Macrophage autophagy protects against acute kidney injury by inhibiting renal inflammation through the degradation of TARM1.

Macroautophagy/autophagy activation in renal tubular epithelial cells protects against acute kidney injury (AKI). However, the role of immune cell autophagy, such as that involving macrophages, in AKI remains unclear. In this study, we discovered that macrophage autophagy was an adaptive response during AKI as mice with macrophage-specific autophagy deficiency (atg5-/-) exhibited higher serum creatinine, more severe renal tubule injury, increased infiltration of ADGRE1/F4/80+ macrophages, and elevated expression of inflammatory factors compared to WT mice during AKI induced by either LPS or unilateral ischemia-reperfusion. This was further supported by adoptive transfer of atg5-/- macrophages, but not WT macrophages, to cause more severe AKI in clodronate liposomes-induced macrophage depletion mice. Similar results were also obtained in vitro that bone marrow-derived macrophages (BMDMs) lacking Atg5 largely increased pro-inflammatory cytokine expression in response to LPS and IFNG. Mechanistically, we uncovered that atg5 deletion significantly upregulated the protein expression of TARM1 (T cell-interacting, activating receptor on myeloid cells 1), whereas inhibition of TARM1 suppressed LPS- and IFNG-induced inflammatory responses in atg5-/- RAW 264.7 macrophages. The E3 ubiquitin ligases MARCHF1 and MARCHF8 ubiquitinated TARM1 and promoted its degradation in an autophagy-dependent manner, whereas silencing or mutation of the functional domains of MARCHF1 and MARCHF8 abolished TARM1 degradation. Furthermore, we found that ubiquitinated TARM1 was internalized from plasma membrane into endosomes, and then recruited by the ubiquitin-binding autophagy receptors TAX1BP1 and SQSTM1 into the autophagy-lysosome pathway for degradation. In conclusion, macrophage autophagy protects against AKI by inhibiting renal inflammation through the MARCHF1- and MARCHF8-mediated degradation of TARM1.Abbreviations: AKI, acute kidney injury; ATG, autophagy related; Baf, bafilomycin A1; BMDMs, bone marrow-derived macrophages; CCL2/MCP-1, C-C motif chemokine ligand 2; CHX, cycloheximide; CQ, chloroquine; IFNG, interferon gamma; IL, interleukin; IR, ischemia-reperfusion; MAP1LC3/LC3, microtubule-associated protein 1 light chain 3; LPS, lipopolysaccharide; MARCHF, membrane associated ring-CH-type finger; NC, negative control; NFKB, nuclear factor of kappa light polypeptide gene enhancer in B cells; NLRP3, NLR family, pyrin domain containing 3; NOS2, nitric oxide synthase 2, inducible; Rap, rapamycin; Wort, wortmannin; RT-qPCR, real-time quantitative polymerase chain reaction; Scr, serum creatinine; SEM, standard error of mean; siRNA, small interfering RNA; SYK, spleen tyrosine kinase; TARM1, T cell-interacting, activating receptor on myeloid cells 1; TAX1BP1, Tax1 (human T cell leukemia virus type I) binding protein 1; TECs, tubule epithelial cells; TNF, tumor necrosis factor; WT, wild type.

Selenium nanoparticles alleviate renal ischemia/reperfusion injury by inhibiting ferritinophagy via the XBP1/NCOA4 pathway.

Acute kidney injury (AKI) is closely related to lysosomal dysfunction and ferroptosis in renal tubular epithelial cells (TECs), for which effective treatments are urgently needed. Although selenium nanoparticles (SeNPs) have emerged as promising candidates for AKI therapy, their underlying mechanisms have not been fully elucidated. Here, we investigated the effect of SeNPs on hypoxia/reoxygenation (H/R)-induced ferroptosis and lysosomal dysfunction in TECs in vitro and evaluated their efficacy in a murine model of ischemia/reperfusion (I/R)-AKI. We observed that H/R-induced ferroptosis was accompanied by lysosomal Fe2+ accumulation and dysfunction in TECs, which was ameliorated by SeNPs administration. Furthermore, SeNPs protected C57BL/6 mice against I/R-induced inflammation and ferroptosis. Mechanistically, we found that lysosomal Fe2+ accumulation and ferroptosis were associated with the excessive activation of NCOA4-mediated ferritinophagy, a process mitigated by SeNPs through the upregulation of X-box binding protein 1 (XBP1). Downregulation of XBP1 promoted ferritinophagy and partially counteracted the protective effects of SeNPs on ferroptosis inhibition in TECs. Overall, our findings revealed a novel role for SeNPs in modulating ferritinophagy, thereby improving lysosomal function and attenuating ferroptosis of TECs in I/R-AKI. These results provide evidence for the potential application of SeNPs as therapeutic agents for the prevention and treatment of AKI.

