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Investigating the effects of progestins and adiposity on c-Jun N terminal kinase (JNK) signalling in breast cancer
Thesis (MSc)--Stellenbosch University, 2022.
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| Language: | en_ZA |
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Stellenbosch : Stellenbosch University
2022
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| _version_ | 1867614025304506368 |
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| access_status_str | Open Access |
| author | Simons, Mishkah |
| author2 | Africander, Donita |
| author_browse | Africander, Donita Simons, Mishkah |
| author_facet | Africander, Donita Simons, Mishkah |
| author_sort | Simons, Mishkah |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Thesis (MSc)--Stellenbosch University, 2022. |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/125118 |
| institution | Stellenbosch University (South Africa) |
| language | en_ZA |
| last_indexed | 2026-06-10T12:45:28.762Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2022 |
| publishDateRange | 2022 |
| publishDateSort | 2022 |
| publisher | Stellenbosch : Stellenbosch University |
| publisherStr | Stellenbosch : Stellenbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/125118 Investigating the effects of progestins and adiposity on c-Jun N terminal kinase (JNK) signalling in breast cancer Simons, Mishkah Africander, Donita Stellenbosch University. Faculty of Science. Dept. of Biochemistry. Breast -- Cancer Progestational hormones Cancer -- Risk factors Mitogen-activated protein kinases -- Inhibitors C-Jun N terminal kinase -- Inhibitors Phosphorylation UCTD Thesis (MSc)--Stellenbosch University, 2022. ENGLISH ABSTRACT: Obesity and progestins which are used in contraception and menopausal hormone therapy are known risk factors for breast cancer. However, the precise mechanisms by which these factors increase breast cancer risk are not known, and whether the use of progestins by obese women exacerbates breast cancer risk is also unclear. Mitogen activated protein kinase (MAPK) pathways are known to be activated by some progestins as well as obesity-related factors. Activation of these pathways may thus be a possible mechanism whereby progestins and obesity increase breast cancer risk. This study focused on the relatively understudied c-Jun N terminal kinase (JNK) subgroup of the MAPK pathways in breast cancer. The aim of this study was to investigate the effects of a progestin, alone or combined with conditioned media prepared from differentiated 3T3-L1 adipocytes (CM), on the proliferation and apoptosis of the ER+ MCF-7 BUS and T47D breast cancer cell lines, and evaluate the contribution of the JNK pathway. CM was used as a model for adiposity and effects of promegestone (R5020), medroxyprogesterone acetate (MPA), norethindrone (NET) and levonorgestrel (LNG), and drospirenone (DRSP), were evaluated. Results show that progestin-induced proliferation was abrogated by CM in T47D cells, but enhanced in the MCF-7 BUS cells. Incubation with the JNK inhibitor, SP600125, showed that while the effects in the MCF-7 BUS cells were dependent on the JNK pathway, this was not the case in the T47D cells. Western blot analysis evaluating p53 expression as a marker of apoptosis showed that all progestins alone or in combination with CM, did not regulate p53 expression in the MCF-7 BUS cells. In contrast, none of the progestins in the absence or presence of CM, except DRSP, regulated p53 expression in the T47D cells. However, CM regulated p53 expression in both the T47D and MCF-7 BUS cells, in a JNK independent manner. We are the first to show that phosphorylation of JNK in T47D cells is induced by NET, that R5020 decreased phosphorylation, and that MPA, LNG and DRSP had no effect. In the MCF-7 BUS cells, DRSP and LNG significantly increased JNK phosphorylation, and although not statistically significant, an apparent increase in JNK phosphorylation was also observed with R5020, MPA and NET. Although CM did not induce phosphorylation of JNK in either cell line, we show for the first time that the effects on JNK activation were modulated by CM in a progestin- and cell line-dependent manner. Lastly, quantitative real-time PCR results showed that the mRNA expression of MAP kinase kinase 4 (MKK4) and dual-specificity phosphatase 1 (DUSP1) was differentially modulated by CM as well as the progestins in the T47D and the MCF-7 BUS breast cancer cells suggesting that progestins target upstream regulators of the JNK pathway. In summary, while the findings of this study do not provide a definitive answer on whether the effects of obesity and progestins are additive, it does contribute to the understanding of both progestins and obesity in breast cancer, and role of the JNK pathway. AFRIKAANSE OPSOMMING: Vetsug en progestiene wat in menopousale hormoonterapie en voorbehoedmiddels gebruik word, is