Inhaled silica nanoparticles cause chronic kidney disease in rats

Abstract

Silica nanoparticles (SiNPs) released during the burning of sugarcane have been postulated to have a role in chronic kidney disease of unknown etiology. We tested the hypothesis that pristine SiNPs of the size present in sugarcane might cause chronic kidney injury when administered through the lung in rats. We administered 200- or 300-nm amorphous SiNPs twice weekly (4 mg/dose), or vehicle by oropharyngeal aspiration for 13 wk to rats followed by euthanasia after an additional 13 wk (26 wk total). Tissues were evaluated for the presence of SiNPs and evidence of histological injury. Both sizes of SiNPs caused kidney damage, with early tubular injury and inflammation (at week 13) that continued to inflammation and chronic fibrosis at week 26 despite discontinuation of the SiNP administration. Both sizes of SiNPs caused local inflammation in the lung and kidney and were detected in the serum and urine at week 13, and the 200-nm particles were also localized to the kidney with no evidence of retention of the 300-nm particles. At week 26, there was some clearance of the 200-nm silica from the kidneys, and urinary levels of SiNPs were reduced but still significant in both 200- and 300 nm-exposed rats. In conclusion, inhaled SiNPs cause chronic kidney injury that progresses despite stopping the SiNP administration. These findings support the hypothesis that human exposure to amorphous silica nanoparticles found in burned sugarcane fields could have a participatory role in chronic kidney disease of unknown etiology.NEW & NOTEWORTHY Inhalation of silica nanoparticles (SiNPs) released during the burning of sugarcane has been postulated to have a role in chronic kidney disease of unknown etiology (CKDu). We administered 200- and 300-nm amorphous SiNPs to rats by aspiration and observed kidney damage with tubular injury and inflammation that persisted even after stopping the SiNP exposure. These findings support the hypothesis that human exposure to SiNPs found in sugarcane ash could have a participatory role CKDu.

Keywords: Mesoamerican nephropathy; chronic kidney disease of unknown etiology; silica nanoparticles; sugarcane.

Graphical abstract

Figure 1.

Silica nanoparticle distribution in sugarcane ash from a sugarcane field in Nicaragua. Shown is a frequency histogram detailing the representative silica nanoparticle population in sugarcane ash from Nicaragua as detected by single particle inductively coupled mass spectrometry. The x-axis represents the size of the nanoparticle detected, and the y-axis represents the frequency that the nanoparticle at that size is counted. The red line represents a Gaussian function that the frequency histogram is fitted showing the representative particle population detected. The mean nanoparticle size in digested sugarcane ash as detected by dynamic light scattering was 180 ± 29 nm, and for single particle inductively coupled mass spectrometry was 221 ± 62 nm.

Figure 2.

Inhaled silica nanoparticles (SiNPs) induce tubular injury in rats. A–F: periodic acid-Schiff-stained kidney tissue of control rats (A and D), 200-nm SiNP-exposed rats (B and E), and 300-nm SiNP-exposed rats (C and F) at 13 wk (A–C) and 26 wk (D–F). Both 200- and 300-nm SiNP-exposed rats showed proximal tubular injury, with vacuolar degeneration, loss of brush borders, and a local inflammatory response in the interstitium (black arrows). The proximal tubular basement membrane was tortuous in association with some tubular atrophy (white arrow). n = 4 or 5 male rats/group. Data were analyzed by ANOVA with Bonferroni correction. Magnification: ×40. Scale bar = 100 µm.

Figure 3.

Immunohistochemistry stain of α-smooth muscle actin (SMC-αA) in the kidney tissue of control and silica nanoparticle (SiNP)-exposed rats. Shown are control rats (A and D), 200-nm SiNP-exposed rats (B and E), and 300-nm SiNP-exposed rats (C and F) at 13 wk (A–C) and 16 wk (D–F). There was a general increase in α-smooth muscle actin-positive myofibroblasts in SiNP-exposed rats. Black arrows show areas of α-actin positivity. Magnification: ×40. Scale bar = 100 µm. G and H show the differences between the control group and each SiNP-exposed group using color saturation. There was a significant difference in both 13- and 26-wk groups. n = 4 or 5 male rats/group. **P < 0.01, ***P < 0.001, and ****P < 0.0001 compared with the indicated group. Data were analyzed by ANOVA with Bonferroni correction.

Figure 4.

Immunohistochemistry stain of collagen type III (Coll-III) in the kidney tissue of control and silica nanoparticle (SiNP)-exposed rats. Shown are control rats (A and D), 200-nm SiNP-exposed rats (B and E), and 300-nm SiNP-exposed rats (C and F) at 13 wk (A–C) and 16 wk (D–F). There was a general increase in interstitial fibrosis (collagen type III-positive staining, black arrows) in silica-injected rats. Magnification: ×40. Scale bar = 100 µm. G and H show the differences between the control group and each SiNP-exposed group using color saturation. There was no significant difference in the 13-wk groups but there was in the 26-wk groups. *P < 0.05 and **P < 0.01 compared with the indicated group. n = 4 or 5 male rats/group. Data were analyzed by ANOVA with Bonferroni correction.

Figure 5.

Silica nanoparticles (SiNP) induce focal lung injury in the rat. A: periodic acid-Schiff staining of the lung of a 200-nm SiNP-exposed rat at 26 wk. The lesions were at the level of the respiratory bronchioles (red star) and were quite focal (yellow arrows). They did not occur in the peripheral lung (red bracket), and they were not associated with the vasculature (blue star). B: the SiNP-exposed rat showed focal areas of inflammation, noted by CD68 staining. The infiltration consisted of vesicle-filled macrophages. Some apoptotic cells were also present near the inflammatory foci and a few alveoli with broken walls (black arrow). Magnification: ×200. Scale bar = 50 µm. n = 4 or 5 male rats/group. C: quantification of F4-80 immunohistochemical positive macrophage lesions. A t test comparison of the groups was used (****P < 0.0001).

Figure 6.

Biodistribution of silica nanoparticles (SiNPs). The biodistribution of SiNPs in rats treated with 200- and 300-nm SiNPs is shown. Black circles represent individual rats treated with 200-nm SiNPs; black x’s represent individual rats treated with 300-nm SiNPs. Bars represent the mean of the data, and error bars represent 1 SD on either side of the mean. A: kidney deposition of 200- and 300 nm-SiNPs at 13 and 26 wk. B: lung deposition of 200- and 300-nm SiNPs at 13 and 26 wk. C: spleen deposition of 200- and 300-nm SiNPs at 13 and 26 wk. D: liver deposition of 200- and 300-nm SiNPs at 13 and 26 wk. E: urine clearance of 200- and 300-nm SiNPs at 13 and 26 wk. F: serum clearance of 200- and 300-nm SiNPs at 13 and 26 wk. *P < 0.05 compared with the indicated group via two-way ANOVA with a Tukey’s post hoc test. ns, not significant.