ABSTRACT
Context: The Da-yuan-yin (DYY) decoction is a classical prescription of traditional Chinese medicine that has antipyretic and anti-inflammatory effects. Network Pharmacology (NP) is an emerging discipline based on system-biology theory and biosystem network analysis that researchers can use to predict drug-action targets and mechanisms. Objective: The study intended to use NP evaluate the protective effects of the fifth eluting fraction of the supernatant of the DYY decoction (DYY-5) for mice induced with acute lung injury (ALI) using lipopolysaccharide (LPS) and to explore DYY-5's mechanisms. Design: The research team performed an animal study. Setting: The study took place at the College of Pharmaceutical Science at Soochow University in Suzhou, China. Animals: The animals were 42 male Balb/c mice, about 20 to 25 g in weight. Intervention: The research team: instilled 2 mg/kg of LPS intratracheally (i.t.) to induce ALI. The team divided the mice into seven groups of six mice: (1) a control group; (2) a negative control group-the DYY-5 group with mice treated only with a high dosage, 60 mg/kg, of DYY-5 to investigate the effects of DYY-5 on normal mice; (3) the positive control group, the LPS group, with induced ALI but no treatments; (4) the LPS+60 mg/kg-DYY-5 group with induced ALI treated with a high dosage of DYY-5; (5) the LPS+30 mg/kg-DYY-5 group with induced ALI treated with a medium dosage of DYY-5; (6) the LPS+15 mg/kg-DYY-5 group with induced ALI treated with a low dosage of DYY-5; and (7) a reference drug control group, the LPS+DXM group, with induced ALI treated with 5 mg/kg of dexamethasone (DXM). Outcome Measures: The research team: (1) determined the chemical components of DYY; (2) identified the anticomplementary activities of DYY-5; (3) took lung specimens, serum, and bronchoalveolar lavage fluid (BALF) from the mice for histopathological examination, Western blot, and biochemical analysis; (4) measured total protein concentrations and lung W/D ratios; (5) measured the expressions of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) messenger RNA (mRNA) using quantitative real-time polymerase chain reaction (PCR); (6) measured the levels of pro-inflammatory and anti-inflammatory factors, the activity of myeloperoxidase (MPO) and superoxide dismutase (SOD), and the levels of complements, including complements 3 (C3), C3c, C5a, C5aR1, and C5b-9, using kits; (7) analyzed the levels of nuclear factor-kappa B (NF-κB) and IkB kinase (IKK) using Western blot; and (8) used network pharmacology (NP) to predict DYY-5's mechanisms and potential targets. Results: The study's results were consistent with the NP analysis, which reflected the multitarget and multipathway characteristics of DYY-5 in alleviating ALI. The LPS+30 mg/kg-DYY-5 group had significantly lower lung wet-to-dry (W/D) ratios and total protein concentrations in BALF than the LPS group did, with P < .01 and P < .0001, respectively as did the LPS+60 mg/kg-DYY-5 group (both P < .0001). The 60 mg/kg of DYY-5 compared to the LPS group: (1) regulated the levels tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), and interleukin-1 beta (IL-1ß), with all P < .0001, anti-inflammatory factors-IL-4 (P < .05), IL-10 (P < .001), and IL-13 (P < .001); (2) increased the activity of SOD (P < .0001) and decreased the activity of MPO (P < .0001) and the expressions of iNOS and COX-2 mRNA (both P < .01); (3) blocked the activation of NF-κB and IKK; and (4) alleviated the pathological changes in the lung tissue, by reducing the depositions of C3c and decreasing the levels of C3, C5a and C5aR1 (all P < .0001), C5b-9 (P < .001) and C3c (P < .01) in serum. Conclusions: The protective effects of DYY-5 on ALI were related to antioxidation, anti-complementary activities, and regulation of inflammatory factors through the IKK/NF-κB signal pathway. DYY-5 may be useful as a potential therapeutic agent for treating ALI in clinics.
Subject(s)
Acute Lung Injury , NF-kappa B , Male , Mice , Animals , NF-kappa B/genetics , NF-kappa B/metabolism , Lipopolysaccharides , Cyclooxygenase 2/adverse effects , Complement Membrane Attack Complex/adverse effects , Acute Lung Injury/chemically induced , Acute Lung Injury/drug therapy , Acute Lung Injury/metabolism , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/metabolism , Tumor Necrosis Factor-alpha/therapeutic use , Anti-Inflammatory Agents/therapeutic use , Mice, Inbred BALB C , RNA, Messenger , Superoxide DismutaseABSTRACT
Carfilzomib has been associated with the development of thrombotic microangiopathy (TMA) in relapsed/refractory multiple myeloma patients, a severe disease with no currently available aetiological treatment. We evaluated the potential role of terminal complement pathway in four patients with carfilzomib-induced TMA. Membrane attack complex (C5b-9) deposition on endothelial cells in culture exposed to plasma from patients during the acute phase of the disease suggests complement overactivation as a mechanism of potential endothelial damage in three out of four patients. If confirmed in larger cohorts, C5b-9 evaluation will allow early identification of patients who could benefit from complement blockade and treatment monitoring.
