Neonatal administration of synthetic estrogen, diethylstilbestrol to mice up-regulates inflammatory Cxclchemokines located in the 5qE1 region in the vaginal epithelium.

A synthetic estrogen, diethylstilbestrol (DES), is known to cause adult vaginal carcinoma by neonatal administration of DES to mice. However, the carcinogenic process remains unclear. By Cap Analysis of Gene Expression method, we found that neonatal DES exposure up-regulated inflammatory Cxcl chemokines 2, 3, 5, and 7 located in the 5qE1 region in the vaginal epithelium of mice 70 days after birth. When we examined the gene expressions of these genes much earlier stages, we found that neonatal DES exposure increased these Cxcl chemokine genes expression even after 17 days after birth. It implies the DES-mediated persistent activation of inflammatory genes. Intriguingly, we also detected DES-induced non-coding RNAs from a region approximately 100 kb far from the Cxcl5 gene. The non-coding RNA up-regulation by DES exposure was confirmed on the 17-day vagina and continued throughout life, which may responsible for the activation of Cxcl chemokines located in the same region, 5qE1. This study shows that neonatal administration of DES to mice causes long-lasting up-regulation of inflammatory Cxcl chemokines in the vaginal epithelium. DES-mediated inflammation may be associated with the carcinogenic process.

Oxaliplatin facilitates tumor-infiltration of T cells and natural-killer cells for enhanced tumor immunotherapy in lung cancer model.

Platinum is reported to have adjuvant immune properties, whether oxaliplatin (OXA) could be utilized to synergize with anti-programmed cell death-1 (PD-1) antibody or anti-NKG2D (natural-killer group 2, member D) antibody is investigated. Subcutaneous A549 lung cancer and murine Lewis lung carcinoma (LLC) models were constructed, which were further intravenously injected with platinum-based drugs or concomitant administrated with anti-PD-1 antibody and or anti-NKG2D antibody. The tumor volume and the proportion of myeloid cells (CD45+CD11b+), CD3+T cells and NK (NK1.1+) cells were detected. The relative expression of chemokine (C-X-C motif) ligand 9 (CXCL9), CXCL10 and CXCL11 and C-X-C motif chemokine receptor 3 (CXCR3) was detected with the ELISA, western blot and flow cytometry. The three platinum drugs (cisplatin, DDP; carboplatin, CBP; OXA) showed similar effects to inhibit A549 tumor growth in immune-deficient mice. While OXA exhibited better antitumor efficacy in wild-type mice bearing LLC with downregulated myeloid cells proportion, upregulated concentration of CXCL9, CXCL10 and CXCL11, and upregulated proportion and CXCR3 expression on T cells and NK cells. OXA combined with anti-PD1 or anti-NKG2D synergistically improved tumor growth inhibition and survival. The combination of OXA to anti-PD1 and anti-NKG2D antibodies will provide the most appropriate treatment benefit. Oxaliplatin promotes T cells and NK cells infiltration through the CXCL9/10/11-CXCR3 axis to enhance anti-PD1 or anti-NKG2D immunotherapy in lung cancer.

Inhibition of Drug-Induced Liver Injury in Mice Using a Positively Charged Peptide That Binds DNA.

Hepatic cell death occurs in response to diverse stimuli such as chemical and physical damage. The exposure of intracellular contents such as DNA during necrosis induces a severe inflammatory response that has yet to be fully explored therapeutically. Here, we sought means to neutralize the ability of extracellular DNA to induce deleterious tissue inflammation when drug-induced liver injury had already ensued. DNA exposure and inflammation were investigated in vivo in drug-induced liver injury using intravital microscopy. The necrotic DNA debris was studied in murine livers in vivo and in DNA debris models in vitro by using a positively charged chemokine-derived peptide (MIG30; CXCL9[74-103]). Acetaminophen-induced liver necrosis was associated with massive DNA accumulation, production of CXC chemokines, and neutrophil activation inside the injured tissue. The MIG30 peptide bound the healthy liver vasculature and, to a much greater extent, to DNA-rich necrotic tissue. Moreover, MIG30 bound extracellular DNA directly in vivo in a charge-dependent manner and independently of glycosaminoglycans and chemokines. Post-treatment of mice with MIG30 reduced mortality, liver damage, and inflammation significantly. These effects were not observed with a control peptide that does not bind DNA. Mechanistically, MIG30 inhibited the interaction between DNA and histones, and promoted the dissociation of histones from necrotic debris. MIG30 also inhibited the pro-inflammatory effect of CpG DNA, as measured by a reduction in CXCL8 production, indicating that MIG30 disturbs the ability of DNA to induce hepatic inflammation. Conclusion: The use of DNA-binding peptides reduces necrotic liver injury and inflammation, even at late timepoints.

