The Neuroprotective Effect of Erythropoietin on the Optic Nerve and Spinal Cord in Rats with Experimental Autoimmune Encephalomyelitis through the Activation of the Extracellular Signal-Regulated Kinase 1/2 Signaling Pathway.

Experimental autoimmune encephalomyelitis is a demyelinating disease that causes paralysis in laboratory rats. This condition lacks treatment that reverses damage to the myelin sheaths of neuronal cells. Therefore, in this study, treatment with EPO as a neuroprotective effect was established to evaluate the ERK 1/2 signaling pathway and its participation in the EAE model. EPO was administered in 5000 U/Kg Sprague Dawley rats. U0126 was used as an inhibitor of the ERK 1/2 pathway to demonstrate the possible activation of this pathway in the model. Spinal cord and optic nerve tissues were evaluated using staining techniques such as H&E and the Luxol Fast Blue myelin-specific technique, as well as immunohistochemistry of the ERK 1/2 protein. The EPO-treated groups showed a decrease in cellular sampling in the spinal cord tissues but mainly in the optic nerve, as well as an increase in the expression of the ERK 1/2 protein in both tissues. The findings of this study suggest that EPO treatment reduces cellular death in EAE-induced rats by regulating the ERK pathway.

Activation of angiotensin II type 2 receptor leads to preservation of primary cilia in tubular cells during renal ischaemia-reperfusion injury.

Ischaemia-reperfusion (IR)-associated acute kidney injury (AKI) is a severe clinical condition that lacks effective pharmacological treatments. Our recent research revealed that pretreatment with the angiotensin II type 2 receptor (AT2R) agonist C21 alleviates kidney damage during IR. Primary cilia are organelles crucial for regulation of epithelial cell homeostasis, which are significantly affected by IR injury. This study aimed to evaluate the impact of AT2R activation on cilia integrity during IR and to identify pathways involved in the nephroprotective effect of C21. Rats were subjected to 40 min of unilateral ischaemia followed by 24 h of reperfusion. Immunofluorescence analysis of the kidneys showed that the nephroprotective effect of C21 was associated with preservation of cilia integrity in tubular cells. AT2R agonists increased α-tubulin acetylation in primary cilia in tubular cells in vivo and in a cell model. Analysis of ERK phosphorylation indicated that AT2R activation led to diminished activation of ERK1/2 in tubular cells. Similar to AT2R agonists, inhibitors of α-tubulin deacetylase HDAC6 or inhibitors of ERK activation ameliorated IR-induced cell death and preserved cilia integrity. Immunofluorescence analysis of tubular cells revealed significant ERK localization at primary cilia and demonstrated that ERK inhibition increased cilia levels of acetylated α-tubulin. Overall, our findings demonstrate that C21 elicits a preconditioning effect that enhances cilia stability in renal tubular cells, thereby preserving their integrity when exposed to IR injury. Furthermore, our results indicate that this effect might be mediated by AT2R-induced inhibition of ERK activation. These findings offer potential insights for the development of pharmacological interventions to mitigate IR-associated AKI. KEY POINTS: The AT2R agonist C21 prevents primary cilia shortening and tubular cell deciliation during renal ischaemia-reperfusion. AT2R activation inhibits ERK1/2 in renal tubular cells. Both AT2R agonists and ERK1/2 inhibitors increase alpha-tubulin acetylation at the primary cilium in tubular cells. AT2R activation, ERK1/2 inhibition or inhibition of alpha-tubulin deacetylation elicit protective effects in tubular cells subjected to ischaemia-reperfusion injury.

Anadenanthera colubrina regulated LPS-induced inflammation by suppressing NF-κB and p38-MAPK signaling pathways.

