Listeria monocytogenesy mecanismo de caballo de Troya a través de la barrera hematoencefálica / isteria monocytogenesand Trojan horse mechanism across the blood-brain barrier

Las infecciones del sistema nervioso central son potencialmente mortales, causadas por patógenos, como bacterias, virus y hongos. Para llegar hasta el cerebro, los microorganismos utilizan diversas vías y formas. Este patogeno es una bacteria grampositiva corta, flagelar e intracelular, con la capacidad de inducir su internalización en células fagocíticas (monocitos/macrófagos) y no fagocíticas (células endoteliales). Al infectar los macrófagos, estos microorganismos se valen de su capacidad de fijación, adhesión y migración transendotelial, para cruzar la barrera hematoencefálica, finalmente, generando meningitis bacteriana. En esta revisión describimos el mecanismo de caballo de Troya usado por Listeria monocytogenespara invadir el cerebro en el desarrollo de enfermedades infecciosas e incorporamos nuevos conocimientos sobre moléculas que intervienen en dicho mecanismo(AU)

Central nervous system infections are life-threatening, caused by pathogens such as bacteria, viruses and fungi. To access the brain, microorganisms use various mechanisms. Listeria monocytogenes is a short, flagellar and intracellular gram-positive bacterium, with the ability to induce its internalization in phagocytic (monocytes/macrophages) and non-phagocytic (endothelial cells) cells. By infecting macrophages, these microorganisms take advantage of their binding, adhesion, and transendothelial migrationcapacity to cross the blood-brain barrier, finally generating bacterial meningitis. In this review we describe the Trojan horse mechanism used by Listeria monocytogenesto invade the brain in the development of infectious diseases and we incorporate new knowledge about molecules that intervene in this mechanism(AU)

Evaluation of the immunomodulatory effect of Bougainvillea xbuttiana (Var. Orange) on the cytokine production induced by Botrhops jararaca venom in macrophages / Evaluación del efecto inmunomodulador de Bougainvillea xbuttiana (var. naranja) sobre la producción de citoquinas de macrófagos inducida pelo veneno de Bothrops jararaca

To study the effect of 50% ethanol extract of Bougainvillea xbuttiana on the enzymatic activity, cell via bility and cytokine production provoked by the venom of Bothrops jararaca in macro - phages. Three assays were used to study the effects of B. xbuttiana extract on the damage pro - duced by B. jararaca : Enzymatic activity was detected by measuring the proteoly tic and phos - pholipase A2; macrophages cytotoxicity was determined by the MTT method; levels of cytokine were evaluated using ELISA and a biological assay. After treatment with 300 µg/mL B. xbuttiana extract for 30 min, the proteolytic and phospholipase A2 activities of the venom were reduced to 95 and 61%, respectively. In macrophages cultures treated with B. xbuttiana extract combined with venom, the production of TNF - α, IL - 6 and IFN - γ was reduced, whereas IL - 10 was potenti - ated. Our results support the potential effect of the B. xbuttiana extract as a complementary therapy against the toxicity caused by the venom of B . jararaca snakes

Estudiar el efecto del extracto etanólico al 50% de Bougainvillea xbuttiana sobre la actividad enzimática viabilidad celular y producci ón de citoquinas provocada por el veneno de Bothrops jararaca en macrófagos Se utilizaron tres ensayos para estudiar los efectos del extracto de B. xbuttiana sobre el daño producido por B. jararaca : Se detectó actividad enzimática mediante la medición del proteolítico y fosfolipasa A2; la citotoxicidad de los macrófagos se determinó por el método MTT; Los niveles de citoquinas se evaluaron utilizando ELISA y un ensayo biológico. Después del tratamiento con 300 µg/mL de extracto de B. xbuttiana durante 30 mi n, las actividades proteolíticas y de fosfolipasa A2 del veneno se redujeron a 95 y 61%, respectivamente. En cultivos de macrófagos tratados con extracto de B. xbuttiana combinado con veneno, la producción de TNF - α, IL - 6 e IFN - γ se redujeron, mientras que IL - 10 se potenció. Nuestros resultados apoyan el efecto potencial del extracto de B. xbuttiana como terapia complementaria frente a la toxicidad provocada por el veneno de B. jararaca .

The role and impact of macrophages in the pathogenesis of primary biliary cholangitis / 临床肝胆病杂志

Primary biliary cholangitis （PBC） is a chronic autoimmune disease of cholestasis in which immune factors lead to progressive small bile duct destruction， cholestasis， and eventually liver fibrosis， liver cirrhosis， and even liver failure. Macrophages， as a group with functional heterogeneity， play different roles in the whole disease process of PBC. This article summarizes the possible ways by which macrophages are involved in the pathogenesis of PBC and discusses their impact on the disease and the potential therapeutic targets of macrophages. It is pointed out that macrophages are mainly involved in innate immunity in PBC injury and are associated with gut microbiota dysbiosis， and they are also associated with cholestasis， liver fibrosis， and liver cirrhosis in the later stages of the disease.

