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Comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds
Thesis (MSc)--Stellenbosch University, 2025.
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| Format: | Thesis |
| Language: | English |
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Stellenbosch : Stellenbosch University
2025
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| _version_ | 1867614070420537344 |
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| access_status_str | Open Access |
| author | Armstrong, Kayleigh Ann |
| author2 | Van de Vyver, Mari |
| author_browse | Armstrong, Kayleigh Ann Van de Vyver, Mari |
| author_facet | Van de Vyver, Mari Armstrong, Kayleigh Ann |
| author_sort | Armstrong, Kayleigh Ann |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Thesis (MSc)--Stellenbosch University, 2025. |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/134496 |
| institution | Stellenbosch University (South Africa) |
| language | English |
| last_indexed | 2026-06-10T12:46:11.731Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2025 |
| publishDateRange | 2025 |
| publishDateSort | 2025 |
| publisher | Stellenbosch : Stellenbosch University |
| publisherStr | Stellenbosch : Stellenbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/134496 Comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds Armstrong, Kayleigh Ann Van de Vyver, Mari Howard, Kayla Benecke, Rohan Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Medicine. Division of Clinical Pharmacology. Diabetics -- Wounds and injuries Blood -- Coagulation Diabetes -- Complications Foot -- Ulcers Thesis (MSc)--Stellenbosch University, 2025. Armstrong, K. A. 2025. Comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds. Unpublished masters thesis. Stellenbosch: Stellenbosch University [online]. Available: https://scholar.sun.ac.za/items/cf24beb9-8330-420b-9b53-4de972ca0422 ENGLISH ABSTRACT: Introduction: Non-healing diabetic foot ulcers (DFUs) are a major complication of type 2 diabetes mellitus (T2DM). Topical autologous blood clot therapy (TABCT) is a novel cellular-based treatment approach involving the application of an autologous blood clot to a non-healing wound. TABCT provides a temporary extracellular matrix (ECM) scaffold, initiating healing through the release of various chemokines and growth factors. It is, however, uncertain whether TABCT would be efficacious in T2DM patients, due to the multifactorial cellular dysregulation prevalent in the diabetic microenvironment, and how systemic levels of pharmaceuticals would influence its efficacy. This study, therefore, compared the effects of regularly prescribed anti-inflammatory (prednisone), anti-glycaemic (metformin), and antibiotic (amoxicillin and rifampicin) agents on chemokine and growth factor release. The pro-regenerative capacity of the TABCT secretome was subsequently compared by assessing peripheral blood mononuclear cell (PBMC) chemotaxis (ex vivo) and fibroblast migration (in vitro). Methods: Twelve study participants were recruited according to specific inclusion criteria and divided into two groups: self-reported healthy controls (n=6) and T2DM patients (n=6). Anthropometric measurements (height, weight, hip, and waist circumference) and metabolic status (blood pressure, blood glucose, and lipid profile) of each participant were assessed. Whole blood samples were collected in one EDTA tube and two citrate tubes. PBMCs were isolated from the EDTA blood sample. The whole blood in the citrate tubes was individually “spiked-in” with the pharmaceuticals of interest prior to ex vivo clot induction. The clot size and protein concentration within the clot secretome were determined. Three separate ELISAs were performed to quantify CCL5, P-selectin and platelet-derived growth factor-BB (PDGF-BB) within the secretome. The chemotactic effects of the clot-derived secretomes on PBMC migration were assessed ex vivo. An in vitro scratch assay was used to determine the effects of the clot-derived secretome on dermal fibroblast-mediated wound closure. Results: As predicted, the T2DM cohort exhibited a poorer metabolic status compared to the controls. Prednisone treatment increased clot size in the control group (1.45 ± 0.19-fold), and elevated clot-derived CCL5 (no treatment 209 ± 107 pg/mL; prednisone 300 ± 104 pg/mL) and PDGF-BB (no treatment 8.7 ± 6.8 pg/mL; prednisone 13.6 ± 10.5 pg/mL) secretion in the diabetic group. Rifampicin upregulated PDGF-BB (no treatment 14.5 ± 4.2 pg/mL; rifampicin 19.9 ± 3.9 pg/mL) expression in the control cohort. Compared with standard cell culture conditions, exposure to the clot secretome reduced PBMC migration velocity and distance, while increasing directionality, confirming the chemotactic effect of blood clots. This chemotactic effect was not influenced by the pharmaceutical treatments, and it did not alter PBMC chemotaxis. The clot secretome stimulated fibroblast migration, with individual variability observed. Metformin significantly enhanced fibroblast migration within the control cohort (no treatment 15.3 ± 2.9%; metformin 21.1 ± 5.5%), but not in the T2DM cohort (no treatment 16.4 ± 4%; metformin 18 ± 4.3%). Conclusion: This study investigated the effects of regularly prescribed pharmaceuticals on the efficacy of TABCT. Based on the specific patient profiles investigated in this study, pro-regenerative effects of the TABC, with and without pharmaceutical treatment, were demonstrated. No major abnormalities were observed in immune cell chemotaxis and