Changes of lysosomal membrane permeabilization and lipid metabolism in sidt2 deficient mice

Abstract

The SID1 transmembrane family member 2 (sidt2) deficient mouse model was used to investigate the function of sidt2 in lysosomal membrane permeabilization and lipid metabolism of liver tissue. The mouse model was established by Cre/LoxP technology. Enzymatic methods were used to analyze the sidt2^-/- mouse serum lipids, aspartate transaminase, alanine transaminase and serum bilirubin, compared with sidt2^+/+ mice. Defective lipid metabolism and damaged liver functions were observed in the sidt2^-/- mice. By using hematoxylin and eosin and Oil Red O staining, changes of morphology were observed in sidt2^-/- mice with optical microscopy. Transmission electron microscopy was also used. Hepatic steatosis and partial liver tissue apoptosis were observed. The tissue distribution of sidt2 protein and mRNA was measured in knockout mice. The results indicated that negligible sidt2 mRNA and protein expression were observed in sidt2^-/- mice, and that sidt2^-/- mice had abnormal liver functions. Transmission electron microscopy revealed membrane lipid droplets in the liver cell cytoplasm, and some apoptotic body formation. These results demonstrated that absence of the lysosomal membrane protein sidt2 led to changes in lysosomal membrane permeabilization and lipid metabolism.

Keywords: SID1 transmembrane family member 2−/− mice; lipid metabolism; lysosomal membrane permeabilization.

Figure 1.

The tissue distribution of sidt2 protein in sidt2 knockout mice. (A and B) Analysis of tissue distribution of sidt2 protein by western blotting. The (C) mRNA and (D) protein expression levels of sidt2 were examined via reverse transcription and western blotting, respectively. Sidt2, SID1 transmembrane family member 2. Data are presented as the mean ± standard error of the mean (n=8).

Figure 2.

Changes of serum biochemical parameters in sidt2^−/− mice. (A) T-CHOL, (B) TG, (C) AST, (D) ALT, (E) HDL-C, (F) LDL-C, (G) T-BIL, (H) I-BIL and (I) D-BIL concentrations were analyzed by enzymatic methods. Data are expressed as the mean ± standard error of the mean (n=8). **P<0.01 vs. sidt2^+/+. Sidt2, SID1 transmembrane family member 2; T-CHOL, total cholesterol; TG, total triglycerides; AST, aspartate transaminase; ALT, alanine transaminase; HDL-C, high density lipoprotein cholesterol; LDL-C, low density lipoprotein cholesterol; T-BIL, total bilirubin; I-BIL, indirect bilirubin; D-BIL, direct bilirubin.

Figure 3.

Morphological changes of the liver in sidt2^+/+ and sidt2^−/− mice. General morphology of (A) sidt2^+/+ and (B) sidt2^−/− mice liver samples. Hematoxylin and eosin staining of (C) sidt2^+/+ and (D) sidt2^−/− mice liver samples. Oil Red O staining of (E) sidt2^+/+ and (F) sidt2^−/− mice liver samples. Ultrastructure of (G) sidt2^+/+ and (H) sidt2^−/− mice liver samples by transmission electron microscopy. Sidt2, SID1 transmembrane family member 2.

Figure 4.

Mitochondrial damage and lysosomal membrane permeabilization-related indices in sidt2^−/− mice. The black arrows indicate where mitochondrial edema and destruction occurred. (A) Sidt2^+/+ mouse liver samples observed via TEM indicating the mitochondrial integrity. Ab is a magnification of Aa. (B) The sidt2^−/− mouse liver samples observed via TEM. (Ba) Edema. (Bb) Disrupted mitochondria. Bb and Bd are amplifications of Ba and Bc. Magnification, ×12,000. (C) The total Fe, Fe²⁺, and Fe³⁺ levels of the sidt2^+/+ and sidt2^−/− mouse lysosome samples. (D) H₂O₂ and NO levels in sidt2^+/+ and sidt2^−/− mouse liver tissue homogenates. Concentrations were analyzed via enzymatic methods. Data are expressed as the mean ± standard error of the mean (n=8). **P<0.01. Sidt2, SID1 transmembrane family member 2; TEM, transmission electron microscopy.