Therapeutic effect of transplantation of bone marrow mesenchymal stem cells co-cultured with bone marrow M2 macrophages on a rat model of liver cirrhosis / 临床肝胆病杂志

ObjectiveTo investigate the effect of transplantation of bone marrow mesenchymal stem cells （BMSCs） co-cultured with bone marrow-derived M2 macrophages （M2-BMDMs）， named as BMSCM2， on a rat model of liver cirrhosis induced by carbon tetrachloride （CCl4）/2-acetaminofluorene （2-AAF）. MethodsRat BMDMs were isolated and polarized into M2 phenotype， and rat BMSCs were isolated and co-cultured with M2-BMDMs at the third generation to obtain BMSCM2. The rats were given subcutaneous injection of CCl4 for 6 weeks to establish a model of liver cirrhosis， and then they were randomly divided into model group （M group）， BMSC group， and BMSCM2 group， with 6 rats in each group. A normal group （N group） with 6 rats was also established. Since week 7， the model rats were given 2-AAF by gavage in addition to the subcutaneous injection of CCl4. Samples were collected at the end of week 10 to observe liver function， liver histopathology， and hydroxyproline （Hyp） content in liver tissue， as well as changes in the markers for hepatic stellate cells， hepatic progenitor cells， cholangiocytes， and hepatocytes. A one-way analysis of variance was used for comparison of continuous data between multiple groups， and the least significant difference t-test was used for further comparison between two groups. ResultsCompared with the N group， the M group had significant increases in the activities of serum alanine aminotransferase （ALT） and aspartate aminotransferase （AST） （P<0.01）； compared with the M group， the BMSC and BMSCM2 groups had significant reductions in ALT and AST （P<0.01）， and the BMSCM2 group had significantly better activities than the BMSC group （P<0.05）. Compared with the N group， the M group had significant increases in Hyp content and the mRNA and protein expression levels of alpha-smooth muscle actin （α-SMA） in the liver （P<0.01）； compared with the M group， the BMSC and BMSCM2 groups had significant reductions in Hyp content and the expression of α-SMA （P<0.05）， and the BMSCM2 group had a significantly lower level of α-SMA than the BMSC group （P<0.01）. Compared with the N group， the M group had significant increases in the mRNA expression levels of the hepatic progenitor cell markers EpCam and Sox9 and the cholangiocyte markers CK7 and CK19 （P<0.01） and significant reductions in the expression levels of the hepatocyte markers HNF-4α and Alb （P<0.01）； compared with the M group， the BMSC and BMSCM2 groups had significant reductions in the mRNA expression levels of EpCam， Sox9， CK7， and CK19 （P<0.05） and significant increases in the mRNA expression levels of HNF-4α and Alb （P<0.05）， and compared with the BMSC group， the BMSCM2 group had significant reductions in the mRNA expression levels of EpCam and CK19 （P<0.05） and significant increase in the expression level of HNF-4α （P<0.05）. ConclusionM2-BMDMs can enhance the therapeutic effect of BMSCs on CCl4/2-AAF-induced liver cirrhosis in rats， which provides new ideas for further improving the therapeutic effect of BMSCs on liver cirrhosis.

Mechanism of action of macrophage efferocytosis in acute and chronic liver diseases and related targeted therapy / 临床肝胆病杂志

Efferocytosis refers to the process by which apoptotic cells are engulfed and cleared by phagocytes， including professional phagocytes， such as macrophages and dendritic cells， and non-professional phagocytes， such as epithelial cells. Liver macrophages are the main cells with the function of efferocytosis in the liver. In recent years， an increasing number of studies have shown that various acute and chronic liver diseases are associated with the efferocytosis function of liver macrophages， including acute liver injury， alcoholic liver disease， nonalcoholic fatty liver disease， autoimmune liver disease， liver fibrosis， and liver cancer. This article elaborates on the expression of molecules associated with the efferocytosis function of macrophages， the process of efferocytosis， and the role of efferocytosis function in different liver diseases， so as to provide new ideas for the treatment of liver diseases.

Therapeutic effect of Xiayuxue decoction on a mouse model of nonalcoholic fatty liver disease induced by high-fat diet and its mechanism / 临床肝胆病杂志

ObjectiveTo investigate the mechanism of action of Xiayuxue decoction in inhibiting nonalcoholic fatty liver disease （NAFLD） induced by high-fat diet in mice by regulating nucleotide binding oligomerization domain like receptor containing pyrin domain protein 6 （NLRP6）. MethodsA total of 15 male C57BL/6 mice were randomly divided into low-fat diet （LFD） group， high-fat diet （HFD） group， and Xiayuxue decoction-HFD group （XYXD group）， with 5 mice in each group. Liver function parameters （alanine aminotransferase ［ALT］ and aspartate aminotransferase ［AST］） and blood lipid metabolic indicators （triglycerides ［TG］ and total cholesterol ［TC］） were measured； HE staining and oil red O staining were performed for liver tissue to observe histomorpholoty and lipid droplet deposition； quantitative real-time PCR was used to measure the expression levels of inflammatory factors （tumor necrosis factor-α ［TNF-α］， interleukin-1β ［IL-1β］， interleukin-18 ［IL-18］， and NLRP6） in liver tissue； Western blot was used to measure the protein expression levels of NLRP6， nuclear factor-kappa B （NF-κB）， and NF-κB p65； immunohistochemistry was used to measure the expression of NLRP6 and CD68. Mouse Raw264.7 cells were treated with palmitic acid （PA）， lipopolysaccharide， and serum containing Xiayuxue decoction to observe inflammation. A one-way analysis of variance was used for comparison of continuous data between multiple groups， and the least significant difference t-test was used for further comparison between two groups. ResultsCompared with the LFD group， the HFD group had significant increases in the serum levels of ALT， AST， TC， and TG （all P<0.05）. Liver histopathological examination showed that the HFD group had marked hepatic steatosis and a signficant increase in NAS score （P<0.05）， and quantitative real-time PCR showed significant increases in the inflammatory factors such as IL1β and IL-18 and a significant reduction in the expression of NLRP6 （all P<0.05）. Immunohistochemistry showed that the expression of NLRP6 showed a similar trend as that of the macrophage marker CD68. Western blot showed that after the downregulation of NLRP6 expression， there was a significant increase in phosphorylated NF-κB p65 （P<0.05）. Compared with the HFD group， Xiayuxue decoction effectively improved liver inflammation， upregulated the expression of NLRP6， and downregulated phosphorylated NF-κB p65 in HFD mice （all P<0.05）. After Raw264.7 cells were treated with PA， NLRP6 was downregulated to promote the progression of inflammation （P<0.05）， and treatment with Xiayuxue decoction could upregulate NLRP6 and inhibit inflammation NF-κB （P<0.05）. ConclusionXiayuxue decoction can effectively improve hepatic steatosis and liver inflammation in a mouse model of NAFLD， possibly by regulating NLRP6/NF-κB to alleviate macrophage activation.

