Neurotrophic Factor Mediated Neurorestorative Effect of Pongamia pinnata Against Transient Cerebral Hypoperfusion and Reperfusion in Rats: Preliminary Histological and Molecular Level Evidence / Efecto Neurorestaurador Mediado por Factor Neurotrófico de Pongamia pinnata Contra la Hipoperfusión y Reperfusión Cerebral Transitoria en Ratas: Evidencia Preliminar de un Estudio a Nivel Histológico y Molecular

SUMMARY: Stroke is the leading cause of acquired physical disability in adults and second leading cause of mortality throughout the world. Treatment strategies to curb the effects of stroke would be of great benefit. Pongamia pinnata is a recent attraction in medicine, owing to its abundant medicinal benefits with minimal side effects. The present study aimed to examine acute and subacute effect of Pongamia pinnata leaf extract on transient cerebral hypoperfusion and reperfusion (tCHR) in Wistar rats. 24 adult Wistar rats (12 each for acute and subacute study) were divided in to four groups each viz normal control group, tCHR + NS group, tCHR + 200mg/kg bw and tCHR + 400mg/kg bw groups. Cerebral ischemia induction was carried out by bilateral common carotid artery occlusion and reperfusion. Ethanolic extract of Pongamia pinnata leaves were orally administered for 7 days and 21 days after the surgical procedure for acute and subacute study respectively. Behavioural analysis, histological assessment, and estimation of mRNA levels of HIF-1, GDNF, BDNF and NF-kB were performed. In both acute and subacute study, there was significant improvement in the beam walking assay, neuronal count, decreased neuronal damage in histological sections and higher mRNA expression of BDNF and GDNF in the treatment groups. There was no significant difference in the expression of HIF1 and NF-kB. Thus, Pongamia pinnata has excellent neurorestorative property reversing many of the effects of ischemic stroke induced by tCHR in rats with the underlying mechanism being an improvement in the expression of neurotrophic factors GDNF and BDNF.

El ataque cerebrovascular es la principal causa de discapacidad física adquirida en adultos y la segunda causa de mortalidad en todo el mundo. Las estrategias de tratamiento para frenar los efectos del ataque cerebrovascular serían de gran beneficio. Pongamia pinnata es una atracción reciente en la medicina, debido a sus abundantes beneficios medicinales con mínimos efectos secundarios. El presente estudio tuvo como objetivo examinar el efecto agudo y subagudo del extracto de hoja de Pongamia pinnata sobre la hipoperfusión y reperfusión cerebral transitoria (tCHR) en ratas Wistar. Se dividieron 24 ratas Wistar adultas (12 cada una para el estudio agudo y subagudo) en cuatro grupos, el grupo control normal, el grupo tCHR + NS, los grupos tCHR + 200 mg/kg de peso corporal y tCHR + 400 mg/kg de peso corporal. La inducción de la isquemia cerebral se llevó a cabo mediante oclusión y reperfusión bilateral de la arteria carótida común. El extracto etanólico de hojas de Pongamia pinnata se administró por vía oral durante 7 días y 21 días después del procedimiento quirúrgico para estudio agudo y subagudo respectivamente. Se realizaron análisis de comportamiento, evaluación histológica y estimación de los niveles de ARNm de HIF-1, GDNF, BDNF y NF-kB. Tanto en el estudio agudo como en el subagudo, hubo una mejora significativa en el ensayo de desplazamiento del haz, el recuento neuronal, una disminución del daño neuronal en las secciones histológicas y una mayor expresión de ARNm de BDNF y GDNF en los grupos con tratamiento. No hubo diferencias significativas en la expresión de HIF1 y NF-kB. Por lo tanto, Pongamia pinnata tiene una excelente propiedad neurorestauradora que revierte muchos de los efectos del ataque cerebrovascular isquémico inducido por tCHR en ratas, siendo el mecanismo subyacente una mejora en la expresión de los factores neurotróficos GDNF y BDNF.

Columbianadin ameliorates experimental acute reflux esophagitis in rats via suppression of NF-κB pathway

Purpose: Reflux esophagitis is a condition characterized by inflammation and irritation of the esophagus, resulting from the backflow of stomach acid and other gastric contents into the esophagus. Columbianadin is a coumarin derivative that exhibits anti-inflammatory and antioxidant effects. In this study, we tried to scrutinize the protective effect of Columbianadin against acute reflux esophagitis in rats. Methods: RAW 264.7 cells were utilized to assess cell viability and measure the production of inflammatory parameters. The rats received anesthesia, and reflux esophagitis was induced via ligation of pylorus and fore stomach and corpus junction. Rats received the oral administration of Columbianadin (25, 50 and 100 mg/kg) and omeprazole (20 mg/kg). The gastric secretion volume, acidity, and pH were measured. Additionally, the levels of oxidative stress parameters, cytokines, and inflammatory markers were determined. At the end of the study, mRNA expression was assessed. Results: Columbianadin remarkably suppressed the cell viability and production of tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß, IL-6, cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), and prostaglandin (PGE2). Columbianadin treatment remarkably suppressed the secretion of gastric volume, total acidity and enhanced the pH level in the stomach. Columbianadin remarkably altered the level of hydrogen peroxidase, free iron, calcium, and plasma scavenging activity, sulfhydryl group; oxidative stress parameters like malonaldehyde, glutathione, superoxide dismutase, catalase, glutathione peroxidase; inflammatory cytokines viz., TNF-α, IL-6, IL-1ß, IL-10, IL-17, and monocyte chemoattractant protein-1; inflammatory parameters including PGE2, iNOS, COX-2, and nuclear kappa B factor (NF-κB). Columbianadin remarkably (P < 0.001) suppressed the mRNA expression TNF-α, IL-6, IL-1ß and plasminogen activator inhibitor-1. Conclusions: Columbianadin demonstrated a protective effect against acute reflux esophagitis via NF-κB pathway.

E-cadherin and NF-κB expression in the vagina after ovarian ischemia and reperfusion

Purpose: To investigate inflammation and cell adhesion molecules in the vagina after ovarian ischemia-reperfusion (IR) injury. Methods: 20 Wistar albino female rats were divided into two groups: control, and IR groups. In IR group, blood flow was restricted for 2 hours for ovarian ischemia. Then, tissues were re-blood 2 hours for reperfusion. Vagina tissues were excised and processed for histopathological analysis. Histopathological and biochemical follow-ups were performed. Results: Both malondialdehyde and myeloperoxidase values were increased in IR group compared to control group. Glutathione content was decreased in IR group compared to control group. Epithelial degeneration, inflammation, dilatation, and nuclear factor-κB (NF-κB) expression were increased in IR group compared to control group. E-cadherin expression was significantly decreased in IR group. In the IR group, E-cadherin showed a positive reaction in adenomas, gland-like cryptic structures, cellular junctions with clustered inflammatory cells. In the IR group, NF-κB expression was increased in basement membrane, inflammatory cells, in blood vessels. Conclusions: Ovarian ischemia caused degeneration of epithelial cells in the vaginal region and disruptions in the cell junction complex, which leads to activation of E-cadherin and NF-κB signaling pathway and alterations in reproductive and embryonal development in the vaginal region.

