Possible pharmacological targets and mechanisms of sivelestat in protecting acute lung injury.

Acute lung injury/acute respiratory distress syndrome (ALI/ARDS) is a life-threatening syndrome induced by various diseases, including COVID-19. In the progression of ALI/ARDS, activated neutrophils play a central role by releasing various inflammatory mediators, including elastase. Sivelestat is a selective and competitive inhibitor of neutrophil elastase. Although its protective effects on attenuating ALI/ARDS have been confirmed in several models of lung injury, clinical trials have presented inconsistent results on its therapeutic efficacy. Therefore, in this report, we used a network pharmacology approach coupled with animal experimental validation to unravel the concrete therapeutic targets and biological mechanisms of sivelestat in treating ALI/ARDS. In bioinformatic analyses, we found 118 targets of sivelestat against ALI/ARDS, and identified six hub genes essential for sivelestat treatment of ALI/ARDS, namely ERBB2, GRB2, PTK2, PTPN11, ESR1, and CCND1. We also found that sivelestat targeted several genes expressed in human lung microvascular endothelial cells after lipopolysaccharide (LPS) treatment at 4 h (ICAM-1, PTGS2, RND1, BCL2A1, TNF, CA2, and ADORA2A), 8 h (ICAM-1, PTGS2, RND1, BCL2A1, MMP1, BDKRB1 and SLC40A1), and 24 h (ICAM-1). Further animal experiments showed that sivelestat was able to attenuate LPS-induced ALI by inhibiting the overexpression of ICAM-1, VCAM-1, and PTGS2 and increasing the phosphorylation of PTK2. Taken together, the bioinformatic findings and experimentative data indicate that the therapeutic effects of sivelestat against ALI/ARDS mainly focus on the early stage of ALI/ARDS by pharmacological modulation of inflammatory reaction, vascular endothelial injury, and cell apoptosis-related molecules.

Ginsenoside Compound K Reduces Psoriasis-related Inflammation by Activation of the Glucocorticoid Receptor in Keratinocytes.

AIM: To investigate the effects and mechanism of Ginsenoside Compound K (GCK) on psoriasis, focusing on the glucocorticoid receptor (GR) in keratinocytes. METHODS: An imiquimod (IMQ)-induced psoriasis-like dermatitis mouse model was generated to evaluate the anti-inflammatory effect of GCK. Hematoxylin and eosin (H&E) staining was used to assess skin pathological changes. Protein expression of K17 and p-p65 in mice skin was assayed by immunohistochemical. Protein expression and phosphorylation of p65 IκB were assayed by Western blot. Protein expression of K1, K6, K10, K16, K17, and GR were assayed by Western blot and immunofluorescence. Enzyme-linked immunosorbent assay (ELISA) was used to determine cytokine levels of TNF-α, IL-6, CXCL-8, and ICAM-1. Real-time polymerase chain reaction (RT-PCR) was used to quantify TNF-α, IL-6, IL-8, and ICAM-1 mRNA expression. Cell viability was determined by Cell Counting Kit-8(CCK-8) assay. A high-content cell-imaging system was used to assay cell proliferation. Nuclear translocation of p65 and GR was assayed by imaging flow cytometry and immunofluorescence microscopy. Small interfering RNA was used to confirm the role of GR in the anti-inflammatory and immunoregulatory effect of GCK in normal human epidermal keratinecytes (NHEKs). RESULTS: GCK reduced the psoriasis area, severity index, and epidermal thickening in IMQ-induced mice. GCK significantly attenuated the mRNA levels of IL-6, IL-8, TNF-α, and ICAM-1 and reduced epidermal hyperproliferation in the skin of IMQ-induced mice. GCK inhibited in vitro activation of NF-κB, leading to attenuated release of inflammatory mediators (IL-6, IL-8, TNF-α, and ICAM-1) and suppression of NHEK hyperproliferation and abnormal differentiation. These inhibitory effects of GCK were diminished by GR silencing in NHEKs. CONCLUSION: GCK suppressed psoriasis-related inflammation by suppressing keratinocyte activation, which may be related to promoting GR nuclear translocation and inhibiting NF-κB activation. In summary, GCK appears to be a GR activator and a promising therapeutic candidate for antipsoriatic agents.

Phospholipase C-γ as a Potential Therapeutic Target for Graves' Orbitopathy.