BIRC6 Modulates the Protein Stability of Axin to Regulate the Growth, Stemness, and Resistance of Renal Cancer Cells via the ß-Catenin Pathway.

The mechanism underlying the development of renal cell carcinoma (RCC) remains unclear, and effective prevention and therapeutic measures are lacking. BIRC6, a protein inhibitor of apoptosis, has attracted great interest. Our data indicated that overexpression of BIRC6 elevated cell growth, colony formation, migration, and invasion of cultured RCC cells, while siRNA knockdown of BIRC6 suppressed these processes. Additionally, BIRC6 was highly expressed in RCC clinical samples along with a downregulated level of Axin. Immunoprecipitation assays found that BIRC6 interacted with Axin and the two proteins colocalized within the cytoplasm of RCC cells. Overexpression of BIRC6 promoted the ubiquitination modification of Axin, while genetic knockdown of BIRC6 suppressed it. Furthermore, overexpression of BIRC6 significantly promoted the turnover of Axin, suggesting BIRC6's inhibitory effect on Axin protein stability. BIRC6 was also upregulated in cancer stem-like cells of RCC and increased the drug resistance of RCC cells against sunitinib. Western blotting assays showed that the overexpression of BIRC6 upregulated CXCR4 protein expression and activated the ß-catenin pathway. Two cell lines were then constructed with BIRC6 overexpressed by lentiviruses. Pharmacological administration of a Wnt/ß-catenin inhibitor, XAV-939, or genetic knockdown of ß-catenin inhibited cell growth, tumor sphere formation, colony formation, migration, and invasion of BIRC6-overexpressed cells. In vivo administration of XAV-939 markedly suppressed the tumorigenesis of BIRC6-overexpressed RCC cells in nude mice. In conclusion, we propose that BIRC6 activates the ß-catenin signaling pathway via mediating the ubiquitination and degradation of Axin, promoting the growth, stemness, and drug resistance of RCC cells. This project aims to elucidate the role of BIRC6 as a potential therapeutic target and provide new insights into the clinical treatment of RCC.

hUC-MSC transplantation therapy effects on lupus-prone MRL/lpr mice at early disease stages.

BACKGROUND: The efficacy of human umbilical cord mesenchymal stem cell (hUC-MSC) transplantation in treating systemic lupus erythematosus (SLE) has been confirmed by small-scale clinical trials. However, these trials focused on severe or refractory SLE, while few studies focused on mild SLE. Therefore, this study focused on the therapeutic effects of hUC-MSC transplantation in early-stage or mild MRL/lpr lupus model mice. METHODS: Commercially available hUC-MSCs were transplanted into 8-week-old MRL/lpr mice by tail vein injection. Flow cytometry was used to analyze B cells and their subsets in the peripheral blood. Further, plasma inflammatory factors, autoantibodies, and plasma biochemical indices were detected using protein chip technology and ELISA kits. In addition, pathological staining and immunofluorescence were performed to detect kidney injury in mice. RESULTS: hUC-MSC transplantation did not affect the mice's body weight, and both middle and high dose hUC-MSC transplantation (MD and HD group) actually reduced spleen weight. hUC-MSC transplantation significantly decreased the proportion of plasmablasts (PB), IgG1- PB, IgG1+ PB, IgG1+ memory B (MB) cells, IgG1+ DN MB, and IgG1+ SP MB cells. The hUC-MSC transplantation had significantly reduced plasma levels of inflammatory factors, such as TNF-α, IFN-γ, IL-6, and IL-13. Pathological staining showed that the infiltration of glomerular inflammatory cells was significantly reduced and that the level of glomerular fibrosis was significantly alleviated in hUC-MSC-transplanted mice. Immunofluorescence assays showed that the deposition of IgG and IgM antibodies in the kidneys of hUC-MSC-transplanted mice was significantly lower than in the control. CONCLUSION: hUC-MSC transplantation could inhibit the proliferation and differentiation of peripheral blood B cells in the early-stage of MRL/lpr mice, thereby alleviating the plasma inflammatory environment in mice, leading to kidney injury remission. The study provides a new and feasible strategy for SLE treatment.

Podocytes are likely the therapeutic target of IgA nephropathy with isolated hematuria: Evidence from repeat renal biopsy.

