Investigating the immunomodulatory role of mesenchymal stromal cells in primary sclerosing cholangitis

Janmohamed, Ashnila (2022). Investigating the immunomodulatory role of mesenchymal stromal cells in primary sclerosing cholangitis. University of Birmingham. M.D.

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Abstract

Background and Aims: Liver-infiltrating T lymphocytes and high levels of tumour necrosis factor (TNFα) are implicated in the destruction of bile ducts in primary sclerosing cholangitis (PSC). Mesenchymal stromal cells (MSC) possess broad immunomodulatory properties, suggesting a potential therapeutic role in PSC. Herein, we sought to investigate the immunomodulatory role of umbilical cord derived MSC (UC-MSC) on T cells from patients with PSC by studying the effect of UC-MSC and UC-MSC derived conditioned media (UC-MSC-CM) on proliferation and activation of circulating and intrahepatic CD4+ and CD8+ T cells.
Method: Peripheral blood mononuclear cells (PBMC) were isolated from patients with PSC and hereditary haemochromatosis (controls). Intrahepatic CD4+ and CD8+ T cells were acquired by negative selection from explant livers of patients with PSC and primary biliary cholangitis (PBC). Cell trace violet labelled cells were co-cultured with UC-MSC at ratios; 1:1, 1:4, 1:16, 1:64 1:256 (UC-MSC: PBMC/T cells) and anti- CD3/anti-CD28 for either 4 (if intrahepatic T cells used) or 5 (if PBMC used) days. On the last day of culture, the PBMC/T cells were further stimulated with phorbol 12- myristate 13-acetate, ionomycin and brefeldin A for 4 hours. UC-MSC effect on CD4+ and CD8+ T cell proliferation/activation (TNFα, interferon (IFN)-γ, IL2 expression) was studied by flow cytometry. Experiments were repeated using UC-MSC-CM derived from both resting (untreated) and interferon (IFN) γ stimulated UC-MSC at concentrations ranging from 100%-50%.
Results: UC-MSC significantly suppressed proliferation of both circulating and intrahepatic PSC CD4+and CD8+T cells in a dose dependent manner. No significant differences in the UC-MSC inhibitory effect on T cell proliferation was seen between patients with PSC, PBC and hereditary haemochromatosis suggesting that the UCMSC effect is not disease specific. UC-MSC significantly reduced both circulating and intrahepatic PSC CD4+TNFα+ and CD4+IFN-γ +expressing cells. In addition intrahepatic PSC CD4+IL2+ expressing cells were also reduced in the presence of UC-MSC. Conditioned media derived from untreated UC-MSC significantly reduced circulating PSC CD4+ and CD8+ T cell proliferation in a dose dependent manner confirming that cell-cell contact is not mandatory for UC-MSC to inhibit T-cell proliferation and that soluble factors are involved. UC-MSC-CM at 100% concentration reduced circulating PSC CD4+IFNγ+expressing cells however no effect on CD4+TNF-α+, CD4+IL2+, CD8+TNF-α+, CD8+IFNγ+, CD8+IL2+expressing cells was seen suggesting that the effect of UC-MSC-CM on T cell responses may not be as strong as when UC-MSC are in direct contact with T cells. No difference in the effect on T cell responses was seen between UC-MSC-CM derived from resting and IFNγ stimulated UC-MSC.
Conclusion: UC-MSC are able to suppress the proliferation and activation of PSC patient derived circulating and intrahepatic T cells. This effect of UC-MSC on T cells is not disease specific. UC-MSC can act directly and indirectly on circulating T cells. Our results support efforts to assess in vivo the immunomodulatory effects of UC-MSC in patients with PSC.

Type of Work:

Thesis (Doctorates > M.D.)

Award Type:

Doctorates > M.D.

Supervisor(s):