BACKGRUOUND: Phospholipase C-γ (PLC-γ) plays a crucial role in immune responses and is related to the pathogenesis of various inflammatory disorders. In this study, we investigated the role of PLC-γ and the therapeutic effect of the PLC-specific inhibitor U73122 using orbital fibroblasts from patients with Graves' orbitopathy (GO). METHODS: The expression of phospholipase C gamma 1 (PLCG1) and phospholipase C gamma 2 (PLCG2) was evaluated using polymerase chain reaction in GO and normal orbital tissues/fibroblasts. The primary cultures of orbital fibroblasts were treated with non-toxic concentrations of U73122 with or without interleukin (IL)-1ß to determine its therapeutic efficacy. The proinflammatory cytokine levels and activation of downstream signaling molecules were determined using Western blotting. RESULTS: PLCG1 and PLCG2 mRNA expression was significantly higher in GO orbital tissues than in controls (P<0.05). PLCG1 and PLCG2 mRNA expression was significantly increased (P<0.05) in IL-1ß, tumor necrosis factor-α, and a cluster of differentiation 40 ligand-stimulated GO fibroblasts. U73122 significantly inhibited the IL-1ß-induced expression of proinflammatory molecules, including IL-6, IL-8, monocyte chemoattractant protein-1, cyclooxygenase-2, and intercellular adhesion molecule-1 (ICAM-1), and phosphorylated protein kinase B (p-Akt) and p38 (p-p38) kinase in GO fibroblasts, whereas it inhibited IL-6, IL-8, and ICAM-1, and p-Akt and c-Jun N-terminal kinase (p-JNK) in normal fibroblasts (P<0.05). CONCLUSION: PLC-γ-inhibiting U73122 suppressed the production of proinflammatory cytokines and the phosphorylation of Akt and p38 kinase in GO fibroblasts. This study indicates the implications of PLC-γ in GO pathogenesis and its potential as a therapeutic target for GO.

Low-moderate dose whole-brain γ-ray irradiation modulates the expressions of glial fibrillary acidic protein and intercellular adhesion molecule-1 in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinson's disease mouse model.

The anti-inflammatory efficacy of radiation therapy (RT) with single fractions below 1.0 Gy has been demonstrated in Alzheimer's disease mouse models. As neuroinflammation is also a major pathological feature of Parkinson's disease (PD), RT may also be effective in PD treatment. Therefore, this study aimed to investigate the anti-inflammatory effect of low-moderate dose RT (LMDRT, 0.6 Gy/single dose, for 5 days) exposure in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP; 30 mg/kg, intraperitoneally, for 5 consecutive days)-induced PD mouse model. Importantly, LMDRT reduced the levels of glial fibrillary acidic protein and intercellular adhesion molecule-1 (CD54) in the striatum region, which increased following MPTP administration. LMDRT also modulated inflammatory gene expression patterns in the substantia nigra region of the MPTP-treated mice. However, LMDRT had no direct effects on the severe loss of dopaminergic neurons and impaired motor behavior in the rotarod test. These results indicate that LMDRT has anti-inflammatory effects by modulating neuroinflammatory factors, including glial fibrillary acidic protein and intercellular adhesion molecule-1, but showed no behavioral improvements or neuroprotection in the MPTP-induced mouse model of PD.

Ultrasound-guided Shoulder Intraarticular Ozone Injection Versus Pulsed Radiofrequency Application for Shoulder Adhesive Capsulitis: A Randomized Controlled Trial.

BACKGROUND: A diseased shoulder due to pain, stiffness, or weakness negatively affects patients' quality of life and their ability to carry out activities of daily living. Adhesive capsulitis is a disease characterized by shoulder pain and global limitation of movement in the shoulder joint. Many interventions have been proposed for the treatment of primary adhesive capsulitis. The current study compares the effect of ultrasound-guided intraarticular injection of ozone versus steroid versus intraarticular application of pulsed radiofrequency. OBJECTIVES: The primary outcome of the current study was to compare the improvement in the Visual Analog Scale (VAS) after the 3 treatment modalities. The secondary outcomes included functional improvement measured by the Shoulder Pain and Disability Index (SPADI) and level of inflammatory biomarkers measured by serum intercellular adhesion molecule (ICAM-1) and high-sensitivity C-reactive protein(hs-CRP). STUDY DESIGN: The current study is a prospective, double blinded, randomized controlled trial. We employed a double blinding technique for both the patients and the outcome assessors. SETTING: Our study was carried out at the Medical Research Institute, Alexandria University, Egypt, after approval of the local ethical committee (IORG0008812). The study was registered in the "clinical trials library for protocol registration and results system" with number NCT04724317.The study included 45 patients with a diagnosis of primary adhesive capsulitis. METHODS: Patients were randomly assigned to 3 equal groups: steroid group, ozone group, and pulsed radiofrequency group. Pain and global shoulder functions were assessed using the VAS at rest and with movement, range of motion (ROM), and the SPADI. Moreover, ICAM-1 and hs-CRP were measured as inflammatory markers. RESULTS: The results of the current study reveal that all patients in all groups have had a statistically significant improvement after their intervention regarding pain, disability, ROM, and inflammatory markers. Pairwise comparisons revealed that improvement of the VAS during movement had a statistically significant improvement starting from the second week and continuing to the fourth and eighth week. VAS during rest had a significant improvement starting from follow-up week one in the steroid group. Moreover, improvement in the ROM and SPADI scores started from the second week follow-up. Percent improvement was calculated for each group and there was a statistically significant difference between groups in VAS at rest and ROM in the pulsed radiofrequency group compared to the steroid group.Regarding inflammatory markers, both ICAM-1 and hs-CRP had a significant improvement after all 3 interventions with no statistically significant difference among the groups. LIMITATIONS: This study is a single-center study. A shortage of previously published data, and heterogeneity in the published methodology of the 3 interventions limited our discussion data for comparison with the previous literature. CONCLUSION: Ultrasound-guided shoulder joint intraarticular injection of steroid, ozone, or pulsed radiofrequency application all result in a significant improvement in pain, disability, and ROM in primary adhesive capsulitis. They can be used as an effective treatment modality for this condition. Comparing groups statistically, the pulsed radiofrequency group had a more delayed, but statistically better long-term improvement compared to the other 2 groups.

