Expression Of NF Research Articles

Effect of 17β-Estradiol on Endothelial Cell Expression of Inflammation-Related MicroRNA.

Human Umbilical Vein Endothelial cells (HUVECs) were treated with media in the absence (control) and presence of 17β-estradiol (100 nM) for 24 hr. Thereafter, endothelial cell release of cytokines (IL-6 and IL-8), the intracellular expression of the central protein inflammatory mediator NF- B, and the levels of inflammatory-associated miRNAs: miR-126, miR-146a, miR-181b, miR-204, and miR-let-7a, were determined. 17β-estradiol-treated cells released significantly lower levels of IL-6 (47.6±1.5 pg/mL vs 59.3±4.9 pg/mL) and IL-8 (36.3±2.3 pg/mL vs 44.0±2.0 pg/mL). Cellular expression of total NF- B (26.0±2.8 AU vs 21.2±3.1 AU) was not different between groups; however, activated NF- B (Ser536) (12.9±1.7 AU vs 20.2±2.2 AU) was markedly reduced in 17β-estradiol-treated cells as compared to untreated cells. Furthermore, cellular expressions of miR-126 (1.8±0.3 fold), miR-146a (1.7±0.3 fold), miR-181b (2.1±0.4 fold), miR-204 (1.9±0.4 fold), and miR-Let-7a (1.8±0.3 fold) were markedly increased in response to 17β-estradiol treatment. These data suggest that the anti-inflammatory effect of 17β-estradiol in endothelial cells may be mediated by miRNAs.

Increased expression of GLUT4, catalase and nitric oxide by a crystalised fraction from ethanolic extracts of oak leaf lettuce and okra seed in C2C12 cell line

The fractions obtained from low temperature-induced crystallisation of ethanolic extracts of green- and red-oak leaf and okra seed were accounted for total polyphenol content and the DPPH antioxidant activity, resulting in 5.99, 4.44, and 12.35 mg gallic acid equivalent per g sample, and 0.45, 0.35, and 0.99 mg Trolox equivalent per g sample, respectively. Insulin resistance was the result after incubating C2C12 skeletal muscle cells in high glucose DMEM for 20 h. Oxidative stress and inflammation were triggered by re-incubating these resistant cells with insulin. The expression of p-p38 MAPK and NF-B proteins and the NF-B p65 transcription factor activity were enhanced. Such inflammatory protein markers were reduced when the crystalised extracts replaced insulin. Increased catalase activity and NO production were also determined for the incubations using the crystalised extracts. It was suitable to include these vegetables in the daily diets of prediabetic individuals for better management of the patient’s health status by increasing insulin sensitivity and decreasing inflammation.

Exosomes derived from CD271+CD56+ bone marrow mesenchymal stem cell subpopoulation identified by single-cell RNA sequencing promote axon regeneration after spinal cord injury.

Rationale: Spinal cord injury (SCI) results in neural tissue damage. However, the limited regenerative capacity of adult mammals' axons upon SCI leads to persistent neurological dysfunction. Thus, exploring the pathways that can enhance axon regeneration in injured spinal cord is of great significance. Methods: Through the utilization of single-cell RNA sequencing in this research, a distinct subpopulation of bone marrow mesenchymal stem cells (BMSCs) that exhibits the capacity to facilitate axon regeneration has been discovered. Subsequently, the CD271+CD56+ BMSCs subpopulation was isolated using flow cytometry, and the exosomes derived from this subpopulation (CD271+CD56+ BMSC-Exos) were extracted and incorporated into a hydrogel to create a sustained release system. The aim was to investigate the therapeutic effects of CD271+CD56+ BMSC-Exos and elucidate the underlying mechanisms involved in promoting axon regeneration and neural function recovery. Results: The findings indicate that CD271+CD56+ BMSC-Exos share similar physical and chemical properties with conventional exosomes. Importantly, in an SCI model, in situ implantation of CD271+CD56+ BMSC-Exos hydrogel resulted in increased expression of NF and synaptophysin, markers associated with axon regeneration and synapse formation, respectively. This intervention also contributed to improved neural function recovery. In vitro experiments demonstrated that CD271+CD56+ BMSC-Exos treatment significantly enhanced axon extension distance and increased the number of branches in dorsal root ganglion axons. Moreover, further investigation into the molecular mechanisms underlying CD271+CD56+ BMSC-Exos-mediated axon regeneration revealed the crucial involvement of the miR-431-3p/RGMA axis. Conclusion: In summary, the implantation of CD271+CD56+ BMSC-Exos hydrogel presents a promising and effective therapeutic approach for SCI.

Integrating Network Pharmacology and an Experimental Model to Investigate the Effect of Zhenwu Decoction on Doxorubicin-Induced Heart Failure.