Isolation and identification of exosomes from mouse primary peritoneal macrophages / 中国生物制品学杂志

@#Objective To isolate,purify and identify exosomes secreted by mouse primary peritoneal macrophages.Methods Five male C57BL/6 mice were intraperitoneally injected with 3% mercaptoacetate broth respectively,and the primary peritoneal macrophages were obtained by lavage,and then the purity was analyzed by flow cytometry.The exosomes of mouse primary peritoneal macrophages were extracted by ExoQuick TC exosome kit,which were measured for the protein content with BCA kit,observed for the morphology by transmission electron microscopy,detected for the particle size and distribution with nanoparticle tracking analyzer,and determined for the expression of exosome-specific markers(CD9,CD63 and TSG101) by Western blot.Results About 5 × 10~6 peritoneal macrophages with the purity of(99.17±0.65)%were obtained from each mouse.Approximately 869 μg of exosomal protein was extracted from 5 mL of mouse primary peritoneal macrophage culture supernatant.The exosomes of mouse primary peritoneal macrophages were typical tea saucerlike vesicles with strong refraction under electron microscopy,and highly expressed the exosome-specific markers TSG101,CD63 and CD9.The particle size distribution was concentrated between 100 and 200 nm,with an average particle size of175.2 nm.Conclusion Intraperitoneal injection of mercaptoacetate broth can improve the yield of mouse primary peritoneal macrophages.ExoQuick TC.exosome kit can extract sufficient amount of exosomes with high purity from mouse primary peritoneal macrophages.

Role of Macrophage Activation and Polarization in Myocardial Fibrosis and Intervention of Traditional Chinese Medicine / 中国实验方剂学杂志

Myocardial fibrosis （MF） is a common pathological manifestation of various heart diseases. Due to the non-renewable nature of myocardial cells， the occurrence of MF represents irreversible damage to the myocardium. Previous studies have suggested that fibroblast-mediated collagen deposition is the main mechanism of MF. Recent studies have found that there is an immune regulation mechanism in the heart itself， and macrophage activation/polarization plays an important role in MF. With the deepening of traditional Chinese medicine research， scholars have found that traditional Chinese medicine can interfere with MF by regulating the renin-angiotensin-aldosterone system （RAAS） system and the inflammatory process， repairing the extracellular matrix， managing oxidative stress， and maintaining the balance of autophagy. This process is closely related to the activation and M1/M2 polarization of macrophages. Throughout the MF process， macrophage activation is beneficial， but excessive activation will be harmful. In the early stage of MF， appropriate M1 macrophage polarization is conducive to activating immunity and removing harmful substances. In the middle and late stages of MF， appropriate M2 macrophage polarization is conducive to remodeling the damaged myocardium. If macrophage activation is excessive/insufficient， or the balance of M1/M2 macrophage polarization is broken， the effect changes from improvement to destruction. Traditional Chinese medicines that regulate the activation/polarization of macrophages have the effects of replenishing Qi and nourishing Yin， as well as regulating Qi and activating blood， but there are also some heat-clearing， dampness-drying， and detoxification products. Therefore， the occurrence of MF may be caused by Qi and Yin deficiency， damp heat accumulation， and Qi stagnation and blood stasis. By summarizing the biological processes involved in macrophage activation/polarization in MF， this paper expounded on the research progress of traditional Chinese medicine in regulating macrophage activation and M1/M2 polarization from different angles to improve MF， so as to provide a reference for the treatment of MF with traditional Chinese medicine.

Regulatory effect and mechanism of macrophage polarization in liver fibrosis / 临床肝胆病杂志

Liver fibrosis is the healing reaction of chronic liver injury caused by various factors such as viral infection， alcohol， and chemical substances and is a key link in the progression of chronic liver diseases to liver cirrhosis and liver cancer. Liver macrophages are considered important mediators of liver injury and repair， and the polarization trend of macrophages has a bidirectional regulatory effect on liver fibrosis. This article reviews the role of different phenotypes of liver macrophages in the development and progression of liver fibrosis， in order to provide new ideas for the prevention and treatment of fibrosis.