Association Between Soluble Intercellular Adhesion Molecule-1 and Intracerebral Hemorrhage Outcomes in the FAST Trial.

BACKGROUND: Neutrophil-mediated inflammation in the acute phase of intracerebral hemorrhage (ICH) worsens outcome in preclinical studies. sICAM-1 (soluble intercellular adhesion molecule-1), an inducible ligand for integrins and cell-cell adhesion molecules, is critical for neutrophil extravasation. We aimed to determine whether serum levels of sICAM-1 are associated with worse outcomes after ICH. METHODS: We conducted a post hoc secondary analysis of an observational cohort using data from the FAST trial (Factor-VII for Acute Hemorrhagic Stroke Treatment). The study exposure was the admission serum level of sICAM-1. The coprimary outcomes were mortality and poor outcome (modified Rankin Scale score 4-6) at 90 days. Secondary radiological outcomes were hematoma expansion at 24 hours and perihematomal edema expansion at 72 hours. We used multiple linear and logistic regression analyses to test for associations between sICAM-1 and outcomes, after adjustment for demographics, ICH severity characteristics, change in the systolic blood pressure in the first 24 hours, treatment randomization arm, and the time from symptom onset to study drug administration. RESULTS: Of 841 patients, we included 507 (60%) with complete data. Hematoma expansion occurred in 169 (33%), while 242 (48%) had a poor outcome. In multivariable analyses, sICAM-1 was associated with mortality (odds ratio, 1.53 per SD increase [95% CI, 1.15-2.03]) and poor outcome (odds ratio, 1.34 per SD increase [CI, 1.06-1.69]). In multivariable analyses of secondary outcomes, sICAM-1 was associated with hematoma expansion (odds ratio, 1.35 per SD increase [CI, 1.11-1.66]), but was not associated with log-transformed perihematomal edema expansion at 72 hours. In additional analyses stratified by treatment assignment, similar results were noted in the recombinant activated factor-VII arm, but not in the placebo arm. CONCLUSIONS: Admission serum levels of sICAM-1 were associated with mortality, poor outcome, and hematoma expansion. Given the possibility of a biological interaction between recombinant activated factor-VII and sICAM-1, these findings highlight the need to further explore the role of sICAM-1 as a potential marker of poor ICH outcomes.

Endothelium-biomarkers for postthrombotic syndrome: a case-control study.

BACKGROUND: The postthrombotic syndrome (PTS) is a long-term complication of deep venous thrombosis (DVT). Increase knowledge on the PTS pathophysiology and novel biomarkers are needed in order to predict PTS development and to improve treatment results. The aim of this study was to analyze novel endothelium-biomarkers for PTS in patients with DVT out of the acute phase. METHODS: A case-control study was conducted. Inclusion criteria were symptomatic and confirmed DVT patients treated with anticoagulants for at least 3 months. Villalta score was performed at the time of inclusion and used to diagnose and classify the severity of PTS. Plasma inter-cellular adhesion molecule 1 (ICAM-1), P-selectin, fractalkine and vascular endothelial growth factor (VEGF) were quantified using cytometric bead array. Endothelial progenitor cells (EPCs) and circulating endothelial cells (CEC) level were quantified by flow cytometry. RESULTS: Thirty two patients and 61 controls were included. PTS patients showed higher levels of CEC (0.56/µl (0.34-1.5) vs. 0.20/µl (0.11-0.77); P  = 0.04) and EPC (0.75/µl (0.38-1.52) vs. 0.09/µl (0.05-0.82); P  = 0.0021) compared to no PTS patients. Patients with PTS had significantly higher levels of fractalkine (387.60 pg/ml (222.30-597.90) vs. 98.00 pg/ml (82.30-193.02); P  = 0.044) than patients without PTS. Fracktalkine levels showed a strong linear correlation with Villalta score, r  = 0.86, P  < 0.0001. No differences were observed in P-selectin, ICAM-1 and VEGF between studied groups. CONCLUSIONS: The formation and early resolution of DVT are characterized by inflammation and endothelial/platelet activation. We have identified possible novel biomarkers such as CEC, EPC and fractalkine for the development of PTS. These results suggest a possible role of these mediators in the maintenance and worsening of PTS turning them into potential therapeutic targets.