We aimed to determine the chemical profile and unveil Anadenanthera colubrina (Vell.) Brenan standardized extract effects on inflammatory cytokines expression and key proteins from immunoregulating signaling pathways on LPS-induced THP-1 monocyte. Using the RT-PCR and Luminex Assays, we planned to show the gene expression and the levels of IL-8, IL-1ß, and IL-10 inflammatory cytokines. Key proteins of NF-κB and MAPK transduction signaling pathways (NF-κB, p-38, p-NF-κB, and p-p38) were detected by Simple Western. Using HPLC-ESI-MSn (High-Performance Liquid-Chromatography) and HPLC-HRESIMS, we showed the profile of the extract that includes an opus of flavonoids, including the catechins, quercetin, kaempferol, and the proanthocyanidins. Cell viability was unaffected up to 250 µg/mL of the extract (LD50 = 978.7 µg/mL). Thereafter, the extract's impact on the cytokine became clear. Upon LPS stimuli, in the presence of the extract, gene expression of IL-1ß and IL-10 were downregulated and the cytokines expression of IL-1ß and IL-10 were down an upregulated respectively. The extract is involved in TLR-4-related NF-κB/MAPK pathways; it ignited phosphorylation of p38 and NF-κB, orchestrating a reduced signal intensity. Therefore, Anadenanthera colubrina's showed low cytotoxicity and profound influence as a protector against the inflammation, modulating IL-1ß and IL-10 inflammatory cytokines gene expression and secretion by regulating intracellular NF-κB and p38-MAPK signaling pathways.

In vitro effects and mechanisms of Humulus lupulus extract on bone marrow progenitor cells and endothelial cells.

Osteoporosis is the most common metabolic bone disorder and is associated with a high incidence of fractures. Angiogenesis and adequate blood flow are important during bone repair and maintenance. Estrogens play a key role in bone formation, in the prevention of bone resorption and vasculature maintenance. Hormone replacement therapy (HRT) has been used with great benefits for bone fracture prevention but has been linked to the development of serious important side effects, including cancer and stroke. Phytoestrogens are an attractive alternative to HRT because their chemical structure is similar to estradiol but, they could behave as selective modulators: acting as antagonists of estrogen receptors in the breast and endometrium and as agonists in the vascular endothelium and bone. Hops contain a wide variety of phytoestrogens that have individually been shown to possess estrogenic activity by either blocking or mimicking. In this study we have to evaluate the in vitro effects and mechanisms of action of hops extracts on the osteogenic and adipogenic capacity of bone marrow progenitor cells (BMPCs), and the angiogenic potential of EA.hy926 endothelial cells. We show that hops extracts increase the proliferative capacity of BMPCs and promote their osteogenic differentiation while decreasing their pro-osteoclastogenic capacity; and that these effects are mediated by the MAPK pathway. Additionally, hops extracts prevent the adipogenic differentiation of BMPCs and promote endothelial cell activity, by mechanisms also partially mediated by MAPK.

Puerarin inhibits HDAC1-induced oxidative stress disorder by activating JNK pathway and alleviates acrolein-induced atherosclerosis.

OBJECTIVE: Atherosclerosis (AS) is a common pathogenesis of cardiovascular diseases. Puerarin (Pue) is a Chinese herbal remedy used to prevent and treat AS. Here, this research investigated the effect of Pue on AS progression. METHODS: ApoE-/- mice were induced with acrolein. Body weight, blood lipid index, inflammatory factors, mitochondrial oxidative stress, and lipid deposition were detected. IL-6 and TNF-α were detected by ELISA. Oil red staining and H&E staining were used to observe the aortic sinus plaque lesions. Serum expressions of inflammatory factors IL-6, TNF-a, SOD, GSH and MDA were detected by ELISA, the mRNA expression levels of HDAC1 in the aorta were detected by RT-qPCR, and IL-6 and TNF-α in the aorta were detected by immunohistochemistry. JNK, p-JNK, OPA-1, and HDAC1 were detected by Western blotting. RESULTS: Pue administration can effectively reduce lipid accumulation in AS mice induced by acrolein. Pue promoted the activity of SOD, GSH and MDA, and inhibited the formation of atherosclerotic plaques and the process of aortic histological changes. Pue reduced IL-6 and TNF-α. HDAC1 expression was down-regulated and p-JNK-1 and JNK protein expression was up-regulated. CONCLUSION: Pue reduces inflammation and alleviates AS induced by acrolein by mediating the JNK pathway to inhibit HDAC1-mediated oxidative stress disorder.

