Inflammatory Cell Adhesion Research Articles

FAK and Pyk2 activity promote TNF-\u03b1 and IL-1\u03b2-mediated pro-inflammatory gene expression and vascular inflammation

Protein tyrosine kinase (PTK) activity has been implicated in pro-inflammatory gene expression following tumor necrosis factor-α (TNF-α) or interkeukin-1β (IL-1β) stimulation. However, the identity of responsible PTK(s) in cytokine signaling have not been elucidated. To evaluate which PTK is critical to promote the cytokine-induced inflammatory cell adhesion molecule (CAM) expression including VCAM-1, ICAM-1, and E-selectin in human aortic endothelial cells (HAoECs), we have tested pharmacological inhibitors of major PTKs: Src and the focal adhesion kinase (FAK) family kinases - FAK and proline-rich tyrosine kinase (Pyk2). We found that a dual inhibitor of FAK/Pyk2 (PF-271) most effectively reduced all three CAMs upon TNF-α or IL-1β stimulation compared to FAK or Src specific inhibitors (PF-228 or Dasatinib), which inhibited only VCAM-1 expression. In vitro inflammation assays showed PF-271 reduced monocyte attachment and transmigration on HAoECs. Furthermore, FAK/Pyk2 activity was not limited to CAM expression but was also required for expression of various pro-inflammatory molecules including MCP-1 and IP-10. Both TNF-α and IL-1β signaling requires FAK/Pyk2 activity to activate ERK and JNK MAPKs leading to inflammatory gene expression. Knockdown of either FAK or Pyk2 reduced TNF-α-stimulated ERK and JNK activation and CAM expression, suggesting that activation of ERK or JNK is specific through FAK and Pyk2. Finally, FAK/Pyk2 activity is required for VCAM-1 expression and macrophage recruitment to the vessel wall in a carotid ligation model in ApoE−/− mice. Our findings define critical roles of FAK/Pyk2 in mediating inflammatory cytokine signaling and implicate FAK/Pyk2 inhibitors as potential therapeutic agents to treat vascular inflammatory disease such as atherosclerosis.

Evaluation of anti-atherogenic effects of P2Y12 receptor antagonists in Apolipoprotein E-deficient mice

Prasugrel and Ticagrelor, two P2Y12 receptors antagonists commonly used for the treatment of acute coronary syndrome, are suspected to exert anti-inflammatory effects, which may decrease atherosclerotic lesions. Moreover, Ticagrelor, through its interaction with adenosine metabolism, may have supplemental positive effect on inflammation and atherosclerosis. We evaluated in this study the effects of these two molecules on atherosclerotic lesions and inflammation on a murine model of atherosclerosis. Firstly, twelve C57BL/6 mice treated by vehicule ( N = 4), Prasugrel ( N = 4) or Ticagrelor ( N = 4) were used to evaluate the comparability of the anti-platelet effects by aggregometry. Then, twenty-one Apolipoprotein E deficient mice (ApoE -/-) fed a paigen diet (1.25% cholesterol, 0.5% cholic acid, 15% fat) were treated with vehicule ( N = 7), Prasugrel 5 mg/kg/day ( N = 7), or Ticagrelor 180 mg/kg/day ( N = 7) during 6 weeks. Anti-atherosclerotic effects of these molecules were evaluated in vivo using 99mTc-cAbVCAM1-5, an imaging agent evaluating the expression of the inflammatory marker Vascular Cell Adhesion Molecule-1 (VCAM-1). Maximum platelet aggregation was lower in C57BL/6 mice treated by Prasugrel and Ticagrelor (8%), than in mice treated by vehicule (70%). 99mTc-cAbVCAM1-5 uptake in the aortic root evaluated by in vivo Single Photon Emission Computed Tomography (SPECT) imaging (2.1 ± 0.6%ID/mm 3 vs 2.2 ± 0.7ID/mm 3 vs 1.9 ± 0.7ID/mm 3 ), ex-vivo biodistribution (2.9 ± 0.8%ID/g vs 2.5 ± 0.4%ID/g vs 2.6 ± 0.5%ID/g), and autoradiography (3.9 ± 1.0%ID/g vs 4.2 ± 1.0%ID/g vs 4.0 ± 0.6%ID/g) were not different between the control group, Prasugrel group and Ticagrelor group respectively. Ex vivo analysis confirmed that plaque surface and lipid content were identical between the three groups. Neither Prasugrel nor Ticagrelor has anti-atherosclerotic effects in this specific model of ApoE -/- mice fed a Paigen diet.