Oral contraceptive pill (OCP) treatment alters the gene expression of intercellular adhesion molecule-1 (ICAM-1), tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein-1 (MCP-1) and plasminogen activator inhibitor-1 (PAI-1) in polycystic ovary syndrome (PCOS) women compared to drug-naive PCOS women.

BACKGROUND: Polycystic ovary syndrome (PCOS) presents clinical symptoms of menstrual abnormalities, excessive hair growth (hirsutism), scalp hair loss, acne and infertility. Metabolic abnormalities such as obesity, insulin resistance, glucose intolerance and cardiovascular problems constitute an essential part of PCOS, all of which can have significant long-term health consequences. Low-grade chronic inflammation demonstrated by persistent moderately elevated serum levels of inflammatory and coagulatory markers plays a critical role in the pathogenesis of PCOS. Oral contraceptive pills (OCPs) constitute the mainstay of pharmacologic therapy for women with PCOS to regularize cyclicity and ameliorate androgen excess. On the other hand, OCP use is associated with various venous thromboembolic and proinflammatory events in the general population. PCOS women always carriers the increased lifetime risk of these events. The studies on the effect of OCPs on inflammatory, coagulation and metabolic parameters in PCOS are less robust. Therefore in this study, we investigated and compared the messenger RNA (mRNA) expression profiles of genes implicated in inflammatory and coagulation pathways between drug-naive and OCP-treated PCOS women. The selected genes include intercellular adhesion molecule-1 (ICAM-1), tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein-1 (MCP-1) and plasminogen activator inhibitor-1 (PAI-1). Furthermore, the correlation between the selected markers and various metabolic indices in the OCP group has also been explored. METHOD: The relative amounts of ICAM-1, TNF-α, MCP-1 and PAI-1 mRNA in peripheral blood mononuclear cells from 25 drug-naive PCOS subjects (controls) and 25 PCOS subjects who received OCPs containing 0.03 mg-ethinyl-estradiol and 0.15 mg-levonorgestrel for at least six months (cases) were estimated using real-time qPCR. The statistical interpretation was conducted using SPSS version 20.0 (SPSS, Inc, Chicago, IL), Epi Info version 2002 (Disease Control and Prevention Centres, Atlanta, GA) and GraphPad Prism 5 (GraphPad Software, La Jolla, CA) software. RESULT: Six months of OCP therapy enhanced the expression of inflammatory genes viz ICAM-1, TNF-α and MCP-1 mRNA in PCOS women by 2.54, 2.05 and 1.74 folds, respectively, in this study. However, PAI-1 mRNA in the OCP group showed no significant increase. Furthermore, in cases, ICAM-1 mRNA expression positively correlated with body mass index (BMI) (p = 0.01), fasting insulin (p = 0.01), insulin 2 h p = 0.02), glucose 2 h (p = 0.01) and triglycerides (p = 0.01). TNF-α mRNA expression positively correlated with fasting insulin (p = 0.0007). MCP-1 mRNA expression positively correlated with (BMI) (p = 0.002). CONCLUSION: OCPs helped reduce clinical hyperandrogenism and regularise menstrual cycles in women with PCOS. However, OCP use was associated with increased fold expression of inflammatory markers which positively correlated with metabolic abnormalities.

Guanxinping ameliorates atherosclerosis via MAPK/NF-κB signaling pathway in ApoE-/- mice.