JNK signaling and its impact on neural cell maturation and differentiation.

C-Jun-N-terminal-kinases (JNKs), members of the mitogen-activated-protein-kinase family, are significantly linked with neurological and neurodegenerative pathologies and cancer progression. However, JNKs serve key roles under physiological conditions, particularly within the central-nervous-system (CNS), where they are critical in governing neural proliferation and differentiation during both embryogenesis and adult stages. These processes control the development of CNS, avoiding neurodevelopment disorders. JNK are key to maintain the proper activity of neural-stem-cells (NSC) and neural-progenitors (NPC) that exist in adults, which keep the convenient brain plasticity and homeostasis. This review underscores how the interaction of JNK with upstream and downstream molecules acts as a regulatory mechanism to manage the self-renewal capacity and differentiation of NSC/NPC during CNS development and in adult neurogenic niches. Evidence suggests that JNK is reliant on non-canonical Wnt components, Fbw7-ubiquitin-ligase, and WDR62-scaffold-protein, regulating substrates such as transcription factors and cytoskeletal proteins. Therefore, understanding which pathways and molecules interact with JNK will bring knowledge on how JNK activation orchestrates neuronal processes that occur in CNS development and brain disorders.

Coenzyme Q10 prevents RANKL-induced osteoclastogenesis by promoting autophagy via inactivation of the PI3K/AKT/mTOR and MAPK pathways.

Coenzyme Q10 (CoQ10) is a potent antioxidant that is implicated in the inhibition of osteoclastogenesis, but the underlying mechanism has not been determined. We explored the underlying molecular mechanisms involved in this process. RAW264.7 cells received receptor activator of NF-κB ligand (RANKL) and CoQ10, after which the differentiation and viability of osteoclasts were assessed. After the cells were treated with CoQ10 and/or H2O2 and RANKL, the levels of reactive oxygen species (ROS) and proteins involved in the PI3K/AKT/mTOR and MAPK pathways and autophagy were tested. Moreover, after the cells were pretreated with or without inhibitors of the two pathways or with the mitophagy agonist, the levels of autophagy-related proteins and osteoclast markers were measured. CoQ10 significantly decreased the number of TRAP-positive cells and the level of ROS but had no significant impact on cell viability. The relative phosphorylation levels of PI3K, AKT, mTOR, ERK, and p38 were significantly reduced, but the levels of FOXO3/LC3/Beclin1 were significantly augmented. Moreover, the levels of FOXO3/LC3/Beclin1 were significantly increased by the inhibitors and mitophagy agonist, while the levels of osteoclast markers showed the opposite results. Our data showed that CoQ10 prevented RANKL-induced osteoclastogenesis by promoting autophagy via inactivation of the PI3K/AKT/mTOR and MAPK pathways in RAW264.7 cells.

NADPH oxidase-generated reactive oxygen species are involved in estradiol 17ß-d-glucuronide-induced cholestasis.

The endogenous metabolite of estradiol, estradiol 17ß-D-glucuronide (E17G), is considered the main responsible of the intrahepatic cholestasis of pregnancy. E17G alters the activity of canalicular transporters through a signaling pathway-dependent cellular internalization, phenomenon that was attributed to oxidative stress in different cholestatic conditions. However, there are no reports involving oxidative stress in E17G-induced cholestasis, representing this the aim of our work. Using polarized hepatocyte cultures, we showed that antioxidant compounds prevented E17G-induced Mrp2 activity alteration, being this alteration equally prevented by the NADPH oxidase (NOX) inhibitor apocynin. The model antioxidant N-acetyl-cysteine prevented, in isolated and perfused rat livers, E17G-induced impairment of bile flow and Mrp2 activity, thus confirming the participation of reactive oxygen species (ROS) in this cholestasis. In primary cultured hepatocytes, pretreatment with specific inhibitors of ERK1/2 and p38MAPK impeded E17G-induced ROS production; contrarily, NOX inhibition did not affect ERK1/2 and p38MAPK phosphorylation. Both, knockdown of p47phox by siRNA and preincubation with apocynin in sandwich-cultured rat hepatocytes significantly prevented E17G-induced internalization of Mrp2, suggesting a crucial role for NOX in this phenomenon. Concluding, E17G-induced cholestasis is partially mediated by NOX-generated ROS through internalization of canalicular transporters like Mrp2, being ERK1/2 and p38MAPK necessary for NOX activation.