In vivo Gonadoprotective Effects of Myricetin on Cisplatin- Induced Testicular Damage via Suppression of TLR4/NF-kB Inflammation Pathway and Heat-Shock Response / Efectos Gonadoprotectores in vivo de la Miricetina sobre el Daño Testicular Inducido por Cisplatino Mediante la Supresión de la Vía de Inflamación TLR4/NF-kB y la Respuesta al Choque Térmico

SUMMARY: The aim of this study is to reveal the gonadoprotective effects of myricetin (MYC), which has many biological properties, on cisplatin (CP)-induced testicular damage in rats. For this purpose, 40 male Wistar albino rats were divided into 4 groups as Control (group given no treatment), MYC (group given 5 mg/kg/i.p myricetin for 7 days), CP (group given 7 mg/kg/i.p cisplatin at 7th day) and MYC + CP (group given 5 mg/kg/i.p myricetin for 7 days before 7 mg/kg/i.p cisplatin injection). After administrations, testicular tissues of animals were extracted and processed according to tissue processing protocol. Hematoxylin & Eosin staining were performed to evaluate the histopathological changes and Johnsen'sTesticular Biopsy Score (JTBS) was applied and mean seminiferous tubule diameters (MSTD) were measured to compare experimental groups in terms of histopathological changes. Moreover, TLR4, NF-kB, HSP70 and HSP90 expression levels were detected by immunohistochemical staining and the density of immunoreactivity were measured to determine the difference in the expression levels of these factors among groups. Additionally, testicular apoptosis was detected via TUNEL assay. JTBS and MSTD data were significantly lower in CP group compared to other groups and MYC administrations significantly protects testicular tissue against CP-induced damage. Moreover, TLR4, NF-kB, HSP70 and HSP90 expressions and apoptotic cells significantly increased in the CP group (p<0.05). However, MYC administrations exerted a strong gonadoprotective effect on testicular tissue in terms of these parameters in MYC+CP group (p<0.05). According to our results, we suggested that MYC can be considered as a protective agent against cisplatin-induced testicular damage.

El objetivo de este estudio es revelar los efectos gonadoprotectores de la miricetina (MYC), que tiene muchas propiedades biológicas, sobre el daño testicular inducido por cisplatino (CP) en ratas. Para este propósito, se dividieron 40 ratas albinas Wistar macho en 4 grupos: Control (grupo que no recibió tratamiento), MYC (grupo que recibió 5 mg/kg/i.p de miricetina durante 7 días), CP (grupo que recibió 7 mg/kg/i.p de cisplatino al séptimo día) y MYC + CP (grupo que recibió 5 mg/ kg/i.p de miricetina durante 7 días antes de la inyección de 7 mg/ kg/i.p de cisplatino). Después de las administraciones, se extrajeron y procesaron tejidos testiculares de animales según el protocolo de procesamiento de tejidos. Se realizó tinción con hematoxilina y eosina para evaluar los cambios histopatológicos y se aplicó la puntuación de biopsia testicular de Johnsen (JTBS) y se midieron los diámetros medios de los túbulos seminíferos (MSTD) para comparar los grupos experimentales en términos de cambios histopatológicos. Además, los niveles de expresión de TLR4, NF-kB, HSP70 y HSP90 se detectaron mediante tinción inmunohistoquímica y se midió la densidad de inmunorreactividad para determinar la diferencia en los niveles de expresión de estos factores entre los grupos. Además, se detectó apoptosis testicular mediante el ensayo TUNEL. Los datos de JTBS y MSTD fueron significativamente más bajos en el grupo CP en comparación con otros grupos y las administraciones de MYC protegen significativamente el tejido testicular contra el daño inducido por CP. Además, las expresiones de TLR4, NF-kB, HSP70 y HSP90 y las células apoptóticas aumentaron significativamente en el grupo CP (p<0,05). Sin embargo, las administraciones de MYC ejercieron un fuerte efecto gonadoprotector sobre el tejido testicular en términos de estos parámetros en el grupo MYC+CP (p<0,05). Según nuestros resultados, sugerimos que MYC puede considerarse como un agente protector contra el daño testicular inducido por cisplatino.

Diacerein mitigates renal ischemia/reperfusion injury via inhibition of renal inflammation, dendritic cells maturation and apoptosis: the role of TLR4/NF-kB/NLRP3/IL-1beta signaling pathway / La diacereína mitiga la lesión por isquemia/reperfusión renal a través de la inhibición de la inflamación renal, la maduración de las células dendríticas y la apoptosis: el papel de la vía de señalización TLR4/NF-kB/NLRP3/IL-1beta

SUMMARY: One of the reasons for acute kidney damage is renal ischemia. Nevertheless, there are limited protective and therapeutic approaches for this problem. Diacerein is an anti-inflammatory drug characterized by numerous biological activities. We aimed to determine the ameliorative impact of diacerein on renal ischemia/reperfusion injury (I/R) condition, exploring the underlying mechanisms. Twenty-four male rats were allotted into four groups (n= 6): sham group; Diacerein (DIA) group; I/R group, in which a non-crushing clamp occluded the left renal pedicle for 45 min, and the right kidney was nephrectomized for 5 min before the reperfusion process; I/R + diacerein group, injected intraperitoneally with 50 mg diacerein/kg i.m 30 minutes prior to I/R operation. Ischemia/ reperfusion was found to affect renal function and induce histopathological alterations. The flow cytometry analysis demonstrated an elevated expression of innate and mature dendritic cells in I/R renal tissues. Moreover, upregulation in the expression of the inflammatory genes (TLR4, Myd88, and NLRP3), and overexpression of the pro-inflammatory cytokines (IL-1β), apoptotic (caspase-3) and pyroptotic (caspase-1) markers were observed in I/R-experienced animals. The aforementioned deteriorations were mitigated by pre-I/R diacerein treatment. Diacerein alleviated I/R-induced inflammation and apoptosis. Thus, it could be a promising protective agent against I/R.

La isquemia renal es una de los motivos del daño renal agudo. Sin embargo, los enfoques protectores y terapéuticos para este problema son limitados. La diacereína es un fármaco antiinflamatorio caracterizado por numerosas actividades biológicas. Nuestro objetivo fue determinar el impacto de mejora de la diacereína en la condición de lesión por isquemia/ reperfusión renal (I/R), explorando los mecanismos subyacentes. Veinticuatro ratas macho se distribuyeron en cuatro grupos (n= 6): grupo simulado; grupo de diacereína (DIA); grupo I/R, en el que una pinza no aplastante ocluyó el pedículo renal izquierdo durante 45 min, y el riñón derecho fue nefrectomizado durante 5 min antes del proceso de reperfusión; Grupo I/R + diacereína, inyectado por vía intraperitoneal con 50 mg de diacereína/kg i.m. 30 min antes de la operación I/R. Se encontró que la isquemia/ reperfusión afecta la función renal e induce alteraciones histopatológicas. El análisis de citometría de flujo demostró una expresión elevada de células dendríticas innatas y maduras en tejidos renales I/R. Además, se observó una regulación positiva en la expresión de los genes inflamatorios (TLR4, Myd88 y NLRP3) y una sobreexpresión de las citoquinas proinflamatorias (IL-1β), marcadores apoptóticos (caspasa-3) y piroptóticos (caspasa-1) en animales con experiencia en I/R. Los deterioros antes mencionados fueron mitigados por el tratamiento previo a la diacereína I/R. La diacereína alivió la inflamación y la apoptosis inducidas por I/R. Por lo tanto, podría ser un agente protector prometedor contra I/R.

