OBJECTIVETo investigate the expression of the granulocyte-macrophage colony-stimulating factor (GM-CSF) and GM-CSF/IL-3/IL-5 receptor common beta chain (beta c receptor) in an adult patient with idiopathic pulmonary alveolar proteinosis (PAP), so as to demonstrate the possible association of the GM-CSF and beta c receptor with the pathogenesis of human PAP.

METHODSThe GM-CSF levels were measured with a commercial ELISA kit (sensitivity 5 pg/ml) and the beta c receptor expression on the cell surface was detected by flow cytometry analysis. Reverse transcription polymerase chain reaction (RT-PCR) analysis was employed to detect the expression of the GM-CSF mRNA and the beta c receptor mRNA in peripheral blood mononuclear cells and alveolar macrophages. The entire coding regions of the GM-CSF cDNA and the beta c receptor cDNA were sequenced by the Sanger dideoxy-mediated chain termination method to detect possible mutations.

RESULTSThe patient with PAP failed to release the GM-CSF protein either from circulating mononuclear cells or from alveolar macrophages. The expression of the GM-CSF mRNA was normal after the stimulation of lipopolysaccharide, whereas a point mutation at position 382 of the GM-CSF cDNA from "T" to "C" was revealed by cDNA sequencing, which caused a change in amino acid 117 of the protein from isoleucine to threonine. The beta c receptor expression on the cell surface was normal, and the beta c receptor mRNA expression and the sequence of the entire coding region of the beta c receptor were also normal.

CONCLUSIONSThe decreased GM-CSF production is associated with the pathogenesis of human PAP. A point mutation of the GM-CSF cDNA may contribute to the decreased GM-CSF production in our adult PAP patient. The mutation of the beta c receptor in some of paediatric patients with PAP may not be a common problem in adult patients.

DNA, Complementary ; chemistry ; Granulocyte-Macrophage Colony-Stimulating Factor ; analysis ; biosynthesis ; Humans ; Male ; Middle Aged ; Pulmonary Alveolar Proteinosis ; etiology ; metabolism ; RNA, Messenger ; analysis ; Receptors, Cytokine ; biosynthesis ; genetics