ERK1/2 modulation in the central nervous system of male and female thiamine-deficient mice with amprolium.

Thiamine deficiency experimental models focus on using the pyrithiamine analog in male rodents, making the thiamine deficiency effects in females and the use of other thiamine antagonists, such as amprolium, unknown. We investigated the impact of thiamine deficiency with amprolium in the cerebral cortex and thalamus of male and female mice by evaluating the modulation of ERK1/2 phosphorylation. The animals were exposed for 20 days to thiamine-deficient chow with different doses of amprolium (20, 40, 60 and 80 mg/kg) and at different treatment periods (five, 10, 15 or 20 days) at a dose of 60 mg/kg. After treatments, ERK1/2 phosphorylation was analyzed by western blot. In male mice, we observed a progressive increase in ERK1/2 phosphorylation in both the cerebral cortex and thalamus in response to the dose of amprolium. In females, ERK1/2 phosphorylation did not progressively increase in response to the amprolium dosage. However, an increase in phosphorylation at the higher doses of 60 and 80 mg/kg was observed. We observed a more intense increase in ERK1/2 phosphorylation in males' cerebral cortex and thalamus from 10 days onwards. In females, the ERK1/2 modulation profiles were similar. The results show that thiamine deficiency induction with amprolium is efficient, compatible with other recognized models that use pyrithiamine, showing changes in cell signaling in the nervous system. The study showed differences in response to thiamine deficiency with amprolium between male and female mice in relation to ERK1/2 phosphorylation and demonstrated that females respond positively to the method and can also be used as model animals.

Acute In Vivo Administration of Compound 21 Stimulates Akt and ERK1/2 Phosphorylation in Mouse Heart and Adipose Tissue.

The angiotensin II type 2 (AT2) receptor has a role in promoting insulin sensitivity. However, the mechanisms underlying the AT2 receptor-induced facilitation of insulin are still not completely understood. Therefore, we investigated whether acute in vivo administration of AT2 receptor agonist compound 21 (C21) could activate insulin signaling molecules in insulin-target tissues. We report that, in male C57BL/6 mice, an acute (5 min, 0.25 mg/kg; i.v.) injection of C21 induces the phosphorylation of Akt and ERK1/2 at activating residues (Ser473 and Thr202/Tyr204, respectively) in both epididymal white adipose tissue (WAT) and heart tissue. In WAT, the extent of phosphorylation (p) of Akt and ERK1/2 induced by C21 was approximately 65% of the level detected after a bolus injection of a dose of insulin known to induce maximal activation of the insulin receptor (IR). In the heart, C21 stimulated p-Akt to a lesser extent than in WAT and stimulated p-ERK1/2 to similar levels to those attained by insulin administration. C21 did not modify p-IR levels in either tissue. We conclude that in vivo injection of the AT2 receptor agonist C21 activates Akt and ERK1/2 through a mechanism that does not involve the IR, indicating the participation of these enzymes in AT2R-mediated signaling.

Epiisopiloturine, an Alkaloid from Pilocarpus microphyllus, Attenuates LPS-Induced Neuroinflammation by Interfering in the TLR4/NF-κB-MAPK Signaling Pathway in Microglial Cells.