Doxorubicin-induced heart failure is a clinical problem that needs to be solved urgently. Previous studies have confirmed that Zhenwu Decoction, a traditional Chinese medicine compound, can effectively improve chronic heart failure. However, its interventional effect on Doxorubicin-induced heart failure has not yet been investigated. In this study, we investigated the therapeutic effect and potential mechanism of Zhenwu Decoction on Doxorubicin-induced heart failure through animal experiments and network pharmacology. The study aimed to investigate the therapeutic effect and potential mechanism of Zhenwu Decoction (ZWD) on Doxorubicin-induced heart failure. A heart-failure mouse model was established in 8-week-old male C57/BL6J mice using Doxorubicin, and the mice were then treated with ZWD for a 4-week period. Firstly, network pharmacology was conducted to explore the potential active components and molecular mechanisms of ZWD on Doxorubicin-induced heart failure. Next, we conducted an in vivo study on the effect of ZWD on Doxorubicin-induced heart failure. After the intervention, the cardiac function and levels of cardiac function injury marker in serum were measured to evaluate the therapeutic effect of ZWD on cardiac function. Then HE staining and Masson staining were used to evaluate the effect of ZWD on myocardial pathology, and biochemical method was used to detect the effect of ZWD on total antioxidant capacity and inflammation, and finally, Western blot was used to detect TGFβ, Smad-3, and collagen I protein expression levels to evaluate its effect on myocardial fibrosis. In Doxorubicin-induced heart failure mice, ZWD improved cardiac function and reduced the levels of CK-MB, NT-proBNP, and BNP in the serum, improved myocardial pathology, and reduced TGFβ, Smad-3 and collagen I protein expression levels to improve myocardial fibrosis. Network pharmacological analysis showed that ZWD has 146 active ingredients and 248 candidate targets. Moreover, 2,809 genes were found to be related to Doxorubicin-induced heart failure, and after screening, 74 common targets were obtained, mainly including IL-6, AKT1, caspase-3, PPARG, PTGS2, JUN, HSP90AA1, and ESR1. KEGG analysis confirmed that PI3K/AKT and IL-6/NF-κB signaling pathways were the two main pathways underlying the cardioprotective effects of ZWD. Finally, in vivo experiments showed that ZWD improved the total antioxidant capacity, reduced the SOD level, increased the protein expression of PI3K, Akt, Bcl-2, Bax, and caspase-3, reduced the levels of TNF-α, IL-6, and IL-1β, and decreased the NF-κB p65, IL-6, and TNF-α protein expression levels. In Doxorubicin-induced heart-failure mice, Zhenwu Decoction improved the cardiac function and myocardial pathology, and improved myocardial fibrosis through the TGFβ/Smad-3 signaling pathway. According to the prediction of network pharmacology, in vivo experiments demonstrated that Zhenwu Decoction can improve the oxidative stress response, improve myocardial cell apoptosis through the PI3K/AKT signaling pathway, and improve myocardial inflammation by reducing the levels of inflammatory factors and by reducing the protein expression of NF-κB p65, IL-6, and TNF-α.

Expression of NF-κB, Bcl-2 and Caspase-8 apoptosis markers in endometrium in external genital endometriosis and chronic endometritis: A retrospective study

Aim. To study the regulation of apoptosis in the endometrium of women with chronic endometritis (CE) and external genital endometriosis (EGE) based on the study of the expression of its regulators such as transcription factor (NF-), Bcl-2 and Caspase-8.
 Materials and methods. The study evaluated the expression of apoptosis factors such as Caspase-8, NF-B transcription factor, and Bcl-2, in patients with chronic endometritis (CE, n=23), external genital endometriosis (EGE, n=20), and intact endometrium (control group; n=7). Statistically significant differences were found for all test markers (p0.05).
 Results. The increased expression of apoptosis factors is probably related to high endometrial pro-inflammatory activity in both CE and EGE. For EGE, this effect is more pronounced at the transcription level in the form of an increase in the expression of the transcription factor NF-B in both glandular and stromal endometrial cells. It is probably due to the systemic nature of the chronic inflammatory process. For CE, despite the antiapoptotic activity of Bcl-2, an increase in Caspase-8 expression in the endometrium may also indicate a predominance of innate immunity activation.
 Conclusion. The imbalance of pro- and antiapoptotic factors can be an inducer of both cytotoxic and cytoprotective mechanisms of EGE and CE. However, it requires further research.