Impact of edaravone on serum CXC chemokine ligand-13 levels and perioperative neurocognitive disorders in elderly patients with hip replacement.

BACKGROUND: Perioperative neurocognitive disorders (PND) are a series of severe complications in the perioperative and anesthetic periods with a decline in memory, execution ability, and information processing speed as the primary clinical manifestation. This study aimed to evaluate the impact of edaravone (EDA) on PND and peripheral blood C-X-C motif chemokine ligand 13 (CXCL13) levels in elderly patients with hip replacement. METHODS: A total of 160 elderly patients undergoing hip arthroplasty in Affiliated Dongguan People's Hospital of Southern Medical University (from March 2016 to March 2018) were randomly and double-blindly categorized into an EDA group and a control group (CON). Group EDA was administered intravenously EDA 30 min before surgery, and group CON was administered intravenously saline. The cognitive function of the two groups was evaluated 1-day before the operation and at 1 and 12 months after surgery, and the incidence of post-operative delirium was tested on days 1, 3, and 7 after surgery using the Chinese version of the confusion assessment method. Serum CXCL13 and interleukin (IL)-6 concentrations were measured before anesthesia, during surgery (30 min after skin incision), and on days 1, 3, and 7 after surgery. The continuous variables in accordance with normal distribution were tested using the Student's t test, the continuous variables without normal distribution using the Mann-Whitney U test, and categorical variables by the χ2 test or Fisher exact test. RESULTS: The incidence of post-operative delirium within 7 days after surgery was significantly higher in group CON than that in group EDA (31.3% vs. 15.0%, t = -5.6, P < 0.001). The modified telephone interview for cognitive status and activities of daily life scores were significantly higher in the group EDA than those in the group CON at 1 month (39.63 ±â€Š4.35 vs. 33.63 ±â€Š5.81, t = -2.13, P < 0.05 and 74.3 ±â€Š12.6 vs. 61.2 ±â€Š13.1, t = -1.69, P < 0.05) and 12 months (40.13 ±â€Š5.93 vs. 34.13 ±â€Š5.36, t = -3.37, P < 0.05 and 79.6 ±â€Š11.7 vs. 65.6 ±â€Š16.6, t = -2.08, P < 0.05) after surgery; and the incidence of neurocognitive dysfunction was significantly lower in the group EDA than that in the group CON (P < 0.05). Serum CXCL13 and IL-6 concentrations were significantly lower in the group EDA than those in the group CON during and after surgery (P < 0.05). CONCLUSION: EDA can significantly reduce the serum concentrations of CXCL13 and IL-6 and improve the PND of patients.

Small molecule Y-320 stimulates ribosome biogenesis, protein synthesis, and aminoglycoside-induced premature termination codon readthrough.

Premature termination codons (PTC) cause over 10% of genetic disease cases. Some aminoglycosides that bind to the ribosome decoding center can induce PTC readthrough and restore low levels of full-length functional proteins. However, concomitant inhibition of protein synthesis limits the extent of PTC readthrough that can be achieved by aminoglycosides like G418. Using a cell-based screen, we identified a small molecule, the phenylpyrazoleanilide Y-320, that potently enhances TP53, DMD, and COL17A1 PTC readthrough by G418. Unexpectedly, Y-320 increased cellular protein levels and protein synthesis, measured by SYPRO Ruby protein staining and puromycin labeling, as well as ribosome biogenesis measured using antibodies to rRNA and ribosomal protein S6. Y-320 did not increase the rate of translation elongation and it exerted its effects independently of mTOR signaling. At the single cell level, exposure to Y-320 and G418 increased ribosome content and protein synthesis which correlated strongly with PTC readthrough. As a single agent, Y-320 did not affect translation fidelity measured using a luciferase reporter gene but it enhanced misincorporation by G418. RNA-seq data showed that Y-320 up-regulated the expression of CXC chemokines CXCL10, CXCL8, CXCL2, CXCL11, CXCL3, CXCL1, and CXCL16. Several of these chemokines exert their cellular effects through the receptor CXCR2 and the CXCR2 antagonist SB225002 reduced cellular protein levels and PTC readthrough in cells exposed to Y-320 and G418. These data show that the self-limiting nature of PTC readthrough by G418 can be compensated by Y-320, a potent enhancer of PTC readthrough that increases ribosome biogenesis and protein synthesis. They also support a model whereby increased PTC readthrough is enabled by increased protein synthesis mediated by an autocrine chemokine signaling pathway. The findings also raise the possibility that inflammatory processes affect cellular propensity to readthrough agents and that immunomodulatory drugs like Y-320 might find application in PTC readthrough therapy.