Over-expression of gastrin inhibits apoptosis of gastric cancer cells via reactive oxygen species and annexin A2-mediated mitochondrial dysfunction / La sobreexpresión de gastrina inhibe la apoptosis de las células de cáncer gástrico a través de especies reactivas de oxígeno y disfunción mitocondrial mediada por anexina A2

SUMMARY: Gastrin plays a vital role in the development and progression of gastric cancer (GC). Its expression is up-regulated in GC tissues and several GC cell lines. Yet, the underlying mechanism remains to be investigated. Here, we aim to investigate the role and mechanism of gastrin in GC proliferation. Gastrin-overexpressing GC cell model was constructed using SGC7901 cells. Then the differentially expressed proteins were identified by iTRAQ analysis. Next, we use flow cytometry and immunofluorescence to study the effect of gastrin on the mitochondrial potential and mitochondria-derived ROS production. Finally, we studied the underlying mechanism of gastrin regulating mitochondrial function using Co-IP, mass spectrometry and immunofluorescence. Overexpression of gastrin promoted GC cell proliferation in vitro and in vivo. A total of 173 proteins were expressed differently between the controls and gastrin- overexpression cells and most of these proteins were involved in tumorigenesis and cell proliferation. Among them, Cox17, Cox5B and ATP5J that were all localized to the mitochondrial respiratory chain were down-regulated in gastrin-overexpression cells. Furthermore, gastrin overexpression led to mitochondrial potential decrease and mitochondria-derived ROS increase. Additionally, gastrin-induced ROS generation resulted in the inhibition of cell apoptosis via activating NF-kB, inhibiting Bax expression and promoting Bcl-2 expression. Finally, we found gastrin interacted with mitochondrial membrane protein Annexin A2 using Co-IP and mass spectrometry. Overexpr ession of gastrin inhibits GC cell apoptosis by inducing mitochondrial dysfunction through interacting with mitochondrial protein Annexin A2, then up-regulating ROS production to activate NF-kB and further leading to Bax/Bcl-2 ratio decrease.

La gastrina juega un papel vital en el desarrollo y progresión del cáncer gástrico (CG). Su expresión está regulada al alza en tejidos de CG y en varias líneas celulares de CG. Sin embargo, el mecanismo subyacente aun no se ha investigado. El objetivo de este estudio fue investigar el papel y el mecanismo de la gastrina en la proliferación de CG. El modelo de células CG que sobre expresan gastrina se construyó usando células SGC7901. Luego, las proteínas expresadas diferencialmente se identificaron mediante análisis iTRAQ. A continuación, utilizamos la citometría de flujo y la inmunofluorescencia para estudiar el efecto de la gastrina en el potencial mitocondrial y la producción de ROS derivada de las mitocondrias. Finalmente, estudiamos el mecanismo subyacente de la gastrina que regula la función mitocondrial utilizando Co-IP, espectrometría de masas e inmunofluorescencia. La sobreexpresión de gastrina promovió la proliferación de células CG in vitro e in vivo. Un total de 173 proteínas se expresaron de manera diferente entre los controles y las células con sobreexpresión de gastrina y la mayoría de estas proteínas estaban implicadas en la tumorigenesis y la proliferación celular. Entre estas, Cox17, Cox5B y ATP5J, todas localizadas en la cadena respiratoria mitocondrial, estaban reguladas a la baja en las células con sobreexpresión de gastrina. Además, la sobreexpresión de gastrina provocó una disminución del potencial mitocondrial y un aumento de las ROS derivadas de las mitocondrias. Por otra parte, la generación de ROS inducida por gastrina resultó en la inhibición de la apoptosis celular mediante la activación de NF-kB, inhibiendo la expresión de Bax y promoviendo la expresión de Bcl-2. Finalmente, encontramos que la gastrina interactuaba con la proteína de membrana mitocondrial Anexina A2 usando Co-IP y espectrometría de masas. La sobreexpresión de gastrina inhibe la apoptosis de las células CG al inducir la disfunción mitocondrial a través de la interacción con la proteína mitocondrial Anexina A2, luego regula el aumento de la producción de ROS para activar NF-kB y conduce aún más a la disminución de la relación Bax/Bcl-2.

Quercetin and resveratrol are associated with the downregulation of TNFalpha/NF-kB/inos axis-mediated acute liver injury in rats induced by paracetamol poisoning / La quercetina y el resveratrol están asociados con la regulación decreciente de la lesión hepática aguda mediada por el eje TNF-alfa/NF-kB/inos en ratas inducida por envenenamiento con paracetamol

SUMMARY: Paracetamol (known as acetaminophen, or APAP) poisoning causes acute liver damage that can lead to organ failure and death. We sought to determine that APAP overdose can augment tumor necrosis factor-alpha (TNF-α)/ nuclear factor kappa B (NF-kB)/induced nitic oxide synthase (iNOS) axis-mediated hepatotoxicity in rats, and the anti-inflammatory polyphenolic compounds, quercetin (QUR) plus resveratrol (RES) can ameliorate these parameters. Therefore, we induced acute hepatotoxicity in rats using APAP overdose (2 g/kg, orally) and the protective group of rats were treated with 50 mg/kg QUR plus 30 mg/kg RES for one week before APAP ingestion. Animals were killed at day 8. APAP poisoning caused the induction of hepatic tissue levels of TNF-α, NF-kB, and iNOS, which were significantly (p<0.05) decreased by QUR+RES. QUR+RES, also inhibited liver injury biomarkers, alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Additionally, a link between liver injury and TNF-α /NF-kB / iNOS axis mediated hepatotoxicity was observed. Thus, the presented data backing the conclusion that intoxication by paracetamol increases TNF-α / NF-kB / iNOS axis -mediated hepatotoxicity, and is protected by a combination of quercetin and resveratrol.