Background: The present study aimed to prove the progression of immunoglobulin A nephropathy (IgAN) patients with isolated hematuria based on repeat renal biopsy data for the first time. Methods: 29 IgAN patients with isolated hematuria who received repeat renal biopsies were analyzed retrospectively, while 29 non-isolated hematuria IgAN patients with similar age and background were randomly selected as the control group. Clinical parameters were collected at the time of biopsy. The treatment strategies (conservative treatment with RASS blocker or immunosuppressive treatment) were choosen according to the pathological results at the first renal biopsy. The activity and chronicity indexes of renal lesions were evaluated. Markers of cell inflammation and proliferation were tseted by immunochemistry. The ultrastructure of podocytes was observed by transmission electron microscopy (TEM). Podocyte and oxidative stress marker (NPHS2 and 4-HNE) were detected by immunofluorescence. Results: The IgAN patients with isolated hematuria had better clinical indicators than those with no-isolated hematuria, such as better renal function, higher albumin and lower uric acid. The interval between two biopsies in IgAN patients with isolated hematuria was 630 (interquartile range, 409.5-1,171) days. The hematuria of the patients decreased significantly from 30 (IQR, 4.00-35.00) RBC/ul in the first biopsy to 11 (IQR, 2.50-30.00) RBC/ul in the repeated biopsy (p < 0.05). The level of triglyceride decreased significantly (p < 0.05). The other clinical indicators were not statistically significant (p > 0.05). Deposits of IgA and C3 in the glomerulus were persistent. The activity index decreased, especially cellular crescent formation, while the chronicity index increased. The ultrastructure of podocytes was improved after treatment. The oxidative stress products of podocytes reduced after treatment. Conclusion: Although the clinical indicators of the IgAN patients with isolated hematuria were in the normal range, various acute and chronic pathological changes have occurred, and irreversible chronic changes have been progressing. Cell inflammation and proliferation persisted. Oxidative stress of podocytes was likely to be the therapeutic target. This study provided a strong basis for the progress of IgAN with isolated hematuria through pathological changes before and after treatment. This study will help clinicians recognize the harm of hematuria, change the traditional treatment concept, and help such patients get early treatment.

Efficacy and safety of sequential immunosuppressive treatment for severe IgA nephropathy: A retrospective study.

Background: This study compared the efficacy and safety of sequential immunosuppressive therapy in patients with non-end-stage IgA nephropathy (IgAN) with Lee's classification of IV ∼ V and provided evidence for the use of immunotherapy in patients with severe IgAN. Methods: We retrospectively analyzed the clinical data of patients with Lee's IV ∼ V non-end-stage IgA nephropathy. Results: 436 patients were diagnosed with IgAN, and 98 patients who met the inclusion criteria were included in this retrospective study. Of these, 17 were in the supportive care group, 20 in the P group (prednisone-only), 35 in P + CTX group (the prednisone combined with cyclophosphamide followed by mycophenolate mofetil), and 26 in the P + MMF group (prednisone combined with mycophenolate mofetil). The four groups showed differences in the segmental glomerulosclerosis score and the proportion of patients with Lee's grade IV (p < 0.05), but no differences in other indicators. Compared with the baseline values, urine protein-to-creatinine ratio (PCR) significantly decreased and serum albumin increased (p < 0.05), but there was no significant difference between the groups. The estimated Glomerular Filtration Rate (eGFR) of the P, P + MMF, and P + CTX groups were higher than that of the supportive care group at the 6th and 24th month after treatment (all p < 0.05). At the 24th month, the eGFR in the P + CTX group was higher than that in the P + MMF group (p < 0.05). The effective remission rate of the P + CTX group was higher than that of the supportive care group (p < 0.05). At 12 months, the effective remission rate of the P group was higher than that of the supportive care group (p < 0.05). At the 24th month, there was no significant difference in the effective remission rates among the three groups (P, P + MMF, and P + CTX). Nine patients with severe IgA nephropathy reached the endpoint. Conclusion: This study showed that immunosuppressive therapy insevere IgAN patient scan effectively reduce urinary protein, increase albumin, and protect renal function in the early stages of IgAN. P + CTX is the most commonly used, which has a high effective remission rate of urine protein and a low incidence of end-point events.

The development and improvement of immunodeficient mice and humanized immune system mouse models.

