Thesis (MMedSc )--University of Stellenbosch, 2011. / ENGLISH ABSTRACT: The present study investigated in particularly the specific nature of the supporting stromal layer
located between the implanted human cyst and host murine tissue, which has yet to be reported.
During an initial phase of this study, the particular light microscopic properties of the existing
hematoxylin and eosin (H&E) stained experimental cyst was investigated, with regards to the
presence or absence of specific morphological features, namely spongiosis, exocytosis, epithelial
keratinization, epithelial thickness and hyperplasia, and the vascularity and fibrosis present in the
stroma of these experimental sections. Subsequent analysis reported significant spongiosis, in
addition to increased exocytosis of immune cells and epithelial keratinization in a number of
cysts. Additionally, increased epithelial thickness and hyperplasia was reported in only 2 / 10
experimental tissues, whereas increased vascularity was observed in the stroma following
analysis of H&E and Special staining, such as Verhoeff-von Gieson and Masson trichrome
results.
During the second phase of the study, immunohistochemical analysis with a particularly wide
array of antibodies raised against specific human and mouse antigens had been applied. This
involved automated immunohistochemical staining with mouse anti-human primary antibodies,
in addition to manual staining with rabbit anti-mouse primary antibodies. Subsequent
visualization was achieved by means of linking to biotinylated secondary antibodies, and
Streptavidin-HRP incubation for standard visualization, followed by counterstaining with
Hematoxylin.
Maintained positive expression of cytokeratins 5, 13, and 14 was demonstrated in both control
human vaginal mucosa and experimental cysts, whereas similar findings were not reported for
cytokeratin 1, given the vast keratinization which was observed. Human collagen type IV and
laminin of the basement membrane reported positive expression in 9 / 10 and 6 / 10 control
human vaginal mucosa tissues respectively. In comparison, negative mouse collagen type IV and
laminin was reported in most experimental cysts compared to positive staining in positive control
mouse tissues.
Immunohistochemical staining for human elastin, fibronectin, von Willebrand factor, and
fibroblasts revealed maintained positive staining in all control human vaginal mucosa and
experimental cysts. However, maintained expression of CD34 (endothelial marker), CD1a
(langerhans cells), and human VEGFR-3 in experimental cysts was not demonstrated, compared
to positive expression in control human vaginal mucosa.
Subsequent analysis of murine antigens illustrated uniformly negative staining for mouse
fibronectin, langerhans cells (CD207), and fibroblasts, in addition to negative staining in positive
control mouse tissue sections. Furthermore, negative staining for mouse VEGFR-2 was reported
in all experimental cysts; however strong positive staining of this marker in mouse kidney tissue
had been reported.
The findings of this study suggested that the exact nature of the stromal layer is of both human
and murine origin. Furthermore, the tissue region located beneath the human vaginal epithelium
is suggested to be of human nature, whereas the second distinct region located at the periphery of
experimental cyst tissues, is suggested to be murine origin; however the findings of
immunohistochemical analysis could not illustrate definitively the exact nature of the
intermediate stromal layer, but could in fact demonstrate a mixture of human and murine tissue. / AFRIKAANSE OPSOMMING: Die huidige studie het die spesifieke molekulêre en histologiese eienskappe van die stromale laag
geleë tussen menslike sist- en muis velweefsel bestudeer, wat tans nog nie bekend is nie.
Gedurende die eerste fase van hierdie studie is die besondere lig-mikroskopiese eienskappe van
die bestaande hematoksilien en eosien (H&E) eksperimentele siste bestudeer, met betrekking tot
die aan- of afwesigheid van spesifieke morfologiese eienskappe, naamlik spongiose, eksositose
van immuunselle, epiteel keratinisasie, epiteel dikte en hiperplasie, en laastens die stromale
vaskulariteit en fibrose. Gevolglike analise het daarop gedui dat beduidende spongiose,
eksositose en epiteel keratinisasie gevind word in die eksperimentele siste in vergelyking met
kontrole vaginal weefsel. Hierteenoor is die verdikking van die epiteel en hiperplasie in slegs 2 /
10 eksperimentele siste gevind, terwyl vermeerderde vaskulariteit aangedui is na gevolglike
H&E en spesiale (soos byvoorbeeld Verhoeff-von Gieson en Masson trichrome)
kleuringsresultate.
Die tweede fase van die studie het die immunokleuring met verskeie mens- en muis spesifieke
antiliggame behels, waarby die uitdrukking van verskeie mens antigene vergelyk is met dié van
muis. As sulks is ge-automatiseerde immunohistochemie toegepas met muis primêre
antiliggame, tesame met fisiese kleuring met konyn primêre antiliggame toegepas. Gevolglike
visualisasie is aangedui deur middel van binding met sekondêre antiliggaam en Streptavidin-
HRP, gevolg deur teenkleuring met Hematoksilien.
Algehele behoud van positiewe uitdrukking van sitokeratien 5, 13, en 14 is bevind, terwyl
sitokeratien 1 uitdrukking nie daarwerklik vergelykbaar is met dié van kontrole mens vaginale
weefsel nie. Die uitdrukking van mens kollageen IV en laminien van die basaal membraan is
verder bestudeer, en het egter positiewe kleuring in 9 / 10 en 6 / 10 van kontrole mens vaginale
mukosa aangedui. In vergelykking hiermee kon die huidige bevindings egter net positiewe
kleuring in 4 / 10 en 3 / 10 eksperimentele siste vir kollageen IV en laminien onderskeidelik,
illustreer.
Immunohistochemiese analise van menslike elastien, fibronektien, von Willebrand (vW) faktor
en fibroblaste het op deurgaans positiewe uitdrukking van hierdie merkers aangedui in beide
eksperimentele en kontrole menslike weefsel. In teenstelling hiermee is volgehoue uitdrukking
van CD34 (endoteel merker), CD1a (Langerhans sel merker) en mens VEGFR-3 in
ekperimentele siste egter nie illustreerbaar nie, in vergelykking met deurgaans positiewe
uitdrukking van hierdie antigene in kontrole mens vaginale mukosa.
In opvolging is deurgaans negatiewe uitdrukking van muis fibronektien, langerhans sel (CD207)
en fibroblaste bevestig, terwyl negatiewe kleuring ook deurgaans in positiwe kontrole muis
weefsel, bekom deur die disseksie van ‘n naakte muis, gevind is. Verder is ook negatiewe
kleuring vir VEGFR-2 in alle eksperimentele siste gevind, terwyl egter sterk positiewe kleuring
in muis nierweefsel as positiewe weefsel gevind is.
Die resultate van die huidige studie het daarop gedui dat die stromale laag onderliggend tot mens
vaginale epiteel van menslike oorsprong is, terwyl die periferale stroma onderliggend tot muis
velweefsel, ongetwyfeld van muis oorsprong is. Laastens kon die spesifieke oorsprong van die
tussenliggende stroma nie aangedui word nie, maar dat dit moontlik uit beide menslike- en
muisweefsel bestaan.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/6879 |
| Date | 03 1900 |
| Creators | Kok, Cornelius Wilhelmus |
| Contributors | Schneider, J. W., Hille, J. J., University of Stellenbosch. Faculty of Health Sciences. Dept. of Pathology. Anatomical Pathology. |
| Publisher | Stellenbosch : University of Stellenbosch |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | xxiv, 160 p. : ill. |
| Rights | Stellenbosch University |
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