BAFF Promotes FLS Activation Through BAFFR-Mediated Non-canonical NF-κB Pathway and the Effects of CP-25.

B cell activating factor (BAFF) has been shown to play a key role in regulating B cell function, but little is known about whether BAFF affects the function of fibroblast-like synoviocyte (FLS), an effector cell of rheumatoid arthritis (RA). CP-25, a new ester derivative of paeoniflorin, could alleviate the arthritis symptoms of collagen-induced arthritis (CIA) mice by inhibiting BAFF-mediated abnormal activation of B cells. In this study, we aimed to understand the mechanism by which BAFF activates FLS and the effect of CP-25 on FLS function. Therefore, the proliferation and migration abilities of FLS and key proteins on the non-canonical NF-κB pathway were examined. The results showed that compared with the FLS of normal rats/OA patients, the expression of BAFF-R, TRAF2, NIK, p-IKKα, P100, and P52 was higher in the FLS of AA rats/RA patients, while the expression of TRAF3 was lower. And, BAFF promotes FLS activation by activating the non-canonical NF-κB signaling pathway. Meanwhile, BAFFR-siRNA inhibited the proliferation of FLS and the activation of non-canonical NF-κB signaling in FLS induced by BAFF. Additionally, CP-25 could inhibit abnormal proliferation and migration of FLS by regulating non-canonical NF-κB signaling. We concluded that BAFF may act as an important role in facilitating the function of FLS through the BAFFR-mediated non-canonical NF-κB pathway, which would be useful for revealing the pathological mechanism of RA. And CP-25 may become a potential new drug for the treatment of RA, providing a scientific basis for the development of new drugs to treat RA.

Advance in bone destruction participated by JAK/STAT in rheumatoid arthritis and therapeutic effect of JAK/STAT inhibitors.

Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic joint inflammation and bone erosion. The bones in the human body are constantly undergoing bone remodeling throughout their lives, which is the process of bone resorption by osteoclasts to damaged bone tissue and new bone formation by osteoblasts. Osteoblasts (OBs) are the main functional cells in bone formation, responsible for the synthesis, secretion and mineralization of the bone matrix. On the contrary, osteoclasts (OCs) mediate bone breakdown during natural bone turnover, but excessive breakdown occurs in RA. Under the condition of RA inflammation, many molecules, such as IL-1ß, IL-6, TNF-α, IL-17 and hypoxia-inducible factor-1α (HIF-1α) are produced that could mediate bone loss. Studies have shown that cytokines mainly promote the formation of OCs and play a role in bone resorption by stimulating OBs to express receptor activator of NF-κB ligand (RANKL). JAK/STAT plays a crucial role in the process of bone destruction. And JAK/STAT pathway mediates the RANKL/receptor activator of NF-κB (RANK)/osteoprotegerin (OPG) axis. Tofacitinib, Baricitinib, Peficitinib and Filgotinib are now being used in patients with moderate to severe RA, as well as in patients with RA who have an inadequate response to methotrexate therapy and bone destruction. Currently, Tofacitiniband Baritinib areapprovedfor thetreatmentof moderate-to-severely active RA. JAK inhibitors have been reported to have better efficacy and lower adverse effects compared with methotrexate and adalimumab. In addition, two JAK inhibitors are currently in development: the JAK1 selective Upadacitinib, and the JAK3 selective inhibitor Decernotinib. In addition to the above JAK inhibitors, some small molecular compounds inhibit bone destruction by inhibiting the Phosphorylation of STAT3. In this paper, the research progress of bone destruction participated by JAK/ STAT in rheumatoid arthritis and therapeutic effect of JAK/STAT inhibitors were reviewed.

IgD promotes pannus formation by activating Wnt5A-Fzd5-CTHRC1-NF-κB signaling pathway in FLS of CIA rats and the regulation of IgD-Fc-Ig fusion protein.

Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by joint inflammation, synovial hyperplasia, cartilage degeneration, bone erosion, and pannus. Immunoglobulin D (IgD) plays an important role in autoimmune diseases although the content of it in vivo is low. Increased concentrations of anti-IgD autoantibodies have been detected in many RA patients. IgD-Fc-Ig fusion protein is constructed by connecting human IgD Fc domain and IgG1 Fc domain, which specifically block the IgD/ IgDR pathway and regulate the function of cells expressing IgDR to treat RA. The expression levels of Wnt5A and Frizzled 5 are higher in RA synovial tissue specimens. The complex of Wnt5A-Fzd5-LRP5/6-CTHRC1 promotes the expression of hypoxia inducible factor-1α by activating nuclear factor kappa-B (NF-κB), leading to high expression of VEGF and participating in angiogenesis. VEGF is the strongest angiogenic factor found so far. Here, we aimed to explore whether IgD participates in synovitis by binding to IgDR and regulating the activation of Wnt5A-Fzd5-CTHRC1-NF-κB signaling pathway in fibroblast synovial cells (FLSs), whether IgD-Fc-Ig fusion protein inhibits VEGF production in FLS of CIA and explore mechanism. We found that IgDR is expressed on MH7A and FLS. IgD promotes VEGF expression by activating Wnt5A-Fzd5-CTHRC1-NF-κB signaling pathway in MH7A and FLS. After activation of Fzd5 with Wnt5A, IgD-Fc-Ig reduced VEGF-A level in the culture supernatant of MH7A stimulation by IgD. The expressions of CTHRC1, Fzd5, p-P65 and VEGF in MH7A and FLSs were down-regulated after IgD-Fc-Ig treatment. IgD-Fc-Ig suppressed the combination of CTHRC1 and Fzd5 as well. By using the animal model, we demonstrated that IgD-Fc-Ig suppress ankle CTHRC1 and Fzd5 production resulted in inhibition of index of joint inflammation of CIA rats, which were consistent with vitro results. Conclusively, IgD-Fc-Ig inhibits IgD and Wnt5A-induced angiogenesis and joint inflammation by suppressing the combination of CTHRC1 and Fzd5. Our results show that IgD-Fc-Ig exerts its suppressive effect on IgD and Wnt5A by Wnt5A-Fzd5-CTHRC1-NF-κB signaling pathway.

Lactate up-regulates the expression of PD-L1 in kidney and causes immunosuppression in septic Acute Renal Injury.

BACKGROUND: This study aims to explore the mechanism of immunosuppression in septic Acute Renal Injury (AKI) and the role of programmed death-1 (PD-1/PD-L1) pathway in septic AKI. METHODS: This study established a septic AKI model by Cecal ligation and puncture (CLP) in C57/B6 mice, ELISA was used to test the level of lactate and creatinine in serum, blood was collected for flow cytometry and kidney samples for Western blot analyses. This study further analyzed the expression of PD-L1 in kidney and the expression of PD-1 in CD4+, CD8+ T cell, and the number of CD3+ T cells to identify apoptosis in T cells in the blood. RESULTS: The CLP sepsis model induced AKI in C57/B6 mice; The expression of PD-1 and PD-L1 were increased in septic AKI mice; PD-1/PD-L1 induced apoptosis in T cells: the number of lymphocytes decreased by 64%, while the number of CD3+ T cells decreased by 27% compared with the sham group; Results also indicated that lactate up-regulates expression of PD-L1 in the kidney. CONCLUSIONS: Lactate activated PD-1/PD-L1 pathway can induce immunosuppression by inducing apoptosis in lymphocytes in septic AKI. Moreover, blocking the receptor of lactate or PD-1/PD-L1 might be a new therapy for septic AKI.

Uncoupling Protein 2 Drives Myocardial Dysfunction in Murine Models of Septic Shock.