BACKGROUND: Atherosclerosis (AS), one of the leading causes of deaths and disabilities, is a kind of vascular disease of lipid disorders and chronic inflammation. Guanxinping (GXP) has been administrated in the treatment of AS for nearly 20 years with satisfying clinical response. This study aimed to explore its underlying mechanisms of anti-atherosclerotic effect in AS. METHODS: Male ApoE-/- mice were randomized into five groups and fed with either standard diet (control group, CON) or high-fat diet (HFD) for 12 weeks. HFD mice were further divided randomly and either fed continually with HFD as a model group, or atorvastatin (ATO), or low-dose GXP (LGXP), or high-dose GXP (HGXP). After 12 weeks, the body weight, serum triglyceride (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-c), and low-density lipoprotein cholesterol (LDL-c) were detected. Moreover, serum inflammation cytokines including tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and interleukin-1ß (IL-1ß) concentrations were measured. The structure of aortic tissues was examined by hematoxylin-eosin staining. The mRNA expression of TNF-α, IL-6, and IL-1ß were assessed by qPCR. The protein expressions of ICAM-1, VCAM-1, MCP-1, IL-6, IL-1ß, p38MAPK, ERK1/2, JNK, IκB-α, and NF-κBp65 in the aorta were also detected. RESULTS: GXP treatment reduced serum TG, TC, and LDL-c levels in ApoE-/- mice. Moreover, GXP reduced lipid accumulation in the aorta of ApoE-/- mice, induced by HFD. Furthermore, GXP ameliorated the aorta morphological damage and reduced the serum TNF-α, IL-6, and IL-1ß levels. GXP also attenuated the protein expression of ICAM-1, VCAM-1, MCP-1, IL-6, IL-1ß, p38MAPK, ERK1/2, JNK, and NF-κBp65, whereas it increased the IκBα level in aortic tissues of ApoE-/- mice. CONCLUSIONS: Our results show that GXP could ameliorate atherosclerosis, which is mediated by inhibition of the MAPK/NF-κB signaling pathway in ApoE-/- mice. This study provides evidence that GXP might be a promising drug for the treatment of AS.

Triangular-shaped homologous heterostructure as photocatalytic H2S scavenger and macrophage modulator for rheumatoid arthritis therapy.

Rheumatoid arthritis (RA) is a chronic arthropathy causing cartilage destruction, bone erosion, and even disability. Although some advances in RA treatment have been made based on inflammatory cytokine inhibition, long-term treatment and drug effect have been restrained by severe side effects. Herein, we developed a resveratrol (RSV)-loaded Ag/Ag2S triangular-shaped homologous heterostructure with polyethylene glycol/folic acid (PEG/FA) modification (Ag/Ag2S-PEG-FA/RSV NTs) to simultaneously suppress inflammatory cytokine over-expression through photocatalytic H2S scavenging and macrophage polarization stimulation. On one hand, the over-expressed H2S, which acted as a pro-inflammatory mediator to activate the MAPK/ICAM-1 pathway and exacerbate inflammation, was eliminated through photocatalysis. The homologous Ag and Ag2S of the heterostructure enhanced electron separation and transfer by acting as a charge acceptor and electron generator, respectively, which restrained electron/hole recombination and promoted photocatalysis efficiency. Additionally, the intrinsic superoxide dismutase (SOD) and catalase (CAT) activity of Ag decomposed the reactive oxygen species (ROS) over-expressed in the RA microenvironment, which supplied O2 for the photocatalytic H2S scavenging progress. On the other hand, RSV, a natural product with anti-inflammatory activity, could be delivered to the inflammatory joint by the targeting effect of PEG-FA, thus inhibiting the IκB/NF-κB pro-inflammatory pathway to induce macrophage interconversion balance from M1 to M2. As expected, the Ag/Ag2S-PEG-FA/RSV NTs exhibited H2S scavenging capacity and modulated macrophage polarization to reduce the inflammatory cytokine level and halt RA progression in vitro and in vivo. Overall, this study revealed a therapeutic strategy with high efficacy, which opens broad prospects for RA treatment.

VEGF promotes diabetic retinopathy by upregulating the PKC/ET/NF-κB/ICAM-1 signaling pathway.

