Gadolinium Chloride Rescues Niemann⁻Pick Type C Liver Damage

Abstract

Niemann⁻Pick type C (NPC) disease is a rare neurovisceral cholesterol storage disorder that arises from loss of function mutations in the NPC1 or NPC2 genes. Soon after birth, some patients present with an aggressive hepatosplenomegaly and cholestatic signs. Histopathologically, the liver presents with large numbers of foam cells; however, their role in disease pathogenesis has not been explored in depth. Here, we studied the consequences of gadolinium chloride (GdCl₃) treatment, a well-known Kupffer/foam cell inhibitor, at late stages of NPC liver disease and compared it with NPC1 genetic rescue in hepatocytes in vivo. GdCl₃ treatment successfully blocked the endocytic capacity of hepatic Kupffer/foam measured by India ink endocytosis, decreased the levels CD68-A marker of Kupffer cells in the liver-and normalized the transaminase levels in serum of NPC mice to a similar extent to those obtained by genetic Npc1 rescue of liver cells. Gadolinium salts are widely used as magnetic resonance imaging (MRI) contrasts. This study opens the possibility of targeting foam cells with gadolinium or by other means for improving NPC liver disease. Synopsis: Gadolinium chloride can effectively rescue some parameters of liver dysfunction in NPC mice and its potential use in patients should be carefully evaluated.

Keywords: Niemann–Pick type C disease; cholesterol; gadolinium chloride; liver disease; therapeutic option.

Figure 1

Progressive increase of hepatic CD68 positive cells in Niemann–Pick type C (NPC) mice. (A) CD68 staining of livers from 6-, 7-, and 9-week-old (wk) wild-type (WT) and Npc1^−/− mice. Scale bars are 100 µm. (B) Three-dimensional reconstructions of liver sections from 9-week-old WT and Npc1^−/− mice.

Figure 2

Foam-cell-associated NPC liver pathology is corrected by genetic NPC1 rescue. (A) Timeline of the duration and age of doxycycline (DOX) treatment in mice. (B) Expression of the NPC1-YFP transgene, driven by Rosa-rtTA in ROSA26-rtTA-M2 and TRE-Npc1-YFP (R; N), WT, or Npc1^−/− mice, was induced with DOX on day 49. After 1 week of treatment, animals were sacrificed and livers were processed for histological analysis by Hematoxylin and eosin staining. Scale bars are 100 µm. (C) Quantification of the number of foam cells in each condition. *** p < 0.0001.

Figure 3

GdCl₃ treatment decreases Kupffer/foam cells endocytosis. (A) Timeline of the duration and age of GdCl₃ treatment in mice. The time of India ink injection is indicated (postnatal day (P)56). As a control, animals were treated with a saline solution (NaCl). (B) India ink deposits were visualized with light microcopy. Scale bars are 100 µm. (C) CD68 and filipin staining in GdCl₃- and NaCl-treated Npc1^−/− mice. Scale bars are 100 µm. Bottom images show merged CD68 and filipin staining and the areas marked with yellow squares were magnified below. The arrowheads show cells positive for CD68 and filipin. (D) Quantification of % filipin/CD68 positive cells. (E) Quantification of the area covered by CD68 cells.

Figure 4

Foam-cell-associated NPC liver pathology is improved by GdCl₃ treatment. (A) Timeline of the duration and age of different treatments in mice. (B) Alanine transaminase (ALT) levels in the different groups. Data are presented as means ± SD. (n = 3–5 animals/group, two-way ANOVA, and Tukey’s multiple comparison post-test). *** p < 0.0001.