MGSEA \u2013 a multivariate Gene set enrichment analysis

BackgroundGene Set Enrichment Analysis (GSEA) is a powerful tool to identify enriched functional categories of informative biomarkers. Canonical GSEA takes one-dimensional feature scores derived from the data of one platform as inputs. Numerous extensions of GSEA handling multimodal OMIC data are proposed, yet none of them explicitly captures combinatorial relations of feature scores from multiple platforms.ResultsWe propose multivariate GSEA (MGSEA) to capture combinatorial relations of gene set enrichment among multiple platform features. MGSEA successfully captures designed feature relations from simulated data. By applying it to the scores of delineating breast cancer and glioblastoma multiforme (GBM) subtypes from The Cancer Genome Atlas (TCGA) datasets of CNV, DNA methylation and mRNA expressions, we find that breast cancer and GBM data yield both similar and distinct outcomes. Among the enriched functional categories, subtype-specific biomarkers are dominated by mRNA expression in many functional categories in both cancer types and also by CNV in many functional categories in breast cancer. The enriched functional categories belonging to distinct combinatorial patterns are involved different oncogenic processes: cell proliferation (such as cell cycle control, estrogen responses, MYC and E2F targets) for mRNA expression in breast cancer, invasion and metastasis (such as cell adhesion and epithelial-mesenchymal transition (EMT)) for CNV in breast cancer, and diverse processes (such as immune and inflammatory responses, cell adhesion, angiogenesis, and EMT) for mRNA expression in GBM. These observations persist in two external datasets (Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) for breast cancer and Repository for Molecular Brain Neoplasia Data (REMBRANDT) for GBM) and are consistent with knowledge of cancer subtypes. We further compare the characteristics of MGSEA with several extensions of GSEA and point out the pros and cons of each method.ConclusionsWe demonstrated the utility of MGSEA by inferring the combinatorial relations of multiple platforms for cancer subtype delineation in three multi-OMIC datasets: TCGA, METABRIC and REMBRANDT. The inferred combinatorial patterns are consistent with the current knowledge and also reveal novel insights about cancer subtypes. MGSEA can be further applied to any genotype-phenotype association problems with multimodal OMIC data.

Interrelationship of endothelial function parameters in children with bronchial asthma in exacerbation and remission.

In children with asthma, endothelial dysfunction signs are observed, and their extent depends on the severity of the disease. These changes are also present in remission. High level of soluble Vascular Cell Adhesion Molecule-1 (sVCAM-1) expression causes active adhesion of inflammatory cells and can indicate direct endothelium participation in development and supporting of chronic inflammation. Bronchial asthma (BA) is characterised by airways chronic inflammation. A special role in this inflammatory process formation is played by development of endothelial dysfunction. The aim of the study was to evaluate endothelial state in children with clinically stable and exacerbated asthma. 91 children with persistent asthma were examined. Among them there were 40 patients with mild persistent (group I), 34 subjects with moderate persistent (group II) and 17 individuals with severe persistent (group III) asthma. 20 healthy children were selected as controls. The serum levels of sVCAM-1 were determined by enzyme-linked immunosorbent assay (ELISA). The ultrasound assessment of endothelium-dependent flow-mediated dilation of the brachial artery (FMD%) has been made. Ultrasonography has been used for investigation of the intima-media thickness (I-M) complex. Data analysis was performed with the Statsoft Statistica Version 8 (Tulsa, OK). The serum levels of sVCAM-1 were significantly increased in the patients with asthma exacerbation (p < 0.001) and remission (p < 0.001), compared with the controls. The index of FMD% was significantly diminished in the patients of I, II, III group with exacerbation (p < 0.001) and stayed lower in the subjects with asthma in remission (p < 0.001), compared with the controls. The thickness of I-M complex was significantly increased in the patients of I, II, III group, compared with the controls (p < 0.001). The endothelium parameter levels: sVCAM-1 (H = 56.11, p = 0.0001), FMD% (H = 43.20, p = 0.0000), the thickness of I-M complex (H = 49.37, p = 0.0000) depend on the severity of the disease. Correlations between the endothelium and pulmonary function parameters were proved (p < 0.05). Endothelial dysfunction in children with asthma was determined. Dependence of severity of the disease on functional state of the vascular endothelium was proved.

Icariin Attenuates High Glucose-Induced Apoptosis, Oxidative Stress, and Inflammation in Human Umbilical Venous Endothelial Cells.

Endothelial dysfunction is closely associated with diabetic complications. Icariin, a flavonoid glycoside isolated from the Epimedium plant species, exhibits antidiabetic properties. However, its impact on endothelial function remains poorly understood, particularly under hyperglycemia. In this study, we investigated the potential protective effect of icariin on high glucose-induced detrimental effects on vascular endothelial cells. Human umbilical venous endothelial cells were incubated in media containing 5.5 mM glucose (normal glucose) or 25 mM glucose (high glucose) in the presence or absence of 50 µM icariin for 72 h. We found that high glucose markedly induced cell apoptosis, enhanced reactive oxygen species generation, and elevated expression levels of inflammatory factors and cell adhesion molecules, which were greatly subdued by icariin supplementation. In conclusion, icariin exerted a beneficial effect on high glucose-induced endothelial dysfunction. This new finding provides a promising strategy for future treatment of diabetic vascular complications.

Comparison of acute thrombogenicity for magnesium versus stainless steel stents in a porcine arteriovenous shunt model.