GRK2 inhibits Flt-1+ macrophage infiltration and its proangiogenic properties in rheumatoid arthritis

Rheumatoid arthritis (RA) is an autoimmune disease with a complex etiology. Monocyte-derived macrophages (MDMs) infiltration are associated with RA severity. We have reported the deletion of G-protein-coupled receptor kinase 2 (GRK2) reprograms macrophages toward an anti-inflammatory phenotype by recovering G-protein-coupled receptor signaling. However, as more GRK2-interacting proteins were discovered, the GRK2 interactome mechanisms in RA have been understudied. Thus, in the collagen-induced arthritis mouse model, we performed genetic GRK2 deletion using GRK2f/fLyz2-Cre+/- mice. Synovial inflammation and M1 polarization were improved in GRK2f/fLyz2-Cre+/- mice. Supporting experiments with RNA-seq and dual-luciferase reporter assays identified peroxisome proliferator-activated receptor γ (PPARγ) as a new GRK2-interacting protein. We further confirmed that fms-related tyrosine kinase 1 (Flt-1), which promoted macrophage migration to induce angiogenesis, was inhibited by GRK2-PPARγ signaling. Mechanistically, excess GRK2 membrane recruitment in CIA MDMs reduced the activation of PPARγ ligand-binding domain and enhanced Flt-1 transcription. Furthermore, the treatment of mice with GRK2 activity inhibitor resulted in significantly diminished CIA pathology, Flt-1+ macrophages induced-synovial inflammation, and angiogenesis. Altogether, we anticipate to facilitate the elucidation of previously unappreciated details of GRK2-specific intracellular signaling. Targeting GRK2 activity is a viable strategy to inhibit MDMs infiltration, affording a distinct way to control joint inflammation and angiogenesis of RA.

Actividad fagocítica y toxicidad del extracto acuoso de Schinus molle L. sobre Mus musculus BALB/c

Introducción: La medicina tradicional podría ser una alternativa segura para potenciar la inmunidad de pacientes inmunocomprometidos propensos a infecciones recurrentes. Objetivo: Evaluar la actividad fagocítica y toxicidad in vivo del extracto acuoso de Schinus molle L. en Mus musculus BALB/c. Material y Métodos: Estudio experimental que utilizó una dosis única de extracto acuoso de hojas de Schinus molle de 2000 mg/kg. La fagocitosis in vivo se determinó en 10 ejemplares de M. musculus BALB/c que cumplieron criterios de inclusión y exclusión, distribuidos aleatoria y equitativamente en los grupos control y experimental. Los especímenes del grupo experimental fueron inoculados vía intraperitoneal con 0,5 ml de suspensión de Staphylococcus aureus y 0,5 ml del extracto acuoso. Los del grupo control con 0,5 ml del mismo inóculo bacteriano y 0,5 ml de solución salina estéril. La toxicidad del extracto se evaluó por el método de las clases de toxicidad aguda en 12 ejemplares de ratones con las mismas características y cumpliendo los mismos criterios aplicados en la evaluación de la fagocitosis in vivo. Resultados: El 57,1 % de los macrófagos expuestos al extracto acuoso de S. molle presentaron importante actividad fagocítica, encontrándose una media de 21 bacterias fagocitadas por macrófago. No se evidenciaron significativamente signos ni síntomas de toxicidad en los especímenes durante los 14 días de experimentación. Conclusiones: El extracto acuoso de S. molle incrementó significativamente la fagocitosis in vivo de los macrófagos peritoneales de M. musculus BALB/c, sin evidencia de clínica de toxicidad y en ausencia de mortalidad.

Introduction: Traditional medicine could be a safe alternative to enhance the immunity of immunocompromised patients prone to recurrent infections. Objective: To evaluate the phagocytic activity and in vivo toxicity of the aqueous extract of Schinus molle L. on Mus musculus BALB/c. Material and Methods: Experimental study that used a single dose of aqueous extract of Schinus molle leaves of 2000 mg/kg. In vivo phagocytosis was determined in 10 specimens of M. musculus BALB/c that met the inclusion and exclusion criteria, which were randomly and equally distributed in the control and experimental groups. The specimens of the experimental group were inoculated intraperitoneally with 0.5 ml of Staphylococcus aureus suspension and 0.5 ml of the aqueous extract. Those of the control group were inoculated with 0.5 ml of the same bacterial inoculum and 0.5 ml of sterile saline solution. The toxicity of the extract was evaluated by the method of the acute toxicity classes in 12 specimens of mice with the same characteristics that fulfilled the same criteria applied in the evaluation of phagocytosis in vivo. Results: The results demonstrate that 57.1% of the macrophages exposed to the aqueous extract of S. molle showed significant phagocytic activity, finding an average of 21 phagocytosed bacteria per macrophage. No significant signs or symptoms of toxicity were evidenced in the specimens during the 14 days of experimentation. Conclusions: The aqueous extract of S. molle significantly increased in vivo phagocytosis of peritoneal macrophages from M. musculus BALB/c, without clinical evidence of toxicity and in the absence of mortality.

