ABSTRACT
Inhibition of immunocyte infiltration and activation has been suggested to effectively ameliorate nonalcoholic steatohepatitis (NASH). Paired immunoglobulin-like receptor B (PirB) and its human ortholog receptor, leukocyte immunoglobulin-like receptor B (LILRB2), are immune-inhibitory receptors. However, their role in NASH pathogenesis is still unclear. Here, we demonstrate that PirB/LILRB2 regulates the migration of macrophages during NASH by binding with its ligand angiopoietin-like protein 8 (ANGPTL8). Hepatocyte-specific ANGPTL8 knockout reduces MDM infiltration and resolves lipid accumulation and fibrosis progression in the livers of NASH mice. In addition, PirB-/- bone marrow (BM) chimeras abrogate ANGPTL8-induced MDM migration to the liver. And yet, PirB ectodomain protein could ameliorate NASH by sequestering ANGPTL8. Furthermore, LILRB2-ANGPTL8 binding-promoted MDM migration and inflammatory activation are also observed in human peripheral blood monocytes. Taken together, our findings reveal the role of PirB/LILRB2 in NASH pathogenesis and identify PirB/LILRB2-ANGPTL8 signaling as a potential target for the management or treatment of NASH.
Subject(s)
Non-alcoholic Fatty Liver Disease , Animals , Humans , Mice , Angiopoietin-Like Protein 8 , Macrophages , Membrane Glycoproteins , Monocytes , Receptors, Immunologic/geneticsABSTRACT
The activity of the mTOR pathway is frequently increased in acute myeloid leukemia, and is tightly related with cellular proliferation. Leucine is tightly linked to the mTOR pathway and can activate it, thereby stimulating cellular proliferation. LAT3 is a major transporter for leucine, and suppression of its expression can reduce cell proliferation. Here, we show that suppression of LAT3 expression can reduce proliferation of the acute leukemia cell line, K562. We investigated the mRNA and protein expression of LAT3 in several leukemia cell lines and normal peripheral blood mononuclear cells (PBMNCs) using RT-PCR and Western blotting. We also evaluated cell viability using a methyl thiazolyl tetrazolium (MTT) assay after blocking LAT3 expression with either shRNA targeted to LAT3 or a small molecular inhibitor BCH (2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid). LAT3 mRNA and protein expression was detected in leukemia cell lines, but not in normal PBMNCs. Using K562 cells, it was found that cellular proliferation and mTOR pathway activity were significantly reduced when LAT3 was blocked with either shRNA or BCH. Our results suggest that leukemia cell proliferation can be significantly suppressed by blocking LAT3. This finding may lead to a new strategy to develop clinical therapy for the treatment of acute myeloid leukemia.
Subject(s)
Amino Acid Transport Systems, Basic/genetics , Cell Proliferation , Gene Expression Regulation, Leukemic/genetics , RNA Interference , Amino Acid Transport Systems, Basic/antagonists & inhibitors , Amino Acid Transport Systems, Basic/metabolism , Amino Acids, Cyclic/pharmacology , Blotting, Western , Cell Line, Tumor , Cell Survival/drug effects , Cell Survival/genetics , Cells, Cultured , Dose-Response Relationship, Drug , HL-60 Cells , Humans , Jurkat Cells , K562 Cells , Leukemia, Erythroblastic, Acute/genetics , Leukemia, Erythroblastic, Acute/metabolism , Leukemia, Erythroblastic, Acute/pathology , Phosphorylation/drug effects , Reverse Transcriptase Polymerase Chain Reaction , TOR Serine-Threonine Kinases/metabolismABSTRACT
This study was purposed to establish a retrovirus-mediated murine model with MLL-AF9 leukemia, so as to provide a basis for further investigation of the pathogenesis and therapeutic strategy of MLL associated leukemia. Murine (CD45.2) primary hematopoietic precursor positively selected for expression of the progenitor marker c-Kit by means of MACS were transduced with a retrovirus carrying MLL-AF9 fusion gene. After cultured in vitro, the transduced cells were injected intravenously through the tail vein into the lethally irradiated mice (CD45.1). PCR, flow cytometry and morphological observation were employed to evaluate the murine leukemia model system. The results showed that MLL-AF9 fusion gene was expressed in the infected cells, and the cells had a dramatically enhanced potential to generate myeloid colonies with primitive and immature morphology. Flow cytometric analysis revealed that the immortalized cells highly expressed myeloid lineage surface markers Gr-1 and Mac-1. Moreover, the expression levels of Hoxa9 and Meis1 mRNA were significantly higher in the MLL-AF9 cells than that in control. The mice transplanted with MLL-AF9 cells displayed typical signs of leukemia within 6-12 weeks. Extensive infiltration leukemic cells was observed in the Wright-Giemsa stained peripheral blood smear and bone marrow, and also in the histology of liver and spleen. Flow cytometric analysis of the bone marrow and spleen cells demonstrated that the CD45.2 populations expressed highly myeloid markers Gr-1 and Mac-1. The leukemic mice died within 12 weeks. It is concluded that the retrovirus-mediated murine model with MLL-AF9 leukemia is successfully established, which can be applied in the subsequent researches.