The present study aimed to investigate whether the Magmaris resorbable magnesium scaffold (RMS) has platelet-repelling properties by comparing its acute thrombogenicity with an equivalent stainless steel stent in an arteriovenous shunt model. An ex vivo porcine carotid jugular arteriovenous shunt was established and connected to Sylgard tubing containing the Magmaris RMS with sirolimus-eluting PLLA coating and an equivalent 316L stainless steel stent with sirolimus-eluting PLLA coating. Six shunts (two shunt runs per pig) were run comparing the two scaffolds (n=9) in alternating order. Nested generalised linear mixed models were employed to compare variables between scaffold groups. Confocal fluorescent microscopy containing CD61/CD42b demonstrated that the 316L equivalent stent had significantly greater platelet coverage of the total scaffold compared with Magmaris (5.8% vs. 2.8%, adjusted rate ratio 2.21 [1.41, 3.47], p=0.012). Scanning electron microscopy demonstrated significantly greater thrombus deposition on the 316L equivalent stent as a percentage of the total scaffold compared with Magmaris (8.0% vs. 5.3%, p=0.009). Magmaris also had significantly less CD14 positive monocyte deposition and a trend towards less PM-1 positive neutrophil compared with the 316L equivalent stent. Magmaris has less thrombogenicity and inflammatory cell deposition compared with the equivalent 316L stainless steel (in geometry and design) stent in a porcine arteriovenous shunt model. These data suggest that resorbable magnesium scaffolds may have inherent properties that reduce adhesion of platelets and inflammatory cells.

Red Blood Cell Adhesion in Adult Patients with Sickle Cell Disease, at Baseline and with Pain, Measured on SCD Biochip Microfluidic Assay

Red Blood Cell Adhesion in Adult Patients with Sickle Cell Disease, at Baseline and with Pain, Measured on SCD Biochip Microfluidic AssayAlexandra Boye-Doe1, Erina Quinn1, Charlotte Yuan1, Umut A. Gurkan, PhD1, and Jane A. Little, MD21Case Western Reserve University, Cleveland, OH; 2Division of Hematology/Oncology, Case Western Reserve University/ University Hospitals Seidman Cancer Center, Cleveland, OHBackground:Despite being monogenic, sickle cell disease (SCD) has a variable phenotype, in which clinical complications and manifestations evolve as patients age. In children, pain generally resolves between crises, whereas adults may experience acute, chronic, or acute-on-chronic pain. We have taken a multifaceted approach to characterize adhesion and inflammation during self-identified pain episodes in adults with SCD in order to better understand pain syndromes in adults and the potential for more specifically targeted therapies. In this pilot study, we assessed changes in red blood cell (RBC) adhesion to the subendothelial protein laminin (LN) during crisis in adults with SCD self-reporting for pain crisis on whom we had baseline adhesion data from a routine clinic visit.Methods:Surplus blood from patients' routine bloodwork was used. Crisis samples were collected from patients at the Acute Care Clinic or Emergency Department at University Hospitals Cleveland Medical Center (UHCMC) when patients presented for management of pain. Baseline samples were collected during routine visits to the Sickle Cell Clinic. This study was approved by the IRB at UHCMC. Within a 24-hour period, RBC adhesion to LN was quantitated by microscopy after passage of unprocessed whole blood through a LN-coated microfluidic adhesion assay, the SCD biochip [1]. Samples were analyzed for hemoglobin (Hb) phenotype by high-performance liquid chromatography (HPLC) in the clinical lab. Correlative clinical data, including, baseline lab values, and medical history, were obtained from the patients' medical records and used to characterize our results. Data from people with multiple samples were used as median values.Results:Blood samples from 19 unselected patients with sickle cell hemoglobin SS (HbSS) were obtained at crisis, and compared with baseline samples obtained from 2014 to 2018 (n = 67 samples). 2 groups were identified: Group 1 with increased adhesion (>25% rise from baseline, n= 10) during crisis, and Group 2 with decreased adhesion (>25% fall from baseline, n=8) or no change (<25% change) (n=1) during crisis (Fig. 1). Time between a patient's initial crisis event and when they presented for pain management varied, possibly affecting observed adhesion. Nonetheless, patients showing an increased adhesion in crisis also showed a decrease in Hb (p = 0.039) between their baseline analyses and the crisis visit. A decrease in Hb may associate with increased adhesion, if the latter contributes to crisis-related hemolysis. However, other markers of hemolysis did not change. By contrast, patients with decreasing RBC adhesion showed a decrease in absolute reticulocyte count (ARC, p = 0.019) at their crisis visit. Statistical analysis of HbS, HbA, and HbF levels n = 64 showed no significant change between baseline or crises.Discussion:A decrease in adhesion during crisis may reflect the presence of sickled RBCs that adhere preferentially to other basement membrane proteins such as fibronectin or thrombospondin. In addition, inflammatory white blood cell adhesion, possibly central to vaso-occlusion, is not evaluated in this study. We are currently examining cytokine and monocyte profiles from people with SCD presenting for management of pain, so that we may better understand inflammatory mediators of pain. Our data are the first to try to understand RBC adhesion in people with SCD who present for management of pain, whether this as an isolated or common event for that person. We found that unselected adults with SCD who presented for management of pain had heterogeneous changes in RBC adhesion, which may reflect differences in underlying mechanism. The pathophysiologic heterogeneity of pain in adults with SCD will be important to understand as anti-adhesion therapies are being developed and adopted clinically. [Display omitted] DisclosuresLittle:Doris Duke Charitable Foundations: Research Funding; NHLBI: Research Funding; Hemex: Patents & Royalties: Patent, no honoraria; PCORI: Research Funding.

