www.medenosrce.net/arhimed   arhimed(a-t>medenosrce.net   [23/04/2026 12:15:25] http://www.medenosrce.net/arhimed/poglej.asp?id=155 MORFOLOŠKA ANALIZA PRESADKOV STROMALNIH MATICNIH CELIC KOSTNEGA MOZGA V MOŽGANIH PODGANE Avtor: Jure Zupan, Lojze Šmid ml. Mentor: doc. dr. Mara Bresjanac Somentor: prof. dr. Borut Peterlin IZHODIŠCE: Izvorne celice imajo zaradi svoje zmožnosti diferenciacije v razlicne celicne vrste velik terapevtski potencial, saj bi lahko v prihodnosti z njimi nadomestili bolezensko okvarjena ali poškodovana tkiva. Poleg zarodnih poznamo tudi odrasle izvorne celice, med katere štejemo tudi stromalne celice kostnega mozga (SCKM). Uporabnost SCKM v zdravljenju sloni na njihovi dostopnosti, enostavnem gojenju in vitro, možnosti avtologne presaditve brez nevarnosti zavrnitve in sposobnosti diferenciacije v razlicne celicne vrste. NAMEN: Namen naše raziskave je bil preveriti (1) preživetje presajenih cloveških SCKM, njihovo (2) prostorsko razporejanje in (3) diferenciacijo v smer celic živcevja po 28 dneh od presaditve v podganji striatum. V raziskavi smo primerjali usodo SCKM, presajenih v vnet možganski parenhim s presadki, vstavljenimi v nevneto tkivo kontrolnih podgan. HIPOTEZI: I. V podganji striatum presajene cloveške SCKM preživijo cas opazovanja, migrirajo skozi prejemnikove možgane in se diferencirajo v smer celic živcevja. II. Vnetje na mestu presadka izboljša preživetje, migracijo in diferenciacijo presajenih SCKM. METODE: Samice podganjega soja Wistar smo razdelili v pet skupin (n1-5 = 6): A - presadek SCKM v vnet striatum, B - presadek SCKM v nevnet striatum, C - okvara striatuma brez presadka, C - presadek dvojno oznacenih SCKM in D - presadek mrtvih SCKM. Po 28 dneh smo iste živali v globoki anesteziji žrtvovali, možgane izolirali, narezali 20 mcm debele koronalne rezine, ki smo jih oznacevali z imunohistokemijskimi (IHK) in imunofluorescentnimi (IF) metodami na antigene, znacilne za razlicne vrste celic živcevja, ter pod mikroskopom analizirali preživetje presajenih SCKM, njihovo migracijo in diferenciacijo. Rezultate smo statisticno analizirali z eksaktnim permutacijskim testom z Bonferonijevo korekcijo in eksaktno permutacijsko analizo variance. REZULTATI: Vse živali so opazovalno obdobje preživele. Presadke smo našli v striatumih vseh prejemnikov, ocenjeno najmanjše število preživelih celic (skupina A 1777 ± 957, skupina B 2096 ± 360, kar znaša 16% oz. 22% vsajenih celic) se med skupinama ni znacilno razlikovalo (p = 0,201). Povprecna mediolateralna oddaljenost celic od mesta presaditve je znašala v skupini A 83 ± 30 mcm in se je statisticno pomembno (p = 0,019) razlikovala od skupine B, kjer je bila 56 ± 13 mcm. Povprecna rostrokavdalna migracija v skupini A je bila 80 ± 30 mcm, v skupini B pa 70 ± 45 mcm (p = 0,72). Živali skupin, ki so prejele kvinolinsko kislino, so imele mocneje izražen vnetni in glijalni odziv v striatumu od ostalih skupin (p < 0,01). Pomembnega števila diferenciranih presajenih celic nismo našli v možganih nobenega prejemnika. ZAKLJUCKI: Nobene hipoteze nismo v celoti potrdili. V naših poskusnih pogojih so številne cloveške SCKM preživele 28 dni v podganjih možganih in obsežno migrirale, nismo pa ugotovili pomembne diferenciacije SCKM v celice živcevja. Vnetje na mestu presadka ni povzrocilo pomembnih razlik v preživetju in diferenciaciji SCKM, ugotovili pa smo pomembno zvecanje migracije SCKM v primerjavi s kontrolno skupino. «» [Abstract / English version] MORFOLOŠKA ANALIZA PRESADKOV STROMALNIH MATICNIH CELIC KOSTNEGA MOZGA V MOŽGANIH PODGANE Author: Jure Zupan, Lojze Šmid ml. Mentor: doc. dr. Mara Bresjanac Co-mentor: prof. dr. Borut Peterlin BACKGROUND: Stem cells have a great therapeutic potential for future replacement of diseased or damaged tissue, due to their ability to differentiate into different cell lines. Beside embryonic it is known that there are also adult stem cells, which include bone marrow stromal cells (BMSC). The therapeutic potential of BMSC is due to their characteristics: they are easily accessible, easily grown in vitro, their low immunogenicity allows autologous transplantation without a danger of rejection potential, and they can differentiate into different cell types. AIM: The aim of our study was to evaluate (1) the survival of grafted human BMSC, their (2) distribution and (3) differentiation into neural tissue cell types 28 days after grafting into rat striatum. In the study we compared the outcome of BMSC grafting into the previously inflamed brain parenchyma to their grafting into the non-inflamed brains of the control group. HYPOTHESES: I. Human BMSC grafted into the rat striatum survive the observation period, they migrate through the recipient brain and differentiate into different neural tissue cell types. II. Inflammation at the site of graft placement enhances survival, migration and differentiation of grafted BMSC. METHODS: Female albino Wistar rats were divided into 5 groups (n1-5=6): A - BMSC grafted into inflamed striatum, B - BMSC grafted into intact striatum, C - striatal inflammation without a graft, C -double labelled BMSC graft, D - dead BMSC graft. The rats were euthanized 28 days post-grafting in deep anaesthesia, their brains were removed, cut into 20 mcm thick coronal sections, and processed with imunohistochemical (IHC) and imunofluorescent (IF) methods to reveal various neural cell antigens. Using a microscope we analyzed survival of grafted BMSC, their migration and differentiation. Results were statistically analyzed using an exact permutation test with Bonferoni's correction, and an exact permutation analysis of variance. RESULTS: All animals survived the evaluation period. Grafts were found in the striata of all grafted animals. Analysis of graft survival did not reveal any statistically significant (p=0.201) difference between groups (group A 1777 ± 957, group B 2096 ± 360; which accounts for 16% and 22% of transplanted cells, respectively). Mean mediolateral cell migration showed statistically significant difference and was 83 ± 30 mcm in group A and in 56 ± 13 mcm in group B (p=0.019). Mean rostrocaudal migration did not show any statistically significant differences between group A (80 ± 30 mcm) and group B (70 ± 45 mcm; p=0.72). Animals in the lesioned group had a more pronounced inflammatory and glial response compared to the other groups (p<0.01). We did not find a significant number of differentiated transplanted cells in any of the recipients' brains. CONCLUSIONS: None of the hypothesis was confirmed in its entirety. Grafted human BMSC survived the evaluation period of 28 days in great numbers; they migrated extensively but failed to differentiate into neural tissue cell types. Inflammation at the site of graft placement did not enhance BMSC survival or differentiation, but a significant increase in BMSC migration was found compared to the control group.