University of Twente Student Theses
The effect of nitric oxide on macrophage polarization in the synovial membrane
Dalenoord, F.R.A. (2021) The effect of nitric oxide on macrophage polarization in the synovial membrane.
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| Abstract: | Osteoarthritis (OA) and rheumatoid arthritis (RA) are two common chronic joint disorders that cause major disability and high healthcare costs worldwide. Due to aging and increase in obesity, the number of people affected by these diseases and its healthcare costs will increase in the coming years. The most affected joint for OA patients is the knee joint, while the metacarpophalangeal and proximal interphalangeal joints are most affected in RA. Within these synovial joints, the synovial membrane, which provides synovial fluid, is influenced both by OA and RA. A characteristic feature of these diseases is inflammation in the synovial membrane and a thicker synovial membrane, which is due to an influx of immune cells such as macrophages. In OA and RA these macrophages are polarized to an inflammatory state, which degrade the synovial joint in the whole. One of the recently discussed messenger molecules that seems to affect the polarization of macrophages, is nitric oxide. It is hypothesized to be a stimulator of polarization to pro-inflammatory macrophages. As this has not been investigated yet, the results could potentially provide crucial information on the development of OA and RA. In this report, the effect of nitric oxide on the polarization of macrophages in the synovial membrane is investigated. A synovial membrane-on-chip system was used to mimic the synovial membrane. The device consists of two distinctive compartments, seperated by a PDMS membrane. The top chamber was filled with human synovial fibroblasts and macrophages and the bottom compartment with human umbilical vein endothelial cells (HUVECs). As controls, this culture is compared to a double culture (only synovial fibroblasts and macrophages) and a single culture (only synovial fibroblasts). To measure the concentration of nitric oxide (NO) a Griess assay was performed. Furthermore, both immunofluorescence and qPCR were performed to determine the polarization of the seeded macrophages and other extracellular matrix markers, such as collagen 1 (COL1) and hyaluronic acid synthase 1 (HAS1). Partial to no integration of macrophages was visible 3 hours after seeding of macrophages in double culture samples and worsens over the days, while full integration was visible for triple culture samples (fibroblasts, macrophages and HUVECs). The Griess assay showed an increasing difference between double and triple culture samples. Double culture samples were high in NO, while triple culture samples were significantly low. Immunofluorescence showed clusters of collagen 1 in low concentrations of NO, which suggests higher amounts of tissue-regenerative macrophages. Moreover, more CD163 accentuated macrophages were visible in triple culture samples. qPCR results showed a higher gene expression of M2 markers for low NO levels, than high NO levels. IL1R, a M1 marker, showed a decrease in gene expression at low NO levels, while an increase was seen at high NO levels. While more research is needed to prove causality, it seems that low levels are coupled with a polarization to M2 macrophages and high levels of nitric oxide are coupled with polarization to M1 macrophages. |
| Item Type: | Essay (Bachelor) |
| Faculty: | TNW: Science and Technology |
| Subject: | 42 biology, 50 technical science in general |
| Programme: | Biomedical Technology BSc (56226) |
| Link to this item: | https://purl.utwente.nl/essays/87768 |
| Export this item as: | BibTeX EndNote HTML Citation Reference Manager |
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