Diabetic retinopathy (DR) is a common microvascular complication in patients with diabetes mellitus. DR is caused by chronic hyperglycemia and is characterized by progressive loss of vision because of damage to the retinal microvasculature. In this study, we investigated the regulatory role and clinical significance of the vascular endothelial growth factor (VEGF)/protein kinase C (PKC)/endothelin (ET)/nuclear factor-κB (NF-κB)/intercellular adhesion molecule 1 (ICAM-1) signaling pathway in DR using a rat model. Intraperitoneal injections of the VEGF agonist, streptozotocin (STZ) were used to generate the DR model rats. DR rats treated with the VEGF inhibitor (DR+VEGF inhibitor) were used to study the specific effects of VEGF on DR pathology and the underlying mechanisms. DR and DR+VEGF agonist rats were injected with the PKCß2 inhibitor, GF109203X to determine the therapeutic potential of blocking the VEGF/PKC/ET/NF-κB/ICAM-1 signaling pathway. The body weights and blood glucose levels of the rats in all groups were evaluated at 16 weeks. DR-related retinal histopathology was analyzed by hematoxylin and eosin staining. ELISA assay was used to estimate the PKC activity in the retinal tissues. Western blotting and RT-qPCR assays were used to analyze the expression levels of PKC-ß2, VEGF, ETs, NF-κB, and ICAM-1 in the retinal tissues. Immunohistochemistry was used to analyze VEGF and ICAM-1 expression in the rat retinal tissues. Our results showed that VEGF, ICAM-1, PKCß2, ET, and NF-κB expression levels as well as PKC activity were significantly increased in the retinal tissues of the DR and DR+VEGF agonist rat groups compared to the control and DR+VEGF inhibitor rat groups. DR and DR+VEGF agonist rats showed significantly lower body weight and significantly higher retinal histopathology scores and blood glucose levels compared to the control and DR+VEGF inhibitor group rats. However, treatment of DR and DR+VEGF agonist rats with GF109203X partially alleviated DR pathology by inhibiting the VEGF/ PKC/ET/NF-κB/ICAM-1 signaling pathway. In summary, our data demonstrated that inhibition of the VEGF/ PKC/ET/NF-κB/ICAM-1 signaling pathway significantly alleviated DR-related pathology in the rat model. Therefore, VEGF/PKC/ET/NF-κB/ICAM-1 signaling axis is a promising therapeutic target for DR.

Pharmacological Characterization of a Novel Neuroactive Steroid Prodrug, NTS-104, and Its Neuroprotective Activity in Experimental Stroke Models.

BACKGROUND: Ischemic stroke affects about 700 000 patients per year in the United States, and to date, there are no effective pharmacological agents that promote recovery. Here, we studied the pharmacokinetics, pharmacodynamics, and efficacy of NTS-105, a novel neuroactive steroid, and NTS-104, a prodrug of NTS-105, in 2 models of ischemic stroke. METHODS: The pharmacodynamics and pharmacokinetics of NTS-104/105 were investigated in naive and stroke rats, and models of embolic and transient middle cerebral artery occlusion were used to investigate the dose-related effects of NTS-104. All rats were randomly assigned into the experimental groups, and all outcome measurements were performed blindly. RESULTS: Blood plasma and brain pharmacokinetic analysis revealed that NTS-104 rapidly converted to NTS-105, which reached peak concentration at ≈1 hour after dosing and distributed similarly to normal and ischemic brains. NTS-104 administration 4 hours after embolic middle cerebral artery occlusion led to a dose-dependent improvement of neurological outcomes and a dose-dependent reduction of infarct volumes relative to vehicle-treated animals. A single dose level study confirmed that NTS-104 administered 4 hours after transient middle cerebral artery occlusion was also neuroprotective. Quantitative ELISA revealed that NTS-104 treatment resulted in time- and dose-dependent changes in AKT activation and cytokine levels within the ischemic brain, which included reductions of IL-6, VEGF, ICAM-1, IL-1ß, MCP-1, RAGE, and GM-CSF. Time- and dose-dependent reductions in IL-6 and GM-CSF were also observed in the plasma along with an elevation of galectin-1. CONCLUSIONS: NTS-104 is a novel prodrug that converts to a novel neuroactive steroid, NTS-105, which improves functional outcomes in experimental ischemic stroke models.

RNA-seq-based transcriptome analysis of the anti-inflammatory effect of artesunate in the early treatment of the mouse cerebral malaria model.

BACKGROUND: cerebral malaria (CM) is an important complication of malaria with a high mortality rate. Artesunate is recommended as the first-line artemisinin compound treatment for severe malaria. Due to the difficulty of obtaining brain tissue samples clinically, the use of animals to research host responses to CM parasite infections is necessary. Rodent malaria models allow for detailed time series studies of host responses in multiple organs. To date, studies on the transcriptome of severe malaria are only limited to the parasites in the peripheral blood of patients, and there is little data on the transcriptional changes in brain tissue in mice with CM treated with artesunate. METHOD AND RESULT: in this study, fresh tissue samples (three biological replicates per mouse) from the same area of the brain in each animal were collected from the uninfected, Plasmodium berghei ANKA-infected and artesunate-treated C57BL/6 mice, and then transcriptome research was performed by the RNA-seq technique. Differentially expressed genes (DEGs) included Il-21, Tnf, Il-6, Il-1ß, Il-10, Ifng, and Icam-1. Among which, Il-6, Il-10, Tnf-α and Il-1ß were further verified and validated via qRT-PCR and ELISA. This revealed that Il-1ß (p < 0.0001), Il-10 (p < 0.05) and Tnf-α (p < 0.05) were significantly up-regulated in the Pb ANKA-infected versus uninfected group, while Il-1ß (p < 0.0001) and Tnf-α (p < 0.05) were significantly down-regulated after artesunate treatment. All DEGs were closely related to the top 3 artesunate treatment pathways, including the JAK-STAT signaling pathway, apoptosis, and Toll-like receptor signaling pathway. CONCLUSION: the mechanism of improving the prognosis of cerebral malaria by artesunate may not only involve the killing of plasmodium but also the inhibition of a cytokine storm in the host. This study provides new insights into the molecular mechanism by which artesunate improves the prognosis of cerebral malaria.