Impact of edaravone on serum CXC chemokine ligand-13 levels and perioperative neurocognitive disorders in elderly patients with hip replacement / 中华医学杂志(英文版)

BACKGROUND@#Perioperative neurocognitive disorders (PND) are a series of severe complications in the perioperative and anesthetic periods with a decline in memory, execution ability, and information processing speed as the primary clinical manifestation. This study aimed to evaluate the impact of edaravone (EDA) on PND and peripheral blood C-X-C motif chemokine ligand 13 (CXCL13) levels in elderly patients with hip replacement.@*METHODS@#A total of 160 elderly patients undergoing hip arthroplasty in Affiliated Dongguan People's Hospital of Southern Medical University (from March 2016 to March 2018) were randomly and double-blindly categorized into an EDA group and a control group (CON). Group EDA was administered intravenously EDA 30 min before surgery, and group CON was administered intravenously saline. The cognitive function of the two groups was evaluated 1-day before the operation and at 1 and 12 months after surgery, and the incidence of post-operative delirium was tested on days 1, 3, and 7 after surgery using the Chinese version of the confusion assessment method. Serum CXCL13 and interleukin (IL)-6 concentrations were measured before anesthesia, during surgery (30 min after skin incision), and on days 1, 3, and 7 after surgery. The continuous variables in accordance with normal distribution were tested using the Student's t test, the continuous variables without normal distribution using the Mann-Whitney U test, and categorical variables by the χ2 test or Fisher exact test.@*RESULTS@#The incidence of post-operative delirium within 7 days after surgery was significantly higher in group CON than that in group EDA (31.3% vs. 15.0%, t = -5.6, P < 0.001). The modified telephone interview for cognitive status and activities of daily life scores were significantly higher in the group EDA than those in the group CON at 1 month (39.63 ± 4.35 vs. 33.63 ± 5.81, t = -2.13, P < 0.05 and 74.3 ± 12.6 vs. 61.2 ± 13.1, t = -1.69, P < 0.05) and 12 months (40.13 ± 5.93 vs. 34.13 ± 5.36, t = -3.37, P < 0.05 and 79.6 ± 11.7 vs. 65.6 ± 16.6, t = -2.08, P < 0.05) after surgery; and the incidence of neurocognitive dysfunction was significantly lower in the group EDA than that in the group CON (P < 0.05). Serum CXCL13 and IL-6 concentrations were significantly lower in the group EDA than those in the group CON during and after surgery (P < 0.05).@*CONCLUSION@#EDA can significantly reduce the serum concentrations of CXCL13 and IL-6 and improve the PND of patients.

CXCL14 is a candidate biomarker for Hedgehog signalling in idiopathic pulmonary fibrosis.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is associated with aberrant expression of developmental pathways, including Hedgehog (Hh). As Hh signalling contributes to multiple pro-fibrotic processes, Hh inhibition may represent a therapeutic option for IPF. However, no non-invasive biomarkers are available to monitor lung Hh activity. METHODS: We assessed gene and protein expression in IPF and control lung biopsies, mouse lung, fibroblasts stimulated in vitro with sonic hedgehog (SHh), and plasma in IPF patients versus controls, and cancer patients before and after treatment with vismodegib, a Hh inhibitor. RESULTS: Lung tissue from IPF patients exhibited significantly greater expression of Hh-related genes versus controls. The gene most significantly upregulated in both IPF lung biopsies and fibroblasts stimulated in vitro with SHh was CXCL14, which encodes a soluble secreted chemokine whose expression is inhibited in vitro by the addition of vismodegib. CXCL14 expression was induced by SHh overexpression in mouse lung. Circulating CXCL14 protein levels were significantly higher in plasma from IPF patients than controls. In cancer patients, circulating CXCL14 levels were significantly reduced upon vismodegib treatment. CONCLUSIONS: CXCL14 is a systemic biomarker that could be used to identify IPF patients with increased Hh pathway activity and monitor the pharmacodynamic effects of Hh antagonist therapy in IPF. TRIAL REGISTRATION NUMBER: Post-results, NCT00968981.

Alcoholic liver disease: pathogenesis and new targets for therapy.