in vitro wound closure. However, given the wide variability in disease advancement amongst diabetic patients, pharmaceutical administration should be taken into consideration with the use of TABCT. AFRIKAANSE OPSOMMING: Inleiding: Diabetiese voetsere wat nie genees nie, is ‘n hoofkomplikasie van tipe 2 diabetes mellitus (T2DM). Oppervlakkige outologiese bloedklontterapie (OOBKT) is ‘n nuwe sellulêrgebaseerde benadering vir behandeling en behels die aanwending van ‘n outologiese bloedklont aan ‘n nie-genesende wond. OOBKT bied ‘n tydelike buite-sellulêre matriks (ECM) steier wat genesing begin danksy die vrystelling van verskeie chemokine en groeifaktore. Dit is egter onseker of OOBKT doeltreffend is in T2DM pasiënte, as gevolg van die multifaktoriëlesellulêre wanregulasie wat in die diabetiese mikro-omgewing heers, en hoe farmaseutiese middels se sistemiese vlakke die doeltreffendheid sal beïnvloed. Hierdie studie het gevolglik die uitwerking van algemeen voorgeskrewe farmakologiese middels nl. anti-inflammatories (prednisoon), anti-glisemies (metformien) en antibiotiese middels (amoksisillien en rifampisien), op chemokine en die vrystelling van groeifaktore vergelyk. OOBKT sekretome se pro-generatiewe kapasiteit is daarna vergelyk deur die omliggende bloed mono-kernsel (PBMS) chemotaksis (ex vivo) en fibroblast migrasie (in vitro) te bepaal. Metodes: Twaalf deelnemers is vir die studie gewerf volgens spesifieke insluitingskriteria en in twee groepe verdeel: self-gerapporteerde gesonde kontroles (n=6) en T2DM-pasiënte (n=6). Antropometriese mates (lengte, gewig en middel-tot-heup ratio) en metaboliese status (bloeddruk, bloedglukose, en lipiedprofiel) van elke deelnemer is vasgestel. Heel bloedmonsters is geneem in een EDTA buis en twee sitraatbuise. PBMS was van die EDTA bloedmonster geïsoleer. Die sitraatbuise se heel bloed was individueel met die farmaseutiese middels van belang gestimuleer, voor die ex vivo klont induksie. Die klontgrootte en proteïenkonsentrasie binne die klont sekretome was bepaal. Drie afsonderlike ELISAs was uitgevoer om die CCL5, P-selektien en plaatjie-afgeleide groeifaktor-BB (PAGF-BB) binne die sekretome te kwantifiseer. Die chemotaktiese effekte van die klontafgeleide sekretome op PBMS migrasie was ex vivo vasgestel. ‘n In vitro kraptoets was gebruik om die effekte van die klontafgeleide sekretome op dermis-fibroblast-bemiddelde wondsluiting te bepaal. Resultate: Soos voorspel, het die T2DM-groep ‘n swakker metaboliese status getoon in vergelyking met die kontrolegroep. Prednisoon-behandeling het die klontgrootte in die kontrolegroep laat toeneem (1.45 ± 0.19-fold), en het klont-afgeleide CCL5 verhoog (geen behandeling 209 ± 107 pg/mL; prednisoon 300 ± 104 pg/mL) en PAGF-BB (geen behandeling 8.7 ± 6.8 pg/mL; prednisone 13.6 ± 10.5 pg/mL) sekresie in die diabetiese groep. Rifampisien het PAGF-BB (geen behandeling 14.5 ± 4.2 pg/mL; 19.9 ± 3.9 rifampisien pg/mL) se uitdrukking in die kontrolegroep verhoog. In vergelyking met standaard selkultuur toestande, het blootstelling aan die klontsekretome PBMS migrasie snelheid en afstand verminder, terwyl dit rigtinggevoeligheid vermeerder het en die chemotaktiese effek van bloedklonte bevestig. Hierdie chemotaktiese effek was nie deur die farmaseutiese behandelings beïnvloed nie en dit het nie die PBMS chemotaksis verander nie. Die klontsekretome het fibroblast migrasie gestimuleer en individuele wisselvalligheid is waargeneem. Metformien het fibroblast migrasie opvallend in die kontrolegroep verhoog (geen behandeling 15.3 ± 2.9%; metformien 21.1 ± 5.5%), maar nie in die T2DM-groep nie (geen behandeling 16.4 ± 4%; metformien 18 ± 4.3%). Afsluiting: Hierdie studie het die uitwerking van algemeen voorgeskrewe farmaseutiese middels op die doeltreffendheid van OOBKT ondersoek. Gebaseer op die spesifieke pasiëntprofiele wat in hierdie studie ondersoek is, was pro-regeneratiewe effekte van die OOBK met en sonder farmaseutiese behandeling gedemonstreer. Geen groot abnormaliteite is in die immuun sel-chemotaksis en in vitro wondsluiting waargeneem nie. Maar gegewe die wye veranderlikhede in siektetoename onder diabetiese pasiënte, moet farmaseutiese toediening egter in ag geneem word met die gebruik van OOBKT. Masters 2025-12-11T09:12:34Z 2025-12-11T09:12:34Z 2025-12 Thesis https://scholar.sun.ac.za/handle/10019.1/134496 en Stellenbosch University 104 pages : illustrations application/pdf Stellenbosch : Stellenbosch University |
| spellingShingle | Diabetics -- Wounds and injuries Blood -- Coagulation Diabetes -- Complications Foot -- Ulcers Armstrong, Kayleigh Ann Comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds |
| title | Comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds |
| title_full | Comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds |
| title_fullStr | Comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds |
| title_full_unstemmed | Comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds |
| title_short | Comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds |
| title_sort | comparing the influence of regularly prescribed pharmaceuticals on the chemotactic effects of autologous blood clots in the context of diabetic wounds |
| topic | Diabetics -- Wounds and injuries Blood -- Coagulation Diabetes -- Complications Foot -- Ulcers |
| url | https://scholar.sun.ac.za/handle/10019.1/134496 |
| work_keys_str_mv | AT armstrongkayleighann comparingtheinfluenceofregularlyprescribedpharmaceuticalsonthechemotacticeffectsofautologousbloodclotsinthecontextofdiabeticwounds |