C12 DAPAGLIFLOZIN INCREASES PAMPK AND REDUCES MYOCARDIAL AND RENAL NF–KB EXPRESSION IN PRECLINICAL MODELS OF SHORT–TERM DOXORUBICIN CARDIOTOXICITY THROUGH MYD–188 AND NLRP3 PATHWAYS

Abstract Background Doxorubicin–mediated adverse cardiovascular events are among the leading causes of morbidity and mortality in breast cancer patients. Sodium–glucose cotransporter 2 (SGLT2) inhibitors have recently been shown to be of therapeutic value in patients with type 2 diabetes, chronic kidney disease, and heart failure with reduced ejection fraction (HFrEF), conditions that commonly coexist and are interrelated pathophysiologically. Purpose: We hypothesized that Dapagliflozin (an SGLT2i), administered during doxorubicin, could improve cardiac function in preclinical models Methods Female C57Bl/6 mice were untreated (Sham, n=6) or treated for 10 days with doxorubicin i.p at 2.17 mg/kg (DOXO, n=6), DAPA at 12 mg/kg (DAPA, n=6) or doxorubicin combined to DAPA (DOXO–DAPA, n=6). Ejection fraction, radial and longitudinal strain were analyzed through transthoracic echocardiography (Vevo 2100). Cardiac tissue expression of NLRP3 inflammasome, Myd88, DAMPs (galectine 3 and calgranulinS100), pAMPK, NF–kB, and 13 chemokines (IL–1α, IL–1β, IL–2, IL–4, IL–6, IL–10, IL–12, IL17–α, IL–18, IFN–γ, TNF–α, G–CSF, and GM–CSF) were quantified through ELISA and western blot methods. Results DAPA improved significantly the EF and prevented the reduction of radial and longitudinal strain after 10 days of treatment with doxorubicin. A reduced expression of NLRP3, MyD88, DAMPs and NF–kB in cardiac tissues was seen in DOXO–DAPA group compared to DOXO mice (p<0.001). Cardiac expression of IL–1β, IL–6, TNF–α, G–CSF and GM–CSF were significantly reduced after treatment with DAPA indicating anti–inflammatory properties. Expression of pAMPK was strongly enhanced in DAPA–DOXO compared to DOXO group. Levels of Calgranulin S100 and galectine–3 were strongly enhanced in DOXO group; on the other hand their expression were reduced by 47.7 and 52.3% in DAPA–DOXO group vs DOXO (p<0.005). Hystology evaluation indicate reduction NF–kB expression in myocardial and renal tissue in DOXO–DAPA vs DOXO. Radial strain (RS) is 38.8% in DAPA–DOXO vs 14.1% in DOXO groups (P < 0.05); longitudinal strain (LS) is – 23.4 % in DAPA–DOXO vs – 10.5% in DOXO groups (P < 0.001) Conclusion In this preclinical study, DAPA is able to improve cardiac function and reduce biomarkers involved in heart failure and fibrosis. The overall picture of the study pushes the use of DAPA in prevention of cardiomyopathies induced by anthracyclines in cancer patients.

C7 COMBINATION OF SPIRULINA, GANODERMA LUCIDUM AND MORINGA OLEIFERA IMPROVES CARDIAC FUNCTIONS AND REDUCES PRO–INFLAMMATORY BIOMARKERS IN PRECLINICAL MODELS OF SHORT–TERM DOXORUBICIN–MEDIATED CARDIOTOXICITY

Abstract Background Anthracyclines are essential adjuvant therapies for a variety of cancers, particularly breast, and gastric and esophageal cancers. Whilst prolonging cancer–related survival, these agents can in–duce drug–related cardiotoxicity. Spirulina, Reishi (Ganoderma Lucidum) and Moringa are three nutaceuticals with anti–inflammatory effects that are currently used in cancer patients to improve quality of life and fatigue. Methods Female C57Bl/6 mice were untreated (Sham, n=6) or treated for 10 days with short–term doxorubicin i.p at 2.17 mg/kg (DOXO, n=6), Singo at 12 mg/kg (Singo, n=6) or doxorubicin combined to Singo (DOXO–Singo, n=6). Ejection fraction, radial and longitudinal strain were analyzed through transthoracic echocardiography (Vevo 2100, Fujifilm). Myocardial expression of NLRP3, DAMPs (galectine 3 and calgranulin S100) and 13 cytokines (IL–1α, IL–1β, IL–2, IL–4, IL–6, IL–10, IL–12, IL17–α, IL–18, IFN–γ, TNF–α, G–CSF, and GM–CSF) were quantified through selective mouse ELISA methods. Myocadial fibrosis, necrosis and hypertrophy were analyzed through Immunohistochemistry (IHC). Human cardiomyocytes (HFC cell line) were exposed to subclinical concentration of doxorubicin (200 nM) alone or in combination to Singo ( at 10, 25 and 50 µg/ml) for 24 and 48h. Cell viability studies were performed through MTS assay. Expression of NLRP3 and p65/NF–kB) were made through selective ELISA methods. Results In preclinical models of short–term DOXO cardiotoxicity, Singo improved significantly EF and FS and prevented the reduction of radial and longitudinal strain after 10 days of treatment with DOXO. A reduced expression of myocardial NLRP3 and NF–kB levels in cardiac tissues were seen in DOXO–Singo group compared to DOXO mice (p<0.05). Myocardial expression of pro–inflammatory cytokines were significantly reduced after treatment with Singo indicating anti–inflammatory properties. Myocardial levels of Cal–granulin S100 and galectine–3 were strongly enhanced in DOXO group; on the other hand their expression were reduced in Singo–DOXO group vs DOXO (p<0.05). Immunohistochemistry analysis indicates that Singo reduces fibrosis and hypertrophy in myocardial tissue of mice during exposure to DOXO. Conclusion Singo is able to improve cardiac function and reduce biomarkers involved in heart failure and fibrosis. Cardioprotective properties of Singo are mediated by the reduction of lipid peroxidation products and expression of NLRP3–NF–kB –cytokine pathways.