Alcoholic liver disease (ALD) is a major cause of morbidity and mortality worldwide. The spectrum of disease ranges from fatty liver to hepatic inflammation, necrosis, progressive fibrosis and hepatocellular carcinoma. In developed countries, ALD is a major cause of end-stage liver disease that requires transplantation. The most effective therapy for ALD is alcohol abstinence. However, for patients with severe forms of ALD (that is, alcoholic hepatitis) and for those who do not achieve abstinence from alcohol, targeted therapies are urgently needed. The development of new drugs for ALD is hampered by the scarcity of studies and the drawbacks of existing animal models, which do not reflect all the features of the human disease. However, translational research using liver samples from patients with ALD has identified new potential therapeutic targets, such as CXC chemokines, osteopontin and tumor necrosis factor receptor superfamily member 12A. The pathogenetic roles of these targets, however, remain to be confirmed in animal models. This Review summarizes the epidemiology, natural history, risk factors and current knowledge of the pathogenetic mechanisms of ALD. In addition, this article provides a detailed description of the findings of these translational studies and of the animal models used to study ALD.

Activation of innate immunity accelerates sialoadenitis in a mouse model for Sjögren's syndrome-like disease.

OBJECTIVE: Sjögren's syndrome is a chronic autoimmune disorder characterized by progressive lymphocytic infiltration within the salivary and lacrimal glands. This study was undertaken to investigate the effects of innate immunity activation on sialoadenitis in a mouse strain genetically susceptible for development of SS-like disease. METHODS: Female New Zealand Black X New Zealand White F1 mice were repeatedly treated with toll-like 3 receptor agonist poly(I:C). Submandibular glands were investigated at different time points for sialoadenitis by immunohistochemistry and for gene expression of different chemokines by quantitative PCR. Submandibular gland-infiltrating cells were characterized by flow cytometry. RESULTS: Poly(I:C) treatment significantly upregulated the expression of multiple chemokines within the submandibular glands. The severity and incidence of sialoadenitis was considerably higher in poly(I:C)-treated mice. There was a preponderance of dendritic cells and NK cells in the initial inflammatory cell infiltrates, and these were followed by CD4+ T cells. CONCLUSIONS: Our data clearly demonstrate that systemic activation of innate immunity accelerates sialoadenitis in a mouse model for SS-like disease. These findings suggest that chronic activation of innate immunity can influence certain features of SS.

LPS responsiveness and neutrophil chemotaxis in vivo require PMN MMP-8 activity.

We identify matrix metalloproteinase (MMP)-8, the polymorphonuclear (PMN) leukocyte collagenase, as a critical mediator initiating lipopolysaccharide (LPS)-responsiveness in vivo. PMN infiltration towards LPS is abrogated in Mmp8-null mice. MMP-8 cleaves LPS-induced CXC chemokine (LIX) at Ser(4)-Val(5) and Lys(79)-Arg(80). LIX bioactivity is increased upon N-terminal cleavage, enhancing intracellular calcium mobilization and chemotaxis upon binding its cognate receptor, CXCR2. As there is no difference in PMN chemotaxis in Mmp8-null mice compared with wild-type mice towards synthetic analogues of MMP-8-cleaved LIX, MMP-8 is not essential for extravasation or cell migration in collagenous matrices in vivo. However, with biochemical redundancy between MMPs 1, 2, 9, and 13, which also cleave LIX at position 4 approximately 5, it was surprising to observe such a markedly reduced PMN infiltration towards LPS and LIX in Mmp8-/- mice. This lack of physiological redundancy in vivo identifies MMP-8 as a key mediator in the regulation of innate immunity. Comparable results were found with CXCL8/IL-8 and CXCL5/ENA-78, the human orthologues of LIX. MMP-8 cleaves CXCL8 at Arg(5)-Ser(6) and at Val(7)-Leu(8) in CXCL5 to activate respective chemokines. Hence, rather than collagen, these PMN chemoattractants are important MMP-8 substrates in vivo; PMN-derived MMP-8 cleaves and activates LIX to execute an in cis PMN-controlled feed-forward mechanism to orchestrate the initial inflammatory response and promote LPS responsiveness in tissue.

Anti-tumor necrosis factor-alpha antibody treatment reduces serum CXCL16 levels in patients with rheumatoid arthritis.

The aim of this study was to analyze the change of serum chemokins levels of CXCL16, CX3CL1/Fractalkine, and CXCL10/interferon-gamma inducible protein-10 (IP-10) with rheumatoid arthritis (RA), by infliximab treatment. The effects of infliximab treatment were studied in 23 patients with RA, over a period of 30 weeks. The serum levels of CXCL16, Fractalkine, and IP-10, were measured at the baseline, just before initial treatment, and at 14 and 30 weeks after the initial treatment, with infliximab by ELISA. The higher levels of serum CXCL16 in the RA patients before treatment with infliximab significantly decreased at 14 and 30 weeks after the initial treatment with infliximab, but the serum Fractalkine and IP-10 levels did not decrease significantly. Infliximab treatment significantly lowered the serum levels of CXCL16 in patients with RA. CXCL16 is one of the crucial chemokines regulated by infliximab treatment.