Animal models play an indispensable role in the study of human diseases. However, animal models of different diseases do not fully mimic the complex internal environment of humans. Immunodeficient mice are deficient in certain genes and do not express these or show reduced expression in some of their cells, facilitating the establishment of humanized mice and simulation of the human environment in vivo. Here, we summarize the developments in immunodeficient mice, from the initial nude mice lacking T lymphocytes to NOD/SCID rgnull mice lacking T, B, and NK cell populations. We describe existing humanized immune system mouse models based on immunodeficient mice in which human cells or tissues have been transplanted to establish a human immune system, including humanized-peripheral blood mononuclear cells (Hu-PBMCs), humanized hematopoietic stem cells (Hu-HSCs), and humanized bone marrow, liver, thymus (Hu-BLT) mouse models. The different methods for their development involve varying levels of complexity and humanization. Humanized mice are widely used in the study of various diseases to provide a transitional stage for clinical research. However, several challenges persist, including improving the efficiency of reconstructing the human B cell immune response, extending lifespan, improving the survival rate of mice to extend the observation period, and improving the development of standardized commercialized models and as well as their use. Overall, there are many opportunities and challenges in the development of humanized immune system mouse models which can provide novel strategies for understanding the mechanisms and treatments of human disease.

Life factors acting on systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a highly heterogeneous autoimmune disease that primarily affects women. Currently, in the search for the mechanisms of SLE pathogenesis, the association of lifestyle factors such as diet, cigarette smoking, ultraviolet radiation exposure, alcohol and caffeine-rich beverage consumption with SLE susceptibility has been systematically investigated. The cellular and molecular mechanisms mediating lifestyle effects on SLE occurrence, including interactions between genetic risk loci and environment, epigenetic changes, immune dysfunction, hyper-inflammatory response, and cytotoxicity, have been proposed. In the present review of the reports published in reputable peer-reviewed journals and government websites, we consider the current knowledge about the relationships between lifestyle factors and SLE incidence and outline directions of future research in this area. Formulation of practical measures with regard to the lifestyle in the future will benefit SLE patients and may provide potential therapy strategies.

The LncRNA CASC11 Promotes Colorectal Cancer Cell Proliferation and Migration by Adsorbing miR-646 and miR-381-3p to Upregulate Their Target RAB11FIP2.

BACKGROUND: We previously reported that the long non-coding RNA (lncRNA) CASC11 promotes colorectal cancer (CRC) progression as an oncogene by binding to HNRNPK. However, it remains unknown whether CASC11 can act as a competitive endogenous RNA (ceRNA) in CRC. In this study, we focused on the role of CASC11 as a ceRNA in CRC by regulating miR-646 and miR-381-3p targeting of RAB11FIP2. METHODS: We identified the target microRNAs (miRNAs) of CASC11 and the target genes of miR-646 and miR-381-3p using bioinformatic methods. A dual-luciferase reporter assay was performed to validate the target relationship. Quantitative real-time PCR (qRT-PCR), western blotting (WB), and immunohistochemistry (IHC) were used to measure the RNA and protein expression levels. Rescue experiments in vitro and in vivo were performed to investigate the influence of the CASC11/miR-646 and miR-381-3p/RAB11FIP2 axis on CRC progression. RESULTS: We found that CASC11 binds to miR-646 and miR-381-3p in the cytoplasm of CRC cells. Moreover, miR-646 and miR-381-3p inhibitors reversed the suppressive effect of CASC11 silencing on CRC growth and metastasis in vitro and in vivo. We further confirmed that RAB11FIP2 is a mutual target of miR-646 and miR-381-3p. The expression levels of CASC11 and RAB11FIP2 in CRC were positively correlated and reciprocally regulated. Further study showed that CASC11 played an important role in regulating PI3K/AKT pathway by miR-646 and miR-381-3p/RAB11FIP2 axis. CONCLUSION: Our study showed that CASC11 promotes the progression of CRC as a ceRNA by sponging miR-646 and miR-381-3p. Thus, CASC11 is a potential biomarker and a therapeutic target of CRC.

Humanized Mouse Models of Systemic Lupus Erythematosus: Opportunities and Challenges.

Animal models have played a crucial role in the understanding of the mechanisms and treatments of human diseases; however, owing to the large differences in genetic background and disease-specific characteristics, animal models cannot fully simulate the occurrence and progression of human diseases. Recently, humanized immune system mice, based on immunodeficient mice, have been developed that allow for the partial reconstruction of the human immune system and mimic the human in vivo microenvironment. Systemic lupus erythematosus (SLE) is a complex disease characterized by the loss of tolerance to autoantigens, overproduction of autoantibodies, and inflammation in multiple organ systems. The detailed immunological events that trigger the onset of clinical manifestations in patients with SLE are still not well known. Two methods have been adopted for the development of humanized SLE mice. They include transferring peripheral blood mononuclear cells from patients with SLE to immunodeficient mice or transferring human hematopoietic stem cells to immunodeficient mice followed by intraperitoneal injection with pristane to induce lupus. However, there are still several challenges to be overcome, such as how to improve the efficiency of reconstruction of the human B cell immune response, how to extend the lifespan and improve the survival rate of mice to extend the observation period, and how to improve the development of standardized commercialized models and use them. In summary, there are opportunities and challenges for the development of humanized mouse models of SLE, which will provide novel strategies for understanding the mechanisms and treatments of SLE.

