Altered CD73-Adenosine Signaling Linked To Cardiovascular And Infection In Patients Undergoing Hemodialysis

In the past decade, T cell dysfunction observed in patients undergoing hemodialysis (HD) has been linked to the extremely high morbidity of cardiovascular events (CVEs) and infections. The cell-surface 5′-nucleotidase CD73 sets the balance between pro-inflammatory nucleotides and anti-inflammatory adenosine. As an integral component of the purinergic system, CD73 functions as a rate-limiting enzyme that catalyzes extracellular ATP metabolism to form adenosine. When exposed to inflammatory, hypoxic, metabolic, and other types of stress, increased ATP release from stressed or dying cells represents a pro-inflammatory signal. This signal is terminated when ATP is metabolized to the anti-inflammatory mediator adenosine, thus preventing excessive activation of tissue defense mechanisms. Therefore, CD73-adenosine signaling is crucial to maintaining equilibrium within the immune system, exerting potent anti-inflammatory effects to protect against cellular damage and facilitate tissue repair. Here, we investigated the correlation between CD73-adenosine signaling and complications of CVEs and infection in patients with ESRD and explored the immunological mechanisms underlying the tendency for severe complications in these patients.

A total of 395 patients who had been receiving HD for at least six months were evaluated for relative levels of CD73+ T cells and followed for one year to document CVEs and infections. Thirty age-and sex-matched healthy individuals served as controls for the immunological test validations. Differences in the levels of CD73-expressing T cells between healthy controls and patients undergoing HD were compared. The relation between CD73+ T cells and clinical outcomes was analyzed by the Kaplan–Meier curve and univariate Cox regression.In addition, a 5/6 nephrectomy (5/6 Nx) rat model and cecal ligation and puncture (CLP) were used to verify the effect of chronic kidney disease (CKD) and sepsis on CD73-adenosine signaling.

1. Advanced age and HD are associated with lower CD73+T cell fraction

T cell flow cytometry was performed on samples of 395 patients in November 2020. Values of percentages of T cell subset were listed in Table 1. Thirty age-and sex-matched healthy individuals served as controls for the immunological test validations. Linear regression analysis showed that the percentage of the CD8+CD73+ T cells negatively correlated with age in both patients and healthy controls, whereas the fraction of the CD4+CD73+T cells correlated with age in healthy controls only (Figure 1). The uremia effect on CD73 expression was more evident in the CD4+ T cells, and the proportion of CD4+CD73+ cells was significantly lower in patients on HD than in healthy controls (p < 0.05; Figure 2). The fractions of CD8+CD73+ T cells were not significantly different between the HD group and healthy controls.

Table1. Percentages of T cell subsets in patients on HD

1Mann–Whitney U test was used to investigate differences in T cell subsets between genders. *p<0.05. **p<0.01. NS: non-significant, p> 0.1.

HD: hemodialysis.

Figure 1. Correlations between the percentages of CD73+T cells and age.

Figure 2. Correlations between the percentages of CD73+T cells and hemodialysis (HD) condition.

2. Lower CD73+T cell fraction is associated with systemic inflammation and T cell terminal differentiation in patients on HD

We examined the association between CD73+T cell subsets and circulating inflammatory markers at enrolment. As shown in Table 3, the levels of high sensitivity C-reactive protein (hsCRP), tumor necrosis factor-α(TNF-α), and interleukin 6 (IL-6) were significantly and negatively correlated with CD8+CD73+T cell percentages (p < 0.05), whereas soluble interleukin-2 receptor (sIL-2R) levels were significantly and negatively associated with fractions of both CD4+CD73+ and CD8+CD73+T cells (p < 0.05). The percentages ofCD28−T cells, as an important marker of T cell terminal differentiation, were significantly and negatively correlated with fractions of both CD4+CD73+ and CD8+CD73+T cells (p < 0.01).

Table 2. Correlations between the percentages of CD73+T cells and inflammatory markers in patients on HD

HD: hemodialysis; hsCRP: high sensitivity-C reactive protein;IL-6: interleukin 6;sIL-2R:soluble interleukin-2 receptor; TNF-α: tumor necrosis factor-α.

Spearman rank analysis was used to investigate the relationship between inflammatory markers and percentages of CD4+CD73+and CD8+CD73+T cells. *p<0.05. **p<0.01. NS: non-significant, p> 0.1.

3. Lower fraction of CD4+CD73+ T cells predicts CVEs in patients on HD

Percentages of CD4+CD73+ and CD8+CD73+T cells significantly and negatively correlated with CVE incidence. Patients with a lower CD4+CD73+ or CD8+CD73+ T cell percentage had a significantly higher CVE incidence, especially in the younger age group (Figure 3). In the univariate Cox proportional hazards model, lower percentages of both CD4+CD73+and CD8+CD73+T cells were CVE predictors, in addition to older age, history of CVD or diabetes mellitus, lower serum prealbumin and creatinine levels, and higher levels of hsCRP and N-terminal pro-brain natriuretic peptide (NT-proBNP) [Table 4]. In the multivariate Cox hazard model, a lower log-transformed percentage of CD4+CD73+ T cells was independently associated with CVEs (HR 0.530, 95% CI 0.287‒0.979; p = 0.043).

Figure 3. Cardiovascular event (CVE)-free survival curves for different age-CD73+ Tcell percentage groups.

Table 3. Cox hazard model for CVEs in patients on HD

Lower CD73+T cell levels are strongly associated with CVEs and infection complications in patients undergoing HD. Altered CD73-adenosine signaling likely plays a substantial role in immune dysfunction in CKD.