How KEAP1 Deletion in Single Cell Mitigates Sickle Cell Pathology

Sickle cell disease (SCD) is one of the most frequent blood disorders caused by a point mutation of β-globin gene. In the pathology of SCD, we often encounter contributions of vascular and cellular components. The activation of vascular endothelial cells promotes adhesion of inflammatory cells and vaso-occlusion responsible for low blood flow, hypoxemia and organ damages. Hemolysis triggers activation and recruitment of myeloid cells to the inflammation site, further sustaining the inflammatory process. Recent progress has created the need to examine targeted induction of the KEAP1-NRF2 system as a potential therapeutic tool for the SCD. We have demonstrated that genetic as well as pharmacological induction of NRF2 in SCD prevent organ damages and suppress inflammation, although it still remains unclear how the NRF2 induction modifies the pathology or which type of cells contribute to the NRF2-mediated improvement of SCD. To address these questions, we conducted targeted deletion of Keap1 gene and induced expression of Nrf2 gene in endothelial cells and macrophages/monocytes using Tie1-Cre and LysM-Cre system, respectively. We found that SCD::Keap1F/F::Tie1-Cre mice (NRF2-increase in endothelial cells) showed low hematocrit and low platelet counts. While severe infiltration of monocytes, lymphocytes and macrophages was observed in the lungs of SCD mice, such phenotype was diminished in the SCD::Keap1F/F::Tie1-Cre mice. In the compound mutant mice, livers showed fewer necrotic zones and less to none accumulation of free iron. On the contrary, imaging MS (iMScope) analyses revealed decrease of free heme in livers and spleens in SCD::Keap1F/F::LysM-Cre mice (NRF2-increase in macrophages/monocytes). Furthermore, upregulations of most known Nrf2-dependent pathways and other yet-to-be-defined pathways are found in an RNA-sequencing analysis of the mice. Importantly, NRF2 induction enhanced heme catabolism and decreases the plasma level of free heme and the quantity of accumulated free heme in the SCD::Keap1F/F::LysM-Cre mouse tissues, whereas the change is not obvious in SCD::Keap1F/F::Tie1-Cre mice. On the contrary, vascular leakage of fluids was decreased in the SCD::Keap1F/F::Tie1-Cre mice. Thus, our results demonstrate that cell-lineage-specific activation of Nrf2 does inhibit the inflammation and organ injury with some particularities to each genotype. DisclosuresNo relevant conflicts of interest to declare.

Transplantation of Endothelial Progenitor Cells Attenuates Lipopolysaccharide-Induced Lung Injury via Inhibiting the Inflammatory Secretion of Neutrophils in Rats

BackgroundEndothelial progenitor cells (EPCs) are special types of stem cells and are a potential novel therapeutic approach in acute lung injury (ALI). Transplantation of EPCs can ameliorate the inflammatory state by reducing adhesion and exudation of inflammatory cells. However, the mechanism underlying the effect of EPCs on inflammatory response modulation remains unclear. The aim of the present study was to investigate the effect of EPCs on the modulation of neutrophils in vitro and in vivo. Materials and MethodsEPCs were cocultured with neutrophils after lipopolysaccharide stimulation in vitro or transplanted into ALI rats, and neutrophil inflammatory mediators including tumor necrosis factor-α, interleukin-1β, neutrophil elastase, myeloperoxidase and matrix metalloproteinases-9 were detected by enzyme-linked immunosorbent assay, an myeloperoxidase detection kits, reverse transcription-polymerase chain reaction and western blotting. ResultsThe results showed that EPCs significantly downregulated the expression of inflammatory mediators when cocultured with neutrophils in vitro or in vivo. ConclusionsThese findings demonstrated that EPCs contributed to lung injury in ALI rats by downregulating neutrophil inflammatory mediators.

Detection of biomarkers using a novel proximity extension assay in patients with ST-elevation myocardial infarction

BackgroundST-elevation myocardial infarction (STEMI) involves inflammation, activation of platelets, coagulation and changes in fibrinolysis as well as remodelling of myocardial tissue after STEMI. Recently, new proximity extension assays including panels of biomarkers for cardiovascular disease have been developed. This study investigates a wide range of cardiovascular protein biomarkers in the acute phase of STEMI compared with the stable phase three months after STEMI.We hypothesized that major changes in inflammation, haemostasis, tissue remodelling, and proteolysis are prevalent. Materials and methodsThis was a prospective, observational study including 48 STEMI patients (mean age 60 ± 12 years, 79% men) treated with primary percutaneous coronary intervention (PPCI). Blood samples were obtained immediately prior to PPCI and again three months later. Levels of 92 biomarkers reflecting inflammation, immune response, cell adhesion, haemostasis, fibrinolysis, tissue remodelling, and proteolysis were assessed using a proximity extension assay (Olink® CARDIOVASCULAR III). ResultsWhen comparing the acute phase of STEMI with the stable phase three months later, a total of 29 biomarkers differed significantly after Bonferroni correction (p < 0.0005). In the acute phase of STEMI, we found an overall increase of biomarkers reflecting immune and inflammatory response, cell adhesion, and haemostasis. Biomarkers reflecting tissue remodelling and proteolysis were increased at three months follow-up compared with the acute phase. Out of the 29 biomarkers, six biomarkers did not confirm our predefined hypotheses. ConclusionsUsing a novel proximity extension assay technique, we detected changes in several biomarkers when comparing the acute phase with three months follow-up in patients with STEMI. These biomarkers may play important roles in the pathogenesis of STEMI.

