Guizhong Zhang

Roles of TIPE2 in hepatitis B virus-induced hepatic inflammation in humans and mice.

Hepatitis B virus (HBV)-induced hepatic inflammation afflicts hundreds of millions of people worldwide and is a leading cause of hepatic cancer. While the deleterious effect of the chronic hepatitis is well recognized, the molecular mechanisms underlying the pathogenesis of HBV-induced hepatic inflammation are not well understood. We report here that the tumor necrosis factor-alpha-induced protein-8 like-2 (TIPE2 or TNFAIP8L2), a newly identified regulator of immune receptor signaling, plays an important role in controlling HBV-induced hepatitis. Patients with chronic hepatitis B had significantly reduced levels of TIPE2 expression in their peripheral blood mononuclear cells (PBMCs) as compared to healthy individuals. The TIPE2 expression negatively correlated with the blood levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total bilirubin (Tbil) as well as the HBV load of the patients. Importantly, using a murine model of HBV-induced hepatitis, we found that TIPE2-deficient mice developed significantly more severe hepatic inflammation than wild type mice. These results indicate that TIPE2 plays an important role in taming HBV-induced hepatic inflammation.

Tissue-specific expression of TIPE2 provides insights into its function

Tumor necrosis factor-alpha-induced protein-8 like-2 (TNFAIP8L2, TIPE2) is a newly discovered negative regulator of innate immunity and cellular immunity. TIPE2 deficiency in mice causes fetal inflammatory diseases and TIPE2 downregulation in humans is associated with systemic autoimmunity. However, TIPE2 deficiency leads to a selective defect in humoral immunity. Due to the lack of a suitable antibody, the nature of cells and tissues that express TIPE2 protein has not been determined. In this study, we generated a highly specific antibody to TIPE2 and examined TIPE2 expression in various murine tissues by immunohistochemistry and RT-PCR. We found that TIPE2 was a cytoplasmic protein expressed preferentially in lymphoid tissues and a small group of non-lymphoid tissues. Within the lymphoid compartment, T cells appear to express high level of TIPE2 protein, while B cells and B cell zones of lymphoid organs were devoid of TIPE2. Within most of the non-lymphoid tissues, TIPE2 was not detected. However, several endocrine tissues and skeletal muscle expressed detectable TIPE2 protein and mRNA. Furthermore, high levels of TIPE2 were detected in monocyte/macrophage derived cell lines and ovarian adenocarcinoma cells, but not detectable or weakly expressed in most human carcinoma cell lines. These results indicate that TIPE2 may perform tissue-specific functions in both lymphoid and non-lymphoid compartments. They may also explain why TIPE2 deficiency enhanced cellular but not humoral immunity.

TIPE2 negatively regulates inflammation by switching arginine metabolism from nitric oxide synthase to arginase.

TIPE2, the tumor necrosis factor (TNF)-alpha-induced protein 8-like 2 (TNFAIP8L2), plays an essential role in maintaining immune homeostasis. It is highly expressed in macrophages and negatively regulates inflammation through inhibiting Toll-like receptor signaling. In this paper, we utilized RAW264.7 cells stably transfected with a TIPE2 expression plasmid, as well as TIPE2-deficient macrophages to study the roles of TIPE2 in LPS-induced nitric oxide (NO) and urea production. The results showed that TIPE2-deficiency significantly upregulated the levels of iNOS expression and NO production in LPS-stimulated macrophages, but decreased mRNA levels of arginase I and urea production. However, TIPE2 overexpression in macrophages was capable of downregulating protein levels of LPS-induced iNOS and NO, but generated greater levels of arginase I and urea production. Furthermore, TIPE2−/− mice had higher iNOS protein levels in lung and liver and higher plasma NO concentrations, but lower levels of liver arginase I compared to LPS-treated WT controls. Interestingly, significant increases in IκB degradation and phosphorylation of JNK, p38, and IκB were observed in TIPE2-deficient macrophages following LPS challenge. These results strongly suggest that TIPE2 plays an important role in shifting L-arginase metabolism from production of NO to urea, during host inflammatory response.