Neuroinflammation is present in the pathophysiological mechanisms of several diseases that affect the central nervous system (CNS). Microglia have a prominent role in initiating and sustaining the inflammatory process. Epiisopiloturine (EPI) is an imidazole alkaloid obtained as a by-product of pilocarpine extracted from Pilocarpus microphyllus (jaborandi) and has shown promising anti-inflammatory and antinociceptive properties. In the present study, we investigated the effects of EPI on the inflammatory response in microglial cells (BV-2 cells) induced by lipopolysaccharide (LPS) and explored putative underlying molecular mechanisms. Cell viability was not affected by EPI (1-100 µg/mL) as assessed by both LDH activity and the MTT test. Pretreatment with EPI (25, 50, and 100 µg/mL) significantly reduced the proinflammatory response induced by LPS, as observed by a decrease in nitrite oxide production and iNOS protein expression. EPI (25 µg/mL) reduced IL-6 and TNF-α production, by 40% and 34%, respectively. However, no changes were observed in the anti-inflammatory IL-10 production. Mechanistically, EPI inhibited the TLR4 expression and phosphorylation of NF-κB p65 and MAPKs (JNK and ERK1/2) induced by LPS, but no changes were observed in TREM2 receptor expression in LPS-stimulated cells. In conclusion, our data demonstrated the potent anti-inflammatory properties of EPI in microglial cells. These effects are associated with the reduction of TLR4 expression and inhibition of intracellular signaling cascades, including NF-κB and MAPKs (JNK and ERK1/2).

The involvement of PDIA2 gene in the progression of renal cell carcinoma is potentially through regulation of JNK signaling pathway.

Renal cell carcinoma (RCC) with poor prognosis and high incidence rate is a common malignant disease. Current therapies could bring little benefit for the patients with advanced-stage RCC. PDIA2 is an isomerase responsible for protein folding and its role in cancer including RCC is under investigation. In this study, we found that PDIA2 was expressed much higher in RCC tissues than the control but the methylation level of PDIA2 promoter was lower based on the TCGA data. Patients with higher PDIA2 expression exerted worse survival. In clinical specimen, PDIA2 expression was correlated to patients' clinical factors such as TNM stage (I/II vs III/IV, p = 0.025) and tumor size (≤ 7 cm vs > 7 cm, p = 0.004). Moreover, K-M analysis showed that PDIA2 was associated with patients' survival in RCC. PDIA2 was expressed much higher in cancer cells A498 than 786-O than that in 293 T cells. After PDIA2 was knocked down, cell proliferation, migration and invasion was potently inhibited. But cell apoptotic rate increased reversely. Furthermore, the efficacy of Sunitinib on RCC cells was strengthened after PDIA2 knockdown. In addition, knockdown of PDIA2 gene leaded to downregulation of levels of JNK1/2, phosphorylated JNK1/2, c-JUN, and Stat3. But this inhibition was partially released when JNK1/2 was overexpressed. In consistent, cell proliferation was also partially recovered. In summary, PDIA2 plays important role in progression of RCC and JNK signaling pathway might be regulated by PDIA2. This study suggests PDIA2 as a candidate target for therapy of RCC.

Asparaginyl endopeptidase contributes to cetuximab resistance via MEK/ERK signaling in RAS wide-type metastatic colorectal cancer.

BACKGROUND: Cetuximab, a monoclonal antibody targeting epidermal growth factor receptor (EGFR), is effective for RAS wild-type metastatic colorectal cancer (mCRC) patients. However, cetuximab resistance often occur and the mechanism has not been fully elucidated. The purpose of this study was to investigate the role of asparaginyl endopeptidase (AEP) in cetuximab resistance. METHODS: Differentially expressed genes between cetuximab responders and non-responders were identified by analyzing the gene expression profile GSE5851, retrieved from Gene Expression Omnibus (GEO). The potential genes were further validated in cetuximab-resistant CRC cell lines. The expression of AEP in the peripheral blood and tumor tissues of mCRC patients in our hospital were detected by enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry, respectively. The survival analysis was carried out by Kaplan-Meier method. The function and associated pathways of AEP were further investigated by lentivirus transfection, CCK8 assay, colony formation assay, real-time polymerase chain reaction (qPCR) and western blot. RESULTS: Through bioinformatics analysis, we found that the expression of AEP gene was related to progress free survival (PFS) of mCRC patients treated with cetuximab alone (P = 0.00133). The expression of AEP was significantly higher in the cetuximab-resistant CRC cell lines, as well as in mCRC patients with shorter PFS treated with cetuximab-containing therapy. Furthermore, AEP could decrease the sensitivity of CRC cells to cetuximab in vitro. And the phosphorylation level of MEK and ERK1/2 was increased in AEP overexpression cells. The downregulation of AEP using specific inhibitors could partially restore the sensitivity of CRC cells to cetuximab. CONCLUSION: The higher expression of AEP could contribute to the shorter PFS of cetuximab treatment in mCRC. The reason might be that AEP could promote the phosphorylation of MEK/ERK protein in the downstream signal pathway of EGFR.