Subject(s)
Complement System Proteins/drug effects , Multiple Myeloma/drug therapy , Oligopeptides/adverse effects , Thrombotic Microangiopathies/chemically induced , Ubiquitin/antagonists & inhibitors , Adult , Aged , Antibodies, Monoclonal, Humanized/administration & dosage , Antibodies, Monoclonal, Humanized/adverse effects , Antibodies, Monoclonal, Humanized/pharmacology , Antibodies, Monoclonal, Humanized/therapeutic use , Complement Membrane Attack Complex/adverse effects , Complement Membrane Attack Complex/metabolism , Complement System Proteins/metabolism , Endothelial Cells/immunology , Endothelial Cells/metabolism , Female , Humans , Male , Middle Aged , Multiple Myeloma/complications , Oligopeptides/therapeutic use , Prospective Studies , Proteasome Inhibitors/adverse effects , Proteasome Inhibitors/therapeutic use , Thrombotic Microangiopathies/drug therapy , Thrombotic Microangiopathies/etiology , Thrombotic Microangiopathies/metabolism , Ubiquitin/metabolismABSTRACT
PURPOSE: Podocytes, terminal differentiation cell in glomerulu, are crucial to kidney-related diseases such as membranous nephropathy (MN). MN is characterized by podocyte injury and glomerular basement membrane thickening. This paper focused to investigate the expression of chemokine (C-X-C motif) ligand 12 (CXCL12) in MN patients and its possible role in podocyte injury. METHODS: Through the enzyme-linked immunosorbent assay, CXCL12 level in the serum and urine of MN patients was examined. Further, several assays of cell viability, apoptosis, quantitative real-time PCR and western blot were applied to explore the effects of CXCL12 in the model of podocyte injury. RESULTS: We found a significant increase of CXCL12 in serum and urine of MN patients, which indicated that CXCL12 may be involved in the progression of MN. And in vitro C5b-9-induced podocyte injury model, the proliferation of podocytes was inhibited whereas CXCL12/CXCR4 and phosphorylated STAT3 (p-STAT3) were increased. Silencing of CXCL12 remarkably promoted cell proliferation, inhibited cell apoptosis and suppressed CXCL12/CXCR4, p-STAT3 and caspase 3. Consistently, STAT3 inhibitor and berberine (a CXCL12 antagonist) also reduced CXCL12 treatment-induced apoptosis. CONCLUSIONS: All data suggested that silencing of CXCL12 had a protective effect on podocyte injury, which may be through inhibiting CXCL12/STAT3 signaling pathway.
Subject(s)
Chemokine CXCL12/genetics , Complement Membrane Attack Complex/adverse effects , Gene Silencing , Glomerulonephritis, Membranous/genetics , Podocytes/pathology , RNA/genetics , Apoptosis , Blotting, Western , Cell Survival , Cells, Cultured , Chemokine CXCL12/biosynthesis , Enzyme-Linked Immunosorbent Assay , Flow Cytometry , Glomerulonephritis, Membranous/metabolism , Glomerulonephritis, Membranous/pathology , Humans , Immunologic Factors/adverse effects , Kidney Glomerulus/drug effects , Kidney Glomerulus/metabolism , Kidney Glomerulus/pathology , Podocytes/drug effects , Podocytes/metabolism , Real-Time Polymerase Chain Reaction , Signal TransductionABSTRACT
The sublytic C5b-9 complexes can result in glomerular mesangial cells (GMCs) apoptosis, which involved in the initiation and development of rat Thy-1 nephritis. Activating transcription factor 3 (ATF3) is an immediate early gene for cells to cope with a variety of stress signals, and our previous study revealed that ATF3 could promote GMCs apoptosis attacked by sublytic C5b-9. But the mechanism of ATF3 promoting GMCs apoptosis triggered by sublytic C5b-9 attack has not been elucidated. In this study, the data showed that the expression of ATF3, growth arrest and DNA damage-45 alpha (Gadd45α), Krüppel-like factor 6 (KLF6) and proliferating cell nuclear antigen (PCNA) in the GMCs in response to sublytic C5b-9 stimulation for the indicated time was significantly increased, and ATF3 expression could lead to GMCs apoptosis through up-regulation of Gadd45α and KLF6, but not up-regulation of PCNA. Furthermore, Gadd45α was identified as a downstream target gene regulated by ATF3 directly, and KLF6 might be regulated by ATF3 in an indirect manner.