El envenenamiento por paracetamol (conocido como acetaminofeno o APAP) causa daño hepático agudo que puede provocar una insuficiencia orgánica y la muerte. El objetivo de este trabajo fue determinar si la sobredosis de APAP puede aumentar la hepatotoxicidad mediada por el eje del factor de necrosis tumoral alfa (TNF-α)/factor nuclear kappa B (NF-kB)/óxido nítico sintasa inducida (iNOS) en ratas, y si el polifenólico antiinflamatorio compuesto por quercetina (QUR) más resveratrol (RES) pueden mejorar estos parámetros. Por lo tanto, inducimos hepatotoxicidad aguda en ratas usando una sobredosis de APAP (2 g/kg, por vía oral). El grupo protector de ratas se trató con 50 mg/ kg de QUR más 30 mg/kg de RES durante una semana antes de la ingestión de APAP. Los animales se sacrificaron el día 8. El envenenamiento con APAP en el tejido hepático provocó la inducción de niveles de TNF-α, NF-kB e iNOS, que se redujeron significativamente (p<0,05) con QUR+RES. QUR+RES, también inhibió los biomarcadores de daño hepático, la alanina aminotransferasa (ALT) y el aspartato aminotransferasa (AST). Además, se observó una relación entre la lesión hepática y la hepatotoxicidad mediada por el eje TNF-α /NF-kB/iNOS. Por lo tanto, los datos presentados respaldan la conclusión de que la intoxicación por paracetamol aumenta la hepatotoxicidad mediada por el eje TNF-α /NF-kB / iNOS, y está protegida por una combinación de quercetina y resveratrol.

The role of TNF-α and NFkß in an experimental model of intestinal carcinogenesis with 1, 2-dimethyhydrazine

Purpose: To analyze the potential of tumor necrosis factor-α (TNF-α) and factor nuclear kappa B (NF-κB) as colorectal cancer (CRC) biomarkers in an experimental model of intestinal carcinogenesis with 1,2-dimethyhydrazine (1,2-DMH). Methods: Twenty-four male Wistar rats were divided into two groups: sham and 1,2-DMH. First, 1,2-DMH (20 mg/kg/week) was administered for 15 consecutive weeks. In the 25th week, proctocolectomy was conducted. Histopathological analysis, immunohistochemistry, and gene expression of TNF-α and NF-κB were performed. Statistical analysis was performed using GraphPad Prism. The location of aberrant crypt foci (ACF) was analyzed by Kruskal-Wallis' test. For analyses with two groups with parametric data, the t-test was used; for non-parametric data, the Mann-Whitney's test was used. P < 0.05 was considered significant. Results: The number of ACF and macroscopic lesions was significantly higher (p < 0.5) in the 1,2-DMH group compared to the sham group, and most ACF were concentrated in the distal segment of the colon. There was a statistically significant increase (p < 0.5) in protein and gene expression of TNF-α and NF-κB in the 1,2-DMH group compared to the sham group. Conclusions: Our results provide supportive evidence that TNF-α and NF-κB pathways are strongly involved in CRC development in rats and might be used as early biomarkers of CRC pathogenesis in experimental studies.

Alpha-kinase1 promotes tubular injury and interstitial inflammation in diabetic nephropathy by canonical pyroptosis pathway

BACKGROUND: Alpha-kinase 1 (ALPK1) is a master regulator in inflammation and has been proved to promote renal fibrosis by promoting the production of IL-1ß in diabetic nephropathy (DN) mice. Pyroptosis is involved in high glucose (HG)-induced tubular cells injury, characterized by activation of Gasdermin D (GSDMD) and the release of IL-1ß and IL-18, resulting in inflammatory injury in DN. It is reasonable to assume that ALPK1 is involved in pyroptosis-related tubular injury in DN. However, the mechanism remains poorly defined. METHODS: Immunohistochemistry (IHC) staining was performed to detect the expression of pyroptosis- and fibrosis-related proteins in renal sections of DN patients and DN mice. DN models were induced through injection of streptozotocin combined with a high-fat diet. Protein levels of ALPK1, NF-κB, Caspase-1, GSDMD, IL-1ß, IL-18 and α-SMA were detected by Western blot. HK-2 cells treated with high-glucose (HG) served as an in vitro model. ALPK1 small interfering RNA (siRNA) was transfected into HK-2 cells to down-regulate ALPK1. The pyroptosis rates were determined by flow cytometry. The concentrations of IL-1ß and IL-18 were evaluated by ELISA kits. Immunofluorescence staining was used to observe translocation of NF-κB and GSDMD. RESULTS: The heat map of differentially expressed genes showed that ALPK1, Caspase-1 and GSDMD were upregulated in the DN group. The expression levels of ALPK1, Caspase-1, GSDMD and CD68 were increased in renal biopsy tissues of DN patients by IHC. ALPK1expression and CD68+ macrophages were positively correlated with tubular injury in DN patients. Western blot analysis showed increased expressions of ALPK1, phospho-NF-κB P65, GSDMD-NT, and IL-1ß in renal tissues of DN mice and HK-2 cells, accompanied with increased renal fibrosis-related proteins (FN, α-SMA) and macrophages infiltration in interstitial areas. Inhibition of ALPK1 attenuated HG-induced upregulation expressions of NF-κB, pyroptosis-related proteins Caspase-1, GSDMD-NT, IL-1ß, IL-18, α-SMA, and pyroptosis level in HK-2 cells. Also, the intensity and nuclear translocation of NF-κB and membranous translocation of GSDMD were ameliorated in HG-treated HK-2 cells after treatment with ALPK1 siRNA. CONCLUSIONS: Our data suggest that ALPK1/NF-κB pathway initiated canonical caspase-1-GSDMD pyroptosis pathway, resulting in tubular injury and interstitial inflammation of DN.

WD repeat domain 5 promotes the development of late-onset preeclampsia by activating nuclear factor kappa B

Purpose: Over-activation of nuclear factor kappa B (NF-κB) was proven to be involved in the pathogenesis of preeclampsia. However, its regulation mechanism is not clear yet. This paper explored the role of WD repeat domain 5 (WDR5) in the development of late-onset preeclampsia and its relationship with NF-κB. Methods: WDR5 expression was detected in normal placentas and placentas from late-onset preeclampsia patients. CCK-8 and colony formation assays were conducted to appraise the proliferative ability of trophoblast. Migration and invasion were observed by wound healing and transwell assays. The interaction between WDR5 and NF-κB inhibitor I-kappa-B-alpha (IkBa) was verified by Co-immunoprecipitation analysis. Immunofluorescence was used to analyze the activation of NF-κB. Finally, we tested the role of WDR5 using the mice late-onset preeclampsia model. Results: WDR5 was highly expressed in the placentas of late-onset preeclampsia patients. WDR5 overexpression suppressed cell proliferation, migration, and invasion in trophoblast. WDR5 could interact with IkBa to activate NF-κB. Knockdown of NF-κB counteracted the anti-proliferative and anti-metastatic effects of WDR5 overexpression in trophoblast. In-vivo studies suggested that targeting WDR5 combated late-onset preeclampsia development. Conclusions: Our finding provides new insights into the role of WDR5 in late-onset preeclampsia development.

