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. 2020 Apr 25:2020:2347827.
doi: 10.1155/2020/2347827. eCollection 2020.

CTLA4-Ig Abatacept Ameliorates Proteinuria by Regulating Circulating Treg/IL-17 in Adriamycin-Induced Nephropathy Rats

Lanjun Shuai  1   2 , Qia Cheng  1   2 , Tian Shen  1   2 , Zhuwen Yi  1   2 , Xiaochuan Wu  1   2
Affiliations

CTLA4-Ig Abatacept Ameliorates Proteinuria by Regulating Circulating Treg/IL-17 in Adriamycin-Induced Nephropathy Rats

Lanjun Shuai et al. Biomed Res Int. .

Abstract

Objective: This study is aimed at investigating the efficacy of CTLA4-Ig abatacept in normalizing proteinuria and its possible mechanism in adriamycin-induced nephropathy (AIN) rats.

Methods: A total of 32 healthy male Sprague-Dawley rats were randomly divided into a normal group, an AIN group, an abatacept group, and a prednisone group. Adriamycin (6.5 mg/kg) was injected once via the tail vein of rats to induce nephrotic syndrome. After adriamycin treatment, the abatacept group rats were given abatacept (0.5 mg/kg) once by intraperitoneal injection on day 14. In addition, the prednisone group rats were given prednisone (12.5 mg/kg) daily consecutively by gavage from day 14 to day 21. Blood, urine, and kidney tissue specimens were collected when sacrificed on day 21. The 24-hour urinary protein, serum albumin, cholesterol, creatinine, and urea nitrogen were then detected. An enzyme-linked immunosorbent assay was used to determine the level of urine CD80 and serum IL-17. Flow cytometry was used to investigate the prevalence of circulating Treg. Hematoxylin-eosin staining and electron microscopy were used for a renal histological study. Immunofluorescence staining was performed to confirm the CD80 expression of renal tissue.

Results: The 24-hour urinary protein of the abatacept group was significantly lower than that of the prednisone group and the AIN group. The level of urine CD80 of the abatacept group was significantly lower than that of the AIN group. Compared with the AIN group and the prednisone group, the circulating Treg prevalence of the abatacept group was significantly higher, while the level of serum IL-17 was lower. A negative kidney staining of CD80 expression was demonstrated in each group in this study. The 24-hour urinary protein had a negative correlation with the circulating Treg prevalence and Treg/IL-17 and a positive correlation with the urine CD80 and serum IL-17. Urinary CD80 had a positive correlation with serum IL-17 and no correlation with the circulating Treg prevalence.

Conclusions: CTLA4-Ig abatacept can reduce proteinuria of adriamycin-induced nephropathy rats, possibly at least partially as a result of regulating circulating Treg/IL-17. CTLA4-Ig abatacept could be a promising regimen for idiopathic nephrotic syndrome.

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Conflict of interest statement

No conflicts of interest, financial of otherwise, are declared by the authors.

Figures

Figure 1
Figure 1
AIN was induced by tail vein injection of adriamycin in rats. It is characterized by proteinuria (a), hypoalbuminemia (b), and hypercholesterolemia (c). Proteinuria was reduced using prednisone (d). Data are shown as means ± SDs, n = 8. ∗∗∗P < 0.001vs. the Nor group. Nor: normal group; AIN: adriamycin-induced nephropathy group. HE staining showed glomerular morphology with minimal alterations ((d) original magnification ×100; (e) original magnification ×400); electron microscopy showed foot-process effacement ((g) original magnification ×2500; (f) original magnification ×12000).
Figure 2
Figure 2
Abatacept reduced proteinuria of AIN rats. ∗∗∗P < 0.001vs. the AIN group and Pre group. ###P < 0.001vs. the AIN group. Nor: normal group; AIN: adriamycin-induced nephropathy group; Pre: prednisone group; Aba: abatacept group.
Figure 3
Figure 3
Abatacept decreased urinary CD80 of AIN rats. ∗∗P < 0.01vs. the AIN group. Nor: normal group; AIN: adriamycin-induced nephropathy group; Pre: prednisone group; Aba: abatacept group.
Figure 4
Figure 4
Abatacept regulated circulating Treg and IL-17 of AIN Rats. (a) CD3/CD45/SSC double-gating method was used for absolute count of CD3+CD45+FOXP3+ Treg cells. (b1) Treg of the normal group. (b2) Treg of the AIN group. (b3) Treg of the prednisone group. (b4) Treg of the abatacept group. (c1) Abatacept increased circulating Treg prevalence. ∗∗P < 0.01vs. the AIN group and P < 0.05vs. the Pre group. (c2) Abatacept decreased serum IL-17. ∗∗∗P < 0.001vs. the AIN group and Pre group. (c3) Abatacept regulated Treg/IL-17. ∗∗∗P < 0.001vs. the AIN group and Pre group. Nor: normal group; AIN: adriamycin-induced nephropathy group; Pre: prednisone group; Aba: abatacept group.
Figure 5
Figure 5
(a) Abatacept preserved renal function of AIN rats. ∗P < 0.05vs. the AIN group. Nor: normal group; AIN: adriamycin-induced nephropathy group; Pre: prednisone group; Aba: abatacept group. (b–e): abatacept maintained AIN rat kidney morphology. (b). Renal HE staining of the normal group. (c). Renal HE staining of the AIN group. (d). Renal HE staining of the prednisone group. (e). Renal HE staining of the abatacept group (original magnification ×400).
Figure 6
Figure 6
Negative CD80 staining of the kidneys in each group of rats. (a). CD80 immunofluorescence staining of the kidneys in the normal group. (b). CD80 immunofluorescence staining of the kidneys in the AIN group. (c). CD80 immunofluorescence staining of the kidneys in the prednisone group. (d). CD80 immunofluorescence staining of the kidneys in the abatacept group (original magnification ×400).
Figure 7
Figure 7
24-hour urinary protein showed a negative correlation with circulating Treg prevalence and Treg/IL-17 and a positive correlation with serum IL-17 and urinary CD80.
Figure 8
Figure 8
Urinary CD80 showed a positive correlation with serum IL-17 and Treg/IL-17, whereas there was no correlation with the circulating Treg prevalence.
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