(J-381) The Fate of Monocytes Under Various Inflammatory Conditions

Myelofibrosis is a rare bone marrow cancer that changes the extracellular matrix (ECM). 

Cancer is known to be linked to chronic inflammation which results in the thickening, hardening, and scarring of tissues, also known as fibrosis. In the diseased state, the ECM becomes hardened and inflamed leading to an overproduction of monocytes (Vining, 2022). It is known that monocytes can become inflammatory and differentiate into dendritic cells and macrophages. Interleukin-6 (IL-6) is a pro-inflammatory cytokine that is produced by monocytes during an immune response. In the experiment, we are interested in comparing the concentrations of IL-6 that are present in different stages during the progression of the disease. We will fabricate a gradient of various hydrogel environments, ranging from viscous soft, which represents the most healthy ECM environment, to elastic stiff, which mimics the most diseased. This will help get us closer to understanding how the viscoelasticity of the matrix affects the differentiation fate of monocytes and the associated inflammation. We are interested in this relationship because it can potentially lead to a treatment for this incurable cancer along with other inflammatory diseases.

The level of  IL-6 production by the cells is an indicator of the phenotype of the monocytes. It is known that dendritic cells show enhanced production of IL-6 by GM-CSF activation. (Sonderegger et al., 2008) Classically, IL-4 is an anti-inflammatory cytokine but previous studies have shown that the effect of GM-CSF and IL-4 directs monocytes to a dendritic cell phenotype by up-regulating tumor necrosis factor alpha (TNF-alpha), another pro-inflammatory cytokine. (Du et al., 2022),(Sonderegger et al.,2008) We will be using an enzyme-linked immunoassay (ELISA), which is a test used to detect and quantify the levels of antigens, antibodies, and other proteins. This is how we will determine if the cells have differentiated. We will be testing for the presence of IL-6 and quantifying its concentration. We will use the supernatant produced during the cell culture of the monocytes conditioned with granulocyte-macrophage colony-stimulating factor (GM-CSF) and of monocytes conditioned with GM-CSF/IL-4. The values produced from the test will be used to validate if the combination of the GM-CSF and IL-4 will result in a marked increase of IL-6 in vitro. Future work will recreate these conditioned cell fates in hydrogels representative of various tissue environments.

The reason that the differential fate of monocytes is relevant to myelofibrosis is because tumor cells are largely fibrous and unable to be infiltrated by certain immune cell types.(Salmon et al., 2012) Their mechanical nature also makes them more susceptible to attack by other immune cell types.(Basu et al., 2016) Understanding ways to direct immune cell differentiation in cancer and other diseased environments can lead to more effective treatments.

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   Du, H., Bartleson, J. M., Butenko, S., Alonso, V., Liu, W. F., Winer, D. A., & Butte, M. J. (2022). Tuning immunity through tissue mechanotransduction. Nature Reviews Immunology, 23(3), 174–188. https://doi.org/10.1038/s41577-022-00761-w  

   
   

2. 
   

   Hiasa, M., Abe, M., Nakano, A., Oda, A., Amou, H., Kido, S., Takeuchi, K., Kagawa, K., Yata, K., Hashimoto, T., Ozaki, S., Asaoka, K., Tanaka, E., Moriyama, K., & Matsumoto, T. (2009). GM-CSF and IL-4 induce dendritic cell differentiation and disrupt osteoclastogenesis through M-CSF receptor shedding by up-regulation of TNF-alpha converting enzyme (TACE). Blood, 114(20), 4517–4526. https://doi.org/10.1182/blood-2009-04-215020

   
   


3. 
   

   Sonderegger, I., Iezzi, G., Maier, R., Schmitz, N., Kurrer, M., & Kopf, M. (2008). GM-CSF mediates autoimmunity by enhancing IL-6-dependent Th17 cell development and survival. The Journal of Experimental Medicine, 205(10), 2281–2294. https://doi.org/10.1084/jem.20071119 

   
   


4. 
   

   Salmon, H., Franciszkiewicz, K., Damotte, D., Dieu-Nosjean, M.-C., Validire, P., Trautmann, A., Mami-Chouaib, F., & Donnadieu, E. (2012). Matrix architecture defines the preferential localization and migration of T cells into the stroma of human lung tumors. The Journal of Clinical Investigation, 122(3), 899–910. https://doi.org/10.1172/JCI45817 

   
   


5. 
   

   Basu, R., Whitlock, B. M., Husson, J., Le Floc’h, A., Jin, W., Oyler-Yaniv, A., Dotiwala, F., Giannone, G., Hivroz, C., Biais, N., Lieberman, J., Kam, L. C., & Huse, M. (2016). Cytotoxic T cells use mechanical force to potentiate target cell killing. Cell, 165(1), 100–110. https://doi.org/10.1016/j.cell.2016.01.021 

   
   


6. Vining, K.H., Marneth, A.E., Adu-Berchie, K. et al. Mechanical checkpoint regulates monocyte differentiation in fibrotic niches. Nat. Mater. 21, 939–950 (2022). https://doi.org/10.1038/s41563-022-01293-3