Lidocaine inhibited migration of NSCLCA549 cells via the CXCR4 regulation.

BACKGROUND: Lidocaine is a local anesthetic that wildly used in surgical treatment and postoperative medical care for lung cancers. We hypothesized that lidocaine at clinical plasma concentration can inhibit CXCL12/CXCR4 axis-regulated cytoskeletal remodeling thereby reduce the migration of Non-small-cell lung cancers (NSCLC) cells. METHODS: We determined the effect of lidocaine at clinical plasma concentration on CXCL12-induced cell viability, apoptosis, cell death, monolayer cell wound healing rate, individual cell migration indicators, expression of CXCR4, CD44, and ICAM-1, intracellular Ca2+ level, and filamentous actin level alteration of NSCLC cells A549 and CXCR4-knocked down A549 cells using CCK-8, Bcl-2 ELISA, Cell death ELISA, wound healing assay, chemotaxis assay, western blotting, QPCR, Fura-2-based intracellular Ca2+ assay, and Fluorescein Phalloidin staining respectively. RESULTS: Lidocaine did not affect cell viability, apoptosis, and cell death but inhibited CXCL12-induced migration, intracellular Ca2+ releasing, and filamentous actin increase. Lidocaine decreased expression of CXCR4, increased CD44, but had no effect on ICAM-1. CXCL12 induced the increase of CD44 and ICAM-1 but did not affect CD44 in the presence of lidocaine. The knockdown of CXCR4 eliminated all the effects of lidocaine. The overexpression of CXCR4 promoted migration but the migration was inhibited by lidocaine. CONCLUSION: Lidocaine at clinical plasma concentrations inhibited CXCL12-induced CXCR4 activation, thereby reduced the intracellular Ca2+-dependent cytoskeleton remodeling, resulting in slower migration of A549 cells.

Effects of sea buckthorn puree on risk factors of cardiovascular disease in hypercholesterolemia population: a double-blind, randomized, placebo-controlled trial.

To investigate the effect of sea buckthorn puree consumption on reducing blood lipid and other risk factors of CVD. A total of 111 patients with hypercholesteromia who were treated with 90 ml sea buckthorn puree or placebo for 90 days were enrolled in this trial. Physical examination and analysis of lipid markers, hsCRP concentrations and cell adhesion protein concentrations with fasting blood samples were performed at 0-day, 45-day and 90-day after treatment. We found that the sea buckthorn puree did not affect the levels of serum total cholesterol (TC), low density lipoprotein-cholesterol (LDL-C) and triglyceride (TG). However, the level of HDL-C was decreased by sea buckthorn puree in a short period (first 6 weeks), which was increased in the last 6 weeks (p < 0.05). Compared with placebo, diastolic blood pressure (DBP) was decreased after taking sea buckthorn puree (p > 0.05). There was moderately decreased in hsCRP concentration in sea buckthorn group. Nevertheless, there was no correlation between changes in ICAM-1 and VCAM-1 concentration. In conclusion, long term consumption of sea buckthorn puree has anti-inflammatory and anti-hypertensive effects on hypercholesterolemia in hypercholesterolemic patients. However, it did not translate into an effect on the concentration of lipid markers in the circulation.

Innate Priming of Neutrophils Potentiates Systemic Multiorgan Injury.