To systematically clarify the effects of apolipoprotein E (aopE) and low-density lipoprotein receptor (LDLR) gene mutant on hyperlipidemia, vascular inflammation impairment and pathogenesis of atherosclerosis (AS), total RNA was isolated from fresh aortas of young apoE/LDLR double knockout (apoE(-/-)/LDLR(-/-)) and wild type (WT) mice using TRIzol reagent. Then RNA was reversely transcribed to first-strand cDNA by reverse transcriptase for reverse transcription polymerase chain reaction (RT-PCR) and real-time RT-PCR. Primer pairs were designed using primer design software according to the gene sequences available in GenBank. &beta;-actin was used as an internal control. Then RT-PCR assay was used to analyze the expression patterns of interleukin-1&beta; (IL-1&beta;), tumor necrosis factor-&alpha; (TNF-&alpha;), nuclear factor-κB (NF-κB), granulocyte-macrophage colony-stimulating factor (GM-CSF), CD36, endothelin-1 (ET-1), toll-like receptor 2 (TLR2), monocyte chemoattractant protein-1 (MCP-1), vascular adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1) and platelet-derived growth factor-&alpha; (PDGF-&alpha;). SYBR Green quantitative real-time RT-PCR was used to validate gene expressions identified by RT-PCR. Blood samples were taken from the retro-orbital venous plexus, and serum levels of total cholesterol (TC), triglyceride (TG), low-density lipoprotein (LDL) and high-density lipoprotein (HDL) were measured by using biochemical techniques. Serum concentrations of circulating TNF-&alpha;, IL-1&beta; and oxidized LDL (ox-LDL) were determined by ELISA. Frozen sections of aortic sinus were stained with Sudan IV to visualize intimal fatty lesions. The results showed that the relative expressions of IL-1&beta;, GM-CSF, ET-1, TLR2, CD36, MCP-1, ICAM-1 and VCAM-1 in apoE(-/-)/LDLR(-/-) mice at the age of 1 month were higher than those in age-matched WT mice (P<0.05, P<0.01), respectively. The expressions of PDGF-&alpha; and TNF-&alpha; in apoE(-/-)/LDLR(-/-) mice at the age of 2 months were up-regulated compared to those in age-matched WT mice (P<0.05). All the expressions of target genes continued to be up-regulated (P<0.05, P<0.01) except that ET-1 expression at the age of 2 months, TLR2, VCAM-1 and ICAM-1 expressions at the age of 3 months were down-regulated to that in WT mice. NF-κB expression had no significant changes between two genotype mice at different ages. All the gene expressions kept unchanged in WT mice at different ages, except that IL-1b expressions were slightly up-regulated at the ages of 2 and 3 months. Serum levels of TC, TG, LDL, HDL, TNF-&alpha;, IL-1&beta; and ox-LDL in apoE(-/-)/LDLR(-/-) mice at different ages were higher than those in age-matched WT mice (P<0.05, P<0.01), and were increasing with age. Primary atherosclerotic lesions were observed in 1-month old apoE(-/-)/LDLR(-/-) mice and were progressing with age. There were no lesions observed in all WT mice at different ages. The data suggest that hyperlipidemia due to apoE and LDLR gene mutant may stimulate the temporal expressions of AS-related genes and contribute to primary atherogenetic lesions and vascular inflammation impairment.
Animals ; Aorta ; metabolism ; Apolipoproteins E ; genetics ; Atherosclerosis ; genetics ; CD36 Antigens ; metabolism ; Chemokine CCL2 ; metabolism ; Endothelin-1 ; metabolism ; Gene Expression ; Granulocyte-Macrophage Colony-Stimulating Factor ; metabolism ; Hyperlipidemias ; metabolism ; Intercellular Adhesion Molecule-1 ; metabolism ; Interleukin-1beta ; blood ; metabolism ; Lipoproteins, LDL ; blood ; Mice ; Mice, Knockout ; NF-kappa B ; metabolism ; Platelet-Derived Growth Factor ; Receptors, LDL ; genetics ; Toll-Like Receptor 2 ; metabolism ; Tumor Necrosis Factor-alpha ; blood ; metabolism ; Vascular Cell Adhesion Molecule-1 ; metabolism

The hematopoietic system of the mouse arises from extraembryonic mesoderm that migrate through primitive streak to the presumptive yolk sac at day 7.0 of gestation. However, the mechanisms regulating mesoderm commitment to hematopoietic lineages remain poorly understood. Previous studies demonstrated that the development kinetics and growth factor responsiveness of hematopoietic precursors derived from embryonic stem cells (ES cells) is similar to that found in the yolk sac, indicating that the onset of hematopoiesis within the embryoid bodies (EBs) parallels that found in the embryo. Furthermore, in vitro differentiation of ES cells to hematopoietic cells is valuable for establishment of therapeutic clone against a variety of hematological disorders. Despite the identification of multipotential hematopoietic progenitors in EBs, a subset of more primitive progenitors, identical to the high proliferative potential colony-forming cells (HPP-CFC) derived from human and murine hematopoietic tissues, have not been clearly identified regarding particular their replating potential in vitro. HPP-CFC is among the most primitive hematopoietic multipotent precursors cultured in vitro. In this study, our aim was to investigate the in vitro and in vivo hematopoietic capacity of HPP-CFC within the day 12 EBs, rather than the expansion of more committed progenitors. In this study the HPP-CFC could be detected within EBs differentiated for 5 to 14 days of murine ES cells, but the development dynamics of the HPP-CFC differed greatly among distinct serum lots. Qualitatively HPP-CFC is capable of forming secondary colonies. As to our expectation the ES cells-derived HPP-CFC demonstrated similar regeneration capacity to those from yolk sac, giving rise to secondary granulocyte, erythrocyte, macrophage and mast cells, however largely differed from the counterparts of adult bone marrow. In addition, by RT-PCR ES cells-derived HPP-CFC were found to express transcription factors associated closely with stem cell proliferation including SCL, GATA-2 and AML1 as well as various receptors of hematopoietic growth factors such as c-kit, GM-CSF receptor and interleukin 3 receptor et al. Finally, in order to understand the in vivo hematopoietic capacity of the ES cells-derived HPP-CFC, spleen colony-forming unit (CFU-S) assay was performed. Nevertheless, typical CFU-S was not observed after transplantation of the day 12 EB cells or HPP-CFC colonies into lethally irradiated adult murine. In conclusion the HPP-CFC differentiated from murine ES cells displayed robust hematopoietic activity in vitro, however their in vivo reconstitution ability was not detected. The difference between in vitro and in vivo hematopoietic activities of ES cells-derived primitive hematopoietic precursors deserves further investigation.
Animals ; Basic Helix-Loop-Helix Transcription Factors ; genetics ; Cell Differentiation ; genetics ; physiology ; Colony-Forming Units Assay ; Core Binding Factor Alpha 2 Subunit ; genetics ; Embryonic Stem Cells ; cytology ; GATA2 Transcription Factor ; genetics ; Hematopoietic Stem Cells ; cytology ; metabolism ; Humans ; Mice ; Proto-Oncogene Proteins ; genetics ; Proto-Oncogene Proteins c-kit ; genetics ; Receptors, Granulocyte-Macrophage Colony-Stimulating Factor ; genetics ; Receptors, Interleukin-3 ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; T-Cell Acute Lymphocytic Leukemia Protein 1