Enhanced Atherosclerosis in TIPE2-Deficient Mice Is Associated with Increased Macrophage Responses to Oxidized Low-Density Lipoprotein

Atherosclerosis has been widely recognized as an inflammatory disease of the arterial wall in which macrophages play a major role. Yet, how macrophage-mediated pathology is regulated during atherosclerosis is poorly understood. TNF-α–induced protein 8–like 2 (TIPE2, also known as TNFAIP8L2) is highly expressed in resting macrophages and can negatively regulate inflammation through inhibiting immune receptor signaling. We report in this article that TIPE2 plays a crucial atheroprotective role likely by regulating macrophage responses to oxidized low-density lipoprotein (ox-LDL). TIPE2-deficient macrophages treated with ox-LDL produced more oxidative stress and proinflammatory cytokines, and exhibited heightened activation of the JNK, NF-κB, and p38 signaling pathways. As a consequence, TIPE2 deficiency in bone marrow–derived cells exacerbated atherosclerosis development in Ldlr−/− mice fed a high-fat diet. Importantly, ox-LDL markedly downregulated TIPE2 mRNA and protein levels in macrophages, suggesting that ox-LDL mediates atherosclerosis by TIPE2 inhibition. These results indicate that TIPE2 is a new inhibitor of atherosclerosis and a potential drug target for treating the disease.

TIPE2 deficiency accelerates neointima formation by downregulating smooth muscle cell differentiation.

Phenotypic switching of vascular smooth muscle cells (VSMCs) is known to play a key role in the development of atherosclerosis. However, the mechanisms that mediate VSMC phenotypic switching are unclear. We report here that TIPE2, the tumor necrosis factor (TNF) α-induced protein 8-like 2 (TNFAIP8L2), plays an atheroprotective role by regulating phenotypic switching of VSMCs in response to oxidized low-density lipoprotein (ox-LDL) stimuli. TIPE2-deficient VSMCs treated with ox-LDL expressed lower levels of contractile proteins such as SMαA, SM-MHC and calponin, whereas the proliferation, migration and the synthetic capacity for growth factors and cytokines were increased remarkably. Furthermore, TIPE2 inhibited VSMCs proliferation by preventing G1/S phase transition. Interestingly, these effects of TIPE2 on VSMCs were dependent on P38 and ERK1/2 kinase signals. As a result, neointima formation was accelerated in the carotid arteries of TIPE2-deficient mice. These results indicate that TIPE2 is a potential inhibitor of atherosclerosis.

The expression of TIPE1 in murine tissues and human cell lines

Members of the tumor necrosis factor-alpha-induced protein-8 (TNFAIP8 or TIPE) family play important roles in immune homeostasis and cancer. TIPE1 (TNFAIP8-like 1) is a new member of the TIPE family that may regulate cell death. However, due to the lack of a suitable antibody, the nature of cells and tissues that express TIPE1 protein has not been determined. In this study, we generated a highly specific antibody to TIPE1 and examined TIPE1 expression in various murine tissues and human cell lines by immunohistochemistry, reverse transcription real-time PCR, and Western blot. We found that TIPE1 protein was detected in a wide variety of tissues in C57BL/6 mice, such as neurons in brain, hepatocytes, germ cells of female and male reproductive organs, muscular tissues, and a variety of cells of the epithelial origin, particularly those with secretory functions. TIPE1 protein was not expressed in mature T or B lymphocytes, but detectable in human B lymphoblast cell line HMy2.CIR and murine T cell line EL4. Furthermore, high levels of TIPE1 mRNA were detected in most human carcinoma cell lines, especially in cells transformed with viral genomes. These results indicate that TIPE1 may perform functions in cell secretion and carcinogenesis, but not in immunity.

Identical Expression Profiling of Human and Murine TIPE3 Protein Reveals Links to Its Functions

Tumor necrosis factor-alpha-induced protein-8 like-3 (TNFAIP8L3, TIPE3) is a newly discovered member of TNFAIP8 family and regarded as a lipid second messenger transfer protein that promotes cancer. Yet the nature of the cells and tissues that express TIPE3 protein has not been determined. In this study, we examined TIPE3 expression in various murine and human tissues by immunohistochemistry and quantitative PCR. We found that TIPE3 expression was almost identical in most organs from human and mice. TIPE3 is a cytoplasmic protein expressed preferentially in epithelial-derived cells with secretory functions. Furthermore, TIPE3 protein is highly expressed in most human carcinoma cell lines. These results suggest that TIPE3 may play important roles in carcinogenesis and cell secretion.