Changes of intestinal wall barrier function and its correlation with susceptibility to infection in patients with cirrhotic portal hypertension / 中华肝脏病杂志

Objective: To investigate the changes of intestinal wall barrier function and its correlation with infection occurrence in patients with cirrhotic portal hypertension. Methods: 263 patients with cirrhotic portal hypertension were split into: the clinically evident portal hypertension (CEPH) combined with infection group (n = 74); CEPH group (n = 104); and Non-CEPH group (n = 85). Among them, 20 CEPH patients and 12 non-CEPH patients in non-infection status were subjected to sigmoidoscopy. Immunohistochemical staining was used to detect the expression of trigger receptor-1 (TREM-1), CD68, CD14, the inducible nitric oxide synthase molecule, and Escherichia coli (E.coli) in the medullary cells of the colon mucosa. An enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of soluble myeloid cell trigger receptor-1 (sTREM-1), soluble leukocyte differentiation antigen-14 subtype (sCD14-ST) and intestinal wall permeability index enteric fatty acid binding protein (I-FABP). Fisher's exact probability method, one-way ANOVA, Kruskal-Wallis-H test, Bonferroni method, and Spearman correlation analysis were used for statistical analysis. Results: The serum sTREM-1 and I-FABP levels were higher in CEPH patients than those of non-CEPH patients in the non-infectious state (P < 0.05), but the difference in blood sCD14-ST levels was not statistically significant (P > 0.05). Serum levels of sTREM-1, sCD14-ST, and I-FABP in infected patients were higher than those in patients without a concurrent infection (P < 0.05). Serum sCD14-ST levels were positively correlated with serum sTREM-1, C-reactive protein (CRP), and procalcitonin (PCT), and sTREM-1 levels were also positively correlated with CRP and PCT (r > 0.5, P < 0.001). The rates of CD68, inducible nitric oxide synthase, CD14-positive cells, and E.coli-positive glands were higher in the intestinal mucosa of the CEPH group than those of the control group (P < 0.05). Spearman's correlation analysis showed that the rate of E.coli-positive glands in CEPH patients was positively correlated with the expression of molecular markers CD68 and CD14 in the lamina propria macrophages. Conclusion: Patients with cirrhotic portal hypertension have increased intestinal permeability and inflammatory cells, accompanied by bacterial translocation. Serum sCD14-ST and sTREM-1 can be used as indicators to predict and evaluate the occurrence of infection in patients with cirrhotic portal hypertension.

Protective mechanism of salvianolic acid B on blood vessels / 中国中药杂志

Salvianolic acid B(Sal B) is the main water-soluble component of Salvia miltiorrhiza Bunge. Studies have found that Sal B has a good protective effect on blood vessels. Sal B can protect endothelial cells by anti-oxidative stress, inducing autophagy, inhibiting endoplasmic reticulum stress(ERS), inhibiting endothelial inflammation and adhesion molecule expression, inhibiting endothelial cell permeability, anti-thrombosis, and other ways. In addition, Sal B can alleviate endothelial cell damage caused by high glucose(HG). For vascular smooth muscle cell(VSMC), Sal B can reduce the synthesis and secretion of inflammatory factors by inhibiting cyclooxygenase. It can also play a vasodilatory role by inhibiting Ca~(2+) influx. In addition, Sal B can inhibit VSMC proliferation and migration, thereby alleviating vascular stenosis. Sal B also inhibits lipid deposition in the subendothelium, inhibits macrophage conversion to foam cells, and reduces macrophage apoptosis, thereby reducing the volume of subendothelial lipid plaques. For some atherosclerosis(AS) complications, such as peripheral artery disease(PAD), Sal B can promote angiogenesis, thereby improving ischemia. It should be pointed out that the conclusions obtained from different experiments are not completely consistent, which needs further research. In addition, previous pharmacokinetics showed that Sal B was poorly absorbed by oral administration, and it was unstable in the stomach, with a large first-pass effect in the liver. Sal B had fast distribution and metabolism in vivo and short drug action time. These affect the bioavailability and biological effects of Sal B, and the development of clinically valuable Sal B non-injectable delivery systems remains a great challenge.

Identification of M1 macrophage-related genes in rejection after kidney transplantation based on weighted gene co-expression network analysis / 器官移植

Objective To identify M1 macrophage-related genes in rejection after kidney transplantation and construct a risk prediction model for renal allograft survival. Methods GSE36059 and GSE21374 datasets after kidney transplantation were downloaded from Gene Expression Omnibus (GEO) database. GSE36059 dataset included the samples from the recipients with rejection and stable allografts. Using this dataset, weighted gene co-expression network analysis (WGCNA) and differential analysis were conducted to screen the M1 macrophage-related differentially expressed gene (M1-DEG). Then, GSE21374 dataset (including the follow-up data of graft loss) was divided into the training set and validation set according to a ratio of 7∶3. In the training set, a multivariate Cox's model was constructed using the variables screened by least absolute shrinkage and selection operator (LASSO), and the ability of this model to predict allograft survival was evaluated. CIBERSORT was employed to analyze the differences of infiltrated immune cells between the high-risk group and low-risk group, and the distribution of human leukocyte antigen (HLA)-related genes was analyzed between two groups. Gene set enrichment analysis (GSEA) was used to further clarify the biological process and pathway enrichment in the high-risk group. Finally, the database was employed to predict the microRNA (miRNA) interacting with the prognostic genes. Results In the GSE36059 dataset, 14 M1-DEG were screened. In the GSE21374 dataset, Toll-like receptor 8 (TLR8), Fc gamma receptor 1B (FCGR1B), BCL2 related protein A1 (BCL2A1), cathepsin S (CTSS), guanylate binding protein 2(GBP2) and caspase recruitment domain family member 16 (CARD16) were screened by LASSO-Cox regression analysis, and a multivariate Cox's model was constructed based on these 6 M1-DEG. The area under curve (AUC) of receiver operating characteristic of this model for predicting the 1- and 3-year graft survival was 0.918 and 0.877 in the training set, and 0.765 and 0.736 in the validation set, respectively. Immune cell infiltration analysis showed that the infiltration of rest and activated CD4+ memory T cells, γδT cells and M1 macrophages were increased in the high-risk group (all P < 0.05). The expression level of HLA I gene was up-regulated in the high-risk group. GSEA analysis suggested that immune response and graft rejection were enriched in the high-risk group. CTSS interacted with 8 miRNA, BCL2A1 and GBP2 interacted with 3 miRNA, and FCGR1B interacted with 1 miRNA. Conclusions The prognostic risk model based on 6 M1-DEG has high performance in predicting graft survival, which may provide evidence for early interventions for high-risk recipients.