The management of asthma focuses on the reduction of airway inflammation accompany with symptomatic care after recognition. Glucocorticosteroid is the most important drug to reduce airway inflammation, and it has been used inhaled, orally and systemically. New knowledge about the pathogenesis of allergy and asthma has made the development and clinical trial of target or immunomodulator therapy. It includes cytokine, cytokine blockers, specific cytokine receptor blocker, and immunostimulatory oligodeoxynucleotides. These agents are thought to hold the promise for more beneficial outcomes in the future, although it showed limited therapeutic benefits only for patients, especially intractable or severe asthma, until now.
Asthma ; Humans ; Hypersensitivity ; Immunomodulation ; Inflammation ; Oligodeoxyribonucleotides ; Receptors, Cytokine

Chimeric antigen receptor T cell (CAR-T) therapy was awarded as the largest research breakthrough in 2017 by the American Society of Clinical Oncology, at present, it is rapidly becoming the most promising new treatment for hematological malignancies. However, this therapy also produces a new challenge: toxic adverse events such as cytokine release syndrome (CRS) and neurotoxicity, partial of them can bring death to the patients. The incidence and severity of the above toxic events in different multi-center trial reports are also different, which may be attributed to the different in the considerably variable assessment and grading of toxicities between clinical trials and across institutions. The ASTCT published at 2018 advanced the consensus grading for cytokine release syndrome and neurologic toxicity associated with immune effector cells, it was focusing on CRS and neurotoxicity associated with immune effector cells. In order to provide reference for the development of relevant work in this field and the formulation of security strategies in our country, the main content of the consensus was summarized briefly.
Cell- and Tissue-Based Therapy ; Consensus ; Cytokine Release Syndrome ; Humans ; Receptors, Antigen, T-Cell ; Receptors, Chimeric Antigen

OBJECTIVETo investigate the biological activity and molecular mechanism of interferon alpha (IFNalpha) on human myeloma cell line Sko-007.

METHODSThe effect of IFNalpha on the growth of Sko-007 cells was measured by MTT assay. Cells cycle distribution and the expression of two IL-6 receptor chains (IL-6R and gp130) on Sko-007 cell surface in the absence or presence of IFNalpha were monitored by FACS analysis. The activation state of protein kinase ERK, which is involved in Ras/MAPK signal transduction pathway mediating cell survival and proliferation, and the expression of anti-apoptotic Bcl-2 family proteins-Bcl-2, Bcl-x(L) and Mcl-1 in Sko-007 cells with or without IFNalpha were determined by immunoblot assay.