Small Molecule Mediated Intervention of NF-kB or TGF-β Signaling Reverses Age-Related Phenotypes in Satellite Cells and Enhances Muscle Repair

Skeletal muscle normally exhibits robust regenerative potential in youth; however, myogenic activity declines with age, resulting in defective muscle repair after injury. In this study, we identified signaling pathways in mouse muscle stem cells that are altered with aging and contribute to age-dependent decline in muscle regenerative potential. By analyzing the transcriptional profile of highly purified muscle satellite cells, we identified a unique set of genes whose expression changes with chronological time. This set contains genes regulating inflammatory processes, cell adhesion and matrix interactions, proteolysis, extracellular matrix remodeling, skeletal muscle development, cancer and connective tissue disorders. Ingenuity Pathway Analysis (IPA) of these transcripts indicated a central role for the NF-κB pathway in muscle satellite cell dysfunction. In addition, we noted altered expression of extracellular matrix genes, a signature that could be linked to changes in TGF-β-like signaling activity. Pharmacological modulation of either the NF-κB or TGF-β-like signaling pathways with small molecule inhibitors targeting IKK2 kinase or ALK4/5 receptor-like kinase enhanced regenerative potential in aged muscle, altering both the kinetics and magnitude of the response. These results identify key pathways underlying age-related muscle dysfunction and demonstrate the potential therapeutic relevance of small molecules that target them.

Abstract 1265: Protective efficacy of gut microbiome and microbial metabolites after rice bran consumption against colon tumorigenesis

Abstract Colorectal cancer (CRC) is the 2nd leading cause of cancer-related deaths in US. Therefore, practical interventions that reduce incidence of adenomatous polyps and/or prevent their progression to CRC are urgently needed. In this regard, several studies have reported the chemopreventive potential of dietary rice bran [a wasted ‘by-product' of white rice (Oryza sativa) processing] against CRC. Given that high heat can stabilize rice-bran for safe human consumption, our research efforts are evaluating ‘stabilized-rice bran', as a functional food for CRC control and prevention. Rice bran has been previously shown to increase colonization of native gut probiotics and undergo microbiota fermentation. However, little is known about rice bran constituents as prebiotics, microbial substrates that promote healthy gut microbiota, and how these microbes produce novel, bioactive microbial metabolites. In this regard, our studies show that native gut probiotic - fermented rice bran extracts have strong in vitro activity against CRC cells and significantly decrease their colonosphere formation (a stem cell characteristic). Rice bran (fermented and non-fermented) also strongly inhibits TNF alpha-induced NF-kappa B activation and interleukin (IL)- 4-induced expression of CD44, which along with its variant form (CD44 v3-v6) plays an essential role in the adhesion and infiltration of inflammatory cells in CRC. Notably, even the stool extracts from rice bran-fed CRC survivors showed increased inhibitory effects on CRC cell viability compared to the stools collected from the same individuals prior to rice bran consumption. Next, extensive in vivo investigations were performed to determine the relative efficacy of dietary rice bran-modified gut microbiota and microbiota-fermented rice bran on inflammation associated-colon tumorigenesis. First, Bifidobacterium Longum -fermented (fermented with selected rice bran responsive microbe that was induced during rice bran feeding clinical trials) and non-fermented dietary rice bran were fed to conventionally-raised or germ-free mice and evaluated for their comparative efficacy against azoxymethane (AOM)/ dextran sodium sulphate (DSS)-induced CRC. In second set of studies, we ‘humanized' gut of germ-free mice with stool inoculum from rice bran feeding human clinical trials and evaluated their efficacy against AOM/DSS-induced CRC. In conventional mice study, both the rice bran diets showed protective efficacy against CRC. Interestingly, germ-free mice having undergone transfaunation with rice-bran modified stool inoculum showed significantly less cancerous lesions compared to mice humanized with control stool inoculums, indicating that rice bran has the potential to re-structure the ‘high risk' gut microflora of CRC patients towards the colonization of populations with protective effects against CRC growth and progression. Citation Format: Akhilendra K. Maurya, Sushil Kumar, Rama Kant, Dileep Kumar, Bupinder Raina, Anita Yadav, Elizabeth P. Ryan, Komal Raina. Protective efficacy of gut microbiome and microbial metabolites after rice bran consumption against colon tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1265.

Implication of the Significance of Dietary Compatibility: Based on the Antioxidant and Anti-Inflammatory Interactions with Different Ratios of Hydrophilic and Lipophilic Antioxidants among Four Daily Agricultural Crops.