Animalexperiment on regulating effect of stored red blood cells transfusion on BMDMs in inflammatory conditions / 中国输血杂志

【Objective】 To discuss the regulating effect of stored red blood cells (RBCs) transfusion on BMDMs in inflammatory conditions, and the relationship between stored RBCs transfusion and inflammatory response induced by bacterial infection. 【Methods】 Forty C57BL/6 male mice of 6-8 weeks (18-22 g/mouse) were randomly divided into experimental group and control group. Each mouse was infected with 200 µL Pseudomonas aeruginosa injecting into the tail vein, and 400 µL fresh (storage >14 d) and stored RBCs (storage 0.05). F4/80 of experimental group and control group 2, 4 and 8 hours after RBCs infused were 1.83±0.11 vs 0.75±0.06, 0.46±0.06 vs 0.33±0.06 (P0.05), respectively. iNOS, TNF-α, MCP1 of M1 in liver of experimental group and control group 2, 4 and 8 hours after RBCs infused were respectively: iNOS 3.44±0.20 vs 2.46±0.08, 9.25±0.55 vs 2.67±0.12, 2.80±0.08 vs 2.39 ±0.01; TNF-α 1.69±0.22 vs 1.13±0.03, 1.44±0.24 vs 0.96±0.09, 1.31±0.05 vs 0.96±0.06; MCP1 4.96±0.08 vs 4.28±0.27, 4.63±0.04 vs 2.07±0.09, 2.28±0.19 vs 1.33±0.03 (P0.05). 【Conclusion】 Stored RBCs infusion can greatly promote the M1 polarization of BMDMs in liver.

Effect of histone H4 on polarization of alveolar macrophages in mice with acute respiratory distress syndrome / 中国职业医学

Objective To investigate the role of histone H4 in the polarization of alveolar macrophages (AM) in lipopolysaccharide (LPS)-induced acute respiratory distress syndrome (ARDS) in mice. Methods i) The specific pathogen free male C57BL/6 mice were randomly divided into control group and 2, 4, 6 and 8 mg/kg LPS groups, with six mice in each group. The mice in the LPS groups were intratracheally administered LPS according to their respective doses, while the mice in the control group received an equivalent volume of 0.9% saline. After 12 hours, the arterial blood gas was analyzed, and the pulmonary edema and histopathological changes in lung tissues of mice in each group were observed. The level of histone H4 in bronchoalveolar lavage fluid (BALF) of mice was detected using enzyme-linked immunosorbent assay , and mice AMs of the five group were isolated using adherent method. ii) AMs from normal mice were isolated using adherent method and randomly divided into control group, histone H4 injury group, BALF injury group and anti-histone H4 antibody (anti-H4) intervention group. In the histone H4 injury group, AMs were treated with histone H4 at a final concentration of 20 mg/L. In the BALF injury group and anti-H4 intervention group, AMs were treated with 200 μL BALF supernatant from mice intratracheally administered 6 mg/kg body weight LPS, with the latter group treated with 25 mg/L anti-H4 antibody. The control group AMs were treated with phosphate-buffered saline. iii) After 12 hours of stimulation, the cells were collected, and the relative expression of tumor necrosis factor-α (Tnfa), interleukin-1β (Il1b), differentiation antigen 206 (Cd206) and arginase 1 (Arg1) in AMs was detected using real-time quantitative polymerase chain reaction. Results i) Compared with the control group, mice in all four LPS groups exhibited rapid breathing, inflammatory reaction and lung edema in lung tissues, which were aggravated in a dose-dependent manner. The ratio of partial pressure of arterial oxygen to fraction of inspired oxygen in mice decreased with the increase of LPS dose (P<0.05). The wet/dry weight ratio of lung, the level of histone H4 in BALF and the relative expression of Tnfa and Il1b mRNA in AMs increased with the increase of LPS dose (all P<0.05). The mice in the 6 and 8 mg/kg LPS groups developed ARDS. The level of histone H4 in BALF and the relative expression of Tnfa and Il1b mRNA in AMs of mice in 6 and 8 mg/kg LPS groups were higher than those in the other three groups (all P<0.05). ii) The relative expression of Tnfa and Il1b mRNA increased (both P<0.05), and the relative expression of Cd206 and Arg1 mRNA decreased (both P<0.05) in AMs of histone H4 injury group and BALF injury group compared with the control group. Compared with BALF injury group, the relative mRNA expression of Tnfa and Il1b in AMs of anti-H4 intervention group decreased (both P<0.05), while the relative expression of Arg1 mRNA increased (P<0.05). Conclusion LPS can induce a dose-dependent increase in histone H4 levels in BALF in mice. Histone H4 drives the development of ARDS by activating AMs to M1 polarization. Antagonizing histone H4 to interfere with AM polarization to M1 could be a target for the treatment of ARDS.