P128 CTLA–4 AND PD–1 BLOCKING AGENTS AFFECTS LONGITUDINAL AND RADIAL STRAIN IN PRECLINICAL MODELS, INCREASES SYSTEMIC CXCL12, CARDIAC FIBRONECTIN EDA, S–100 CALGRANULIN, GALECTINE–3 AND NLRP–3/MYD–88/CHEMOKINE PATHWAYS

Abstract Background Immune checkpoint inhibitors (ICIs) have significantly changed the oncology clinic in recent years, improving survival expectations in cancer patients. ICI therapy have a broad spectrum of side effects from endocrinopathies to cardiovascular diseases. In this study, pro–inflammatory and pro–fibrotic effects of short–term ICIs therapy in preclinical models were analyzed. Methods Firstly, in a human in vitro model, human cardiomyocytes co–cultured with hPBMC were exposed to ICIs (with CTLA–4 or PD–1 blocking agents, at 200 nM) for 72h. After treatment, production of DAMPs and 12 cytokines were analyzed in the supernatant through colorimetric and enzymatic assays. C57/Bl6 mice were treated with CTLA–4 or PD–1 blocking agents (15 mg/kg) for 10 days. Before (T0), after three days (T3) and after treatments (T10), ejection fraction, fractional shortening, radial and longitudinal strain were calculated by using bidimensional echocardiography (Vevo 2100,Fujfilm). Fibrosis, necrosis, hypertrophy and vascular NF–kB expression were analyzed through Immunohistochemistry. Myocardial expression of DAMPs (S100– Calgranulin, Fibronectin and Galectine–3), MyD88, NLRP3 and twelve cytokines have been analyzed. Systemic levels of SDF–1, IL–1β and IL–6 were analyzed before, during and after ICIs therapy. Results Radial and longitudinal strain were decreased after 10 days of ICIs therapy. Histological analysis of NF–kB expression shows that short–term anti–CTLA–4 or anti–PD–1 treatment increased vascular and myocardial inflammation. No myocardial hypertrophy was seen with the exception of the pembrolizumab group. Myocardial fibrosis and expression of galectin–3, pro–collagen 1–α and MMP–9 were increased after treatment with all ICIs. Both anti–CTLA–4 or anti–PD–1 treatments increased the expression of DAMPs, NLRP3 inflammasome and MyD88 and induced both in vitro and in vivo the secretion of IL–1β, TNF–α and IL–6. Systemic levels of SDF–1, IL–1β and IL–6 were increased during and after treatment with ICIs. Conclusions Short therapy with PD–1 and CTLA–4 blocking agents increases vascular expression of NF–kB, systemic SDF–1, IL–1β, IL–6 levels and myocardial NLRP3, MyD88 and DAMPs expression in preclinical models. A pro–inflammatory cytokine storm was induced in myocardial tissues and in cultured cardiac cells after ICIs therapy.

CHRONOLOGICAL AGE AFFECTS MSC SENESCENCE IN VITRO: A SYSTEMATIC REVIEW

The use of mesenchymal stromal cells (MSCs) in regenerative medicine and tissue engineering is well established, given their properties of self-renewal and differentiation. However, several studies have shown that these properties diminish with age, and understanding the pathways involved are important to provide regenerative therapies in an ageing population.In this PRISMA systematic review, we investigated the effects of chronological donor ageing on the senescence of MSCs. We identified 3023 studies after searching four databases including PubMed, Web of Science, Cochrane, and Medline. Nine studies met the inclusion and exclusion criteria and were included in the final analyses.These studies showed an increase in the expression of p21, p53, p16, ROS, and NF- B with chronological age. This implies an activated DNA damage response (DDR), as well as increased levels of stress and inflammation in the MSCs of older donors. Additionally, highlighting the effects of an activated DDR in cells from older donors, a decrease in the expression of proliferative markers including Ki67, MAPK pathway elements, and Wnt/ -catenin pathway elements was observed. Furthermore, we found an increase in the levels of SA- -galactosidase, a specific marker of cellular senescence.Together, these findings support an association between chronological age and MSC senescence. The precise threshold for chronological age where the reported changes become significant is yet to be defined and should form the basis for further scientific investigations. The outcomes of this review should direct further investigations into reversing the biological effects of chronological age on the MSC senescence phenotype.