Subject(s)
Activating Transcription Factor 3/genetics , Cell Cycle Proteins/genetics , Complement Membrane Attack Complex/immunology , Kruppel-Like Transcription Factors/genetics , Mesangial Cells/immunology , Mesangial Cells/metabolism , Nephritis/immunology , Nuclear Proteins/genetics , Proto-Oncogene Proteins/genetics , Activating Transcription Factor 3/immunology , Activating Transcription Factor 3/metabolism , Animals , Apoptosis/immunology , Cell Cycle Proteins/immunology , Cell Cycle Proteins/metabolism , Cells, Cultured , Complement Membrane Attack Complex/adverse effects , Complement Membrane Attack Complex/metabolism , Complement Membrane Attack Complex/pharmacology , DNA Damage , Gene Expression , Gene Silencing/drug effects , Kruppel-Like Factor 6 , Kruppel-Like Transcription Factors/immunology , Kruppel-Like Transcription Factors/metabolism , Luciferases/analysis , Mesangial Cells/cytology , Nephritis/genetics , Nephritis/metabolism , Nephritis/pathology , Nuclear Proteins/immunology , Nuclear Proteins/metabolism , Plasmids , Proliferating Cell Nuclear Antigen/genetics , Proliferating Cell Nuclear Antigen/immunology , Proliferating Cell Nuclear Antigen/metabolism , Proto-Oncogene Proteins/immunology , Proto-Oncogene Proteins/metabolism , RNA, Small Interfering/pharmacology , Rats , Signal Transduction/genetics , Signal Transduction/immunology , Transcriptional Activation/immunology , Transfection , Up-RegulationABSTRACT
The activation of complement system is important factor in inflammatory, neurodegenerative and cerebrovascular diseases. CNS cells are able to synthesize complement components, and myelin and oligodendrocytes (OLG) are known to activate the classical pathway of complement in vitro in the absence of antibodies. Although activation of the complement system is known to promote tissue injury, recent evidence has also indicated that this process can have neuroprotective effects. In particular, terminal C5b-9 complexes enhance OLG survival both in vitro and in vivo. Complement activation may also reduce the accumulation of amyloid and degenerating neurons by promoting their clearance and suggest that certain inflammatory defense mechanisms in the brain may be beneficial in neurodegenerative disease. Complement system activation plays also an important role in brain damage after ischemic injury or head trauma. These findings strongly suggest that complement activation and membrane assembly of C5b-9 can play a role in injury but can also provide neuroprotection depending on the pathophysiological context.
Subject(s)
Autoimmune Diseases of the Nervous System/immunology , Complement Activation/immunology , Complement Membrane Attack Complex/immunology , Nervous System Autoimmune Disease, Experimental/immunology , Animals , Apoptosis/immunology , Brain Ischemia/immunology , Complement Membrane Attack Complex/adverse effects , Demyelinating Diseases/immunology , Demyelinating Diseases/pathology , Humans , Neurodegenerative Diseases/immunology , Oligodendroglia/immunologyABSTRACT
El sistema del complemento es un complejo formado por más de 28 proteínas plasmáticas, cuya activación secuencial en cascada tiene como resultado un producto final, el complejo de ataque de membrana (MAC ó C5b-9) y unas fracciones protéicas derivadas de la fragmentación de los diferentes componentes del complemento. Estos productos de activación del complemento, además de desarrollar diferentes funciones biológicas, se han implicado en la patogenia de diversas enfermedades autoinmunes, incluso en algunas de ellas se ha demostrado que el grado de activación del sistema del complemento está relacionado con la actividad del proceso. Objetivo de estudio: Hasta el momento no se han realizado estudios similares en la alopecia areata (AA), por lo cual el objetivo de este trabajo fue detectar la posible existencia de depósitos de MAC en biopsias cutáneas de AA. Material y métodos: Para ello dispusimos de 16 muestras: 9 de ellas pertenecientes al borde de placas alopécicas de cuero cabelludo de pacientes con diferentes formas clínicas de AA, seis, a diferentes áreas pilosas de individuos control aparentemente sanos y una, a cuero cabelludo de otro paciente control afecto de lupus eritamatoso discoide (LED). La técnica utilizada fue el sistema de inmunodetección biotina-estreptavidina (Bio.Genex) y los anticuerpos monoclonales C5b-9 de Dakopatts (Dinamarca). Resultados: En 8 de 9 biopsias de AA, se hallaron depósitos de MAC en la membrana basal (MB) de muchos folículos, fundamentalmente en la porción inferior y en el promontorio. Respecto a los controles, solo en 2 de las 6 biopsias de piel normal se detectaron depósitos de MAC en la zona de la MB adyacente a la glándula sebácea y en el LED, se observaron depósitos densos de MAC a lo largo de la MB de prácticamente todos los folículos pilosos observados. Conclusión: Los depósitos de MAC parecen ser más frecuentes en las lesiones de AA que en piel normal, pero menos intensos que en el LED. Por tanto, podríamos decir que probablemente el MAC está implicado en la patogenia de la AA y quizá también interviene en el desarrollo normal del ciclo folicular (AU)
