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In patients with chronic kidney disease, the abnormal activation of inflammatory pathways is usually an important factor leading to renal fibrosis and further deterioration of renal function. Finding effective intervention targets of the inflammatory signaling pathway is an important way to treat chronic kidney disease. As a newly discovered lysosomal membrane protein, the correlation between SID1 transmembrane family member 2 (Sidt2) and the inflammatory signaling pathway has not been reported. The aim of this study was to investigate the effect of Sidt2 on inflammation by inhibiting the expression of the Sidt2 gene in a mouse mesangial cell line mediated by a lentiviral CRISPR/Cas9 vector. Hematoxylin and eosin staining and microscopy found that the mesangial cells lost their normal morphology after inhibiting the expression of Sidt2, showing that the cell body became smaller, the edge between the cells was unclear, and part of the nucleus was pyknotic and fragmented, appearing blue-black. The expressions of IKK β, p-IKK α/β, NF-κB p65, p-NF-κB p65, p-IκBα, IκBα, and TNF-α in the NF-κB pathway of the Sidt2-/- group were higher than those of the Sidt2+/+ group. p-Jak2 and IL6 increased in the Jak/Stat pathway, and p-ERK and p-P38 increased in the MAPK pathway. The expressions of IKK β, p-IKK α/β, NF-κB p65, p-NF-κB p65, p-IκBα, IκBα, and TNF-α in the NF-κB pathway of the Sidt2+/++LPS group were significantly higher than those in the Sidt2+/+ group. The expressions of IKK β, p-IKK α/β, NF-κB p65, p-NF-κB p65, p-IκBα, IκBα, and TNF-α in the Sidt2-/-+LPS group were higher than those in the Sidt2-/- group. The expressions of p-IKK α/β, NF-κB p65, p-NF-κB p65, p-IκBα, IκBα, and TNF-α in the Sidt2-/-+LPS group were higher than those in the Sidt2+/++LPS group. In the Jak/Stat pathway, the protein expressions of p-Jak2 and IL6 in the Sidt2+/++LPS group were higher than those in the Sidt2+/+ group. The expressions of p-Jak2 and IL6 in the Sidt2-/-+LPS group were higher than those in the Sidt2-/- group. The expressions of p-Jak2 and IL6 in the Sidt2-/-+LPS group were higher than those in the Sidt2+/++LPS group. The expressions of p-JNK, p-ERK, p-P38, and ERK in the MAPK pathway in the Sidt2+/++LPS group were higher than those in the Sidt2+/+ group. The expressions of p-JNK, p-ERK, p-P38, and ERK in the Sidt2-/-+LPS group were higher than those in the Sidt2-/- group. The expressions of p-JNK, p-ERK, p-P38, and ERK in the Sidt2-/-+LPS group were higher than those in the Sidt2+/++LPS group. These data suggested that deletion of the Sidt2 gene changed the three inflammatory signal pathways, eventually leading to the damage of glomerular mesangial cells in mice.

In most living organisms, isocitrate dehydrogenases (IDHs) convert isocitrate into ɑ-ketoglutarate (ɑ-KG). Phylogenetic analyses divide the IDH protein family into two subgroups: types I and II. Based on cofactor usage, IDHs are either NAD+-specific (NAD-IDH) or NADP+-specific (NADP-IDH); NADP-IDH evolved from NAD-IDH. Type I IDHs include NAD-IDHs and NADP-IDHs; however, no type II NAD-IDHs have been reported to date. This study reports a novel type II NAD-IDH from the marine bacterium Congregibacter litoralis KT71 (ClIDH, GenBank accession no. EAQ96042). His-tagged recombinant ClIDH was produced in Escherichia coli and purified; the recombinant enzyme was NAD+-specific and showed no detectable activity with NADP+. The Km values of the enzyme for NAD+ were 262.6±7.4 μM or 309.1±11.2 μM with Mg2+ or Mn2+ as the divalent cation, respectively. The coenzyme specificity of a ClIDH Asp487Arg/Leu488His mutant was altered, and the preference of the mutant for NADP+ was approximately 24-fold higher than that for NAD+, suggesting that ClIDH is an NAD+-specific ancestral enzyme in the type II IDH subgroup. Gel filtration and analytical ultracentrifugation analyses revealed the homohexameric structure of ClIDH, which is the first IDH hexamer discovered thus far. A 163-amino acid segment of CIIDH is essential to maintain its polymerization structure and activity, as a truncated version lacking this region forms a non-functional monomer. ClIDH was dependent on divalent cations, the most effective being Mn2+. The maximal activity of purified recombinant ClIDH was achieved at 35°C and pH 7.5, and a heat inactivation experiment showed that a 20-min incubation at 33°C caused a 50% loss of ClIDH activity. The discovery of a NAD+-specific, type II IDH fills a gap in the current classification of IDHs, and sheds light on the evolution of type II IDHs.

