Humans ; Arthritis, Psoriatic/drug therapy* ; Interleukin-17 ; Interleukin Inhibitors ; Antirheumatic Agents/therapeutic use* ; Tumor Necrosis Factor-alpha/therapeutic use* ; Interleukin-23/therapeutic use*

OBJECTIVE: To observe the effect of electroacupuncture (EA) on vascular endothelial growth factor-C (VEGF-C), vascular endothelial growth factor receptor-3 (VEGFR-3), proinflammatory factors and apoptosis in myocardial tissue in mice with acute myocardial ischemia (AMI), and to explore the mechanism of EA for AMI. METHODS: Fifty male C57BL/6 mice were randomly divided into a sham operation group, a model group, an EA group, an inhibitor group and an inhibitor+EA group, 10 mice in each group. Except for the sham operation group, the mice in the remaining groups were intervented with ligation at the left anterior descending (LAD) coronary artery to establish AMI model. The mice in the sham operation group were intervented without ligation after thoracotomy. The mice in the EA group were intervented with EA at "Shenmen" (HT 7) and "Tongli" (HT 5), disperse-dense wave, 2 Hz/15 Hz in frequency, 1 mA in current intensity, 30 min each time, once a day, for 3 d. The mice in the inhibitor group were treated with intraperitoneal injection of SAR 131675 (12.5 mg•kg^-1•d^-1, once a day for 3 d). The mice in the inhibitor+EA group were injected intraperitoneally with SAR 131675 30 min before EA. The ECG before modeling, 30 min after modeling and 3 d after intervention was detected, and the ST segment displacement was recorded; after the intervention, the ELISA method was applied to measure the contents of serum creatine kinase isoenzyme (CK-MB), aspartate aminotransferase (AST) as well as tumor necrosis factor-α (TNF-α) and interleukin-23 (IL-23) in myocardial tissue; the HE staining method was used to observe the morphological changes of myocardial tissue; the immunofluorescence double labeling method was applied to measure the number of co-expression positive cells of VEGF-C/VEGFR-3 in myocardial tissue; the TUNEL method was used to detect the level of cardiomyocyte apoptosis; the Western blot method was applied to measure the protein expressions of VEGF-C, VEGFR-3, b-lymphoma-2 (Bcl-2), activated caspase-3 (Cleaved Caspase-3) and activated poly adenosine diphosphate ribose polymerase-1 (Cleaved PARP-1). RESULTS: Compared with the sham operation group, in the model group the ST segment displacement was increased (P<0.01); the contents of CK-MB, AST, TNF-α and IL-23 were increased (P<0.01); the arrangement of myocardial fibers was disordered, and interstitial inflammatory cell infiltration was obvious; the number of co-expression positive cells of VEGF-C/VEGFR-3 was decreased (P<0.01); the number of cardiomyocyte apoptosis was increased (P<0.01); the expressions of VEGF-C, VEGFR-3 and Bcl-2 were decreased (P<0.01); the expressions of Cleaved Caspase-3 and Cleaved PARP-1 were increased (P<0.01). Compared with the model group, in the EA group the ST segment displacement was decreased (P<0.01); the contents of CK-MB, AST, TNF-α, IL-23 were decreased (P<0.01); the severity of myocardial pathological injury was reduced; the number of co-expression positive cells of VEGF-C/VEGFR-3 was increased (P<0.01); the number of cardiomyocyte apoptosis was reduced (P<0.01); the expressions of VEGF-C, VEGFR-3 and Bcl-2 were increased (P<0.01); the expressions of Cleaved Caspase-3 and Cleaved PARP-1 were reduced (P<0.01). There was no significant difference in all the indexes between the model group and the inhibitor group (P>0.05). Compared with the model group, the protein expression of VEGF-C was increased in the inhibitor+EA group (P<0.01). Compared with the inhibitor group, in the EA group the ST segment displacement was decreased (P<0.01); the contents of CK-MB, AST, TNF-α, IL-23 were decreased (P<0.01); the severity of myocardial pathological injury was reduced; the number of co-expression positive cells of VEGF-C/VEGFR-3 was increased (P<0.05); the number of cardiomyocyte apoptosis was reduced (P<0.01); the expressions of VEGF-C, VEGFR-3 and Bcl-2 were increased (P<0.01); the expressions of Cleaved Caspase-3 and Cleaved PARP-1 were reduced (P<0.01). Compared with the inhibitor+EA group, all the indexes in the EA group were improved except the protein expression of VEGF-C (P<0.01). CONCLUSION EA could relieve the inflammatory reaction and apoptosis in AMI mice, and its mechanism may be related to activating VEGF-C/VEGFR-3 pathway and promoting lymphangion genesis.
Mice ; Male ; Animals ; Electroacupuncture ; Vascular Endothelial Growth Factor Receptor-3 ; Caspase 3 ; Vascular Endothelial Growth Factor C ; Tumor Necrosis Factor-alpha/genetics* ; Vascular Endothelial Growth Factor A/genetics* ; Poly(ADP-ribose) Polymerase Inhibitors ; Mice, Inbred C57BL ; Myocardial Ischemia/metabolism* ; Apoptosis ; Interleukin-23 ; Proto-Oncogene Proteins c-bcl-2