Cardiac dysfunction is a major component of sepsis-induced multiorgan failure in critical care units. Uncoupling protein 2 (UCP2) involves immune response, regulation of oxidative stress, and maintenance of mitochondrial membrane potential as well as energy production. However, whether and how UCP2 plays roles in the development of septic cardiac dysfunction are largely unknown. Here, intraperitoneal injection of LPS significantly activated UCP2 expression accompanied by a significant decrease of cardiac function and caused a significantly lower survival rate in mice. Of note, knockdown of UCP2 through a cardiotropic adenoassociated viral vector carrying a short hairpin RNA (shRNA) specifically targeting the UCP2 evoked resistance to LPS-triggered septic cardiac dysfunction and lethality in vivo. Moreover, UCP2 deficiency ameliorated the reduced levels of intracellular ATP in the LPS-challenged heart tissues and preserved mitochondrial membrane potential loss in primary adult mouse cardiomyocytes in LPS-challenged animals. Mechanistically, we confirmed that the inhibition of UCP2 promoted autophagy in response to LPS, as shown by an increase in LC3II and a decrease in p62. At last, the autophagy inhibitor 3-MA abolished UCP2 knockdown-afforded cardioprotective effects. Those results indicate that UCP2 drives septic cardiac dysfunction and that the targeted induction of UCP2-mediated autophagy may have important therapeutic potential.

Effect of splenectomy on attenuation of LPS-induced AKI through GTS-21-induced cholinergic anti-inflammatory pathway.

This work was undertaken to explore the role of splenectomy on attenuation of lipopolysaccharide (LPS)-induced acute kidney injury (AKI) through GTS-21-induced cholinergic anti-inflammatory pathway. C57BL/6 mice were used to construct models of sepsis-induced renal injury. HE, Tunel and blood assays were used to determine the success of the model. The animals were examined after splenectomy with or without LPS and GTS-21+LPS treatments. The pathological changes and apoptosis in the renal tissue were detected using HE and Tunel assays. The contents of creatinine (Cr) and cystatin-C (Cys-C) were measured using ELISA. The expression of IL-6, NF-kB p65, Caspase-3, anti-apoptotic protein Bcl-2, apoptotic protein Bax and α7nAChR was quantified using qRT-PCR. The expression of Bcl-2, Bax, Caspase-3, IL-6, NF-kB p65, α7nAChR and p-STAT3 was using assessed using Western blot analysis. HE, Tunel, BUN and serum creatinine (SC) assay showed that renal injury models were successfully established. Compared with the control, the apoptosis in the LPS group was significantly increased and decreased after GTS-21 treatment. However, splenectomy combined with GTS-21 increased the apoptosis, indicating that splenectomy could partially offset the anti-apoptosis effect of GTS-21. In animals treated with LPS, the contents of Cr and Cys-C increased significantly. These contents reduced following GTS-21 treatment, but increased after splenectomy. After LPS treatment, the expression of IL-6, NF-kB p65, p-STAT3, Caspase-3 and Bax was significantly up-regulated, while the expression of α7nAChR and Bcl-2 significantly down-regulated. Compared with LPS treated mice, splenectomy reduced the expression of IL-6, NF-kB p65 and p-STAT3, suggesting that splenectomy inhibits the activation of α7nAChR pathway by the GTS-21. It is clear that GTS-21 effectively attenuates LPS-induced renal injury; splenectomy suppresses the anti-inflammatory and anti-apoptosis activity and renal protective effect of GTS-21. On other hand, splenectomy reduces the production of inflammatory cytokines in the circulation, and has certain protective effect on the kidney. Therefore, the impact of splenectomy on LPS-induced AKI depends on the strength of the two aspects.

Cross-sectional survey on adult acute kidney injury in Chinese ICU: the study protocol (CARE-AKI).