RESULTIFNalpha arrested Sko-007 cell cycle progression. After stimulation with IFNalpha, an obvious increase in G(0)/G(1) phase (41.1%-->84.1%) and decrease in S phase (57.1%-->13.3%) of Sko-007 cell cycle distribution can be observed. Moreover, the proliferation of Sko-007 cells was dramatically inhibited in the presence of IFNalpha, with a maximal inhibitory rate up to 88%. In addition, the expression of gp130 on cell surface, the activation of protein kinase ERK and the expression of Bcl-2 and Bcl-x(L) were all down-regualted in IFNalpha-stimulated Sko-007 cells.

CONCLUSIONThe inhibitory effect of IFNalpha on the proliferation of Sko-007 cells was mediated by gp130 down-regulation, degradation of Bcl-2 family anti-apoptotic proteins and inhibition of ERK activation.

Antigens, CD ; drug effects ; metabolism ; Cell Cycle ; drug effects ; Cell Division ; drug effects ; Cytokine Receptor gp130 ; Dose-Response Relationship, Drug ; Down-Regulation ; Enzyme Activation ; drug effects ; G1 Phase ; drug effects ; Humans ; Immunoblotting ; Interferon-alpha ; pharmacology ; Membrane Glycoproteins ; drug effects ; metabolism ; Mitogen-Activated Protein Kinases ; metabolism ; Multiple Myeloma ; metabolism ; pathology ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Receptors, Interleukin-6 ; drug effects ; metabolism ; Resting Phase, Cell Cycle ; drug effects ; S Phase ; drug effects ; Tumor Cells, Cultured ; drug effects ; metabolism ; bcl-X Protein

To evaluate soluble GM-CSF-Ralpha expression in patients with acute myeloid leukemia (AML) and its clinic significance, plasma concentration of solGM-Ralpha in de novo 66 patients with AML was detected by enzyme-linked immuno-sorbent assay, and the relationship between solGM-Ralpha levels and various clinical parameters was analyzed. The result showed that the levels of solGM-Ralpha in plasma of patients with AML were significantly higher than that in plasma of normal controls; the lowest level of solGM-Ralpha was found in plasma of patients with AML-M3 (3897.75 +/- 2651.43 pg/ml), the highest level of solGM-Ralpha was observed in plasma of patients with AML-M5 (9990.92 +/- 6325.43 pg/ml). Patients with high level of solGM-Ralpha were generally accompanied with a distinct clinical picture, including higher counts of white blood cell and myeloid precursors, as well as higher expression of CD34, CD95 and CD116 antigen. It is concluded that the high level of solGM-Ralpha in plasma of patients may suggest AML poor prognosis and play a role in pathogenesis of leukemia, the GM-CSF and its receptor solGM-Ralpha needs further study.
Adolescent ; Adult ; Aged ; Antigens, CD34 ; blood ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Leukemia, Myeloid, Acute ; blood ; Male ; Middle Aged ; Prognosis ; Receptors, Granulocyte-Macrophage Colony-Stimulating Factor ; biosynthesis ; blood ; fas Receptor ; blood