TIPE2 protein prevents injury-induced restenosis in mice.

Proliferation of vascular smooth muscle cells (VSMCs) plays an important role in restenosis, a disease characterized by smooth muscle cell hyperplasia and neointimal formation. How proliferation signals are controlled to avoid restenosis is not fully understood. Here we report that TIPE2, the tumor necrosis factor (TNF) α-induced protein 8-like 2 (TNFAIP8L2), suppresses injury-induced restenosis by inhibiting VSMCs proliferation. TIPE2 was significantly upregulated in VSMCs in response to PDGF-BB stimuli and injury. Enforced TIPE2 expression significantly suppressed VSMCs proliferation and cell cycle progression, whereas TIPE2 deficiency in VSMCs promoted cell proliferation and upregulated the expression of Cyclins D1 and D3. TIPE2 likely regulated VSMC proliferation via Rac1-STAT3 and ERK1/2 signaling pathways. It blocked STAT3 activation and nuclear translocation in a Rac1-dependent manner. As a result, TIPE2-deficient VSMCs exhibited enhanced proliferation whereas TIPE2-deficient mice developed more severe restenosis in response to vascular injury. Conversely, adenovirus-mediated gene transfer of TIPE2 significantly reduced injury-induced restenosis in mice. These results indicate that TIPE2 plays a suppressive role in injury-induced restenosis and may serve as a new therapeutic target for treating the disease.

DcR3 combined with hematological traits serves as a valuable biomarker for the diagnosis of cancer metastasis

Decoy receptor 3 (DcR3) is abnormally up-regulated in many cancer cells. It may help cancer cells to escape from immune surveillance and establish metastatic lesions. However, whether DcR3 can be used as a biomarker for the diagnosis of cancer metastasis is unclear. In this study, sera from healthy controls and patients with different cancers were collected, and tested for their DcR3 levels by ELISA. Significantly elevated DcR3 levels were observed in the sera of patients with gastric cancer (2.04 ± 1.01, P = 0.0061), lymphoma (1.62 ± 0.75, P = 0.041), and breast cancer (1.53 ± 0.51, P = 0.023). DcR3 was found to be a suitable biomarker for identifying gastric cancer patients. Importantly, DcR3 was positively associated with platelet distribution width (PDW) (P = 2.45 × 10−6, R = 0.63) in metastatic cancers but negatively associated with hemoglobin (HGB) (P = 0.002, R = −0.59) and hematocrit (HCT) (P = 0.001, R = −0.62) in non-metastatic cancers. Combined with PDW, HGB and HCT, serum DcR3 could be used to predict the occurrence of cancer metastasis. These findings indicate that DcR3 could be used as a biomarker for the diagnosis of gastric cancer, and for cancer metastasis in combination with hematological traits.

CD317 Promotes the survival of cancer cells through apoptosis-inducing factor.

BackgroundLow nutrient environment is a major obstacle to solid tumor growth. However, many tumors have developed adaptive mechanisms to circumvent the requirement for exogenous growth factors.MethodsHere we used siRNA interference or plasmid transfection techniques to knockdown or enhance CD317 expression respectively, in mammalian cancer cells, and subjected these CD317-manipulated cells to serum deprivation to study the role of CD317 on stress-induced apoptosis and the underlying mechanism.ResultsWe report that CD317, an innate immune gene overexpressed in human cancers, protected cancer cells against serum deprivation-induced apoptosis. In tumor cells, loss of CD317 markedly enhanced their susceptibility to serum deprivation-induced apoptosis with no effect on autophagy or caspase activation, indicating an autophagy- and caspase-independent mechanism of CD317 function. Importantly, CD317 knockdown in serum-deprived tumor cells impaired mitochondria function and subsequently promoted apoptosis-inducing factor (AIF) release and nuclear translocation but had little effect on mitochondrial and cytoplasmic distributions of cytochrome C, a pro-apoptotic factor released from mitochondria that initiates caspase processing in response to death stimuli. Furthermore, overexpression of CD317 in HEK293T cells inhibits serum deprivation-induced apoptosis as well as the release and nuclear accumulation of AIF.ConclusionOur data suggest that CD317 functions as an anti-apoptotic factor through the mitochondria-AIF axis in malnourished condition and may serve as a potential drug target for cancer therapy.