Caryocar brasiliense peel ethanolic extract has neuroprotective potential and reduces the activation of ERK1/2 in the ischemia and reperfusion brain acute phase in the rat.

Oxidative stress induced by ischemia and reperfusion (I/R) injury results in cell death by necrosis or apoptosis and triggers the activation of different intracellular pathways, such as mitogen-activated protein activated kinases. Pequi (Caryocar brasiliense) peel, residue of a fruit from Brazilian savannah-like vegetation, has phenolic compounds that have been demonstrated to have antioxidant effects in vitro. The present study aimed to evaluate the neuroprotective effects of C. brasiliense peel ethanolic extract (CBPE) against transient global I/R injury in the rat brain. Global ischemia for 5, 20, and 45 min followed by 2 h of reperfusion caused a significant time-dependent increase in the number of ischemic neurons (p ≤ 0.05); increased immunoreactivity of cleaved caspase-3 (CASP3); and activated extracellular signal-regulated kinase (ERK) 1/2. Pretreatment with CBPE (600 mg/kg, oral) or vitamin E (0.6 mg, oral) for 30 days showed significant protection against acute brain injury induced by 20 and 45 min or 5 min of ischemia, respectively, by reducing the cortical ischemic neuron count (p ≤ 0.05) and p-ERK1/2 immunoreactivity. In addition, active c-Jun N-terminal kinase (JNK) immunoreactivity was reduced in animals not subjected to ischemia. Therefore, we suggest an association between vitamin E and CBPE, which may generate a better neuroprotective response. Interestingly, mainly in the hippocampus and oligodendrocytes, high dose CBPE increase the number of isquemic neurons and of CASP3 immunoreactive cells in animals subjected or not to ischemia, which was not verified in the vitamin E group. Therefore, additional studies are recommended to verify the safety of the continuous use of CBPE.

Erk1/2 signaling mediates the HGF-induced protection against ethanol and acetaldehyde-induced toxicity in the pancreatic RINm5F cell line.

Alcohol-induced pancreas damage remains as one of the main risk factors for pancreatitis development. This disorder is poorly understood, particularly the effect of acetaldehyde, the primary alcohol metabolite, in the endocrine pancreas. Hepatocyte growth factor (HGF) is a protective protein in many tissues, displaying antioxidant, antiapoptotic, and proliferative responses. In the present work, we were focused on characterizing the response induced by HGF and its protective mechanism in the RINm5F pancreatic cell line treated with ethanol and acetaldehyde. RINm5F cells were treated with ethanol or acetaldehyde for 12 h in the presence or not of HGF (50 ng/ml). Cells under HGF treatment decreased the content of reactive oxygen species and lipid peroxidation induced by both toxics, improving cell viability. This effect was correlated to an improvement in insulin expression impaired by ethanol and acetaldehyde. Using a specific inhibitor of Erk1/2 abrogated the effects elicited by the growth factor. In conclusion, the work provides mechanistic evidence of the HGF-induced-protective response to the alcohol-induced damage in the main cellular component of the endocrine pancreas.

A nested bistable module within a negative feedback loop ensures different types of oscillations in signaling systems.

