ALPS case study

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Biology

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Jan 9, 2024

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BIO 340 Autoimmunity case activity 4/11/22 (25 pts) This ALPS case study is to be submitted on D2L by each individual, but you may discuss the case with others. The case study below was written using data and images from http://www.cytometry.org/newsletters/eICCS-2-1/article5.php . https://ricfacility.byu.edu/Research/CellSizeGranularity Watch the animation link provided below on flow cytometry. Different fluorescent-labeled antibodies were used for this experiment to detect CD4, CD8 and TCR on T cells. The dot plots will be analyzed in question #1 below (first half of the animation does not discuss this, but talks about forward & side scatter which is also important for blood cells). Watch flow cytometry method: https://www.youtube.com/watch?v=EQXPJ7eeesQ Flow cytometry - difference of cells shape, size, and complexity Forward scatter – is proportional to the size of the cell, computer converts it into a voltage, larger cell = higher pulse Side scatter – is proportional to the shape and internal complexity (presence of granules), A 20 month-old patient named Joey has extensive lymphadenopathy in the cervical and armpit lymph nodes but appears to be handling infections and vaccinations normally. He has some noticeable spotty bruising on legs and arms. Upon analysis of peripheral blood, it was found that he had elevated numbers of lymphocytes. Further flow cytometric analysis revealed the following for CD4 & CD8 (y-axis) and mature TCR (x-axis) cell markers. Note: as you move further up on the y-axis and to the right on the x-axis, the intensity of the fluorescence for that cell marker is higher. For instance, those dots near the origin (B3 quadrant) do not display high levels of either cell marker. Control sample Joey’s sample 1. Interpret the differences between quadrants B2 (upper right) and B4 (bottom right) for both the control and Joey’s sample. (6 pts) For the control, the percentage of TCR and CD4 and CD8 is 90% and are mostly larger cells. In his sample, B4 takes up a larger percent saying that his cells are less complex, he has a lot more DNT cells with TCR expressed than the control, and higher levels of TCR. He also has a decrease in number of cells that are expressing CD4 and CD8. This indicates that he could have ALPS because he has an increased number of DNT cells. He still has a lot of testing to do to confirm whether he has ALPS and that needs to be compared with symptoms for a diagnosis to be given. 2. Further analysis revealed that platelet counts were about 1/3 of what they should be while IgG and IgM levels in the serum were elevated (hypergammaglobulinemia), despite no detectable infections nor cancer.

a. Why is a low platelet count problematic? What symptom in Joey’s case could be connected to this issue? (3 pts) He will be having a decreased levels blood clotting which makes him susceptible to internal and external bleeding. He has been having a lot of bruising recently indicating that his cells have not been able to clot intravascular breaks effectively. b. Joey’s antibodies were tested further to reveal that they are autoantibodies to his platelets, destroying them. He has thrombocytopenia due to autoimmunity. A lymph node biopsy was taken. Describe what area(s) in the lymph nodes you would expect to see enlarged. (3 pts) The paracortex would be enlarged because there are high levels of IgM and IgG from activated B cells. These needs to be activated by t cells which causes proliferation of them causing more t cells to be within the paracortex and in circulation because it stated earlier that he had increased levels of lymphocytes. 3. A plasma cytokine assay showed elevated IL-10 levels in Joey’s blood. Which cells produce IL-10 and what is the action of this cytokine on B cells? (4 pts) IL-10 is produced by macrophages, dendritic cells, T helper cells. It inhibits the expression of Th1 cytokines, MHC class 2 antigens and costimulatory molecules on macrophages. It will also enhance the activation and proliferation of b cells as well as antibody production. It gives pro survival signal to a group of b cells. The b cells that are already activated will be within germinal center and follicle (lymph nodes). Normally IL-10 tells b cells producing antibodies to go through apoptosis but in this case it is not. This allows those b cells to allow for a high number of IgM and IgG. So, since there are high levels of IL-10, it would make sense as to why there are high levels of IgM and IgG from b cells. It can also explain the reason for his autoimmunity because Th1 cells are responsible for preventing autoimmunity. 4. Joey’s blood was also submitted for genetic analysis. It was discovered that he possesses a mutation in the Fas gene. This mutation leads to a defect in apoptosis of both T and B cells. He was diagnosed with Autoimmune Lymphoproliferative Syndrome (ALPS). a. Explain how Fas normally works to promote apoptosis and why blocking this is an issue for both B and T cells. (3 pts) The fas gene normally codes for a protein trimer that is part of the caspase cascade that is responsible for apoptosis. This is bad because t and b cells use it for apoptosis of cells that could be cancerous.

b. What aspects of Joey’s case history along with this genetic information could be linked to a greater potential for him to develop lymphoma in the future? (3 pts) This inactive FAS gene is a big part of it because the caspase cascade is used for apoptosis. If he does end up having ALPS, which there is strong evidence of, there’s cytopenia which can contribute to a lack of fighting tumors. As well as lymphocyte proliferation will also contribute to possible lymphoma. 5. What is the prognosis for Joey’s condition? Will he always have these issues?—describe evidence considered (3 pts) It’s very likely that joey has ALPS based on his DNT/TCR cell count vs CD4+ and CD8+ t cell count as well as the low platelet count coming from the antibodies attacking them which is caused by IL-10 high elevation letting b cells release more IgM and IgG. More testing of the DNT cells will have to be done to confirm. He might not always have these issues as it is cited that some patients might have the disease and symptoms do not affect them at all throughout their life but if not, they can be prescribed immunosuppressants. This will help with treatment of cytopenia and lymphadenopathy.

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