INTRODUCTION: Acute kidney injury (AKI) is one of the most serious syndromes in intensive care unit (ICU) patients, and is a mysterious problem in clinical practice worldwide. Due to unknown aetiology and mechanism, awareness of AKI diagnosis and treatment in China varies, resulting in underestimated incidence and poor prognosis. To solve this problem, we design this national survey of AKI in adult ICUs. Various indexes are included and analysed to classify the epidemiology of adult AKI in Chinese ICUs, including AKI aetiology, risk factors, mortality, prognosis, therapeutic strategies and cognition of ICU medical staff. METHODS: A multicentre, cross-sectional survey, which will involve about 35 hospitals and 6147 patients from 23 provinces, 4 municipalities and 5 autonomous regions, is planned. All patients who meet the inclusion criteria are eligible to apply for enrolment in the study, which cover baseline demographics, clinical performance, and follow-up related to diagnosis and treatment. CONCLUSION: The study is expected to fill the gap between China and developed countries, and to provide a theoretical foundation for developing more scientific and standardised approaches to AKI diagnosis and treatment. ETHICS AND DISSEMINATION: Ethical approval was obtained from the ethics committee of Harbin Medical University Cancer Hospital (registration number KY2017-21). The findings of this review will be communicated through peer-reviewed publications and scientific presentations. TRIAL REGISTRATION NUMBER: ChiCTR-EOC-17013133; Pre-results.

[Deguelin inhibits proliferation and regulates the expression of MCM3-CDC45 in MCF-7 and H1299 cells in vitro].

OBJECTIVE: To observe the effects of deguelin on the proliferation of breast cancer MCF-7 cells and lung cancer H1299 cells in vitro and the expression of minichromosome maintenance protein 3 (MCM3) and CDC45 in the cells. METHODS: MTT assay was used to evaluate the proliferation of MCF-7 and H1299 cells exposed to different concentrations of deguelin for 48, 72 or 96 h. The growth of the cells was observed microscopically and the changes of MCM3 and CDC45 expressions in MCF-7 and H1299 cells following deguelin treatment were detected with fluorescence quantitative PCR. RESULTS: The proliferation of MCF-7 cells was significantly inhibited by exposure to 0.25, 0.5, 1, 5, 10, 30, and 50 µmol/L deguelin for 48, 72, and 96 h in a concentration- and time-dependent manner. In MCF-7 cells, the IC50 of deguelin at 48, 72, and 96 h was 9, 3, and 2 µmol/L, respectively. Deguelin treatments of H1299 cells at 0.5, 1, 5, 10, 30, 50, and 100 µmol/L also resulted in a concentration- and time-dependent inhibition of the cell growth with an IC50 at 96 h of 2 µmol/L. Optical microscopy of the cells revealed a decreased number of viable cells with obvious cell shrinkage following deguelin treatments. The expression of MCM3 and CDC45 were significantly reduced in the cells after deguelin treatments. CONCLUSION: Deguelin can inhibit the proliferation of MCF-7 and H1299 cells in vitro and down-regulate the expression of MCM3 and CDC45 in the cells.

Deguelin-induced blockade of PI3K/protein kinase B/MAP kinase signaling in zebrafish and breast cancer cell lines is mediated by down-regulation of fibroblast growth factor receptor 4 activity.

Deguelin, a natural component derived from leguminous plants, has been used as pesticide in some regions. Accumulating evidence show that deguelin has promising chemopreventive and therapeutic activities against cancer cells. This study shows that low concentrations of deguelin can lead to significant delay in zebrafish embryonic development through growth inhibition and induction of apoptosis. Furthermore, we identified fibroblast growth factor receptor 4 (FGFR4) as the putative target of deguelin. The candidate was initially identified by a microarray approach and then validated through in vitro experiments using hormone-responsive (MCF-7) and nonresponsive (MDA-MB-231) human breast cancer cell lines. The results show that deguelin suppressed cell proliferation and induced apoptosis in both cancer cell lines, but not in Hs 578Bst cells, by blocking PI3K/AKT and mitogen-activated protein kinases (MAPK) signaling. The FGFR4 mRNA and protein level also diminished in a dose-dependent manner. Interestingly, we found that forced FGFR4 overexpression attenuated deguelin-induced proliferative suppression and apoptotic cell death in both zebrafish and MCF-7 cell lines, p-AKT and p-ERK levels were restored upon FGFR4 overexpression. Taken together, our results strongly suggest that deguelin inhibition of PI3K/AKT and MAPK signaling in zebrafish and breast cancer cell lines is partially mediated through down-regulation of FGFR4 activity.