Small GTPase RAB6 deficiency promotes alveolar progenitor cell renewal and attenuates PM2.5-induced lung injury and fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease characterized by chronic non-specific inflammation of the interstitial lung and extensive deposition of collagen fibers leading to destruction of lung function. Studies have demonstrated that exposure to fine particulate matter (PM2.5) increases the risk of IPF. In order to recover from PM2.5-induced lung injury, alveolar epithelial cells need to be repaired and regenerated to maintain lung function. Type 2 alveolar epithelial cells (AEC2) are stem cells in the adult lung that contribute to the lung repair process through complex signaling. Our previous studies demonstrated that RAB6, a RAS family member lowly expressed in lung cancer, inhibited lung cancer stem cell self-renewal, but it is unclear whether or not and how RAB6 may regulate AEC2 cell proliferation and self-renewal in PM2.5-induced pulmonary fibrosis. Here, we demonstrated that knockout of RAB6 inhibited pulmonary fibrosis, oxidative stress, and AEC2 cell death in PM2.5-injured mice. In addition, knockout of RAB6 decreased Dickkopf 1(DKK1) autocrine and activated proliferation, self-renewal, and wnt/ß-catenin signaling of PM2.5-injured AEC2 cells. RAB6 overexpression increased DKK1 autocrine and inhibited proliferation, self-renewal and wnt/ß-catenin signaling in AEC2 cells in vitro. Furthermore, DKK1 inhibitors promoted proliferation, self-renewal and wnt/ß-catenin signaling of RAB6 overexpressing AEC2 cells, and attenuated PM2.5-induced pulmonary fibrosis in mice. These data establish RAB6 as a regulator of DKK1 autocrine and wnt/ß-catenin signal that serves to regulate AEC2 cell proliferation and self-renewal, and suggest a mechanism that RAB6 disruption may promote AEC2 cell proliferation and self-renewal to enhance lung repair following PM2.5 injury.

DNA repair enzyme OGG1 promotes alveolar progenitor cell renewal and relieves PM2.5-induced lung injury and fibrosis.

Fine particulate matter (PM2.5) airborne pollution increases the risk of chronic respiratory diseases, such as idiopathic pulmonary fibrosis (IPF), which is characterized by non-specific inflammation of the interstitial lung and extensive deposition of collagen fibers. Type 2 alveolar epithelial cells (AEC2s) are alveolar stem cells in the adult lung that contribute to the lung repair process through complex signaling. Our previous studies demonstrated that OGG1, a kind of DNA repair enzyme, have a critical role in protecting cells from oxidative damage and apoptosis induced by PM2.5, but the contribution of OGG1 in proliferation and self-renewal of AEC2s is not known. Here, we constructed OGG1-/-mice to test the effect and mechanism of OGG1 on PM2.5-induced pulmonary fibrosis and injury in vivo. We detected proliferation and self-renewal of OGG1 overexpression or OGG1 knockout AEC2s after PM2.5 injury by flow cytometry and clone formation. We observed that knockout of OGG1 aggravated pulmonary fibrosis, oxidative stress, and AEC2 cell death in PM2.5-injured mice. In addition, OGG1 is required for the proliferation and renewal of AEC2s after PM2.5 injury. Overexpression of OGG1 promotes the proliferation and self-renewal of AEC2s by inhibiting PM2.5-mediated oxidative stress and NF-κB signaling hyperactivation in vitro. Furthermore, NF-κB inhibitors promoted proliferation and self-renewal of OGG1-deficient AEC2s cells after PM2.5 injury, and attenuated PM2.5-induced pulmonary fibrosis and injury in mice. These data establish OGG1 as a regulator of NF-κB signal that serves to regulate AEC2 cell proliferation and self-renewal, and suggest a mechanism that inhibition of the NF-κB signaling pathway may represent a potential therapeutic strategy for IPF patients with low-expression of OGG1.

[Enterovirus 71 can induce autophagy and apoptosis of THP-1 macrophages].