The hydrophilic extracts of eggplant peel (HEEP) and purple sweet potato (HEPP) and lipophilic extracts of tomato (LET) and carrot (LEC) were mixed in different ratios to assess the significance of the compatibility of aliments, based on their antioxidant and anti-inflammatory interactions in H9c2 cells. The results indicated that groups of some combinational extracts (HEPP-HEEP F1/10, LEC-HEEP F3/10, LEC-HEPP F3/10) showed stronger synergistic antioxidant and anti-inflammatory effects than individual groups. For example, the glutathione peroxidase (GPx) activity of the LEC-HEEP (F3/10) group (86.71 ± 1.88) was higher than that in the HEEP (79.97 ± 1.68) and LEC (77.31 ± 1.85) groups. The level of reactive oxygen species (ROS) was 30.37 ± 0.25 in the LEC-HEEP (F3/10) group while the levels were 34.34 ± 0.36 and 46.23 ± 0.51 in the HEEP and LEC groups, respectively. And the level of malondialdehyde (MDA) was 1.82 ± 0.24 in the LEC-HEEP (F3/10) group while the levels were 2.48 ± 0.13 and 3.01 ± 0.24 in the HEEP and LEC groups, respectively. The expressions of inflammatory mediators (IL-1β, IL-6, IL-8) and cell adhesion molecules (VCAM-1, ICAM-1) showed similar tendencies. However, some groups (LET-LEC F5/10, LET-LEC F9/10, LET-HEPP F7/10) showed antagonistic effects based on these indicators. The principal component analysis showed that samples could be defined by two principal components: PC1, the main phenolic acids and flavonoids; PC2, carotenoids. Moreover, phenolics and anthoyanins were in the majority in synergistic groups, and carotenoids were in the majority in antagonistic groups. These results indicated that there exist synergistic or antagonistic interactions of aliments on antioxidation and anti-inflammation, which implied the significance of food compatibility.

MOST IMPORTANT CELLULAR CHANGES INVOLVED IN RENAL ISCHEMIA REPERFUSION INJURY AND THE CONSEQUENT IMPACT ON SELECTED REMOTE ORGANS

Because of the high rate of baseline oxygen use by renal cells, kidney is highly influenced by obstruction of arterial blood inflow and subsequent shortage of the received oxygen, this condition is known as Ischemic injury. There are many clinical settings associated with unavoidable ischemic state such as kidney transplantation, partial nephrectomy or suprarenal procedures of the aorta. During ischemia many cellular changes occur including vascular congestion and adhesion of inflammatory cells to the endothelium with subsequent infiltration into the kidney tissue. Following ischemia, a phase known as Reperfusion begins andinvolves a return of blood and oxygen supply to micro vessels. Reperfusion was expected to restore the damage occurred during the ischemic phase, paradoxically, reperfusion leads to more congestion, red cells trapping and excessive generation of reactive oxygen species (ROS), which can oxidatively modify significantly every type of biomolecule, thereby inducing cell dysfunction and induce reperfusion injury. Ischemia reperfusion injury (IRI) is also related to a phenomenon called Remote Organ Injury (ROI) in which the damaging effect induced by I/R is not only restricted to the tissue that undergoing the initial ischemia but alsoit leads to injury to remote organs such as the liver, lung, gut. ROI usually occurs by the same mechanisms seen in the local injury induced by I/R including the generation of ROS, leukocytes, and inflammatory mediators (e.g; TNF-α). These substances are directly released from the primary injured tissue or indirectly from activated leukocytes or other inflammatory cells causing organ dysfunctions in distant organs. Peer Review History: Received 3 March 2018; Revised 8 March; Accepted 11 March, Available online 15 March 2018 UJPR follows the most transparent and toughest ‘Advanced OPEN peer review’ system. The identity of the authors and, reviewers will be known to each other. This transparent process will help to eradicate any possible malicious/purposeful interference by any person (publishing staff, reviewer, editor, author, etc) during peer review. As a result of this unique system, all reviewers will get their due recognition and respect, once their names are published in the papers. We expect that, by publishing peer review reports with published papers, will be helpful to many authors for drafting their article according to the specifications. Auhors will remove any error of their article and they will improve their article(s) according to the previous reports displayed with published article(s). The main purpose of it is ‘to improve the quality of a candidate manuscript’. Our reviewers check the ‘strength and weakness of a manuscript honestly’. There will increase in the perfection, and transparency. Received file: Reviewer's Comments: Average Peer review marks at initial stage: 5.5/10 Average Peer review marks at publication stage: 8.5/10 Reviewer(s) detail: Dr. Francis Adou Yapo, Felix Houphouet Boigny, University of Abidjan, Ivory Coast, fyapo@yahoo.fr Dr. Fatima Morales Marin, University of Murcia, Spain, fatima.morales@um.es This article has been cited by: Dalia Ibrahim Ismail email orcid ; Alshaymaa Gamal Aboulkhair. The Effect of Aliskiren on Renal Cortical Ischemia/Reperfusion Injury in Albino Rats: A Histological and Immunohistochemical Study. Article 6, Volume 42, Issue 4, Autumn 2020, Page 838-848. Pubmed