The mechanisms underlying exercise-induced neuroprotective effects after traumatic brain injury (TBI) remained elusive, and there is a lack of effective treatments for TBI. In this study, we investigated the effects of an integrative approach of exercise and Yisaipu (TNFR-IgG fusion protein, TNF inhibitor) in a mouse TBI model. Male C57BL/6J mice were randomly assigned to a sedentary group or a group that followed a voluntary exercise regimen. The effects of 6-week prophylactic preconditioning exercise (PE) alone or in combination with post-TBI Yisaipu treatment on moderate TBI associated deficits were examined. The results showed that combined treatments of PE and post-TBI Yisaipu were superior to single treatments on reducing sensorimotor and gait dysfunctions in mice. These functional improvements were accompanied by reduced systemic inflammation largely via decreased serum TNF-α, boosted autophagic flux, and mitigated lesion volume after TBI. Given these neuroprotective effects, composite approaches such as a combination of exercise and TNF inhibitor may be a promising strategy for facilitating functional recovery from TBI and are worth further investigation.
Animals ; Brain Injuries, Traumatic/pathology* ; Disease Models, Animal ; Male ; Mice ; Mice, Inbred C57BL ; Neuroprotective Agents/pharmacology* ; Recovery of Function ; Tumor Necrosis Factor Inhibitors

OBJECTIVE: To investigate the effects of autophagy on osteogenic differentiation of stem cells from the apical papilla (SCAPs) in the presence of tumor necrosis factor- (TNF-) stimulation . METHODS: SCAPs treated with TNF- (0, 5, and 10 ng/mL) with or without 5 mmol/L 3-MA were examined for the expression of autophagy marker LC3-Ⅱ using Western blotting. The cells were transfected with GFP-LC3 plasmid and fluorescence microscopy was used for quantitative analysis of intracellular GFP-LC3; AO staining was used to detect the acidic vesicles in the cells. The cell viability was assessed with CCK-8 assays and the cell apoptosis rate was analyzed using flow cytometry. The cells treated with TNF- or with TNF- and 3-MA were cultured in osteogenic differentiation medium for 3 to 14 days, and real- time PCR was used to detect the mRNA expressions of osteogenesis-related genes (ALP, BSP, and OCN) for evaluating the cell differentiation. RESULTS: TNF- induced activation of autophagy in cultured SCAPs. Pharmacological inhibition of TNF--induced autophagy by 3-MA significantly decreased the cell viability and increased the apoptosis rate of SCAPs ( < 0.05). Compared with the cells treated with TNF- alone, the cells treated with both TNF- and 3-MA exhibited decreased expressions of the ALP and BSP mRNA on days 3, 7 and 14 during osteogenic induction ( < 0.05) and decreased expression of OCN mRNA on days 3 and 7 during the induction ( < 0.05). CONCLUSIONS Autophagy may play an important role during the osteogenic differentiation of SCAPs in the presence of TNF- stimulation.
Autophagy ; drug effects ; physiology ; Cell Differentiation ; drug effects ; physiology ; Cell Survival ; drug effects ; Cells, Cultured ; Dental Papilla ; cytology ; Green Fluorescent Proteins ; Humans ; Osteogenesis ; physiology ; Stem Cells ; drug effects ; physiology ; Transfection ; Tumor Necrosis Factor-alpha ; administration & dosage ; antagonists & inhibitors ; pharmacology

