IRGM/Irgm1 facilitates macrophage apoptosis through ROS generation and MAPK signal transduction: Irgm1+/- mice display increases atherosclerotic plaque stability.

Rationale: Atherosclerosis plaque rupture (PR) is the pathological basis and chief culprit of most acute cardiovascular events and death. Given the complex and important role of macrophage apoptosis and autophagy in affecting plaque stability, an important unanswered question include is whether, and how, immunity-related GTPase family M protein (IRGM) and its mouse orthologue IRGM1 affect macrophage survival and atherosclerotic plaque stability. Methods: To investigate whether serum IRGM of ST-segment elevation myocardial infarction (STEMI) patients is related to plaque morphology, we divided 85 STEMI patients into those with and without plaque rupture (PR and non-PR, respectively) based on OCT image analysis, and quantified the patients' serum IRGM levels. Next, we engineered Irgm1 deficient mice (Irgm1+/-) and chimera mice with Irgm1 deficiency in the bone marrow on an ApoE-/- background, which were then fed a high-fat diet for 16 weeks. Pathological staining was used to detect necrotic plaque cores, ratios of neutral lipids and cholesterol crystal, as well as collagen fiber contents in these mice to characterize plaque stability. In addition, immunofluorescence, immunohistochemical staining and western blot were used to detect the apoptosis of macrophages in the plaques. In vitro, THP-1 and RAW264.7 cells were stimulated with ox-LDL to mimic the in vivo environment, and IRGM/IRGM1 expression were modified by specific siRNA (knockdown) or IRGM plasmid (knocked-in). The effect of IRGM/Irgm1 on autophagy and apoptosis of macrophages induced by ox-LDL was then evaluated. In addition, we introduced inhibitors of the JNK/p38/ERK signaling pathway to verify the specific mechanism by which Irgm1 regulates RAW264.7 cell apoptosis. Results: The serum IRGM levels of PR patients is significantly higher than that of non-PR patients and healthy volunteers, which may be an effective predictor of PR. On a high-fat diet, Irgm1-deficient mice exhibit reduced necrotic plaque cores, as well as neutral lipid and cholesterol crystal ratios, with increased collagen fiber content. Additionally, macrophage apoptosis is inhibited in the plaques of Irgm1-deficient mice. In vitro, IRGM/Irgm1 deficiency rapidly inhibits ox-LDL-induced macrophage autophagy while inhibiting ox-LDL-induced macrophage apoptosis in late stages. Additionally, IRGM/Irgm1 deficiency suppresses reactive oxygen species (ROS) production in macrophages, while removal of ROS effectively inhibits macrophage apoptosis induced by IRGM overexpression. We further show that Irgm1 can affect macrophage apoptosis by regulating JNK/p38/ERK phosphorylation in the MAPK signaling pathway. Conclusions: Serum IRGM may be related to the process of PR in STEMI patients, and IRGM/Irgm1 deficiency increases plaque stability. In addition, IRGM/Irgm1 deficiency suppresses macrophage apoptosis by inhibiting ROS generation and MAPK signaling transduction. Cumulatively, these results suggest that targeting IRGM may represent a new treatment strategy for the prevention and treatment of acute cardiovascular deaths caused by PR.

Treatment of the bone marrow stromal stem cell supernatant by nasal administration-a new approach to EAE therapy.

INTRODUCTION: Multiple sclerosis (MS) is one of the most common autoimmune diseases of the central nervous system (CNS). CNS has its own unique structural and functional features, while the lack of precision regulatory element with high specificity as therapeutic targets makes the development of disease treatment in the bottleneck. Recently, the immunomodulation and neuroprotection capabilities of bone marrow stromal stem cells (BMSCs) were shown in experimental autoimmune encephalomyelitis (EAE). However, the administration route and the safety evaluation limit the application of BMSC. In this study, we investigated the therapeutic effect of BMSC supernatant by nasal administration. METHODS: In the basis of the establishment of the EAE model, the BMSC supernatant were treated by nasal administration. The clinical score and weight were used to determine the therapeutic effect. The demyelination of the spinal cord was detected by LFB staining. ELISA was used to detect the expression of inflammatory factors in serum of peripheral blood. Flow cytometry was performed to detect pro-inflammatory cells in the spleen and draining lymph nodes. RESULTS: BMSC supernatant by nasal administration can alleviate B cell-mediated clinical symptoms of EAE, decrease the degree of demyelination, and reduce the inflammatory cells infiltrated into the central nervous system; lessen the antibody titer in peripheral bloods; and significantly lower the expression of inflammatory factors. As a new, non-invasive treatment, there are no differences in the therapeutic effects between BMSC supernatant treated by nasal route and the conventional applications, i.e. intraperitoneal or intravenous injection. CONCLUSIONS: BMSC supernatant administered via the nasal cavity provide new sights and new ways for the EAE therapy.

LncRNA SNHG3 enhances the malignant progress of glioma through silencing KLF2 and p21.

