P21 <em>Insr/Igf1r</em>-DKO miševa, dok supromene ekspresije izostale u ovarijumima ženki istog genotipa. Isti markeri su bili povećani u nabdubrežnim žlezdama oba pola. Markeri mitohondrijalne fuzije/arhitekture (<em>Mfn1 i Mfn2)</em> bili su povećani u Leydig-ovim ćelijama P21 <em>Insr/Igf1r</em>-DKO miševa, što je praćeno i narušenom mitohondrijalnom fazom steroidogeneze (produkcija progesterona), kao i brojem i morfologijom ovim organela. Ekspresija istih markera u ovarijumima bila je nepromenjena. Sumirano, rezultati ovog istraživanja su pokazali da su INSR i IGF1R važni za diferencijaciju i steroidogenu funkciju Leydig-ovih ćelija P21 i P80 miševa. Takođe, ovi receptori su važni regulatori markera mitohondrijalne biogeneze i fuzije/arhiteture u steroidogenim ćelijama muških gonada P21 miševa, ali ne i u steroidogenim ćelijama ovarijuma. </p> / <p>Leydig cells of testes are the primary site of the male sex hormones synthesis. These hormones are indispensable for both reproductive and general health since serious health problems are often associated with their reduced production. Insulin and insulin-like growth factor 1, IGF1 (insulin like growth factor 1), and signaling triggered through their receptors (INSR and IGF1R), are one of the key factors that regulate specific development of tissue including gonads. However, the role and mechanisms of these receptors action in steroidogenic tissues are not known enough. This study was designed to observe the role of INSR and IGF1R in regulating the differentiation and steroidogenic function of Leydig cells by using the model of prepubertal (P21) and adult (P80) male mice with the conditional deletion of the Insr and Igf1r genes in steroidogenic cells (<em>Insr/Igf1r-</em>DKO). In addition, male and female P21 mice with the samedeletion were used to monitor the expression of the main markers of mitochondrial biogenesis and fusion/architecture in Leydig cells, ovaries and adrenal glands. The results confirmed that deletion of <em> Insr</em> and<em> Igf1r </em> in steroidogenic tissues influences differentiation and functional characteristics of Leydig cells isolated from P21 and P80 mice, suggesting an appearance of "feminization". The number of Leydig cells isolated from both P21 and P80 <em>Insr/Igf1</em>r-DKO mice was reduced. Morphology and ultrastructure of Leydig cells were disturbed in P21 <em>Insr/Igf1r-</em>DKO mice. Steroidogenic capacity and activity, as well as expression of the main elements of steroidogenic machinery (<em>Lhcgr, Star, Cyp11a1, Cyp17a1, Hsd3b1 and 6, Hsd17b3, Sf1) </em>were decreased in Leydig cells from P21 and P80 I<em>nsr/Igf1</em>r-DKO mice, while the expression of transcriptional repressors of steroidogenesis (<em>Arr19</em> and <em>Dax1) </em>was increased in the same cells, but not in the rest of the testes. Transcription profile of the male sex markers (<em>Sry, Sox9</em>, <em>Amh</em>) was altered in Leydig cells from P21 and P80 <em>Insr/Igf1</em>r-DKO mice. Transcription of the female sex markers (<em>Rspo1, Wnt4</em>) in the testes, as well as <em>Cyp19a1 </em>expression and estradiol (E2) production in Leydig cells, from P21 and P80 I<em>nsr/Igf1</em>rDKO mice were increased. Transcription of mitochondrial biogenesis markers (<em>Ppargc1a, Tfam, Mtnd1</em>) was declined in Leydig cells from P21<em> Insr/Igf1r-</em>DKO mice, while changes were absent in the ovaries of the same genotype. Transcription of the same markers was increased in the adrenal glands of both sexes. The mitochondrial fusion/architecture markers (<em>Mfn1</em> and <em>Mfn2</em>) were increased in Leydig cells from<em> Insr/Igf1r</em>-DKO mice and followed by disturbedmitochondrial phase of steroidogenesis (progesterone production), as well as decreased number and disturbed morphology of mitochondria. Expression of the same markers in the ovaries was unchanged. In summary, results of this study showed that INSR and IGF1R are important in differentiation and steroidogenic function of Leydig cells from P21 and P80 mice. Also, these receptors are important regulators of mitochondrial biogenesis and fusion/architecture markers in steroidogenic cells of P21 male mice, but not in steroidogenic cells of ovaries.</p>
| Identifer | oai:union.ndltd.org:uns.ac.rs/oai:CRISUNS:(BISIS)110282 |
| Date | 31 May 2019 |
| Creators | Radović Sava |
| Contributors | Andrić Silvana, Kostić Tatjana, Matić Gordana, Kojić Zvezdana, Kaišarević Sonja |
| Publisher | Univerzitet u Novom Sadu, Prirodno-matematički fakultet u Novom Sadu, University of Novi Sad, Faculty of Sciences at Novi Sad |
| Source Sets | University of Novi Sad |
| Language | Serbian |
| Detected Language | English |
| Type | PhD thesis |
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