Psoriasis is a chronic inflammatory disease. Medical therapy is the mainstay of the management of psoriasis, and the main target of psoriasis treatment is immunological dysregulation. Cyclosporine and methotrexate, the main conventional psoriasis treatments, usually lead to a Psoriasis Area and Severity Index (PASI) 75 response in 50% to 60% of patients, but show some organ toxicity. Biologics for psoriasis have recently become the main therapeutic agents for moderate to severe psoriasis unresponsive to conventional treatment. Tumor necrosis factor-α inhibitors were the first anti-psoriatic biologics to be developed, and also show good efficacy for psoriatic arthritis. Ustekinumab, the sole biologic designed for the inhibition of interleukin (IL)-12/23, has been most widely used for psoriasis in Korea. The main strength of ustekinumab is its relatively long treatment interval. IL-17 inhibitors have recently been introduced in Korea for psoriasis treatment. Secukinumab and ixekizumab are currently available IL-17 inhibitors that block the development of psoriasis lesions in the downstream events of psoriasis pathogenesis. They have excellent therapeutic efficacy, with a PASI 90 response in up to 60%–70% of patients. Selective IL-23 inhibitors have been more recently introduced in our country. They have an excellent PASI 90 response, and a longer injection interval than IL-17 inhibitors. New immunological modulators such as phosphodiesterase inhibitors, tyrosine kinase 2 inhibitors, and janus kinase inhibitors are planned to be introduced for psoriasis treatment. These are small molecules that can be administered orally, and some patients who are reluctant to receive injection therapy are expected to favor these therapeutic agents.
Arthritis, Psoriatic ; Biological Products ; Cyclosporine ; Humans ; Interleukin-17 ; Interleukin-23 ; Interleukins ; Korea ; Methotrexate ; Necrosis ; Phosphodiesterase Inhibitors ; Phosphotransferases ; Psoriasis ; Severity of Illness Index ; Tumor Necrosis Factor-alpha ; TYK2 Kinase ; Ustekinumab

OBJECTIVE: To investigate the role of cathepsin B in hepatic Kupffer cells (KCs) in activating Toll-like receptor 4(TLR- 4)-independent inflammatory pathways in mice with lipopolysaccharide (LPS)-induced sepsis. METHODS: Eighteen wild-type (WT) mice and 18 TLR4-knockout (TLR4) mice were both divided into 3 groups for intraperitoneal injections of a lethal dose (54 mg/kg) of LPS, LPS and CA-074(a cathepsin B inhibitor), or normal saline, and the survival of the mice were observed. Another 36 WT mice and 36 TLR4mice were also divided into 3 groups and subjected to intraperitoneal injections of normal saline, 20 mg/kg LPS, or LPS with CA-074 pretreatment.After the treatments, KCs were collected from the mice for assessing the protein level and activity of cathepsin B.The histopathological changes of the liver were observed with HE staining, and the serum levels of IL-1α, IL-1β, TNF-α and IL-18 were detected. RESULTS: Compared with the WT mice,TLR4mice receiving the lethal dose of LPS had significantly longer survival time (up to 84 h) after the injection,but were still unable to fully resist LPS challenge.CA-074 pretreatment prolonged the survival time of WT mice and TLR4mice to 60 h and 132 h,respectively.In the mouse models of sepsis,20 mg/kg LPS induced significantly enhanced activity of cathepsin B without affecting its expression level in the KCs (<0.05) and increased the serum levels of the inflammatory cytokines.CA-074 pretreatment of the mice obviously lessened the detrimental effects of LPS in TLR4mice by significantly lowering cathepsin B activity in the KCs,alleviating hepatocyte apoptosis and reducing the serum levels of inflammatory cytokines. CONCLUSIONS Cathepsin B plays an important role in activating TLR4-independent inflammatory pathways in mice with LPS-induced sepsis.
Animals ; Cathepsin B ; antagonists & inhibitors ; physiology ; Dipeptides ; pharmacology ; Gene Knockout Techniques ; Hepatocytes ; Inflammation ; metabolism ; Interleukin-18 ; blood ; Interleukin-1alpha ; blood ; Interleukin-1beta ; blood ; Kupffer Cells ; metabolism ; Lipopolysaccharides ; Liver ; pathology ; Mice ; Sepsis ; etiology ; metabolism ; Toll-Like Receptor 4 ; genetics ; Tumor Necrosis Factor-alpha ; blood