Effect of Tuina Manipulation on the Expression of CD68 and MMP-2 in Rabbits with Carotid Atherosclerotic Plaque / 中医杂志

ObjectiveTo explore the effect of tuina manipulation with different cervical rotation angle on carotid atherosclerosis. MethodsTwenty-five New Zealand rabbits were randomly selected， 5 of which as the control group， and the other 20 rabbits as the modeling group. The modeling group were made by arterial intimal balloon injury combined with high-fat diet， and were randomly divided into model group， cervical rotation angle of 90° group， cervical rotation angle of 105° group and cervical rotation angle of 120° group （5 rabbits in each group） after successful modeling. After relaxing the neck muscles of rabbits with manipulation of one-finger meditation， rolling and dialing， the cervical vertebrae of each group was rotated by 90°， 105°and 120° respectively for 2 weeks， while the other two groups were not intervened. Then took the entire length of the left common carotid artery from the rabbit， observed the pathological morphology of the carotid artery tissue using HE staining， and observed the expression of CD68 and MMP-2 in the carotid artery tissue using immunohistochemistry staining， and conducted semi quantitative analysis. ResultsHE staining showed that there was no obvious pathological change in the carotid artery in the control group； the model group showed subintimal foam cells gathered， vascular smooth muscle cells increased and arranged disorderly， and some vascular smooth muscle cells entered the intima from the media to form a fibrous cap. Compared with the model group， the number of foam cells increased and the symptoms of lumen stenosis were improved in all groups of cervical rotation； compared with the group with 90°rotation， the endothelium tended to fall off slightly in the group with 105°rotation and 120°rotation. Compared with the control group， the model group and the cervical rotation angle of 90° group ， cervical rotation angle of 105° group and cervical rotation angle of 120° group showed an increase in positive expression of CD68 and MMP-2 （P＜0.05）； compared with the model group， the positive expression of CD68 and MMP-2 in the cervical rotation angle of 90° group， cervical rotation angle of 105° group and cervical rotation angle of 120° group decreased （P＜0.05）； there was no statistically significant difference in CD68 positive expression among cervical rotation angle of 90° group， cervical rotation angle of 105° group and cervical rotation angle of 120° group compared in pairs （P＞0.05）； the positive expression of MMP-2 in the group of cervical rotation angle of 105° group and cervical rotation angle of 120 °group was higher than that in cervical rotation angle of 90° group （P＜0.05）. ConclusionTuina manipulation can promote the stability of plaque by reducing the CD68 content of macrophages and the level of MMP-2 in plaque； but as the rotation angle of the cervical spine increased， the expression of MMP-2 may increase， which may reduce the plaque stability to a certain extent and affect the therapeutic effect of tuina.

Effect of Zhuangyao Tongluo Formula（壮腰通络方） Containing Serum on TNF-α-mediated Vicious Circle of Pyroptosis of Nucleus Pulposus Cells / 中医杂志

ObjectiveTo explore the mechanism of Zhuangyao Tongluo Formula（壮腰通络方，ZTF） in delaying intervertebral disc degeneration. MethodsM1 macrophages were induced from THP-1 cells using LPS， IFN-γ and PMA. The induced M1 macrophages were then co-cultured with nucleus pulposus cells in a transwell system. Fetal bovine serum was used as the control serum， and the effects of different concentrations （5%， 10%， 15%， 20%） of serum from rats treated with ZTF on the activity of M1 macrophages and nucleus pulposus cells were analyzed using MTT assay. Experiment 1 was established， including the nucleus pulposus cell control group， M1 macrophage control group， nucleus pulposus cell + ZTF group， nucleus pulposus cell + TNF control group， nucleus pulposus cell + TNF + ZTF group， co-culture group， and co-culture + ZTF group. The levels of IL-1β， and IL-18 in the culture supernatant were detected using ELISA. The mRNA expression of IL-1β and IL-18 in nucleus pulposus cells was detected using qPCR. Additionally， the expression of GSDMD protein in nucleus pulposus cells was detected using cell immunofluorescence. In experiment 2， co-culture groups were constructed using TNF-α overexpression （OE） or empty vector （EV） plasmids， including co-culture group， TNF-EV + co-culture group， TNF-EV co-culture group + ZTF， co-culture + ZTF group， TNF-OE co-culture group + ZTF， and TNF-OE + co-culture group. The mRNA and protein expression of TNF-α in M1 cells in each group were detected using qPCR and WB. ResultsThe ZTF with 10% serum was selected for subsequent experiments. The results of experiment 1 showed that compared to the control group of nucleus pulposus cells， there was no statistically significant difference in the levels of IL-1β， IL-18， mRNA， and GSDMD expression in the nucleus pulposus cells + ZTF group （P＞0.05）. However， the TNF-α + co-culture group showed a significant increase in IL-1β， IL-18 levels， mRNA， and GSDMD expression （P＜0.01）. When compared to the co-culture group， the ZTF+ co-culture group showed a significant decrease in IL-1β， IL-18 levels， mRNA， and GSDMD expression （P＜0.01）. The results of experiment 2 showed that there was no statistically significant difference in TNF-α mRNA and protein expression between the empty vector plasmids + co-culture group and the co-culture group （P＞0.05）. Compared to the empty vector + co-culture group， the expression of TNF-α mRNA and protein was significantly reduced in the empty vector co-culture + ZTF group （P＜0.01）. Compared to the co-culture group and the empty vector + co-culture group， the expression of TNF-α mRNA and protein was significantly reduced in the co-culture + ZTF group （P＜0.01）. Compared to the co-culture + ZTF group， the expression of TNF-α mRNA and protein significantly increased in the overexpression vector co-culture + ZTF group （P＜0.01）. Compared to the overexpression vector co-culture + ZTF group， the expression of TNF-α mRNA and protein significantly increased in the overexpression vector co-culture group （P＜0.01）. ConclusionZTF serum can inhibit the TNF-α-induced apoptosis of nucleus pulposus cells and delay lumbar disc degeneration by reducing the expression of TNF-α in M1 macrophages.