In situ isolation of nuclei or nuclear proteins from adherent cells: a simple, effective method with less cytoplasmic contamination

BACKGROUND: Isolation of nuclei or nuclear proteins is a prerequisite for western blot, nuclear proteome profiling, and other evaluations of nuclear proteins. Here, we developed a simple method for in situ isolation of nuclei or nuclear proteins by in situ removing the extranuclear part of adherent cells via a classical nonionic detergent triton X-100. RESULTS: First, the feasibility of our method was confirmed by confocal microscopy, atomic force microscopy, scanning electron microscopy, dynamic light scattering, immunofluorescence imaging, and time-lapse dynamic observation. Next, the optimal concentration range (approximately 0.1-1% for ~ 10 min) of triton X-100 and the optimal treatment time (< 30 min) of 0.1-1% Triton X-100 for our method were determined via western blotting of eight extra-/ intra-nuclear proteins. Subsequently, the effectiveness, sensitivity, and cytoplasmic contamination of our method were tested by investigating the levels of phosphorylated p65 (a NF-κB subunit) in the nuclei of endothelial or tumor cells treated with/without lipopolysaccharide (LPS) via western blotting and by comparing with a commercial nuclear protein extraction kit (a classical detergent-based method). The data show that compared with the commercial kit our method obtained a higher yield of total nuclear proteins, a higher pP65 level in both control and LPS groups, and much lower content of GAPDH (as a reference for cytoplasmic contamination) in nuclei. CONCLUSIONS: The in situ isolation of nuclei or nuclear proteins from adherent cells in this study is a simple, effective method with less cytoplasmic contamination. This method/strategy has the potential of improving the quality of downstream evaluations including western blotting and proteomic profiling.

Avaliação da etapa final do sinal insulínico, via inflamatória, estresse oxidativo, mecanismo epigenético por metilação do DNA e expressão de GLUT4 no músculo gastrocnêmio de ratos adultos, proles de ratas com periodontite apical / Assessment of insulin signaling and inflammatory pathways in skeletal muscle and adipose tissues of adult rats, offspring of the rats with periapical lesion

A programação fetal sugere que estímulos adversos quando aplicados durante o início do desenvolvimento fetal podem alterar o metabolismo da prole, aumentando o risco de doenças na sua vida adulta. A metilação do DNA é um dos mecanismos epigenéticos envolvida nesta programação e regula a expressão gênica. Estudos anteriores verificaram que a periodontite apical (AP) materna em ratas promove em sua prole adulta: resistência insulínica (RI), alteração na etapa inicial do sinal insulínico (SI) no músculo gastrocnêmio (MG) e aumento na concentração plasmática de fator de necrose tumoral-α (TNF-α). O TNF-α pode ativar o fator de transcrição nuclear kappa B (NF-kB) que diminui a expressão gênica do transportador de glicose GLUT4. Nesse contexto, mais estudos são necessários para investigar se as alterações no SI observadas em ratos adultos, proles de ratas com AP também estão presentes na continuidade da cascata insulínica. Ademais, sabendo-se que o estresse oxidativo tem sido implicado como fator contribuinte tanto para o início quanto para a progressão do diabetes, torna-se fundamental verificar o grau de estresse oxidativo tecidual nesta prole adulta. Em vista disso, os objetivos deste estudo foram avaliar a RI, estresse oxidativo, etapa final do SI e via inflamatória no MG de ratos adultos, proles de ratas com AP. Para tanto, as 21 ratas Wistar (2 meses de idade) foram distribuídas em 3 grupos: 1) ratas controle; 2) ratas com 1 AP induzida em 1º molar superior direito; 3) ratas com 4 APs induzidas em 1os e 2os molares superiores e inferiores do lado direito. A AP foi induzida empregandose broca em aço carbono dotada de esfera de 0,1 mm na extremidade. Após 30 dias da exposição pulpar, as ratas de todos os grupos foram colocadas para acasalamento. Quando os filhotes machos de todas as ratas completaram 75 dias de idade, foram realizadas as seguintes análises plasmáticas: 1) glicemia; 2) insulinemia; 3) RI. Ademais, foram realizados os seguintes experimentos no MG: 1) grau de fosforilação em serina/treonina da Akt, após o estímulo insulínico; 2) conteúdo proteico de GLUT4; 3) grau de metilação do DNA na região promotora do gene do GLUT4; 4) expressão gênica do GLUT4 e TNF-α; 5) conteúdo proteico de TNF-α e PGC-1α; 6) grau de fosforilação de NF-kB p50 e NF-kB p65; 7) estresse oxidativo (superóxido dismutase ­ SOD e espécies reativas ao tiobarbitúrico ­ TBARS). A análise estatística foi realizada pela análise de variância, seguida do teste de Tukey. O nível de significância adotado foi de 5%. Os resultados mostraram que AP materna (tanto 1 foco de infecção como 4 focos de infecções) promoveu hiperinsulinemia, RI e diminuição na atividade antioxidante da SOD e na concentração de TBARS no MG de sua prole adulta. Entretanto, somente a AP materna em quatro dentes promoveu aumento na massa corpórea, na ingestão alimentar e no grau de fosforilação das subunidades p50 e p65 do NF-kB no MG de proles adultas. Ademais, houve diminuição no grau de fosforilação em serina e treonina da Akt (após estímulo insulínico) e na expressão gênica e conteúdo proteico de GLUT4 no MG em proles adultas de ratas com 4APs. A AP materna não promoveu alteração na glicemia de jejum, conteúdo de PGC-1α, grau de metilação do DNA na região promotora do gene do GLUT4 e expressão gênica e conteúdo proteico de TNF-α no MG de proles adultas. Esses resultados revelam o impacto que a AP materna tem em longo prazo na predisposição às alterações metabólicas na fase adulta da prole. Isso reforça a importância que a manutenção da saúde bucal materna tem sobre a saúde geral da prole(AU)