OBJECTIVE: To investigate enterovirus 71 (EV71)-induced of autophagy, apoptosis and the related signaling pathways in THP-1 macrophages. METHODS: THP-1 macrophages were infected with EV71 at the multiplicity of infection (MOI) of 0.1 for 2, 8 or 16 h, and the cell proliferation and toxicity were analyzed using CCK-8 kit. The intracellular viral nucleic acid in THP-1 macrophages were detected by fluorescence quantitative PCR, and the ultrastructural changes of the cells were observed using transmission electron microscopy. Cell apoptosis induced by EV71 infection was detected using Hoechst 33342 staining and AnnexinV/PI double staining. Western blotting was performed for analysis of changes in autophagy and apoptosis of the cells and in the expressions of the related proteins. The effect of EV71 infection on apoptosis of THP-1 macrophages incubated with 3-MA and Ac-DEVD-CHO inhibitor for 2 h was assessed using Western blotting. RESULTS: EV71 infection significantly lowered the cell survival rate of THP-1 macrophages at 2, 8 h and 16 h after the infection (P < 0.05). The total copy number of viral nucleic acid in THP-1 macrophages incubated with EV71 increased significantly and progressively over time (P < 0.01). Intracellular autophagosomes and virions could be seen in EV71-infected THP-1 macrophages. The total apoptotic rate of the infected cell also increased significantly over time (P < 0.01). EV71 infection significantly increased LC3 conversion (LC3-Ⅱ/ LC3-I) and the expression of cleaved caspase 3 protein and decreased the protein expressions of p62, Bcl-2 and caspase-3 (P < 0.01) without causing obvious changes in cleaved caspase-8 (P>0.05). 3-MA significantly inhibited the EV71-induced autophagy of THP-1 macrophages and reduced LC3 conversion (LC3-Ⅱ/LC3-I) and p62 protein expression at 8 h after EV71 infection (P < 0.01). Compared with DMSO, Ac-DEVD-CHO significantly inhibited EV71-induced apoptosis of THP-1 macrophages (15.5% vs 7.7%, P < 0.01). CONCLUSIONS: EV71 not only can infect and replicate in THP-1 macrophages, but also induces autophagy and cell apoptosis possibly by activating LC3/p62 autophagy pathway and caspase apoptosis pathway.

Altered frequency of peripheral B-cell subsets and their correlation with disease activity in patients with systemic lupus erythematosus: A comprehensive analysis.

Alternations of peripheral B-cell subsets are closely related to disease activity in systemic lupus erythematosus (SLE) and may also predict the relapse of SLE. In this study, we aimed to comprehensively analyse the frequency of peripheral B-cell subsets, and their correlation with disease activity in patients with SLE. The results showed that for B-cell subsets in the antigen-independent differentiation stage, the frequency of the peripheral hematopoietic stem cell (HSC) subset in all patients with SLE was significantly higher than that of control patients. Surprisingly, several significant correlations were noted in newly diagnosed patients with SLE including a positive correlation in the frequency of the common lymphoid progenitor cell (CLP) with cholesterol serum levels. For B-cell subsets in the antigen-dependent differentiation stage, the frequency of naïve B-cell (N-B) subsets in all patients with SLE was significantly higher than that in the control patients. Moreover, the frequency of plasmablasts positively correlated with the SLEDAI score in the newly diagnosed patients. For memory B-cell (M-B) subtypes in the antigen-dependent differentiation stage, the frequency of the class-switched memory B-cell (CSM-B) subsets was positively correlated with the serum levels of complement C3. Notably, the frequency of the CSM-B subset also negatively correlated with the SLEDAI score, whereas the non-class-switched memory B-cell (NSM-B) subset was positively correlated with the serum levels of haemoglobin. Collectively, these findings may contribute to a better understanding of the role played by different B-cell subsets in the pathogenesis of SLE.

8-Oxoguanine DNA glycosylase modulates the cell transformation process in pulmonary fibrosis by inhibiting Smad2/3 and interacting with Smad7.