Excessive inflammatory reactions mediated by first-responder cells such as neutrophils contribute to the severity of multiorgan failure associated with systemic injury and infection. Systemic subclinical endotoxemia due to mucosal leakage may aggravate neutrophil activation and tissue injury. However, mechanisms responsible for neutrophil inflammatory polarization are not well understood. In this study, we demonstrate that subclinical low-dose endotoxemia can potently polarize neutrophils into an inflammatory state in vivo and in vitro, as reflected in elevated expression of adhesion molecules such as ICAM-1 and CD29, and reduced expression of suppressor molecule CD244. When subjected to a controlled administration of gut-damaging chemical dextran sulfate sodium, mice conditioned with subclinical dose LPS exhibit significantly elevated infiltration of neutrophils into organs such as liver, colon, and spleen, associated with severe multiorgan damage as measured by biochemical as well as histological assays. Subclinical dose LPS is sufficient to induce potent activation of SRC kinase as well as downstream activation of STAT1/STAT5 in neutrophils, contributing to the inflammatory neutrophil polarization. We also demonstrate that the administration of 4-phenylbutyric acid, an agent known to relieve cell stress and enhance peroxisome function, can reduce the activation of SRC kinase and enhance the expression of suppressor molecule CD244 in neutrophils. We show that i.v. injection of 4-phenylbutyric acid conditioned neutrophils can effectively reduce the severity of multiorgan damage in mice challenged with dextran sulfate sodium. Collectively, our data, to our knowledge, reveal novel inflammatory polarization of neutrophils by subclinical endotoxemia conducive for aggravated multiorgan damage as well as potential therapeutic intervention.

How Carrier Size and Valency Modulate Receptor-Mediated Signaling: Understanding the Link between Binding and Endocytosis of ICAM-1-Targeted Carriers.

Targeting of drug carriers to endocytic cell receptors facilitates intracellular drug delivery. Carrier size and number of targeting moieties (valency) influence cell binding and uptake. However, how these parameters influence receptor-mediated cell signaling (the link between binding and uptake) remains uncharacterized. We studied this using polymer carriers of different sizes and valencies, targeted to endothelial intercellular adhesion molecule-1 (ICAM-1), a marker overexpressed in many pathologies. Unexpectedly, induction of cell signals (ceramide and protein kinase C (PKC) enrichment and activation) and uptake, were independent of carrier avidity, total number of carriers bound per cell, cumulative cell surface area occupied by carriers, number of targeting antibodies at the carrier-cell contact, and cumulative receptor engagement by all bound carriers. Instead, "valency density" (number of antibodies per carrier surface area) ruled signaling, and carrier size independently influenced uptake. These results are key to understanding the interplay between carrier design parameters and receptor-mediated signaling conducive to endocytosis, paramount for intracellular drug delivery.

Ipilimumab: a potential immunologic agent in the treatment of metastatic castration-resistant prostate cancer.

Immunotherapy has garnered an important place in the therapeutic landscape of treatment in prostate cancer since approval of sipuleucel-T. Ipilimumab is a checkpoint inhibitor that is currently approved for the treatment of advanced melanoma. In the June issue of Lancet Oncology, Kwon and colleagues report the phase III trial using ipilimumab in a post-docetaxel metastatic castration-resistant prostate cancer population. While the primary endpoint of overall survival was not met, several lessons are learned from the analysis of this trial. Perhaps better refinement of a more favorable group of patients who may potentially benefit from an immunologic treatment should be advocated.

Sipuleucel-T immunotherapy for castration-resistant prostate cancer.

BACKGROUND: Sipuleucel-T, an autologous active cellular immunotherapy, has shown evidence of efficacy in reducing the risk of death among men with metastatic castration-resistant prostate cancer. METHODS: In this double-blind, placebo-controlled, multicenter phase 3 trial, we randomly assigned 512 patients in a 2:1 ratio to receive either sipuleucel-T (341 patients) or placebo (171 patients) administered intravenously every 2 weeks, for a total of three infusions. The primary end point was overall survival, analyzed by means of a stratified Cox regression model adjusted for baseline levels of serum prostate-specific antigen (PSA) and lactate dehydrogenase. RESULTS: In the sipuleucel-T group, there was a relative reduction of 22% in the risk of death as compared with the placebo group (hazard ratio, 0.78; 95% confidence interval [CI], 0.61 to 0.98; P=0.03). This reduction represented a 4.1-month improvement in median survival (25.8 months in the sipuleucel-T group vs. 21.7 months in the placebo group). The 36-month survival probability was 31.7% in the sipuleucel-T group versus 23.0% in the placebo group. The treatment effect was also observed with the use of an unadjusted Cox model and a log-rank test (hazard ratio, 0.77; 95% CI, 0.61 to 0.97; P=0.02) and after adjustment for use of docetaxel after the study therapy (hazard ratio, 0.78; 95% CI, 0.62 to 0.98; P=0.03). The time to objective disease progression was similar in the two study groups. Immune responses to the immunizing antigen were observed in patients who received sipuleucel-T. Adverse events that were more frequently reported in the sipuleucel-T group than in the placebo group included chills, fever, and headache. CONCLUSIONS: The use of sipuleucel-T prolonged overall survival among men with metastatic castration-resistant prostate cancer. No effect on the time to disease progression was observed. (Funded by Dendreon; ClinicalTrials.gov number, NCT00065442.)