bekende risikofaktore vir borskanker. Die presiese meganismes waardeur hierdie faktore die risiko van borskanker verhoog, is egter onbekend, en of die gebruik van progestiene deur vetsugtige vroue die risiko vir borskanker vererger, is ook onduidelik. Dit is wel bekend dat mitogeen-geaktiveerde proteïenkinase (MAPK) padweë geaktiveer word deur sommige progestiene asook vetsugverwante faktore. Aktivering van hierdie padweë kan dus 'n moontlike meganisme wees waardeur progestiene en vetsug die risiko van borskanker verhoog. Die huidige studie het gefokus op die relatief onderbestudeerde c-Jun N terminale kinase (JNK) subgroep van die MAPK padweë in borskanker. Die doel van hierdie studie was om die uitwerking van 'n progestien, in die afwesigheid en teenwoordigheid van gekondisioneerde media van gedifferensieerde 3T3-L1 adiposiete (KM), op die proliferasie en apoptose van die ER+ T47D en MCF-7 BUS borskankersellyne te ondersoek, en die bydrae van die JNK padweg te evalueer. KM is gebruik as 'n model vir adipositeit en effekte van promegestone (R5020), medroksieprogesteroon asetaat (MPA), noretisteroon (NET) en levonorgestrel (LNG), en drospirenoon (DRSP), is geëvalueer. Resultate toon dat progestien-geïnduseerde proliferasie opgehef is deur KM in T47D selle, maar versterk is in MCF-7 BUS selle. Inkubasie in die teenwoordigheid van die JNK inhibeerder, SP600125, het getoon dat alhoewel die effekte in die MCF-7 BUS selle afhanklik was van die JNK padweg, dit nie die geval was in die T47D selle nie. Westernklad analise wat p53 uitdrukking as 'n merker van apoptose ge-evalueer het, het getoon dat alle progestiene alleen of in kombinasie met KM, nie p53 uitdrukking in die MCF-7 BUS borskankerselle reguleer nie. In teenstelling hiermee het geen van die progestiene in die afwesigheid of teenwoordigheid van KM, behalwe DRSP, p53 uitdrukking in die T47D selle gereguleer nie. KM het egter p53 uitdrukking in beide die MCF-7 BUS en T47D selle gereguleer, op 'n JNK onafhanklike wyse. Ons is die eerste om te wys dat NET fosforilering van JNK in T47D selle geïnduseer het, dat R5020 fosforilering verlaag het, en dat MPA, LNG en DRSP geen effek het nie. In die MCF-7 BUS selle het DRSP en LNG, JNK- fosforilering aansienlik verhoog, en alhoewel dit nie statisties betekenisvol is nie, is 'n oënskynlike toename in JNK-fosforilering ook waargeneem met toevoeging van R5020, MPA en NET. Alhoewel KM nie fosforilering van JNK in enige van die sellyne induseer het nie, wys ons vir die eerste keer dat effekte op JNK-aktivering gemoduleer is deur KM op 'n progestien- en sellynafhanklike wyse. Laastens het intydse kwantitatiewe polimerase kettingreaksie resultate getoon dat die mRNA uitdrukking van MAP kinase kinase 4 (MKK4) en dubbel-spesifisiteit fosfatase 1 (DUSP1) differensieel gemoduleer is deur KM sowel as die progestiene, in beide T47D en MCF-7 BUS borskanker selle, wat daarop aandui dat progestiene stroomop-reguleerders van die JNK padweg teiken. Ter samevatting. hoewel die bevindinge van hierdie studie nie 'n definitiewe antwoord verskaf oor of die effekte van vetsug en progestiene byvoegend is nie, dra dit wel by tot die begrip van beide progestiene en vetsug in borskanker, sowel as die rol van die JNK padweg. Masters 2022-03-10T06:32:21Z 2022-04-29T12:54:45Z 2023-03-31T03:00:12Z 2022-03 Thesis http://hdl.handle.net/10019.1/125118 en_ZA Stellenbosch University xii, 77 pages : illustrations application/pdf Stellenbosch : Stellenbosch University |
| spellingShingle | Breast -- Cancer Progestational hormones Cancer -- Risk factors Mitogen-activated protein kinases -- Inhibitors C-Jun N terminal kinase -- Inhibitors Phosphorylation UCTD Simons, Mishkah Investigating the effects of progestins and adiposity on c-Jun N terminal kinase (JNK) signalling in breast cancer |
| title | Investigating the effects of progestins and adiposity on c-Jun N terminal kinase (JNK) signalling in breast cancer |
| title_full | Investigating the effects of progestins and adiposity on c-Jun N terminal kinase (JNK) signalling in breast cancer |
| title_fullStr | Investigating the effects of progestins and adiposity on c-Jun N terminal kinase (JNK) signalling in breast cancer |
| title_full_unstemmed | Investigating the effects of progestins and adiposity on c-Jun N terminal kinase (JNK) signalling in breast cancer |
| title_short | Investigating the effects of progestins and adiposity on c-Jun N terminal kinase (JNK) signalling in breast cancer |
| title_sort | investigating the effects of progestins and adiposity on c jun n terminal kinase jnk signalling in breast cancer |
| topic | Breast -- Cancer Progestational hormones Cancer -- Risk factors Mitogen-activated protein kinases -- Inhibitors C-Jun N terminal kinase -- Inhibitors Phosphorylation UCTD |
| url | http://hdl.handle.net/10019.1/125118 |
| work_keys_str_mv | AT simonsmishkah investigatingtheeffectsofprogestinsandadiposityoncjunnterminalkinasejnksignallinginbreastcancer |