Expression of Lymphoma-2 Gene and NF-κB in Human Adenoid Cystic Carcinoma Tissue and Its Relation with Disease Prognosis

Background: Adenoid cystic carcinoma (ACC) is a common tumor of the head and neck. It is characterized by slow growth, strong invasiveness, easy recurrence and distant organ metastasis. Bcl-2 and NF-κB are indicated to relate to tumor growth, proliferation and apoptosis. This study aims to analyze the relationship between Bcl-2, NF-κB and ACC. Methods: With ELISA method to detect Bcl-2 and NF-κB expression in the blood of patients with ACC, patients with benign adenoma and healthy controls, and the expression of Bcl-2 and NF-κB in tissues was detected by immunohistochemistry, and their relationship with ACC was analyzed. relevance. Relationships between clinicopathological features. Results: The levels of Bcl-2 (4.16±1.34 ng/ml) and NF-κB (4.47 ± 1.37 ng/ml) as well as positive rates of Bcl-2 and NF-κB (87.5% and 85%) were significantly elevated in the ACC group (P <0.05). Bcl-2 was positively correlated with NF-κB (P <0.05). Of note, Bcl-2 and NF-κB expressions were correlated with the incidence frequency, histological type, clinical stage, lymph node metastasis and distant metastasis of ACC patients (P < 0.05) as the distant metastasis is an independent risk factor. Conclusion: In summary, Bcl-2 and NF-κB expression in ACC patients is associated with the clinicopathological features of ACC.

Dracocephalum moldavica L. Extract ameliorates intestinal inflammation by regulating gut microbiota and repairing the intestinal barrier in 2k1c rats

When used in ethnomedicine, Dracocephalum moldavica L. (DML) and its bioactive extract protect against several diseases caused by inflammation and oxidative stress. However, the effect of DML on intestinal inflammation in hypertension has not been thoroughly investigated. This study focused on the effects of DML extract on intestinal inflammation in two-kidney, one clip (2K1C) rats. Male Wistar rats were divided into three groups with daily intragastric administration of either saline (control group and model group) or DML extract (treatment group, 50 mg/kg per day) for 4 weeks. In the model and treatment groups, 2K1C hypertension was induced by clipping the left renal artery. The profiles of stool microbiota, the levels of short-chain fatty acids (SCFAs), the integrity of the gut barrier, oxidative stress biomarkers and inflammatory factors were analyzed in rats supplemented with or without DML extract. This study revealed the alleviation of high blood pressure in 2K1C rats treated with DML extract. The DML extract attenuated gut microbiota dysbiosis in the 2K1C rats by increasing the abundance of the genus Lactobacillus and by decreasing the abundance of Papillibacter, Desulfovibrio and Anaerobiospirillum genera. Treatment with the DML extract increased the levels of SCFAs and repaired the intestinal barrier, it reduced the release of oxidative stress biomarkers and inflammatory factors and suppressed the expression of TLR4 and NF-?B mRNAs. We conclude that the DML extract may alleviate intestinal inflammation by adjusting the profile of gut microbiota and enhancing the intestinal barrier.

Retracted: Dapagliflozin Ameliorates STZ‐Induced Cardiac Hypertrophy in Type 2 Diabetic Rats by Inhibiting the Calpain‐1 Expression and Nuclear Transfer of NF‐κB

Retracted: Dapagliflozin Ameliorates STZ‐Induced Cardiac Hypertrophy in Type 2 Diabetic Rats by Inhibiting the Calpain‐1 Expression and Nuclear Transfer of NF‐<i>κ</i>B

MCC950 Ameliorates Acute Exogenous Lipoid Pneumonia Induced by Sewing Machine Oil in Rats via the NF-κB/NLRP3 Inflammasome Pathway.

Acute exogenous lipoid pneumonia (AELP) is a rare disorder caused by intake of lipid formulations and is often underdiagnosed. Meanwhile, the mechanism of AELP is still underlying. MCC950, was previously found to significantly suppress the release of inflammatory cytokines IL-18 and IL-1β. However, the effect of MCC950 on AELP induced by sewing machine oil has not been reported. The NLRP3, NF-[Formula: see text]B p65, caspase-1 and IL-1β expression in lung tissues were compared between a rat model of AELP and control rats using western blotting and real-time quantitative assay. Moreover, haematoxylin and eosin (H&E) staining was performed to elucidate the mechanisms by which MCC950 ameliorates sewing machine oil-induced AELP in vivo. MCC950 reduced the expression of NF-[Formula: see text]B p65 in the lung samples of the treatment group and further down-regulated the NLRP3 and caspase-1 levels while inhibited the production of IL-1β. Besides, decreases in inflammatory cell infiltration in the lung were shown using H&E staining. MCC950 ameliorates sewing machine oil-induced acute exogenous lipoid pneumonia in rats through inhibition of the NF-[Formula: see text]B/NLRP3 inflammasome pathway.