Fetal programming suggests that adverse stimuli when applied during early fetal development may alter metabolism of offspring, increasing the risk of disease in adulthood. DNA methylation is one of the epigenetic mechanisms involved in this programming and regulates gene expression. Previous studies have shown that maternal apical periodontitis (AP) in rats promotes in their adult offspring: insulin resistance (IR), alteration in the initial steps of the insulin signaling (IS) in the gastrocnemius muscle (GM) and increase in plasma concentration of tumor necrosis factor-α (TNF-α). TNF-α can activate nuclear transcription factor kappa B (NF-kB) thus decreasing gene expression of the GLUT4 glucose transporter. In this context, more studies are needed to investigate whether changes in IS observed in adult rats, offspring of rats with AP are also present in the continuity of the insulin cascade. In addition, since oxidative stress has been implicated as a contributing factor for both the onset and progression of diabetes, it is essential to verify the degree of tissue oxidative stress in this adult offspring. Therefore, the aims of this study were to evaluate IR, oxidative stress, final steps of IS and inflammatory pathway in the GM of adult rats, offspring of rats with AP. For this purpose, 21 female Wistar rats (2 months of age) were distributed into three groups: 1) control rats (CN); 2) group with one AP induced in the upper right first molar (1AP); 3) group with four APs induced in the first and second upper and lower right molars (4AP). AP was induced using a surgical round bur measuring 0.1 mm in diameter. After 30 days of pulp exposure, female rats of all groups were placed for mating. Pregnant rats were separated into individual cages. When the male offspring of all rats reached 75 days of age, the following plasma analyzes were performed: 1) glycemia; 2) insulinemia; 3) IR. Furthermore, the following analyzes were performed in the GM: 1) Akt serine/threonine phosphorylation status, after insulin stimulation; 2) GLUT4 content; 3) degree of DNA methylation in the proximal promoter region of the GLUT4 gene; 4) GLUT4 and TNF- α gene expression; 5) TNF-α and PGC-1α content; 6) p50 and p65 subunits of NF-kB phosphorylation status; 7) oxidative stress (superoxide dismutase ­ SOD and thiobarbiturate reactive species ­ TBARS). Statistical analyzes were performed by analysis of variance, followed by the Tukey test. The level of significance adopted was 5%. Results showed that maternal AP (both 1 infection focus and 4 infection focus) promoted hyperinsulinemia, IR and decrease in SOD antioxidant activity and TBARS concentration in the GM of their adult offspring. However, only maternal AP in four teeth promoted an increase in body weight, food intake and p50 and p65 subunits of NF-kB phosphorylation status in the GM of adult offspring. Furthermore, there was a decrease in Akt serine and threonine phosphorylation status (after insulin stimulation) and in gene expression and protein content of GLUT4 in the GM in adult offspring of rats with 4APs. Maternal AP did not change fasting glycemia, PGC-1α content, degree of DNA methylation in the proximal promoter region of GLUT4, gene expression and protein content of TNF-α in the GM of adult offspring. These results demonstrate that maternal AP is associated with IR and promotes important alterations in IS and inflammation pathways in adult offspring. This reinforces the importance of maternal oral health on the overall health of offspring(AU)

Campsiandra laurifolia reduces oxidative stress and Iiflammation in rats with ulcerative colitis

Ulcerative colitis (UC) affects the mucosa and submucosa of the large intestine. One of the mechanisms involved in its etiology is oxidative stress (OS), directly involved in the inflammatory process characteristic of UC. The Campsiandra laurifolia, known as acapurana, was described as possessing antioxidant properties. We used 24 male Wistar rats, divided into control (CO), control + acapurana (CO + A), colitis (CL), and colitis + acapurana (CL + A) groups. This study performed histological analysis, measuring anal sphincter pressure (ASP) and lipoperoxidation (LPO). The activity of the antioxidant enzyme superoxide dismutase (SOD) and glutathione (GSH) levels were evaluated. The expression of the nuclear factor kappa B (NFkB) and inducible nitric oxide synthase (iNOS) was analyzed by immunohistochemistry. The statistical analysis used was the one-way analysis of variance (ANOVA), followed by the Student-Newman-Keuls test; values were expressed as mean ± standard error, and the significance level was p < 0.05. In the animals of the CL group, we observed the destruction of the crypts and the presence of mucosal ulcers, edema, and submucosal inflammatory infiltrate, as well as increased damage to the intestinal mucosa, reduced ASP, increased LPO and SOD activity, reduced GSH levels, and increased expression of NFkB and iNOS. The administration of C. laurifolia in the CL + A group was shown to cause regeneration of crypts, reduction of inflammatory infiltrate, reduction of damage to the intestinal mucosa, increase in ASP, and reduction in LPO with the restoration of SOD activity and GSH levels. The immunohistochemistry of NFkB and iNOS was significantly reduced. Therefore, the C. laurifolia aqueous extract appears to exert an antioxidant and anti-inflammatory effect in rats with AA-induced colitis. (AU)

A Network Pharmacological Approach to Explore the Mechanisms of TongXieYaoFang in Inflammatory Bowel Disease

Abstract Inflammatory bowel disease (IBD) is a chronic inflammatory disorder of the intestine, demonstrating an increasing incidence every year. TongXieYaoFang (TXYF) has been used widely in China as a complementary therapy to relieve the symptoms of IBD for hundreds of years. In the present research, a network pharmacology-based approach was used to systematically explore the intrinsic mechanisms of TXYF in IBD at the molecular level. Network pharmacology-based methods, which mainly included database mining, screening of bioactive compounds, target prediction, collection of IBD-related targets, gene enrichment analysis, network construction, and molecular docking, were employed in the present study. Network analysis revealed a total of 108 potential targets derived from 22 component compounds of TXYF, among which 34 targets were common with the IBD-related targets. In the protein-protein interaction (PPI) network, 10 key targets were identified. The gene enrichment analysis suggested that anti-inflammatory processes, such as NF-kappa B signaling pathway and Toll-like receptor signaling pathway, could be the core processes involved in the action of TXYF in IBD. Molecular docking results revealed that three compounds present in TXYF exhibited strong binding affinity for PTGS2. The present study provides novel insights into the molecular mechanisms and network approaches of TXYF action in IBD from a systemic perspective. The potential targets and pathways identified in the present study would assist in further research on the clinical application of TXYF in IBD therapy.

Effect of trans-resveratrol on oxidative stress biomarkers associated with atherosclerosis / Efeito do trans-resveratrol em biomarcadores de estresse oxidativo associados à aterosclerose

Growing evidence indicates that oxidative stress plays an important role in the pathophysiology of many cardiovascular diseases, including atherosclerosis. In this context, the use of bioactive compounds with antioxidant action can bring health benefits, especially in the prevention and control of pathophysiological events. Studies suggest that the polyphenol trans-resveratrol can reduce oxidative stress by acting on the nuclear factor erythroid-2-related factor 2 (Nrf2) and this effect would be associated with dosage. Thus, the present study aimed to investigate the effect of different doses of trans-resveratrol on biomarkers related to atherosclerosis and oxidative stress. In the first step, 27 randomized clinical trials, which evaluated the effect of trans-resveratrol on atherosclerosis-related biomarkers, were classified according to their protocol characteristics and profile of each outcome. Biochemical data from 12 biomarkers were selected to calculate the net change (%). Using multivariate analysis, the trials were distributed into 3 clusters. The studies that composed Clusters II and III were more effective in improving blood pressure and reducing dyslipidemia, respectively. These studies were characterized by a longer intervention time (> 2 months) with doses of around 200-500 mg/day. These results showed that the effects of transresveratrol are mainly related to dosage and intervention time. Based on these results, two doses were selected to apply in an experimental protocol to investigate the effect of trans-resveratrol on hepatic oxidative stress biomarkers mediated by Nrf2 pathway. LDLr(-/-) mice were fed for 8 weeks on a standard diet, followed by over 24 weeks on a Western diet, both containing trans-resveratrol at doses of 250 mg/kg diet/day (low dose resveratrol, LRD) or 400 mg/kg diet/day (high dose resveratrol, HRD). A control group (CONT) was maintained without supplementation. In general, both doses of trans-resveratrol did not affect the body weight and lipid profile of the animals. Only the LRD group showed reduced levels of two important biomarkers of oxidative stress in the liver (GSH/GSSG ratio and malonaldehyde), besides to reduced expression of factor nuclear kappa B (NF-kB). However, contrary to our hypothesis, both doses reduced Nrf2 expression in the liver compared to the CONT group. Regarding inflammatory cytokines, no changes were observed in the levels of TNF-α and IL-10. Furthermore, both doses increased the level of the pro-inflammatory cytokine IL-6. Taken together, our results suggest that trans-resveratrol supplementation at doses lower than 500 mg/day may contribute to the reduction of biomarkers related to atherosclerosis and oxidative stress