Icariin Ameliorates Efferocytosis Dysfunction of Alveolar Macrophages Stimulated by Cigarette Smoke Extract via PPARγ Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the mechanism of icariin in ameliorating efferocytosis dysfunction and inflammatory response of alveolar macrophages induced by cigarette smoke extract via the peroxisome proliferator-activated receptor gamma （PPARγ） signaling pathway. MethodThe untreated rat alveolar macrophages （NR8383） were taken as the blank group. The NR8383 cells treated with 10% cigarette smoke extract were divided into model， low-， medium-， and high-dose （10， 20， 40 μmol·L-1） icariin， PPARγ inhibitor， and PPARγ inhibitor + low-， medium-， and high-dose icariin groups. Alamar blue colorimetry was employed to examine the proliferation and toxicity of icariin on NR8383 cells. The efferocytosis rate of NR8383 cells was detected by flow cytometry. Enzyme-linked immunosorbent assay was employed to measure the levels of tumor necrosis factor-alpha （TNF-α）， transforming growth factor-β1 （TGF-β1）， and milk fat globule-epidermal growth factor 8 （MFG-E8）. Western blot and Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） were employed to determine the protein and mRNA levels， respectively， of PPARγ， CD36， and RAS-related C3 botulinum toxin substrate 1 （Rac1）. ResultThe efferocytosis dysfunction model of NR8383 was established with the cigarette smoke extract. Compared with the blank control group， the model group showed decreased efferocytosis rate （P<0.05）， elevated TNF-α level （P<0.05）， lowered TGF-β1 and MFG-E8 levels （P<0.01）， and down-regulated mRNA and protein levels of PPARγ， CD36， and Rac1 （P<0.05， P<0.01）. Compared with the model group， the treatment with icariin increased the efferocytosis rate （P<0.05， P<0.01）， lowered the TNF-α level （P<0.01）， elevated TGF-β1 and MFG-E8 levels （P<0.05）， and up-regulated the protein and mRNA levels of PPARγ， CD36， and Rac1 （P<0.05， P<0.01）. Compared with icariin alone， PPARγ inhibitor + icariin decreased the efferocytosis rate （P<0.05） and down-regulated the protein and mRNA levels of PPARγ （P<0.05， P<0.01）. In addition， PPARγ inhibitor + low-dose icariin down-regulated the protein level of CD36 （P<0.01） and PPARγ inhibitor + low-/medium-dose icariin up-regulated the protein level of Rac1 （P<0.05）. ConclusionIcariin ameliorates the cigarette smoke extract-induced efferocytosis dysfunction of alveolar macrophage by regulating the PPARγ signaling pathway and cytoskeletal structure rearrangement.

Effect of cold stimulation on phenotype of alveolar macrophages in mice / 中国生物制品学杂志

@#Objective To investigate the effect of cold stimulation on the phenotype of alveolar macrophages(MH-S cells) in mice. Methods MH-S cells were cultured at 37 ℃ for 24 h,and cold stimulated at 36,34 and 32 ℃ for 0,0. 5,1,3,6,9 and 12 h respectively. The mRNA transcription levels of interleukin-1β(IL-1β) and interleukin-10(IL-10) genes in MH-S cells were detected by qRT-PCR. MH-S cells were cultured at 37 ℃ for 24 h,and cold stimulated at 34 ℃ for 0. 5 h,which were detected for the mRNA transcription levels of tumor necrosis factor-α(TNF-α),inducible nitric oxide synthase(iNOS)and Arginase1(Arg1)genes by qRT-PCR(MH-S cells with 0 h cold stimulation as control),detected for the expression of iNOS and Arg1 by immunofluorescence assay(MH-S cells cultured at 37 ℃ for 0. 5 h as negative control)and detected for the expression levels of iNOS,TNF-α and nuclear factor-kappa B(NF-κB)by Western blot(MH-S cells cultured at 37 ℃ for 0. 5 h as negative control). Results The mRNA transcription levels of IL-1β and IL-10 genes in MH-S cells were the highest when the cells were cultured at 34 ℃ for 0. 5 h,therefore,the cold stimulation model of MH-S cells was established under this condition. Compared with the cells cultured for 0 h,the mRNA transcription levels of iNOS,TNF-α and Arg1genes in MH-S cells cultured at 34 ℃ for 0. 5 h increased significantly(t = 3. 733,12. 190 and 6. 793,respectively,each P < 0. 05). Compared with the negative control group,the fluorescence expression intensity of iNOS and Arg1 in MH-S cells in the stimulation group increased,especially iNOS,the expression levels of iNOS and TNF-α proteins increased with no significant difference(t = 0. 675 and 1. 514,respectively,each P > 0. 05),and the expression level of NF-κB increased significantly(t = 3. 092,P < 0. 05). Conclusion Cold stimulation at 34 ℃ for 0. 5 h can increase the expression of inflammatory factors such as IL-1β,IL-10,TNF-α,iNOS,Agr1 and NF-κB in MH-S cells,activate NF-κB signaling pathway in MH-S cells,induce the expression of inflammatory proteins and promote cell activation.