Myocardial effects of a low-protein diet in experimental kidney dysfunction

THE AIM: to evaluate the effect of low protein diet supplemented with ketosteril on morphological and epigenomic changes in the myocardium of Wistar rats with simulated kidney dysfunction . MATERIALS AND METHODS. The work was performed on male Wistar rats subjected to 5/6 nephrectomy (NE). The first group after NE received a standard diet (20.16% animal protein ), the second – low protein diet (LPD), including 10% ketosteril . Control rats received a standard diet . After 4 months , blood pressure (BP) and left ventricular mass index (LVMI) were assessed in rats , and a histological examination of the myocardium was performed . In the myocardium , the relative expression levels of NF- kB , miRNA-21, miRNA-133, and miRNA-203 were determined . RESULTS. After 4 months in rats with NE on a standard diet , an increase in blood pressure , an increase in the mass index of the LV myocardium was recorded . MBD with the inclusion of 10% Ketosteril slowed down the growth of systolic blood pressure and the development of LV myocardial hypertrophy in rats with kidney dysfunction . At the histological level , the use of LPD provided a decrease in the degree of hypertrophy of cardiomyocytes and degenerative changes in cardiomyocytes . Animals treated with LPD had less pronounced diffuse and perivascular fibrosis compared with animals fed normal food . The use of MBD slowed down the increase in the relative level of expression of the NFκB gene and miRNA-21 in the myocardium of rats with NE and promoted an increase in the expression level of miRNA-133 and miRNA-203 compared to the indices of animals with NE that received standard food . CONCLUSION: long-term use of LPD with the use of ketoanalogues of essential amino acids can have a potential cardioprotective effect in CKD, slowing down the growth of blood pressure , an increase in LV myocardial mass and the formation of structural changes in the myocardium . It is possible that a decrease in the expression of NF- kB and miRNA-21, as well as an increase in the expression of miRNA-203 and 133 in the myocardium , can play a significant role in this .

EFFECT OF PERIOSTIN ON AUTOPHAGY AND FACTORS RELATED TO BONE METABOLISM IN OSTEOBLASTS

The purpose of this study was to explore the effect of periostin on autophagy and factors related to bone metabolism in osteoblasts. According to experimental design, some osteoblast-like cells (MC3T3-E1) were transfected by lentiviral vector to silence periostin gene. Celigo[Formula: see text] Image Cytometer and Real-Time PCR were used to assess target gene silencing effects. Western blotting was used to evaluate autophagy-associated factors (LC3B,Beclin1) and factors related to bone metabolism (RANKL, OPG, NF-[Formula: see text]B[Formula: see text]P65). Another part of osteoblasts was added with recombinant periostin protein at concentration of 75[Formula: see text]ng/mL. Monodansylcadaverine (MDC) detection was conducted to trace formation of autophagy vesicles. Factors associated with autophagy and bone metabolism were reassessed using western blotting. When periostin gene was silenced, western blotting analysis showed that the expression of LC3B-I, Beclin1, RANKL and NF-[Formula: see text]B[Formula: see text]P65 in experimental group was significantly increased; however, OPG expression was decreased. After adding recombinant periostin protein, MDC detection revealed significantly fewer green fluorescent particles in the experimental group. Western blotting results showed that the expression of LC3B, Beclin1, RANKL and NF-[Formula: see text]B[Formula: see text]P65 was decreased; however, OPG expression was increased. This study examined the regulatory mechanism of periostin in bone metabolism, which will provide a future reference for the treatment of primary teeth abnormal replacement caused by inflammation.

Hepatocyte Nuclear Factor 1α Proinflammatory Effect Linked to the Overexpression of Liver Nuclear Factor-κB in Experimental Model of Chronic Kidney Disease.

Chronic kidney disease (CKD) is associated with low-grade inflammation that activates nuclear factor–κB (NF–κB), which upregulates the expression of numerous NF–κB responsive genes, including the genes encoding IL-6, ICAM-1, VCAM-1, and MCP-1. Herein, we found the coordinated overexpression of genes encoding RelA/p65 (a subunit of NF–κB) and HNF1α in the livers of chronic renal failure (CRF) rats—an experimental model of CKD. The coordinated overexpression of RelA/p65 and HNF1α was associated with a significant increase in IL-6, ICAM-1, VCAM-1, and MCP-1 gene expressions. A positive correlation between liver RelA/p65 mRNA levels and a serum concentration of creatinine and BUN suggest that RelA/p65 gene transcription is tightly related to the progression of renal failure. The knockdown of HNF1α in the HepG2 cell line by siRNA led to a decrease in Rel A/p65 mRNA levels. This was associated with a decrease in IL-6, ICAM-1, VCAM-1, and MCP-1 gene expressions. The simultaneous repression of HNF-1α and RelA/p65 by clofibrate is tightly associated with the downregulation of IL-6, ICAM-1, VCAM-1, and MCP-1 gene expression. In conclusion, our findings suggest that NF–κB could be a downstream component of the HNF1α-initiated signaling pathway in the livers of CRF rats.

