Thesis (MSc)--Stellenbosch University, 2015. / ENGLISH ABSTRACT: In this study, the influence of rooibos on the catalytic activity of enzymes 17β -hydroxysteroid dehydrogenase type 3 (17βHSD3), 17β-hydroxysteroid dehydrogenase type 5 (AKR1C3),
17β-hydroxysteroid dehydrogenase type 2 (17βHSD2), 5α-reductase type 1 (SRD5A1) and
5α-reductase type 2 (SRD5A2), which catalyse prostate androgen metabolism, was investigated.
The activities of both 17βHSD3 and AKR1C3 heterologously expressed in CHO-K1 and HEK293
cells were inhibited significantly by rooibos, with rooibos reducing the conversion of
androstenedione (A4) and 11keto-androstenedione (11KA4) to testosterone (T) and 11ketotestosterone
(11KT), respectively. The catalytic activity of 17βHSD2 towards T, 11hydroxytestosterone
(11OHT) and 11KT was also significantly inhibited by rooibos in transiently
transfected HEK293 cells. In transiently transfected HEK293 cells rooibos did not inhibit SRD5A1
while the rate of T conversion to dihydrotestosterone (DHT) by SRD5A2 was decreased. Analysis
of steroid metabolism in PNT2 cells also suggests that rooibos does not modulate the catalytic
activity of endogenously expressed SRD5A towards A4, however, the conversion of T to DHT was
reduced. In addition, reductive 17βHSD activity towards A4 was inhibited in the presence of
rooibos in both PNT2 and BPH-1 cells. In contrast, the conversion of 11KA4 to 11KT was inhibited
in BPH-1, PC-3 and LNCaP cells, with negligible conversion of 11KA4 in PNT2 cells. Interestingly,
data suggests inhibition of 3α-hydroxysteroid dehydrogenase type 3 (AKR1C2) activity in the
production of androsterone (AST) from 5α–androstenedione (5α-dione), as well as the dehydrogenase reaction of T to A4 in PNT2 cells by rooibos. Androgen metabolism pathways were
subsequently investigated in LNCaP cells to determine androgen metabolism by endogenous
enzymes. Rooibos resulted in the reduced conversion of A4 in LNCaP cells to the same extent as
indomethacin, a known AKR1C3 inhibitor. Rooibos also modulated T, DHT and AST metabolism in
LNCaP cells. Furthermore, uridine diphosphate glucuronosyltransferase (UGT) activity in LNCaP
cells was inhibited by rooibos, decreasing T-, DHT– and AST-glucuronide formation. These data
prompted subsequent investigations into the influence of rooibos at cellular level, and prostatespecific
antigen (PSA) levels were assayed in the presence of rooibos. PSA was significantly
inhibited by rooibos in the absence and presence of DHT, suggesting possible interaction of
rooibos with the mutated androgen receptor (AR) or estrogen receptor-β (ERβ) expressed in
LNCaP cells.
Taken together, rooibos inhibited the catalytic activity of key enzymes that catalyse the activation
of androgens in the prostate, as well as inhibiting enzymes involved in the conjugation of
androgens. At cellular level, PSA levels were also decreased by rooibos, possibly through AR or
ERβ interactions – clearly indicating a modulatory role for rooibos in active androgen production. / AFRIKAANSE OPSOMMING: In hierdie studie was die invloed van rooibos ten opsigte van die katalitiese aktiwiteite van die
ensieme 17β-hidroksi-steroïed-dehidrogenase tipe 2, tipe 3 en tipe 5 (17βHSD2, 17βHSD3,
AKR1C3), asook 5α-reduktase tipe 1 en tipe 2 (SRD5A1, SRD5A2) ondersoek. Hierdie ensieme is
betrokke in die produksie van androgene in die prostaat. Rooibos het die katalitiese aktiwiteit van
17βHSD3 en AKR1C3 in CHO-K1 en HEK293 selle beïnvloed en het vermindere omskakeling van
androstenedioon (A4) en 11keto-androstenedioon (11KA4) na testosteroon (T) en 11-ketotestosteroon
(11KT), afsonderlik, veroorsaak. Die katalitiese aktiwiteit van 17βHSD2 teenoor T,
11-hidroksie-testosteroon (11OHT) en 11KT was ook beïnvloed in die teenwoordigheid van rooibos
in HEK293 selle. Die katalitiese aktiwiteit van SRD5A1 teenoor A4 en T is nie beïnvloed deur
rooibos nie, alhoewel dit voorkom asof rooibos die omsettingstempo van T na dihidrotestosteroon
(DHT) deur SRD5A2, getransfekteer in HEK293 selle, verminder het. Verdere ondersoeke is in
normale prostaat epiteel selle, in die teenwoordigheid van rooibos uitgevoer. Rooibos het geen
invloed op die katalitiese aktiwiteit van SRD5A teenoor A4 gehad nie, alhoewel vermindere
omskakeling van T na DHT aangetoon kon word. Rooibos het ook die omskakeling van A4 na T in
beide PNT2 en BPH-1 selle tot „n mate geïnhibeer. Die omskakeling van 11KA4 na 11KT was ook
verminder in BPH-1, PC-3 en LNCaP selle. Die omskakeling van 11KA4 na 11KT was beduidend
laer in PNT2 selle en kon die invloed van rooibos nie aangetoon word nie. Bykomende data toon
dat rooibos ook die omskakeling van 5α-androstenedioon (5α-dione) na androsteroon (AST),
gekataliseer deur 3α-hidroksi-dehidrogenase tipe 3 (AKR1C2), verminder, gesamentlik met die
vermindere omskakeling van T na A4, deur 17βHSD2, in PNT2 selle. Hierdie studie het ook
ondersoek ingestel, na die metabolisme van androgene in LNCaP selle. Vermindere A4
metabolisme is in die teenwoordigheid van rooibos asook in die teenwoordigheid van
indometasien, „n bekende AKR1C3 inhibitor, gevind. Rooibos verminder dus die aktiwiteit van
reduktiewe 17βHSD in LNCaP selle. Verandering in die metabolisme van T, DHT en AST in
LNCaP selle, in die teenwoordigheid van rooibos, is ook gevind. Verdere ondersoek in LNCaP
selle het gewys dat rooibos „n vermindering in die produksie van gekonjugeerde T, DHT en AST
veroorsaak. Die studie het die invloed van rooibos op prostaat-spesifieke antigeen (PSA) ook
ondersoek. Daar is vasgestel dat rooibos die vlakke van PSA verminder in die afwesigheid en
teenwoordigheid van DHT in LNCaP selle. Hierdie resultaat dui op moontlike interaksie van
rooibos met die androgeen (AR) of estrogeen-reseptor-β (ERβ), teenwoordig in LNCaP selle.
Rooibos het die katalitiese aktiwiteit van ensieme, wat bydra tot androgeen produksie, geïnhibeer,
asook die konjugasie van androgene. Op „n sellulêre vlak, het rooibos die vlakke van PSA-sekresie
verminder, wat moontlike interaksie met die AR en ERβ aandui. Hierdie bevindings dui daarop dat
rooibos wel n rol het om te speel in die modulasie van aktiewe androgene in die prostaat.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/96926 |
| Date | 04 1900 |
| Creators | Du Toit, Therina |
| Contributors | Swart, Amanda C., Storbeck, K-H, Stellenbosch University. Faculty of Science. Dept. of Biochemistry. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | Unknown |
| Type | Thesis |
| Format | xiv, 139 pages : illustrations |
| Rights | Stellenbosch University |
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