Emerging evidence indicates that aldosterone and mineralocorticoid receptors (MRs) are associated with the pathogenesis of erectile dysfunction. However, the molecular mechanisms remain largely unknown. In this study, freshly isolated penile corpus cavernosum tissue from rats was treated with aldosterone, with or without MRs inhibitors. Nuclear factor (NF)-kappa B (NF-κB) activity was evaluated by real-time quantitative PCR, luciferase assay, and immunoblot. The results demonstrated that mRNA levels of the NF-κB target genes, including inhibitor of NF-κB alpha (IκB-α), NF-κB1, tumor necrosis factor-alpha (TNF-α), and interleukin 6 (IL-6), were higher after aldosterone treatment. Accordingly, phosphorylation of p65/RelA, IκB-α, and inhibitor of NF-κB kinase-β was markedly increased by aldosterone. Furthermore, knockdown of MRs prevented activation of the NF-κB canonical pathway by aldosterone. Consistent with this finding, ectopic overexpression of MRs enhanced the transcriptional activation of NF-κB by aldosterone. More importantly, the MRs antagonist, spironolactone blocked aldosterone-mediated activation of the canonical NF-κB pathway. In conclusion, aldosterone has an inflammatory effect in the corpus cavernosum penis, inducing NF-κB activation via an MRs-dependent pathway, which may be prevented by selective MRs antagonists. These data reveal the possible role of aldosterone in erectile dysfunction as well as its potential as a novel pharmacologic target for treatment.
Aldosterone/pharmacology* ; Animals ; Cytokines/biosynthesis* ; Gene Knockdown Techniques ; I-kappa B Kinase/antagonists & inhibitors* ; Interleukin-6/genetics* ; Male ; Mineralocorticoid Receptor Antagonists/pharmacology* ; NF-kappa B/genetics* ; Penis/metabolism* ; Protein Serine-Threonine Kinases/antagonists & inhibitors* ; RNA, Messenger/biosynthesis* ; Rats ; Rats, Inbred WKY ; Receptors, Mineralocorticoid/genetics* ; Signal Transduction/drug effects* ; Spironolactone/pharmacology* ; Transcriptional Activation ; Tumor Necrosis Factor-alpha/biosynthesis* ; NF-kappaB-Inducing Kinase

BACKGROUNDIn addition to neurons, all components of the neurovascular unit (NVU), such as glial, endothelial, and basal membranes, are destroyed during traumatic brain injury (TBI). Previous studies have shown that excessive stimulation of calpain is crucial for cerebral injury after traumatic insult. The objective of this study was to investigate whether calpain activation participated in NVU disruption and edema formation in a mouse model of controlled cortical impact (CCI).

METHODSOne hundred and eight mice were divided into three groups: the sham group, the control group, and the MDL28170 group. MDL28170 (20 mg/kg), an efficient calpain inhibitor, was administered intraperitoneally at 5 min, 3 h, and 6 h after experimental CCI. We then measured neurobehavioral deficits, calpain activity, inflammatory mediator levels, blood-brain barrier (BBB) disruption, and NVU deficits using electron microscopy and histopathological analysis at 6 h and 24 h after CCI.

RESULTSThe MDL28170 treatment significantly reduced the extent of both cerebral contusion (MDL28170 vs. vehicle group, 16.90 ± 1.01 mm΃ and 17.20 ± 1.17 mm΃ vs. 9.30 ± 1.05 mm΃ and 9.90 ± 1.17 mm΃, both P < 0.001) and edema (MDL28170 vs. vehicle group, 80.76 ± 1.25% and 82.00 ± 1.84% vs. 82.55 ± 1.32% and 83.64 ± 1.25%, both P < 0.05), improved neurological scores (MDL28170 vs. vehicle group, 7.50 ± 0.45 and 6.33 ± 0.38 vs. 12.33 ± 0.48 and 11.67 ± 0.48, both P < 0.001), and attenuated NVU damage resulting (including tight junction (TJ), basement membrane, BBB, and neuron) from CCI at 6 h and 24 h. Moreover, MDL28170 markedly downregulated nuclear factor-κB-related inflammation (tumor necrosis factor-α [TNF-α]: MDL28170 vs. vehicle group, 1.15 ± 0.07 and 1.62 ± 0.08 vs. 1.59 ± 0.10 and 2.18 ± 0.10, both P < 0.001; inducible nitric oxide synthase: MDL28170 vs. vehicle group, 4.51 ± 0.23 vs. 6.23 ± 0.12, P < 0.001 at 24 h; intracellular adhesion molecule-1: MDL28170 vs. vehicle group, 1.45 ± 0.13 vs. 1.70 ± 0.12, P < 0.01 at 24 h) and lessened both myeloperoxidase activity (MDL28170 vs. vehicle group, 0.016 ± 0.001 and 0.016 ± 0.001 vs. 0.024 ± 0.001 and 0.023 ± 0.001, P < 0.001 and 0.01, respectively) and matrix metalloproteinase-9 (MMP-9) levels (MDL28170 vs. vehicle group, 0.87 ± 0.13 and 1.10 ± 0.10 vs. 1.17 ± 0.13 and 1.25 ± 0.12, P < 0.001 and 0.05, respectively) at 6 h and 24 h after CCI.