Expression of nuclear factor kappa B in ovine maternal inguinal lymph nodes during early pregnancy

BackgroundPregnancy-induced immunological changes contribute to the maternal immune tolerance. Nuclear factor kappa B (NF-κB) pathway participates in regulating both innate and adaptive immunities, and lymph nodes play key roles in adaptive immune reaction. However, it is unclear whether early pregnancy changes the expression of NF-κB family in maternal lymph node in sheep.MethodsIn this study, the samples of inguinal lymph nodes were collected from ewes on day 16 of the estrous cycle, and on days 13, 16 and 25 of pregnancy, and expression of NF-κB family, including NF-κB p105 (NFKB1), NF-κB p100 (NFKB2), p65 (RELA), RelB (RELB) and c-Rel (REL), were analyzed through real-time quantitative PCR, Western blot and immunohistochemical analysis.ResultsThe expression levels of NF-κB p105 and c-Rel downregulated, but NF-κB p100 upregulated on day 25 of pregnancy. The expression levels of p65, RelB and c-Rel peaked at day 13 of pregnancy, and expression level of RelB was higher during early pregnancy comparing to day 16 of the estrous cycle. In addition, p65 protein was located in the subcapsular sinus and lymph sinuses.ConclusionThis paper reported for the first time that early pregnancy has effects on the expression of NF-κB family, which may contribute to the maternal immunoregulation through blood circulation and lymph circulation during early pregnancy in sheep.

FGF1 alleviates LPS-induced acute lung injury via suppression of inflammation and oxidative stress

BackgroundAcute lung injury (ALI) and its severe form, acute respiratory distress syndrome (ARDS), are devastating clinical disorders with high mortality, and for which more effective therapies are urgently needed. FGF1, the prototype member of the FGF family, is shown to exert protective effects against injurious stimuli in multiple disease models. Here we aimed to evaluate whether FGF1 pretreatment is protective against LPS-induced ALI and elucidate the potential underlying mechanisms.MethodsFor drug-treated groups, C57B/6 mice received a single i.p. injection of FGF1 (1 mg/kg) 1 h before the LPS challenge or not. To induce the ALI model, the mice were treated by intratracheal instillation of LPS (5 mg/kg). Then, histopathological changes in lung tissues were assessed by hematoxylin and eosin staining and transmission electron microscopy. ELISA and qPCR assays were used to detect pro-inflammatory cytokine levels in BALF and lung tissues, respectively. The total number of inflammatory cells (neutrophils and macrophages) in BALF were counted using the Wright-Giemsa method. The expressions of reactive oxygen species (ROS) and malondialdehyde (MDA) were measured using their respective kits. Western blot and immunostaining were used to evaluate the expressions of antioxidants (Nrf-2, HO-1, SOD2, GPX4, and Catalase), as well as the inflammatory and/or apoptosis-related factors (TLR4, NF-κB, and Cleaved- caspase 3).ResultsFGF1 pretreatment significantly ameliorated the LPS-induced histopathological changes, reduced lung wet/dry ratios, ROS and MDA levels, total BALF protein, inflammatory cell infiltration, proinflammatory cytokine levels, and significantly increased the expression of antioxidant proteins (Nrf-2, HO-1, Catalase, and SOD2). In addition, FGF1 pretreatment significantly reduced the expression of TLR4 and cleaved- caspase 3, inhibited NF-κB activation, and reduced LPS-induced cell apoptosis.ConclusionsAltogether, our results suggest that FGF1 pretreatment is protective against LPS-induced ALI through mediating anti-inflammatory and antioxidant effects, which may be attributed to the downregulation of TLR4 expression and inhibition of NF-κB activation, as well as promotion of antioxidant defenses. Therefore, FGF1 administration may prove beneficial in preventative strategies for ALI/ARDS.

P221 DAPAGLIFLOZIN ASSOCIATED TO SACUBITRIL/VALSARTAN EXERTS ADDITIVE CARDIOPROTECTION IN HUMAN CARDIOMYOCYTES EXPOSED TO DOXORUBICIN AND TRASTUZUMAB THROUGH MYD88, NLRP3 MEDIATED PATHWAYS AND IMPROVEMENT OF MYTOGENESIS