Evidências crescentes indicam que o estresse oxidativo desempenha um papel importante na fisiopatologia de muitas doenças cardiovasculares, incluindo a aterosclerose. Nesse contexto, o uso de compostos bioativos com ação antioxidante pode trazer benefícios à saúde, principalmente na prevenção e controle de eventos fisiopatológicos. Estudos sugerem que o polifenol trans-resveratrol pode reduzir o estresse oxidativo atuando na via do fator nuclear eritroide 2 relacionado ao fator 2 (Nrf2) e que esse efeito estaria associado a dosagem. Assim, o presente estudo teve como objetivo investigar o efeito de diferentes doses de trans-resveratrol sobre biomarcadores relacionados à aterosclerose e estresse oxidativo. Na primeira etapa, 27 ensaios clínicos randomizados, que avaliaram o efeito do trans-resveratrol em biomarcadores relacionados à aterosclerose, foram classificados de acordo com suas características de protocolo e perfil de cada resultado. Dados bioquímicos de 12 biomarcadores foram selecionados para calcular a variação líquida (%). Usando análise multivariada, os ensaios foram distribuídos em 3 Clusters. Os estudos que compuseram os Clusters II e III foram mais eficazes na melhora da pressão arterial e na redução da dislipidemia, respectivamente. Esses estudos foram caracterizados por um tempo de intervenção mais longo (> 2 meses) com doses de cerca de 200-500 mg/dia. Esses resultados mostraram que os efeitos do transresveratrol estão relacionados principalmente à dosagem e ao tempo de intervenção. Com base nesses resultados, duas doses foram selecionadas para aplicar em um protocolo experimental para investigar o efeito do trans-resveratrol em biomarcadores de estresse oxidativo hepático mediados pela via do Nrf2. Camundongos LDLr(-/-) foram alimentados por 8 semanas com dieta padrão, seguidos por mais de 24 semanas com Western diet, ambos contendo trans-resveratrol nas doses de 250 mg/kg de dieta/dia (baixa dose de resveratrol, LRD) ou 400 mg/kg de dieta/dia (alta dose de resveratrol, HRD). Um grupo controle (CONT) foi mantido sem suplementação. Em geral, ambas as doses de trans-resveratrol não afetaram o peso corporal e o perfil lipídico dos animais. Apenas o grupo LRD apresentou níveis reduzidos de dois importantes biomarcadores de estresse oxidativo no fígado (razão GSH/GSSG e malonaldeído), além da redução da expressão de fator nuclear kappa B (NF-kB). No entanto, ao contrário da nossa hipótese, ambas as doses reduziram a expressão de Nrf2 no fígado em comparação com o grupo CONT. Em relação às citocinas inflamatórias, não foram observadas alterações nos níveis de TNF-α e IL-10. Além disso, ambas as doses aumentaram o nível da citocina pró-inflamatória IL-6. Em conjunto, nossos resultados sugerem que a suplementação de trans-resveratrol em doses menores de 500 mg/dia podem contribuir para a redução de biomarcadores relacionados à aterosclerose e ao estresse oxidativo

Histological investigation of experimentally induced diabetes effects on the distribution of transforming growth factor (TGF beta), nuclear factor kappa b (Nf-KB), heat schock 90 beta (Hsp beta) and E-cadherin proteins in testicular tissue / Investigación histológica de los efectos de la diabetes inducida experimentalmente en la distribución del factor de crecimiento transformante (TGFß), nuclear factor kappa b (Nf-KB), y proteinas heat schock 90ß (Hsp90ß) y E-cadherina en tejido testicular

SUMMARY: Diabetes is a metabolic disorder characterized by high blood sugar levels and it causes complications in many systems, including the reproductive system. As a result of diabetic conditions, one of the mechanisms that can cause repression of reproductive activity is testicular oxidant stress. The identification of diabetes on the cell signaling molecules axis is still under discussion. The aim of this study was to determine the effect of Transforming Growth Factor (TGFβ), Nuclear Factor kappa B (NF-kB), Heat-schock 90β (HSP90β) signal pathways and E-cadherin cell adhesion molecule on infertility in diabetic rat testicular tissue. In our study, includes histological, molecular and biochemical analysis of testicular tissue removed at the end of the 2 weeks experiment period. A total of 14 adult male rats were divided as control and diabetes. No intervention was given to 7 male rats in the control group. For the diabetic group, 7 male rats were injected by intraperitoneal with a single dose of 55 mg/kg streptozotocin (STZ). TGFβ, NF-kB, HSP90β and E-cadherin proteins were immunohistochemically studied to investigate possible tissue damage, inflammatory process, cell stabilization and integrity due to diabetes. In order to determine oxidant stress, lipid peroxidation product malondialdehyde (MDA), glutathione (GSH) and glutathione peroxidase (GPx) analyzes were performed. Fibrosis, inflammatory changes and loss of spermatogenetic series are prominent findings in the diabetic group. On analysis of all the samples with immunostaining, in the diabetic group, TGFβ and NF-kB immunoexpression significantly increased, while Hsp90β and E-cadherin immunoexpression significantly decreased compared with control groups. Experimental diabetes was found to cause fibrosis, inflammation, disrupting cell adhesion and stabilization in testicular tissue. These results suggest that cellular therapy studies are needed for possible damage.

RESUMEN: La diabetes es una enfermedad metabólica caracterizada por niveles altos de azúcar en sangre y causa complicaciones en muchos sistemas, incluido el sistema reproductivo. Como resultado de las condiciones diabéticas, uno de los mecanismos que puede causar alteraciones en la actividad reproductiva es el estrés oxidativo testicular. La identificación de la diabetes en el eje de las moléculas de señalización celular aún está en discusión. El objetivo de este estudio fue determinar el efecto del factor de crecimiento transformante (TGFβ), el factor nuclear kappa B (NF-kB), las vías de señalización de Heat-Schock 90b (HSP90β) y la molécula de adhesión celular de E-cadherina sobre la infertilidad en testículo de rata diabética. Al término de dos semanas se realizaron análisis histológico, molecular y bioquímico del tejido testicular extraído. Las 7 ratas macho del grupo control no fueron intervenidas. Para el grupo de diabéticos, 7 ratas macho fueron inyectadas por vía intraperitoneal con una dosis única de 55 mg / kg de estreptozotocina (STZ). Se estudiaron inmunohistoquímicamente las proteínas TGFβ, NF-kB, HSP90β y E-cadherina para investigar el posible daño tisular, el proceso inflamatorio, la estabilización celular y la integridad debido a la diabetes. Para determinar el estrés oxidativo, se realizaron análisis del producto de peroxidación lipídica malondialdehído (MDA), glutatión (GSH) y glutatión peroxidasa (GPx). La fibrosis, los cambios inflamatorios y la pérdida de series espermatogenéticas son hallazgos destacados en el grupo de ratas diabéticas. En el análisis de todas las muestras con inmunotinción, en el grupo diabético, la inmunoexpresión de TGFβ y NF-kB aumentó significativamente, mientras que la inmunoexpresión de Hsp90β y e-cadherina disminuyó significativamente en comparación con los grupos control. Se encontró que la diabetes experimental causa fibrosis, inflamación, alteración de la adhesión celular y estabilización en el tejido testicular. Estos resultados sugieren que son necesarios estudios de terapia celular para verificar posibles daños.

