Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, Victoria 3010, Australia; Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute, Parkville, Victoria 3010, Australia.
Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute, Parkville, Victoria 3010, Australia.
Transpl Immunol. 2024 Feb;82:101962. doi: 10.1016/j.trim.2023.101962. Epub 2023 Nov 23.
Human natural killer (NK) cells and gamma delta (γδ) T cells may impact outcomes of solid organ transplantation (SOT) such as lung transplantation (LTx) following the differential engagement of an array of activating and inhibitory receptors. Amongst these, CD16 may be particularly important due to its capacity to bind IgG to trigger antibody-dependent cellular cytotoxicity (ADCC) and the production of proinflammatory cytokines. While the use of immunosuppressive drugs (ISDs) is an integral component of SOT practice, their relative impact on various immune cells, especially γδT cells and CD16-induced functional responses, is still unclear.
The ADCC responses of peripheral blood NK cells and γδT cells from both healthy blood donors and adult lung transplant recipients (LTRs) were assessed by flow cytometry. Specifically, the degranulation response, as reflected in the expression of CD107a, and the capacity of both NK cells and γδT cells to produce IFN-γ and TNF-α was assessed following rituximab (RTX)-induced activation. Additionally, the effect of cyclosporine A (CsA), tacrolimus (TAC), prednisolone (Prdl) and azathioprine (AZA) at the concentration of 1 ng/ml, 10 ng/ml, 100 ng/ml, and 1000 ng/ml on these responses was also compared in both cell types.
Flow cytometric analyses of CD16 expresion showed that its expression on γδT cells was both at lower levels and more variable than that on peripheral blood NK cells. Nevertheless functional analyses showed that despite these differences, γδT cells like NK cells can be readily activated by engagement with RTX to degranulate and produce cytokines such as IFNg and TNF-a. RTX-induced degranulation by either NK cells or γδT cells from healthy donors was not impacted by co-culture with individual ISDs. However, CsA and TAC but not Prdl and AZA did inhibit the production of IFN-γ and TNF-α by both cell types. Flow cytometric analyses of RTX-induced activation of NK cells and γδT cells from LTRs suggested their capacity to degranulate was not markedly impacted by transplantation with similar levels of cells expressing CD107 pre- and post-LTx. However an impairment in the ability of NK cells to produce cytokines was observed in samples obtained post LTx whereas γδT cell cytokine responses were not significantly impacted.
In conclusion, the findings show that despite differences in the expression levels of CD16, γδT cells like NK cells can be readily activated by engagement with RTX and that in vitro exposure to CsA and TAC (calcineurin inhibitors) had a measurable effect on cytokine production but not degranulation by both NK cells and gdT cells from healthy donors. Finally the observation that in PBMC obtained from LTx recipients, NK cells but not γδT cells exhibited impaired cytokine reponses suggests that transplantation or chronic exposure to ISDs differentially impacts their potential to respond to the introduction of an allograft and/or transplant-associated infections.
自然杀伤 (NK) 细胞和γδ (γδ) T 细胞可能通过一系列激活和抑制受体的不同作用来影响实体器官移植 (SOT) 的结果，例如肺移植 (LTx)。在这些受体中，CD16 可能特别重要，因为它能够结合 IgG 触发抗体依赖性细胞毒性 (ADCC) 和产生促炎细胞因子。虽然免疫抑制药物 (ISD) 的使用是 SOT 实践的一个组成部分，但它们对各种免疫细胞的相对影响，尤其是 γδT 细胞和 CD16 诱导的功能反应，仍不清楚。
通过流式细胞术评估来自健康献血者和成人肺移植受者 (LTR) 的外周血 NK 细胞和 γδT 细胞的 ADCC 反应。具体而言，通过利妥昔单抗 (RTX) 诱导激活，评估 NK 细胞和 γδT 细胞的脱颗粒反应，反映在 CD107a 的表达上，以及 NK 细胞和 γδT 细胞产生 IFN-γ 和 TNF-α 的能力。此外，还比较了环孢素 A (CsA)、他克莫司 (TAC)、泼尼松龙 (Prdl) 和硫唑嘌呤 (AZA) 在 1ng/ml、10ng/ml、100ng/ml 和 1000ng/ml 浓度下对这两种细胞类型的这些反应的影响。
CD16 表达的流式细胞术分析表明，γδT 细胞上的 CD16 表达水平较低且更具变异性，而外周血 NK 细胞上的 CD16 表达水平较低且更具变异性。然而，功能分析表明，尽管存在这些差异，γδT 细胞与 NK 细胞一样，很容易通过与 RTX 结合而被激活，从而脱颗粒并产生细胞因子，如 IFNg 和 TNF-a。来自健康供体的 NK 细胞或 γδT 细胞与单个 ISD 共培养不会影响 RTX 诱导的脱颗粒。然而，CsA 和 TAC 但不是 Prdl 和 AZA 抑制了两种细胞类型 IFN-γ 和 TNF-α 的产生。来自 LTR 的 NK 细胞和 γδT 细胞的 RTX 诱导激活的流式细胞术分析表明，移植后细胞表达 CD107 的水平相似，其脱颗粒能力没有明显受到影响。然而，观察到 NK 细胞产生细胞因子的能力受损，而 γδT 细胞细胞因子反应没有受到显著影响。
总之，这些发现表明，尽管 CD16 的表达水平存在差异，但 γδT 细胞与 NK 细胞一样，可以通过与 RTX 结合而被轻易激活，并且在体外暴露于 CsA 和 TAC(钙调神经磷酸酶抑制剂)对健康供体 NK 细胞和 gdT 细胞的细胞因子产生而不是脱颗粒具有可衡量的影响。最后，观察到来自 LTx 受者的 PBMC 中 NK 细胞而非 γδT 细胞表现出受损的细胞因子反应表明，移植或慢性暴露于 ISD 会以不同的方式影响它们对同种异体移植和/或移植相关感染引入的反应潜力。