As a newly discovered long non-coding RNA, small nucleolar RNA host gene 3 (SHNG3) has been reported to be dysregulated in certain cancers. Nevertheless, the details about clinical values and biological effects of SNHG3 on glioma are still covered. In this paper, we determined the expression level of SNHG3 in glioma tissues and cells and evaluated the effect of SNHG3 expression on the prognosis of glioma patients. The functional assays were applied to define the effects of SNHG3 on the biological behaviors in glioma including cell proliferation, cell cycle, and apoptosis. It was revealed that SNHG3 was much more enriched in glioma tissues and cell lines than in normal ones. Furthermore, gain- or loss-of-function experiments indicated that the up-regulation of SNHG3 promoted cell proliferation, accelerate cell cycle progress, and repressed cell apoptosis. The mechanistic assays disclosed that SNHG3 facilitated the malignant progression of glioma through epigenetically repressing KLF2 and p21 via recruiting enhancer of zeste homolog 2 to the promoter of KLF2 and p21. Generally, it was exposed that SNHG3 might function as an oncogene in glioma and could be explored as a potential prognostic biomarker and therapeutic target for glioma.

Functional network analysis reveals biological roles of lncRNAs and mRNAs in MOG35-55 specific CD4+T helper cells.

Long non-coding RNAs have the potential to regulate immune responses. Their impact on multiple sclerosis has remained elusive. For illustrating their roles in experimental autoimmune encephalomyelitis (EAE) pathogenesis, we investigated the differential expression of lncRNAs and mRNAs in CD4+Th cells obtained from myelin oligodendrocytic glycoprotein35-55(MOG35-55)-induced EAE and complete Freund's adjuvant (CFA) controls. We observed differential expression of 1112 lncRNAs and 519 mRNAs in CD4+Th cells. The functional network showed lncRNAs had the capacity to modulate EAE pathogenesis via regulating many known EAE regulators such as Ptpn6. Predicting the function of lncRNAs demonstrated that dysregulated lncRNAs were closely associated with the development of EAE. These dysregulated lncRNAs may have function in EAE and they could be novel biomarkers and therapeutic targets of EAE. However, the precise mechanisms and biological functions of these specific lncRNAs in EAE pathogenesis require further study.

Irgm1 is required for the inflammatory function of M1 macrophage in early experimental autoimmune encephalomyelitis.

The classically activated (M1) macrophage has been shown to play an indispensable role in experimental autoimmune encephalomyelitis (EAE), the animal model of multiple sclerosis (MS). However, most studies focus on the effect of macrophage on CNS demyelination of EAE; whether the M1 macrophage participates in early EAE and the molecular mechanism underlying remains unclear. Here, we showed that the immunity-related GTPase family member 1 (Irgm1), also known as LRG-47, was expressed in M1 macrophages of draining lymph nodes (dLNs) from C57BL/6 mice with early EAE, and the IRGM1 heterozygote substantially reduced M1 macrophage accumulation in dLNs and spleen of the primary EAE stage. In vitro silence of IRGM1 in M1 macrophages impaired NOS2 expression and inflammatory cytokine release. We also found that IRGM1 knockout (Irgm1-/-) in M1 macrophages increased Akt activation but attenuated NF-κB p65 activation, which may reveal Irgm1-mediated mechanisms of action. Interestingly, macrophage depletion in vivo inhibited Th1/Th17 differentiation in the spleen and promoted regulatory T cell (Treg) polarization in dLNs at 7 d postimmunization (dpi). Moreover, we observed that M1 macrophages in vitro promoted Th1/Th17 differentiation, which was reversed by treatment with IRGM1 small interfering RNA (siRNA), anti-TNF-α, or anti-IL-1ß mAb. These results suggest that the M1 macrophage may promote Th1/Th17 cell differentiation during the early EAE, and the proinflammatory function of M1 cells requires Irgm1.

Irgm1 promotes M1 but not M2 macrophage polarization in atherosclerosis pathogenesis and development.

BACKGROUND AND AIMS: Atherosclerosis is a chronic inflammatory vascular disease related to macrophages uptake of low-density lipoprotein and their subsequent transformation into foam cells. M1 (inflammatory)/M2 (anti-inflammatory) balance was suggested to impact disease progression. In this study, we investigated whether the immunity related GTPase (Irgm1) regulates macrophage polarization during atherosclerosis development. METHODS: We used apolipoprotein E (ApoE) knockout and Irgm1 haplodeficient mice and induced atherosclerosis with high-cholesterol diet for the indicated months. Atherosclerotic arteries were collected from patients undergoing vascular surgery, to determine the lesional expression of Irgm1 and distribution of M1/M2 populations. RESULTS: Our results showed that IRGM/Irgm1 expression was increased in atherosclerotic artery samples (1.7-fold, p=0.0045) compared with non-atherosclerotic arteries, which was consistent with findings in the murine experimental atherosclerosis model (1.9-fold, p=0.0002). IRGM/Irgm1 expression was mostly found in lesional M1 macrophages. Haplodeficiency of Irgm1 in ApoE(-/-) mice resulted in reduced infiltrating M1 macrophages in atheroma (94%, p=0.0002) and delayed development of atherosclerotic plaques. In vitro experiments also confirmed that Irgm1 haplodeficiency reduced iNOS expression of polarized M1 macrophages (81%, p=0.0034), with negligible impact on the M2 phenotype. Moreover, we found that Irgm1 haplodeficiency in mice significantly reduced expression level of M1 function-related transcription factors, interferon regulatory factor (Irf) 5 and Irf8, but not Irf4, an M2-related transcription factor. CONCLUSIONS: This study shows that Irgm1/IRGM participates in the polarization of M1 macrophage and promotes development of atheroma in murine experimental atherosclerosis.