Blocking osteopontin-fibronectin interactions reduce extracellular fibronectin deployment and arthritic immunopathology

Elevated levels of a thrombin-cleaved fragment of osteopontin (OPNT) are seen in synovial fluid (SF) and tissues of rheumatoid arthritis (RA) patients. OPNT binds to integrins on cell surfaces, inducing adhesion, migration and survival of inflammatory cells in the synovial joints, where OPNT binds to fibronectin to link fibroblast-like synoviocytes (FLS) with B cells, stimulating the latter to produce inflammatory cytokines. Our aim was to block OPNT-fibronectin interactions and examine whether this reduces inflammation. A human antibody (phage displayed) library was used to select scFv antibodies cognate to OPNT, and a particular scFv antibody (scFv 31) was evaluated. Adhesion, migration and fibronectin polymerization of FLS cells derived from RA patients were monitored, in cultures incorporating scFv 31. Also, scFv 31 was used in mice with CAIA (collagen antibody-induced arthritis), subjected to clinical and histological assessment, analysis of fibronectin and cartilage damage and induction of pro-inflammatory cytokines. The scFv antibody, scFv 31, appeared to cause significantly reduced migration of synovial fibroblasts, altered cell morphology, changes in actin stress fiber arrangement, and marked reduction in fibronectin. In CAIA mice, scFv 31 appeared to prevent arthritic changes through inhibition of synovial hypertrophy and loss of articular cartilage, decrease in fibronectin polymerization and expression of pro-inflammatory cytokines implicated in arthritis. Osteopontin-fibronectin interaction(s) appear to play a role in the expression of key inflammatory molecules by B cells infiltrating the synovial joint. The scFv antibody, scFv 31, provides a potential therapeutic lead for inhibition of some processes implicated in rheumatoid arthritis.

Troxerutin and Cerebroprotein Hydrolysate Injection Protects Neurovascular Units from Oxygen-Glucose Deprivation and Reoxygenation-Induced Injury In Vitro.

Cerebral ischemia/reperfusion (I/R) injury involves complex events of cellular and molecular processes. Previous studies suggest that a neurovascular unit (NVU) acts as an intricate network to maintain the neuronal homeostatic microenvironment. The present study established an NVU model for oxygen-glucose deprivation and reoxygenation (OGD/R) damage, trying to target the major components of the NVU using a coculture of rat neurons, astrocytes, and rat brain microvascular endothelial cells (rBMECs) to investigate the therapeutic effects of troxerutin and cerebroprotein hydrolysate injections (TCHis). The study observed that OGD/R downregulated the expressions of GAP-43, Claudin-5, and AQP-4 obviously detected by Western blotting and immunocytochemical analysis, respectively, while TCHi ameliorated the effect of OGD/R significantly. Meanwhile, TCHi alleviated the abnormalities of ultrastructure of neurons and rBMECs induced by OGD/R. Furthermore, both levels of inflammatory cytokines (IL-1β, IL-6, and TNF-α) and cell adhesion molecules (VCAM-1 and ICAM-1) detected by ELISA in NVU supernatant were found elevated significantly through OGD/R, but TCHi ameliorated the trend. In addition, TCHi also mitigated the increase of proapoptotic factors (Bax, p53, and caspase-3) induced by OGD/R in NVU model statistically. All these findings demonstrated that TCHis played a protective role, which was reflected in anti-inflammation, antiapoptosis, and blood–brain barrier maintenance. The results of the study concluded that the NVU is an ideal target and TCHi acts as a neuroprotective agent against cerebral I/R injuries.

Muscle-derived extracellular superoxide dismutase inhibits endothelial activation and protects against multiple organ dysfunction syndrome in mice

Multiple organ dysfunction syndrome (MODS) is a detrimental clinical complication in critically ill patients with high mortality. Emerging evidence suggests that oxidative stress and endothelial activation (induced expression of adhesion molecules) of vital organ vasculatures are key, early steps in the pathogenesis. We aimed to ascertain the role and mechanism(s) of enhanced extracellular superoxide dismutase (EcSOD) expression in skeletal muscle in protection against MODS induced by endotoxemia. We showed that EcSOD overexpressed in skeletal muscle-specific transgenic mice (TG) redistributes to other peripheral organs through the circulation and enriches at the endothelium of the vasculatures. TG mice are resistant to endotoxemia (induced by lipopolysaccharide [LPS] injection) in developing MODS with significantly reduced mortality and organ damages compared with the wild type littermates (WT). Heterogenic parabiosis between TG and WT mice conferred a significant protection to WT mice, whereas mice with R213G knock-in mutation, a human single nucleotide polymorphism leading to reduced binding EcSOD in peripheral organs, exacerbated the organ damages. Mechanistically, EcSOD inhibits vascular cell adhesion molecule 1 expression and inflammatory leukocyte adhesion to the vascular wall of vital organs, blocking an early step of the pathology in organ damage under endotoxemia. Therefore, enhanced expression of EcSOD in skeletal muscle profoundly protects against MODS by inhibiting endothelial activation and inflammatory cell adhesion, which could be a promising therapy for MODS.

