Background: Neuropsychiatric systemic lupus erythematosus (NPSLE) is a severe complication, which involves pathological damage to the brain and cognitive function. However, its exact mechanism of action still remains unclear. In this study, we explored the role of microglia in the cognitive dysfunction of NPSLE mice. We also analyzed and compared the metabolites in the hippocampal tissues of the lupus model and control mice.Methods: MRL/MpJ-Fas^lpr(MRL/lpr) female mice were used as the NPSLE mouse model. Metabolomics was used to assess hippocampal glycolysis levels. Glucose, lactic acid, IL-6, and IL-1β of the hippocampus were detected by ELISA. Based on the glycolysis pathway, we found that pyruvate kinase isoform M2 (PKM2) in the hippocampus was significantly increased. Thus, the expression of PKM2 was detected by qRT-PCR and Western blotting, and the localization of PKM2 in microglia (IBA-1⁺) or neurons (NeuN⁺) was assessed by immunofluorescence staining. Flow cytometry was used to detect the number and phenotype of microglia; the changes in microglial phagocytosis and the β-catenin signaling pathway were detected in BV2 cells overexpressing PKM2. For in vivo experiments, MRL/lpr mice were treated with AAV9-shPKM2. After 2 months, Morris water maze and conditional fear tests were applied to investigate the cognitive ability of mice; H&E and immunofluorescence staining were used to evaluate brain damage; flow cytometry was used to detect the phenotype and function of microglia; neuronal synapse damage was monitored by qRT-PCR, Western blotting, and immunofluorescence staining.Results: Glycolysis was elevated in the hippocampus of MRL/lpr lupus mice, accompanied by increased glucose consumption and lactate production. Furthermore, the activation of PKM2 in hippocampal microglia was observed in lupus mice. Cell experiments showed that PKM2 facilitated microglial activation and over-activated microglial phagocytosis via the β-catenin signaling pathway. In vivo, AAV9-shPKM2-treated mice showed decreased microglial activation and reduced neuronal synapses loss by blocking the β-catenin signaling pathway. Furthermore, the cognitive impairment and brain damage of MRL/lpr mice were significantly relieved after microglial PKM2 inhibition.Conclusion: These data indicate that microglial PKM2 have potential to become a novel therapeutic target for treating lupus encephalopathy.

Objective: Lupus nephritis (LN) is one of the most severe organ manifestations of systemic lupus erythematosus (SLE). Early identification of renal disease in SLE is important. Renal biopsy is currently recognized as the gold standard for diagnosing LN, however, it is invasive and inconvenient for dynamic monitoring. Urine has been considered more promising and valuable than blood in identifying inflamed kidney tissue. Here, we determine whether the signatures of tRNA-derived small noncoding RNA (tsRNA) in urinary exosomes can serve as novel biomarkers for the diagnosis of LN.Methods: tsRNA sequencing was performed in exosome extracted from pooled urine of 20 LN patients and 20 SLE without LN, and the top 10 upregulated tsRNAs were screened as candidate markers of LN. The candidate urinary exosomal tsRNAs were primarily elected by TaqMan probe-based quantitative reverse transcription-PCR (RT-PCR) in 40 samples (20 LN and 20 SLE without LN) in the training phase. In the validation phase, selected tsRNAs from the training phase were further confirmed in a larger cohort (54 LN patients and 39 SLE without LN). Receiver operating characteristic curve (ROC) analysis was conducted to evaluate the diagnostic efficacy.Results: Upregulated levels of tRF3-Ile-AAT-1 and tiRNA5-Lys-CTT-1 in the urinary exosomes were observed in LN compared with SLE without LN (P < 0.0001 and P < 0.001) and healthy controls (P < 0.01 and P < 0.01), with the area under the curve (AUC) of 0.777 (95% CI: 0.681-0.874, sensitivity 79.63%, specificity 66.69%) and 0.715 (95% CI: 0.610-0.820, sensitivity 66.96%, specificity 76.92%) for discriminating LN from SLE without LN patients. SLE patients with mild activity and moderate to severe activity had higher levels of urinary exosome derived tRF3-Ile AAT-1 (P = 0.035 and P < 0.001) and tiRNA5-Lys-CTT-1 (P = 0.021 and P < 0.001) compared with patients with no activity. Moreover, bioinformatics analysis revealed that both of the tsRNAs regulate the immune process by modulating metabolism and signal pathway.Conclusion: In this study, we demonstrated that urinary exosome tsRNAs can be served as noninvasive biomarkers for the efficient diagnosis and prediction of nephritis in SLE.

Soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) is a soluble form of TREM-1 released during inflammation. Elevated sTREM-1 levels have been found in neuropsychiatric systemic lupus erythematosus (NPSLE) patients; yet, the exact mechanisms remain unclear. This study investigated the role of sTREM-1 in brain damage and its underlying mechanism. The sTREM-1 recombinant protein (2.5 μg/3 μL) was injected into the lateral ventricle of C57BL/6 female mice. After intracerebroventricular (ICV) injection, the damage in hippocampal neurons increased, and the loss of neuronal synapses and activation of microglia increased compared to the control mice (treated with saline). In vitro. after sTREM-1 stimulation, the apoptosis of BV2 cells decreased, the polarization of BV2 cells shifted to the M1 phenotype, the phagocytic function of BV2 cells significantly improved, while the PI3K–AKT signal pathway was activated in vivo and in vitro. PI3K–AKT pathway inhibitor LY294002 reversed the excessive activation and phagocytosis of microglia caused by sTREM-1 in vivo and in vitro, which in turn improved the hippocampus damage. These results indicated that sTREM-1 activated the microglial by the PI3K–AKT signal pathway, and promoted its excessive phagocytosis of the neuronal synapse, thus inducing hippocampal damage. sTREM-1 might be a potential target for inducing brain lesions.

Recent studies indicate that Toll-like receptors (TLRs) and C-type lectin receptors (CLRs) can function as the signal of pattern recognition receptors, which play a pivotal role in the pathogenesis of the autoimmune disease. Systemic lupus erythematosus (SLE) is a classic autoimmune disease. Previous reports mainly focused on the potential role of TLRs in regulating the development of SLE, but little is known about the role of CLRs in the progression of SLE. Our previous studies showed that the inflammation-mediated accumulation of myeloid-derived suppressor cells (MDSCs) including granulocytic (G-MDSCs) and monocytic (M-MDSCs) participated in the pathogenesis of lupus. Mice deficient in Card9 (the downstream molecule of CLRs) were more susceptible to colitis-associated cancer via promoting the expansion of MDSCs. Whether the abnormal activation of CLRs regulates the expansion of MDSCs to participate in the pathogenesis of lupus remains unknown. In the present study, the expressions of CLRs were examined in both SLE patients and mouse models, revealing the expression of Dectin3 was positively correlated with SLEDAI. Dectin3 deficiency retarded the lupus-like disease by regulating the expansion and function of MDSCs. The mechanistic analysis revealed that Dectin3 deficiency promoted FoxO1-mediated apoptosis of MDSCs. Syk-Akt1-mediated nuclear transfer of FoxO1 increased in Dectin3-deficient MDSCs. Notedly, the accumulation of M-MDSCs mainly decreased in Dectin3^−/− lupus mice, and the nuclear transfer of FoxO1 negatively correlated with the expression of LOX-1 on M-MDSCs. The silencing of FoxO1 expression in Dectin3^−/− mice promoted the expansion of LOX-1⁺ M-MDSCs in vivo, and LOX-1⁺ M-MDSCs increased the differentiation of Th17 cells. Both LOX-1 expression on M-MDSCs and Dectin3 expression on MDSCs increased in patients with SLE. These data indicated that increased LOX-1⁺ M-MDSCs were related to the exacerbation of SLE development and might be potential target cells for the treatment of SLE.