Oxidized low-density lipoprotein stimulates dendritic cells maturation via LOX-1-mediated MAPK/NF-κB pathway

Dendritic cells (DCs) play a crucial role as central orchestrators of immune system response in atherosclerosis initiation and progression. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is involved in the immune maturation of DCs, but the underlying mechanisms remain unclear. We isolated mouse bone marrow progenitors and stimulated them with granulocyte-macrophage colony-stimulating factor and interleukin (IL)-4 to induce immature DCs. We then treated DCs with oxidized low-density lipoprotein (oxLDL) to induce maturation. LOX-1 siRNA was used to investigate the modulation of LOX-1 on the development of DCs and the underlying signal pathways. CD11c-positive DCs were successfully derived from mouse bone marrow progenitors. OxLDL promoted the expressions of DCs maturation markers and pro-inflammatory cytokines. OxLDL also upregulated LOX-1 expression and activated MAPK/NF-κB pathways. LOX-1 siRNA could attenuate the expression of MAPK/NF-κB pathways and inflammatory cytokines. In conclusion, oxLDL induced the maturation of DCs via LOX-1-mediated MAPK/NF-κB pathway, which contributed to the initiation and progression of atherosclerosis.

miR-139-5p protects septic mice with acute lung injury by inhibiting Toll-like receptor 4/Myeloid differentiation factor 88/Nuclear factor-&mac_kgr; B signaling pathway

OBJECTIVES: To investigate the role of miR-139-5p and the TLR4/MyD88/NF-κB signaling pathway in acute lung injury in septic mice. METHOD: A total of 140 healthy male SPF C57BL/6 mice were divided into seven groups, i.e., Normal, Control, NC, miR-139-5p mimic, miR-139-5p inhibitor, TAK-242, and miR-139-5p inhibitor+TAK-242 groups. The levels of miR-139-5p, proteins related to the TLR4/MyD88/NF-κB signaling pathway (TLR4, MyD88, and p-NF-κB p50), and MPO, SOD, GSH, and MDA in lung tissue were measured. The lung tissue wet-to-dry mass ratio (W/D), arterial oxygen partial pressure (PaO2), and carbon dioxide partial pressure (PaCO2) were measured. RESULTS: A web-based bioinformatic tool predicted that MyD88 was a target of miR-139-5p, which was verified by a dual luciferase reporter assay. Compared with those in the Normal group, the levels of miR-139-5p, PaO2, SOD, and GSH were significantly lower, while those of TLR4, MyD88, p-NF-κB p50, W/D, PaCO2, IL-1β, TNF-α, IL-6, MPO, and MDA were higher in all other groups. Moreover, compared with their levels in the Control group, these indicators exhibited contrasting results in the miR-139-5p mimic and TAK-242 groups, but were similar in the miR-139-5p inhibitor group. In the miR-139-5p inhibitor+TAK-242 group, acute lung injury, aggravated by miR-139-5p inhibitor, was partially rescued by TAK-242. CONCLUSION: miR-139-5p inhibits the TLR4/MyD88/NF-κB signaling pathway to alleviate acute lung injury in septic mice.

Beneficial role of oleuropein in sepsis-induced myocardial injury. Possible Involvement of GSK-3β/NF-kB pathway

ABSTRACT Purpose The present study explored the potential therapeutic role of oleuropein in sepsis-induced heart injury along with the role of GSK-3β/NF-kB signaling pathway. Methods Sepsis-induced myocardial injury was induced by cecal ligation and puncture (CLP) in rats. The cardiac injury was assessed by measuring the levels of cTnI and creatine kinase-MB (CK-MB). Sepsis-induced inflammation was assessed by measuring interleukin-6 (IL-6), IL-10 and HMGB1 levels. The different doses of oleuropein (5, 10, and 20 mg/kg) were given prior to CLP. Oleuropein (20 mg/kg) was administered after 6 hof CLP. The expressions of GSK-3β, p-GSK-3β (Ser9) and nuclear factor-κB (NF-κB) were measured in heart homogenates. Results Cecal ligation and puncture was associated with myocardial injury, an increase in IL-6, a decrease in IL-10 and an increase in HMGB1. Moreover, it decreased the ratio of p-GSK-3β/GSK-3β and increased the expression of p-NF-kB. Pretreatment with oleuropein attenuated CLP-induced myocardial injury and systemic inflammation in a dose-dependent manner. Administration of oleuropein after the onset of CLP also attenuated cardiac injury and inflammation. It also restored CLP-induced changes in the HMGB1 levels, the ratio of p-GSK-3β/GSK-3β and expression of p- NF-kB. Conclusions Oleuropein attenuates sepsis-induced systemic inflammation and myocardial injury by inhibiting NF-kB and GSK-3β signaling.

Effect of IL-33 on pyroptosis of macrophages in mice with sepsis via NF-κB/p38 MAPK signaling pathway

ABSTRACT Purpose To demonstrate the effect of IL-33 on the macrophage pyroptosis in mice with sepsis through the NF-kB/p38 MAPK signal pathway. Methods In total, 24 C57BL/6 mice were divided into the sham operation group (sham) and the cecal ligation and puncture group (CLP). After CLP, 24 IL-33-/- mice were divided into the IL-33-/- group and the IL-33-/- intervention group. The latter group was intraperitoneally injected with IL-33. Mouse mortality was observed after CLP. Macrophage apoptosis in peritoneal lavage fluid was detected by flow cytometry. Serum inflammatory factor level was detected by ELISA. Apoptotic protein expression and NF-κB/p38 MAKP signaling pathway protein expression were detected by qRT-PCR and Western blot. Results Knocking out IL-33 significantly reduced the mortality of CLP mice, as well as the mRNA expression of IL-33 and the levels of serum inflammatory factors, including IL-33, IL-1β, and IL-18. It also reduced the rate of macrophage apoptosis and the expression of the apoptotic protein caspase-1 p10; increased the expression of IκBα; and reduced the protein expression of NF-κB and p38 MAPK. These effects were reversed after exogenous injection of IL-33. Conclusions IL-33 can increase the level of macrophage pyroptosis in mice with sepsis (by activating the NF-kB/p38MAPK signal pathway) and the mortality of these mice.