The DNA repair enzyme 8-oxoguanine DNA glycosylase-1 (OGG1) is involved in early embryonic development, as well as in multiple conditions, including cardiac fibrosis, diabetes, and neurodegenerative diseases. But, function of OGG1 in pulmonary fibrosis was not entirely clear. In this study, we identified a novel function of OGG1 in the cell transformation process in pulmonary fibrosis. We demonstrated that OGG1 and Smad7 co-localize and interact in A549 cells. Bleomycin-induced pulmonary fibrosis was established in wild-type (WT) and Ogg1-/- mice. Upon treatment with transforming growth factor (TGF)-ß1, increased OGG1 expression was observed in WT mice with pulmonary fibrosis as well as in A549 cells, MRC-5 cells, and primary rat type II alveolar epithelial cells. The increased expression of OGG1 promoted cell migration, while OGG1 depletion decreased migration ability. Expression of the transformation-associated markers vimentin and alpha-smooth muscle actin were also affected by OGG1. We also observed that OGG1 promoted TGF-ß1-induced cell transformation and activated Smad2/3 by interacting with Smad7. The interaction between OGG1 and the TGF-ß/Smad axis modulates the cell transformation process in lung epithelial cells and fibroblasts. Moreover, we demonstrated that Ogg1 deficiency relieved pulmonary fibrosis in bleomycin-treated mice. Ogg1 knockout decreased the bleomycin-induced expression of Smad7 and phosphorylation of Smad2/3 in mice. These findings suggest that OGG1 has multiple biological functions in the pathogenesis of pulmonary fibrosis.

miR-155 increases stemness and decitabine resistance in triple-negative breast cancer cells by inhibiting TSPAN5.

Effective therapeutic targets for triple-negative breast cancer (TNBC), a special type of breast cancer (BC) with rapid metastasis and poor prognosis, are lacking, especially for patients with chemotherapy resistance. Decitabine (DCA) is a Food and Drug Administration-approved DNA methyltransferase inhibitor that has been proven effective for the treatment of tumors. However, its antitumor effect in cancer cells is limited by multidrug resistance. Cancer stem cells (CSCs), which are thought to act as seeds during tumor formation, regulate tumorigenesis, metastasis, and drug resistance through complex signaling. Our previous study found that miR-155 is upregulated in BC, but whether and how miR-155 regulates DCA resistance is unclear. In this study, we demonstrated that miR-155 was upregulated in CD24- CD44+ BC stem cells (BCSCs). In addition, the overexpression of miR-155 increased the number of CD24- CD44+ CSCs, DCA resistance and tumor clone formation in MDA-231 and BT-549 BC cells, and knockdown of miR-155 inhibited DCA resistance and stemness in BCSCs in vitro. Moreover, miR-155 induced stemness and DCA resistance by inhibiting the direct target gene tetraspanin-5 (TSPAN5). We further confirmed that overexpression of TSPAN5 abrogated the effect of miR-155 in promoting stemness and DCA resistance in BC cells. Our data show that miR-155 increases stemness and DCA resistance in BC cells by targeting TSPAN5. These data provide a therapeutic strategy and mechanistic basis for future possible clinical applications targeting the miR-155/TSPAN5 signaling axis in the treatment of TNBC.

Eupatilin Inhibits Renal Cancer Growth by Downregulating MicroRNA-21 through the Activation of YAP1.

Renal cell carcinoma (RCC) is the second most common human urinary tumor. Eupatilin is the main active ingredient of the traditional Chinese medicine Artemisia asiatica. The effect of Eupatilin on RCC and the underlying mechanism remain unknown. Here, we investigated the anticancer effects and mechanisms of Eupatilin in RCC in vivo and in vitro, laying an experimental foundation for the clinical application of Eupatilin in the treatment of RCC. The results showed that Eupatilin significantly inhibited 786-O cell viability and migration and promoted apoptosis. Eupatilin inhibited the expression of miR-21 in 786-O cells, and overexpression of miR-21 suppressed the effect of Eupatilin on viability, apoptosis, and migration in 786-O cells. Eupatilin inhibited the growth of renal tumors in nude mice by downregulating miR-21. YAP1, which was identified as a target of miR-21, showed significantly lower expression in RCC tissues than in healthy tissues. miR-21 significantly inhibited YAP1 protein expression in 786-O cells and tumor tissues isolated from nude mice, and YAP1 attenuated the effect of miR-21 on the viability, apoptosis, and migration of 786-O cells. In conclusion, Eupatilin inhibited the expression of miR-21, which mediated the proapoptotic and antimigratory effects of Eupatilin by suppressing YAP1 in renal cancer cells. These results suggested that Eupatilin could be a potent agent for the treatment of RCC.

Tetraspanin 1 as a mediator of fibrosis inhibits EMT process and Smad2/3 and beta-catenin pathway in human pulmonary fibrosis.