Background: Neuropsychiatric systemic lupus erythematosus (NPSLE), with various morbidities and multiple manifestations in the central nervous system, remains a limited standard for diagnosis. Our study was to discover novel biomarkers for improving the diagnostic efficiency for NPSLE.Methods: We performed a quantitative planar protein antibody microarray to screen 1000 proteins in cerebrospinal fluid from controls, systemic lupus erythematosus (SLE, non-NPSLE) patients, and NPSLE patients. Differentially expressed proteins (DEPs) as candidate biomarkers were developed into a custom multiplexed protein antibody array for further validation in an independent larger cohort. Subsequently, we used least absolute shrinkage and selection operator regression (LASSO) analysis and multivariable logistic regression analysis for optimizing feature selection and constructing a diagnostic model. A receiver operating characteristic curve (ROC) was generated to assess the effectiveness of the models.Results: The expression of 29 proteins in CSF was significantly altered in the comparison of the three groups. We selected 17 proteins as candidate biomarkers in accordance with protein interaction analysis. In the larger cohort, we identified 5 DEPs as biomarkers for NPSLE, including TCN2, CST6, KLK5, L-selectin, and Trappin-2. The diagnostic model included 3 hub proteins (CST6, TCN2, KLK5) and was best at discriminating NPSLE from SLE patients. These CSF biomarkers were also highly associated with disease activity. In addition, there were 6 molecules with remarkable changes in NPSLE CSF and hippocampus, which indicated the consistency of the environment in the brain and the promising molecular targets in the pathogenesis of NPSLE.Conclusions: The dual-chips screening strategy demonstrated KLK5, L-selectin, Trappin-2, TCN2, and CST6 as CSF biomarkers for diagnosing NPSLE.

Background: The aim of this study was to evaluate serum biomarkers of systemic lupus erythematous (SLE) patients, with and without neuropsychiatric (NP) manifestation by high-resolution proteomic analysis. Methods: SLE patients with NP (NPSLE, n=9), SLE patients without NP (non-NPSLE, n=9) and healthy controls (HC, n=9) were prospectively enrolled in this study, and their plasma samples were collected and pooled into 3 NPSLE, 3 non-NPSLE and 3 HC samples for discovery profile. The TMT-LC-MS/MS-based proteomics approach was used to identify the differential proteome among the three matched groups, and the data were analyzed by bioinformatics tools, including Gene Ontology (GO) categories, Kyoto Encyclopedia of Genes and Genomes enrichment analysis, to explore canonical pathways and networks involved in the pathogenesis of NPSLE. To validation of differentially expressed proteomics results, four proteins were measured by ELISA. Results: There were altogether 223 differentially expressed proteins in NPSLE groups compared with healthy controls (HC), of which 96 proteins increased while 127 proteins decreased. Compared with non-NPSLE, there were only 49 differentially expressed proteins in NPSLE groups, of which 37 proteins increased while 12 proteins decreased. The significantly changed pathway that those proteins are involved in was complement and coagulation cascades in NPSLE group compared with health controls. However, we didn’t find significantly changed pathway between NPSLE group and non-NPSLE group. Five proteins were found significantly changed in all group-comparisons with consistent tendencies using Venn analysis, including Vitamin D binding protein (VDBP), C-reactive protein (CRP), KRT16, IGHV4-4 and CTRP3. Four proteins including CTRP3, VDBP, PAPPA and TRYP2 were selected to estimate the validity of the proteomics approach by ELISA. The expression levels of CTRP3 and TRYP2 were significantly changed in NPSLE patients compared with either HC or non-NPSLE patients. Conclusions: Our research has successfully established serum protein profiles of NPSLE and non-NPSLE patients through TMT technology and screened out five proteins significantly changed in group-comparisons with consistent tendencies. The pathway of complement and coagulation cascades may participate in pathogenesis of NPSLE and non-NPSLE.