Wilson's disease (WD) is an autosomal recessive disease caused by mutation of the ATPase copper transporting ß (ATP7B) gene, resulting in abnormal copper metabolism. We aimed to investigate the protective effect of GanDouLing (GDL) on neural stem cell (NSC) function in a mouse model of WD. NSCs were treated with different concentrations of GDL alone or in combination with penicillamine, following which we evaluated cellular growth, apoptosis, and differentiation. Nuclear factor E2-related factor 2 (Nrf2) pathway and NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome activation were analyzed via Western blotting. Treatment with GDL alone or in combination with penicillamine significantly increased proliferation and inhibited apoptosis of NSCs in a dose-dependent manner. In addition, GDL treatment remarkably promoted differentiation of NSCs. Consistently, levels of class III ß-tubulin (Tuj1) and microtubule-associated protein 2 (MAP2) were significantly elevated, whereas glial fibrillary acidic protein (GFAP) levels were obviously suppressed in the presence of GDL or penicillamine. In vivo assays confirmed that GDL increased the ratio of Ki67+, Tuj1+, and MAP2+ cells and suppressed apoptosis in the hippocampal region in WD mice. Behavioral assays revealed that both GDL and penicillamine improved memory ability in WD models. Mechanistically, GDL treatment led to activation of Nrf2 signaling and suppression of the NLRP3 inflammasome in WD mice. Notably, inhibition of Nrf2 signaling reversed the protective effects of GDL on hippocampal NSCs. Collectively, these findings demonstrate that GDL exerts a protective effect on NSCs and promotes neurogenesis by targeting Nrf2 signaling and the NLRP3 inflammasome in WD. [ABSTRACT FROM AUTHOR]

Purpose: Wilson’s disease (WD) is a genetic disorder of copper metabolism with pathological copper accumulation in the brain. The purpose of the present study was to evaluate the relationship between the damaged white matter and the impaired cognitive function in WD patients. Materials and methods: Thirty WD adolescents and thirty age- and sex-matched healthy controls (HC) were enrolled. All subjects had received brain MRI, including conventional and diffusion-tensor imaging (DTI) scans. The DTI parameter of fractional anisotropy (FA) was calculated by diffusion kurtosis estimator software. The t test was used to compare the differences between two groups. The correlation between cognitive function and whiter matter disorders were analyzed by linear regression. The results of FA parameter and MD parameter intergroup analysis were both corrected with False Discovery Rate (FDR) simulations by SPSS. Results: WD adolescents showed significantly lower scores of time-based prospective memory (TBPM) and verbal fluency test (VFT) compared with HC.We found significantly higher FA in the right thalamus, right lentiform nucleus, left thalamus, left lentiform nucleus, and brain stem in WD adolescents. Besides, WD adolescents exhibited significantly lower FA in right cerebellum and cingulum and left middle frontal lobe compared with controls (P<0.05). There were significantly negative correlations between FA in bilateral lentiform and thalamus and cognitive impairment in WD adolescents (P<0.05). Conclusion: The whiter matter of WD adolescents was impaired and mainly distributed in subcortical brain regions. The impaired cognitive function was affected by the damaged whiter matter. The present study may be helpful for recognition and understanding of WD. [ABSTRACT FROM AUTHOR]