Tetraspanin 1(TSPAN1) as a clinically relevant gene target in cancer has been studied, but there is no direct in vivo or vitro evidence for pulmonary fibrosis (PF). Using reanalysing Gene Expression Omnibus data, here, we show for the first time that TSPAN1 was markedly down-regulated in lung tissue of patient with idiopathic PF (IPF) and verified the reduced protein expression of TSPAN1 in lung tissue samples of patient with IPF and bleomycin-induced PF mice. The expression of TSPAN1 was decreased and associated with transforming growth factor-ß1 (TGF-ß1 )-induced molecular characteristics of epithelial-to-mesenchymal transition (EMT) in alveolar epithelial cells (AECs). Silencing TSPAN1 promoted cell migration, and the expression of alpha-smooth muscle actin, vimentin and E-cadherin in AECs with TGF-ß1 treatment, while exogenous TSPAN1 has the converse effects. Moreover, silencing TSPAN1 promotes the phosphorylation of Smad2/3 and stabilizes beta-catenin protein, however, overexpressed TSPAN1 impeded TGF-ß1 -induced activation of Smad2/3 and beta-catenin pathway in AECs. Together, our study implicates TSPAN1 as a key regulator in the process of EMT in AECs of IPF.

lncINS-IGF2 Promotes Cell Proliferation and Migration by Promoting G1/S Transition in Lung Cancer.

Long noncoding RNAs are capable of regulating gene expression at multiple levels. These RNA molecules are also involved in a variety of physiological and pathological processes. Emerging data demonstrate that a series of differentially expressed long noncoding RNAs are implicated in tumorigenesis. In the present study, we used microarray analysis to identify long noncoding RNAs that are dysregulated in non-small-cell lung cancer when compared to normal lung tissues. Accordingly, we performed quantitative real-time polymerase chain reaction to analyze the levels of long noncoding RNA and the cis target gene. We further found the oncogene property of long noncoding RNA that long noncoding RNA downexpression inhibits non-small-cell lung cancer cells proliferation and migration based on 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide and colony formation assays and wound healing as well as transwell assays. The influence of long noncoding RNA on cell cycle of non-small-cell lung cancer cells is also analyzed by flow cytometry. Among the dysregulated long noncoding RNAs, we identified INS-IGF2 readthrough, transcript variant 1, noncoding RNA (NR_003512.3) is upregulated in non-small-cell lung cancer tissues, the cis gene of which is insulin-like growth factor 2 gene hinted by bioinformatics analysis. We also observed that downregulation of INS-IGF2 readthrough, transcript variant 1, noncoding RNA reduces insulin-like growth factor 2 messenger RNA expression. Furthermore, INS-IGF2 readthrough, transcript variant 1, noncoding RNA downregulation suppresses non-small-cell lung cancer cell proliferation and migration. This downregulation results in a concomitant inhibition of the G1/S transition in non-small-cell lung cancer cells. Our findings suggest that INS-IGF2 readthrough, transcript variant 1, noncoding RNA may be an oncogene involved in the development of lung cancer. Therefore, we speculate that INS-IGF2 readthrough, transcript variant 1, noncoding RNA represents a potential therapeutic target for lung cancer.

VP1-binding protein glucose-regulated protein 78 as an important mediator for enterovirus 71 infecting human brain microvascular endothelial cells.

Purpose: Enterovirus 71 (EV71) is one of the main pathogens causing hand, foot and mouth disease, which could even induce severe brain damage in some patients. As the underlying mechanism of the invasion and replication process still remains largely unknown, we investigated the role of candidate proteins expressed during EV71 invasion in human brain microvascular endothelial cells (HBMECs) to delineate the pathophysiological mechanism of EV-71 infection. Materials and Methods: Ninety-one candidate EV71-associated proteins which could bind the major capsid protein (viral protein 1 [VP1]) of EV71 on the HBMEC were identified by applying an analysis of glutathione-S-transferase pull-down coupling with liquid chromatography-electrospray ionisation-tandem mass spectrometry (LC-ESI-MS/MS). Seventy-eight kDa glucose-regulated protein 78 (GRP78) binding to the VP1 protein was further validated by co-immunoprecipitation, immunofluorescence and western blot analysis. To explore the role of GRP78 in EV71 infection, GRP78 was knocked down and overexpressed in HBMEC and was verified by TCID50 assay. Results: LC-ESI-MS/MS-identified 91 proteins were subjected to gene ontology analysis, and on molecular and biological function analysis revealed GRP78 act as an important binding protein in mediating EV71 infection. In addition, immunofluorescence demonstrated the co-localisation of GRP78 and VP1 in cytoplasm of the infected HBMEC. The TCID50 assay showed that knockdown of GRP78 could attenuate the replication capacity of EV71 in HBMEC, and the overexpression could increase the virus titre in HBEMC at 24 h post-infection suggesting that GRP78 was associated with the replication capacity of EV71 in HBMEC. Conclusion: These findings provided evidence that GRP78 plays an important role during the progression of EV71 infection as a mediator in HBMEC.