Examining the Hepatic Immune System in Children With Liver Disease With Fine Needle Aspiration

Yüksel, Muhammed; Demirbaş, Burak; Mizikoglu, Ozlem; Tütüncü, Yıldız; Kanmaz, Turan; Oğuzkurt, Levent; Arıkan, Çiğdem

doi:10.1097/mpg.0000000000003353

Cited by 6 publications

(6 citation statements)

References 13 publications

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“…The biopsy and fine needle aspiration (FNA) procedures were both ultrasound guided under general and local anesthesia. As we recently published, FNA was used to obtain intrahepatic lymphocytes (IHL) ( 14 ). The aspirate was then filtered through a 40μm filter and collected in cold phosphate-buffered saline (PBS) containing 2% FBS (Biowest) ( 14 ).…”

Section: Methodsmentioning

confidence: 99%

“…As we recently published, FNA was used to obtain intrahepatic lymphocytes (IHL) ( 14 ). The aspirate was then filtered through a 40μm filter and collected in cold phosphate-buffered saline (PBS) containing 2% FBS (Biowest) ( 14 ).…”

Section: Methodsmentioning

confidence: 99%

“…To isolate peripheral blood mononuclear cells (PBMC), whole blood was diluted with PBS and layered onto Lymphoprep (GE), followed by density gradient centrifugation ( 15 ). Following a PBS wash, cells were resuspended in PBS, stained with viability dye EF780 (BD-eBioscience, ThermoFisher) before being incubated with cell surface antibodies specific for (regulatory) T-cells, mucosal-associated-invariant-T-cells (MAIT), (regulatory) B cells, and NK/NKT cells as previously described ( 7 , 14 – 16 ). These antibodies are; CD3 (clone HIT3a), CD161 (clone HP-3G10), CD20 (clone 2H7), CD24 (clone ML5), CD25 (clone M-A251), CD38 (clone HIT2), CD4 (clone A161A1), CD45RA (clone HI100), CD56 (clone 5.1H11), CD8 (clone HIT8a), HLA-DR (clone L243), and TCR Valpha7.2 (clone 3C10) ( Figure 1 ), and gating strategy is shown in Supplementary Figure 2 .…”

Section: Methodsmentioning

confidence: 99%

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Standard immunosuppressive treatment reduces regulatory B cells in children with autoimmune liver disease

et al. 2023

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IntroductionAutoimmune hepatitis (AIH) is a chronic liver disease caused by a perturbed immune system. The scarcity of short- and long-term immune monitoring of AIH hampered us to comprehend the interaction between immunosuppressive medication and immune homeostasis.Methods and patientsWe recruited children with AIH at the time of diagnosis and at the 1st, 3rd, 6th, 12th, 18th, and 24th months of immunosuppression (IS). We also enrolled children with AIH being on IS for >2 years. Children with drug-induced liver injury (DILI), and those receiving tacrolimus after liver transplantation (LT), were enrolled as disease/IS control subjects. Healthy children (HC) were also recruited. Peripheral blood mononuclear cells (PBMCs) were isolated from all participants. Healthy liver tissue from adult donors and from livers without inflammation were obtained from children with hepatoblastoma. By using flow cytometry, we performed multi-parametric immune profiling of PBMCs and intrahepatic lymphocytes. Additionally, after IS with prednisolone, tacrolimus, rapamycin, or 6-mercaptopurine, we carried out an in vitro cytokine stimulation assay. Finally, a Lifecodes SSO typing kit was used to type HLA-DRB1 and Luminex was used to analyze the results.ResultsUntreated AIH patients had lower total CD8 T-cell frequencies than HC, but these cells were more naïve. While the percentage of naïve regulatory T cells (Tregs) (CD4+FOXP3lowCD45RA+) and regulatory B cells (Bregs, CD20+CD24+CD38+) was similar, AIH patients had fewer activated Tregs (CD4+FOXP3highCD45RA-) compared to HC. Mucosal-associated-invariant-T-cells (MAIT) were also lower in these patients. Following the initiation of IS, the immune profiles demonstrated fluctuations. Bregs frequency decreased substantially at 1 month and did not recover anymore. Additionally, the frequency of intrahepatic Bregs in treated AIH patients was lower, compared to control livers, DILI, and LT patients. Following in vitro IS drugs incubation, only the frequency of IL-10-producing total B-cells increased with tacrolimus and 6MP. Lastly, 70% of AIH patients possessed HLA-DR11, whereas HLA-DR03/DR07/DR13 was present in only some patients.ConclusionHLA-DR11 was prominent in our AIH cohort. Activated Tregs and MAIT cell frequencies were lower before IS. Importantly, we discovered a previously unrecognized and long-lasting Bregs scarcity in AIH patients after IS. Tacrolimus and 6MP increased IL-10+ B-cells in vitro.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Standard immunosuppressive treatment reduces regulatory B cells in children with autoimmune liver disease