Abstract Introduction The cumulative incidence of cardiac events in breast cancer patients treated with anthracycline and trastuzumab at 1 year after the diagnosis of cancer was 16.4%, at 2 years 23.8 %, and at 3 years 28.2%. Sodium glucose co–transporter 2 inhibitors showed measurable benefits in reduction of HF hospitalization and cardiovascular mortality. LCZ696 (neprilysin inhibitor + valsartan) as able to lower the risk of cardiovascular events in chronic heart failure. Purpose We hypothesize that dapagliflozin associated to LCZ696 could exerts cardioprotective effects in cellular models of doxorubicin and trastuzumab–induced cardiotoxicity Methods Human cardiomyocytes (HL–1 cells) were exposed to subclinical concentration of doxorubicin and trastuzumab (100 nM) alone or in combination with dapagliflozin (50 nM) or LCZ696 (at 100 mM) or both in combination for 48h. Cell viability, apoptosis and necrosis were performed. Quantification of MDA– 4–HNE and Ca2+ were performed through spectrophotometric methods. Anti–inflammatory studies were also performed (expression of NLRP3, TLR4/MyD88 pathways, nuclear expression of NF–kB). Intracellular concentration of IL–1α, IL–1β, IL–2, IL–4, IL–6, IL–8, IL–10, IL–12, IL17–α, IL–18, IFN–γ, TNF–α, G–CSF, and GM–CSF were also performed. Results Dapagliflozin and LCZ696 increased synergistically the cell viability during exposure to doxorubicin and trastuzumab. Combination of dapagliflozin and LCZ696 reduces Ca2+ overload (–68,4% vs cells treated only to anticancer drugs; p &amp;lt; 0,001) and MDA (mean reduction of 57–63,4 % compared to cells exposed only to anticancer drugs; p &amp;lt; 0,05). The expression of MyD88, NLRP3 and NF–kB were strongly reduced after treatment with dapagliflozin and LCZ–696 (–52,5, –43,7 and –57,3 % vs cells exposed only to anticancer drugs, respectively; p &amp;lt; 0.05). Notably, combination of dapagliflozin and LCZ–696 enhanced the expression of IL–10. Expression of IL–1α, IL–1β, IL–6, IL–8, IL17–α and IL–18 was reduced (p &amp;lt; 0.05). Mitogenesis was strictly improved also as demonstrated by confocal microscope analysis Conclusion Dapagliflozin associated to LCZ–696 exerts additive cardioprotective and anti–inflammatory effects compared to each drug alone. Their properties are mediated by the reduction of iCa2+ content that consequently reduces peroxidation and NLRP3– MyD88 expression. These results indicating the potential use of SGLT–2 inhibitors associated to LCZ696 in preclinical models of cardiotoxicity.

C49 SACUBITRIL–VALSARTAN IMPROVES LONGITUDINAL STRAIN AND EJECTION FRACTION IN MICE TREATED WITH DOXORUBICIN THROUGH NLRP3, MYD88 PATHWAYS RESULTING IN A REDUCTION OF MYOCARDIAL IL–1Β, IL–6, TNF–Α, G–CSF AND GM–CSF LEVELS

Abstract Introduction Doxorubicin–mediated adverse cardiovascular events are among the leading causes of morbidity and mortality in breast cancer patients. Sacubitril–valsartan (LCZ 696) is a combination drug, made up of neprilysin inhibitor sacubitril and angiotensin II receptor blocker valsartan, used for the treatment of heart failure in patients with a reduced ejection fraction. Purpose We hypothesized that LCZ 696, administered during doxorubicin, could improve cardiac function and cardiac inflammation in preclinical models Methods Human fetal cardiomyocytes (HFC cell line) were exposed to subclinical concentration of doxorubicin (200 nM) alone or in combination with LCZ–696 (100 mM) for 72 h. After the incubation period, we performed the following tests: cell viability, apoptosis and necrosis; expression of malondialdehyde and 4–hydroxynonenal and concentration of intracellular Ca2+. Moreover, pro–inflammatory studied were also performed (activation of NLRP3 inflammasome; expression of TLR4/MyD88; mTORC1 Fox01/3a; NF–κB). C57Bl/6 mice were untreated (Sham, n = 6) or treated for 10 days with doxorubicin i.p at 2.17 mg/kg (DOXO, n = 6), LCZ–696 at 60 mg/kg (LCZ, n = 6) or doxorubicin combined to LCZ–696 (DOXO–LCZ, n = 6). Ejection fraction, radial and longitudinal strain were analyzed through transthoracic echocardiography (Vevo 2100). Cardiac tissue expression of NLRP3 inflammasome, Myd88, NF–kB and 13 chemokines (IL–1α, IL–1β, IL–2, IL–4, IL–6, IL–10, IL–12, IL17–α, IL–18, IFN–γ, TNF–α, G–CSF, and GM–CSF) were quantified. Results LCZ–696 exerts cardioprotective effects, enhancing cell viability of 48–54.6% compared to only doxorubicin–treated cells (p &amp;lt; 0,001 for all); LCZ 696 decreased NLRP3, MyD88 and NF–kB expression in cardiac cells. In preclinical study, LCZ 696 improved significantly the EF and prevented the reduction of radial and longitudinal strain after 10 days of treatment with doxorubicin. A reduced expression of NLRP3, MyD88 and NF–kB in cardiac tissues was seen in DOXO–LCZ group compared to DOXO mice (p &amp;lt; 0.001). Cardiac expression of IL–1β, IL–6, TNF–α, G–CSF and GM–CSF were significantly reduced after treatment with LCZ–696 indicating anti–inflammatory properties. Conclusion LCZ–696 exerts direct beneficial effects in cardiomyocytes exposed to doxorubicin. In preclinical models, LCZ–696 reduced inflammation and cytokine expression involved in doxorubicin–mediated cardiotoxicity.