Crosstalk between glial and glioblastoma cells triggers the \u201cgo-or-grow\u201d phenotype of tumor cells

BackgroundGlioblastoma (GBM), the most malignant primary brain tumor, leads to poor and unpredictable clinical outcomes. Recent studies showed the tumor microenvironment has a critical role in regulating tumor growth by establishing a complex network of interactions with tumor cells. In this context, we investigated how GBM cells modulate resident glial cells, particularly their paracrine activity, and how this modulation can influence back on the malignant phenotype of GBM cells.MethodsConditioned media (CM) of primary mouse glial cultures unexposed (unprimed) or exposed (primed) to the secretome of GL261 GBM cells were analyzed by proteomic analysis. Additionally, these CM were used in GBM cells to evaluate their impact in glioma cell viability, migration capacity and activation of tumor-related intracellular pathways.ResultsThe proteomic analysis revealed that the pre-exposure of glial cells to CM from GBM cells led to the upregulation of several proteins related to inflammatory response, cell adhesion and extracellular structure organization within the secretome of primed glial cells. At the functional levels, CM derived from unprimed glial cells favored an increase in GBM cell migration capacity, while CM from primed glial cells promoted cells viability. These effects on GBM cells were accompanied by activation of particular intracellular cancer-related pathways, mainly the MAPK/ERK pathway, which is a known regulator of cell proliferation.ConclusionsTogether, our results suggest that glial cells can impact on the pathophysiology of GBM tumors, and that the secretome of GBM cells is able to modulate the secretome of neighboring glial cells, in a way that regulates the “go-or-grow” phenotypic switch of GBM cells.

Orally Active Epoxyeicosatrienoic Acid Analogs.

Biologically active epoxyeicosatrienoic acid (EET) regioisomers are synthesized from arachidonic acid by cytochrome P450 epoxygenases of endothelial, myocardial, and renal tubular cells. EETs relax vascular smooth muscle and decrease inflammatory cell adhesion and cytokine release. Renal EETs promote sodium excretion and vasodilation to decrease hypertension. Cardiac EETs reduce infarct size after ischemia-reperfusion injury and decrease fibrosis and inflammation in heart failure. In diabetes, EETs improve insulin sensitivity, increase glucose tolerance, and reduce the renal injury. These actions of EETs emphasize their therapeutic potential. To minimize metabolic inactivation, 14,15-EET agonist analogs with stable epoxide bioisosteres and carboxyl surrogates were developed. In preclinical rat models, a subset of agonist analogs, termed EET-A, EET-B, and EET-C22, are orally active with good pharmacokinetic properties. These orally active EET agonists lower blood pressure and reduce cardiac and renal injury in spontaneous and angiotensin hypertension. Other beneficial cardiovascular actions include improved endothelial function and cardiac antiremodeling actions. In rats, EET analogs effectively combat acute and chronic kidney disease including drug- and radiation-induced kidney damage, hypertension and cardiorenal syndrome kidney damage, and metabolic syndrome and diabetes nephropathy. The compelling preclinical efficacy supports the prospect of advancing EET analogs to human clinical trials for kidney and cardiovascular diseases.

Development of Human Umbilical Vessels in The Second Trimester of Pregnancy: Histological, Immunohistochemical and Morphometric Study

Introduction: There are many publications describing structure of full term umbilical vessels, however, few studies in early gestation.Aim: To elucidate changes in microstructure of developing umbilical vessels in the second trimester of pregnancy.Material and Methods: Twelve specimens of human umbilical cords were obtained from legal termination of uncomplicated pregnancies at gestational weeks 13, 16, and 20. Two centimeter segments of the cords were cut near the placenta. Specimens were processed for paraffin blocks, sectioned, and stained with haematoxylin & eosin, Masson Trichrome, Orcein, and Periodic Acid Schiff. Immunohistochemistry for alpha smooth muscle actin & laminin antibodies was performed. Morphometry and image analysis for vascular wall thickness & diameter of lumen, and diameter of umbilical cord were done.Results: Umbilical cord sections revealed two arteries (UA) and one vein (UV) embedded in Wharton’s jelly and covered with amniotic epithelium formed of flat then cuboidal epithelium. Umbilical vessels composed of inner intima and outer media. Endothelium demonstrated areas of damage and adhesion of inflammatory cells. Internal elastic lamina was thin, interrupted initially, double layered, and well-developed finally. Smooth muscle cells (SMCs) migrated from vascular lumen or from media by a process of mesenchymal-endothelial transition to replace injured endothelium. The media was composed of immature SMCs initially (thin (morphometry), non-contractile, non-secretory as proved by different stains and collagen content. Wharton’s jelly spindle shaped mesenchymal cells close to vessels revealed positive staining for alpha smooth muscle actin and contributed to the media to compensate for SMC migration. UAs showed thicker wall, narrower lumen than veins and cord diameter increased significantly at the 20 week.Conclusion: Development of umbilical vessels was the result of a continuous remodeling process initiated by secreted endothelial factors from damaged endothelium that influenced; SMCs of media, stem cells of cord blood and Wharton’s jelly simulating early events of atherosclerosis.