In order to analyze the clinical characteristics and biological features of acute leukemia in elderly, 104 acute leukemia patients in elderly were retrospectively analyzed and compared with 71 acute leukemia patients below 60 years old. The results showed that: (1) the proportion of AML in the aged group (73%) was higher than that in the young group (54.9%), and the difference was statistically significant (P < 0.05), but AML (M3) was absent in the aged group; (2) the median of bone marrow blast cell in the aged group was significantly lower than that in the young group (P < 0.05); (3) in AML, the frequently of CD14 expression was higher in the aged group (18.8%) than that in the young group (2.6%), while the expression frequencies of CD15 (37.5%), CD117 (62.5%), and CD38 (59.4%) were respectively lower in the aged group than that in the young group which were (69.2%) for CD15, (89.7%) for CD17, and (84.6%) for CD38 respectively, and the difference was also statistically significant (P < 0.05). (4) CD19 was most frequently expressed in ALL of the aged group and the positive rate was 100%; (5) there was no significant difference in expression of special lineage antigens and overlapping lineage antigens between the aged group and the young group (P > 0.05); (6) the expression frequency of unfavorable karyotypes in the aged group was higher than that in the young group, and the difference was statistically very significant (P < 0.01); (7) the complete remission rate (CR rate) in the aged group was 42.9%, 2-year survival rate in the aged group was 5.4%, and treatment-related mortality rate in the aged group was 26.8%, while the CR rate in the young group was 76.6%, the difference was statistically significant (P < 0.05). It is concluded that the expression frequency of CD14 associated with unfavorable prognosis is higher in the aged group than that in the young group, while the expression frequency of CD15 associated with favorable prognosis is lower in the aged group than that in the young group. The expression frequency of unfavorable karyotypes in the aged group is higher than that in the young group. The CR rate of acute leukemia in elderly is low, thus the patients in elderly often have unfavorable prognosis.
Adult ; Age Factors ; Aged ; Aged, 80 and over ; Female ; Humans ; Immunophenotyping ; Karyotyping ; Leukemia, Myeloid, Acute ; diagnosis ; immunology ; Lipopolysaccharide Receptors ; analysis ; Male ; Middle Aged ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; diagnosis ; immunology ; Prognosis ; Receptor, Macrophage Colony-Stimulating Factor ; analysis ; Remission Induction ; Retrospective Studies

The common gamma chain (gammac) is the central signaling unit for a number of cytokine receptors collectively known as the gammac cytokine receptor family. gammac is critical for ligand binding and signaling by gammac cytokines. gammac cytokine signaling had been thought to be mainly regulated by cytokine-specific receptor alpha chain expression levels with little or no effect by gammac surface levels because gammac expression was presumed to remain unchanged during T-cell activation and development. The extent of gammac cytokine responses is thought to be regulated by cytokine specific receptor subunits and not by the gammac receptor. In contrast to this prevailing view, we have recently reported that gammac itself actively regulates gammac cytokine responses. Interestingly, gammac exerted its regulatory effects not only as a conventional membrane receptor protein but also as a secreted protein whose expression was upregulated upon T-cell stimulation. Here we will review how a soluble form of gammac, which is generated by alternative splicing, regulates gammac cytokine signaling and plays a role in controlling immune activation related to autoimmune disease.
Alternative Splicing ; Autoimmune Diseases* ; Cytokines ; Humans ; Membranes ; Receptors, Cytokine ; T-Lymphocytes

BACKGROUND: Bulla is an air-filled space within the lung parenchyma resulting from deterioration of the alveolar tissue. Molecular mechanism of the formation of the bulla is not well described. Fibroblast growth factor(FGF)-7, bone morphogenetic protein(BMP) receptor, and transforming growth factor(TGF)-beta receptor are known to have a stimulatory or inhibitory role in the lung formation. We investigated to see if these growth factor or cytokine receptors are involved in the bulla formation by immunohistochemical staining of bullous lung tissues from patients with primary spontaneous pneumothorax. MATERIAL AND METHOD: Bullous lung tissues were obtained from 31 patients with primary spontaneous pneumothorax, including 30 males and 1 female from 15 to 39 years old. The bullous tissues were obtained by video-thoracoscopic surgery and/or mini-thoracotomy and fixed in formalin. Blocks of the specimens were embedded with paraffin and cut into 5~6 micrometer thick slices. The sections were deparaffinized and hydrated and then incubated with primary antibodies against FGF-7, BMP-RII, or TGF-RII. RESULT: Of the 31 patients, 24 were TGF-RII positive including 18 strong and 6 weak positives. Observation with high magnification showed that strong immunostaining was detected in the boundary region between bullous and normal lung tissues. In contrast, all of the sections were negative with FGF-7 or BMP-RII antibodies. CONCLUSION: These results suggest that overexpression of TGF-beta RII may be involved in the formation of bulla, although further molecular studies are needed to find out more detailed molecular mechanisms.
Adult ; Antibodies ; Female ; Fibroblasts ; Formaldehyde ; Humans ; Lung ; Male ; Paraffin ; Pneumothorax* ; Receptors, Cytokine ; Transforming Growth Factor beta