et al. 2023

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“…The expression and functions have significant differences between naïve and activated Tregs, suggesting another dimension that the tissue Tregs express higher levels of activation markers compared to blood. For example, activated effector Tregs (CD45RA-FOXP3high) express high levels of HLA-DR compared to naïve Tregs (CD45RA + FOXP3low) [22,23] and activated Treg cells play a major role in fibrogenesis. Although Treg cells only account for 5%-10% of the total CD4 + T lymphocytes in peripheral blood [24], their contribution to maintaining immune tolerance and homoeostasis cannot be ignored.…”

Section: Background and Role Of Tregsmentioning

confidence: 99%

Regulatory T cells (Tregs) in liver fibrosis

Qian

Zhou

et al. 2023

Cell Death Discov.

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The ability of the human liver to both synthesize extracellular matrix(ECM), as well as regulate fibrogenesis, are integral functions to maintaining homoeostasis. Chronic liver injury stimulates fibrogenesis in response to the imbalance between ECM accumulation and fibrosis resolution. Liver disease that induces fibrogenesis is associated with multiple risk factors like hepatitis infection, schistosomiasis, alcohol, certain drugs, toxicants and emerging aetiology like diabetes and obesity. The activation of hepatic stellate cells (HSCs), whose function is to generate and accumulate ECM, is a pivotal event in liver fibrosis. Simultaneously, HSCs selectively promote regulatory T-cells (Tregs) in an interleukin-2–dependent pattern that displays a dual relationship. On the one hand, Tregs can protect HSCs from NK cell attack, while on the other hand, they demonstrate an inhibitory effect on HSCs. This paper reviews the dual role of Tregs in liver fibrogenesis which includes its promotion of immunosuppression, as well as its activation of fibrosis. In particular, the balance between Tregs and the Th17 cell population, which produce interleukin (IL)-17 and IL-22, is explored to demonstrate their key role in maintaining homoeostasis and immunoregulation. The contradictory roles of Tregs in liver fibrosis in different immune microenvironments and molecular pathways need to be better understood if they are to be deployed to manage this disease.

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“…Thus far, liver FNAs have only been evaluated using flow cytometry to identify immune cell types based on conventional markers (Spaan et al, 2015;Pembroke et al, 2015;Gill et al, 2019;Yuksel et al, 2021). Therefore, the full cellular diversity of liver FNAs has not been mapped and, given the fragility of some cell types (hepatocytes) or adhesion to the parenchyma (macrophages, endothelial cells, stellate cells), it was not obvious if we could capture cellular diversity in serially collected FNAs to study changes in activation status by scRNAseq.…”

Section: Scrnaseq Of Liver Fnas Captures a Comprehensive Immunological Picture Of The Human Livermentioning

confidence: 99%

Pathogenic CD8 T cells defined by longitudinal liver sampling in chronic hepatitis B patients starting antiviral therapy

Nkongolo

Mahamed

Kuipery

et al. 2021

Preprint

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SummaryAccumulation of activated immune cells results in non-specific hepatocyte killing in chronic hepatitis B (CHB), leading to fibrosis and cirrhosis. We enrolled 15 CHB patients with active liver damage to receive antiviral therapy, and performed longitudinal liver sampling using fine-needle aspiration to investigate mechanisms of CHB pathogenesis in the human liver. Single-cell sequencing of total liver cells revealed a distinct liver-resident, polyclonal CD8 T cell population that was enriched at baseline and displayed a highly activated immune signature during liver damage. Cytokine combinations, identified by in silico prediction of ligand-receptor interaction, induced the activated phenotype in healthy liver CD8 T cells, resulting in non-specific Fas ligand-mediated killing of target cells. These results define a CD8 T cell population in the human liver that can drive pathogenesis, and a key pathway involved in their function in CHB patients.

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Examining the Hepatic Immune System in Children With Liver Disease With Fine Needle Aspiration

Cited by 6 publications

References 13 publications

Standard immunosuppressive treatment reduces regulatory B cells in children with autoimmune liver disease

Standard immunosuppressive treatment reduces regulatory B cells in children with autoimmune liver disease

Regulatory T cells (Tregs) in liver fibrosis

Pathogenic CD8 T cells defined by longitudinal liver sampling in chronic hepatitis B patients starting antiviral therapy

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