CONCLUSIONSThese findings demonstrate that MDL28170 can protect the structure of the NVU by inhibiting the inflammatory cascade, reducing the expression of MMP-9, and supporting the integrity of TJ during acute TBI.

Animals ; Brain Injuries, Traumatic ; drug therapy ; metabolism ; Calpain ; antagonists & inhibitors ; metabolism ; Dipeptides ; therapeutic use ; Disease Models, Animal ; Glycoproteins ; therapeutic use ; Inflammation ; drug therapy ; metabolism ; Male ; Matrix Metalloproteinase 9 ; metabolism ; Mice ; Mice, Inbred BALB C ; NF-kappa B ; metabolism ; Peroxidase ; antagonists & inhibitors ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo explore the therapeutic effect of enhancer of Zeste homolog 2 (EZH2) inhibitor GSK343 on periodontitis by regulating microphage differentiation.

METHODSMacrophage RAW264.7 cells were divided into the blank (A group), control (B group), lipopolysaccharide (LPS) stimulation (C group), and LPS+GSK343 (D group) groups. Phenotype transformations was determined through Western blot analysis and enzyme-linked immunosorbent assay by detecting the differentiation of phenotypic biological markers, including tumor necrosis factor-α (TNF-α), inducible nitric oxide synthase (iNOS), interleukin-10 (IL-10), and Arginase-1 (Arg-1). Metergasis was identified by performing a phagocytosis test on Escherichia coli (E. coli).

RESULTSMacrophage RAW264.7 cells produced classical phenotypic biomarkers (M1) TNF-α and iNOS under LPS stimulation. The expression levels of IL-10 and Arg-1 increased after adding GSK343 into the culture medium. GSK343 also induced the conversion of M1 macrophages into M2 macrophages. Macrophage RAW264.7 cells exerted a phagocytic effect on E. coli, and this effect was enhanced after adding LPS into the culture medium. GSK343 regulated the macrophage RAW264.7 phagocytosis of E. coli.

CONCLUSIONSGSK343 possibly participates in the regulation of macrophage differentiation and, consequently, in the latent treatment of periodontitis.

Arginase ; Cell Differentiation ; Enhancer of Zeste Homolog 2 Protein ; Enzyme Inhibitors ; pharmacology ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; Indazoles ; pharmacology ; Interleukin-10 ; Lipopolysaccharides ; Macrophages ; Nitric Oxide Synthase Type II ; Periodontitis ; Phagocytosis ; Pyridones ; pharmacology ; Tumor Necrosis Factor-alpha

Lipopolysaccharides (LPS) contamination in herbal crude polysaccharides is inevitable. The present study was performed to explore the effect of polymyxin B on abolishing the influence of LPS contamination in mononuclear cells. LPS was pretreated with polymyxin B sulfate (PB) at different concentrations for 1, 5 or 24 h, and then used to stimulate RAW264.7 and mouse peritoneal macrophages (MPMs). The nitric oxide (NO) and tumor necrosis factor-α (TNF-α) in cell culture supernatant, as the indications of cell response, were assayed. Bupleurum chinensis polysaccharides (BCPs) with trace amount contamination of LPS was treated with PB. 30 μg·mL of PB, treating LPS (10 and 1 000 ng·mL in stimulating RAW264.7 and MPMs respectively) at 37 °C for 24 h, successfully abolished the stimulating effect of LPS on the cells. When the cells were stimulated with LPS, BCPs further promoted NO production. However, pretreated with PB, BCPs showed a suppression of NO production in MPMs and no change in RAW264.7. In the in vitro experiments, LPS contamination in polysaccharide might bring a great interference in assessing the activity of drug. Pretreatment with PB (30 μg·mL) at 37 °C for 24 h was sufficient to abolish the effects of LPS contamination (10 and 1 000 ng·mL).
Animals ; Bupleurum ; chemistry ; Drug Contamination ; Drugs, Chinese Herbal ; analysis ; pharmacology ; Lipopolysaccharides ; analysis ; antagonists & inhibitors ; Macrophages ; drug effects ; metabolism ; Mice ; Nitric Oxide ; metabolism ; Polymyxin B ; analysis ; pharmacology ; Polysaccharides ; analysis ; pharmacology ; Tumor Necrosis Factor-alpha ; metabolism