In this article, we consider a double phosphorylation cycle, a ubiquitous signaling component, having the ability to display bistability, a behavior strongly related to the existence of positive feedback loops. If this component is connected to other signaling elements, it very likely undergoes some sort of protein-protein interaction. In several cases, these interactions result in a non-explicit negative feedback effect, leading to interlinked positive and negative feedbacks. This combination was studied in the literature as a way to generate relaxation-type oscillations. Here, we show that the two feedbacks together ensure two types of oscillations, the relaxation-type ones and a smoother type of oscillations functioning in a very narrow range of frequencies, in such a way that outside that range, the amplitude of the oscillations is severely compromised. Even more, we show that the two feedbacks are essential for both oscillatory types to emerge, and it is their hierarchy what determines the type of oscillation at work. We used bifurcation analyses and amplitude vs. frequency curves to characterize and classify the oscillations. We also applied the same ideas to another simple model, with the goal of generalizing what we learned from signaling models. The results obtained display the wealth of oscillatory dynamics that exists in a system with a bistable module nested within a negative feedback loop, showing how to transition between different types of oscillations and other dynamical behaviors such as excitability. Our work provides a framework for the study of other oscillatory systems based on bistable modules, from simple two-component models to more complex examples like the MAPK cascade and experimental cases like cell cycle oscillators.

Blood orange juice intake modulates plasma and PBMC microRNA expression in overweight and insulin-resistant women: impact on MAPK and NFκB signaling pathways.

Blood orange consumption presents potential health benefits and may modulate epigenetic mechanisms such as microRNAs (miRNAs) expression. MiRNAs are non-coding RNAs responsible for post-transcriptional gene regulation, and these molecules can also be used as biomarkers in body fluids. This study was designed to investigate the effect of chronic blood orange juice (BOJ) intake on the inflammatory response and miRNA expression profile in plasma and blood cells in overweight women. The study cohort was comprised of twenty women aged 18-40 years old, diagnosed as overweight, who consumed 500 mL/d of BOJ for four weeks. Clinical data were collected at baseline and after 4 weeks of juice consumption, e.g., anthropometric and hemodynamic parameters, food intake, blood cell count, and metabolic and inflammatory biomarkers. BOJ samples were analyzed and characterized. Additionally, plasma and blood cells were also collected for miRNA expression profiling and evaluation of the expression of genes and proteins in the MAPK and NFκB signaling pathways. BOJ intake increased the expression of miR-144-3p in plasma and the expression of miR-424-5p, miR-144-3p, and miR-130b-3p in peripheral blood mononuclear cells (PBMC). Conversely, the beverage intake decreased the expression of let-7f-5p and miR-126-3p in PBMC. Computational analyses identified different targets of the dysregulated miRNA on inflammatory pathways. Furthermore, BOJ intake increased vitamin C consumption and the pJNK/JNK ratio and decreased the expression of IL6 mRNA and NFκB protein. These results demonstrate that BOJ regulates the expression of genes involved in the inflammatory process and decreases NFкB-protein expression in PBMC.

Cognitive Deficits Found in a Pro-inflammatory State are Independent of ERK1/2 Signaling in the Murine Brain Hippocampus Treated with Shiga Toxin 2 from Enterohemorrhagic Escherichia coli.

Shiga toxin 2 (Stx2) from enterohemorrhagic Escherichia coli (EHEC) produces hemorrhagic colitis, hemolytic uremic syndrome (HUS), and acute encephalopathy. The mortality rate in HUS increases significantly when the central nervous system (CNS) is involved. Besides, EHEC also releases lipopolysaccharide (LPS). Many reports have described cognitive dysfunctions in HUS patients, the hippocampus being one of the brain areas targeted by EHEC infection. In this context, a translational murine model of encephalopathy was employed to establish the deleterious effects of Stx2 and the contribution of LPS in the hippocampus. The purpose of this work is to elucidate the signaling pathways that may activate the inflammatory processes triggered by Stx2, which produces cognitive alterations at the level of the hippocampus. Results demonstrate that Stx2 produced depression-like behavior, pro-inflammatory cytokine release, and NF-kB activation independent of the ERK1/2 signaling pathway, while co-administration of Stx2 and LPS reduced memory index. On the other hand, LPS activated NF-kB dependent on ERK1/2 signaling pathway. Cotreatment of Stx2 with LPS aggravated the pathologic state, while dexamethasone treatment succeeded in preventing behavioral alterations. Our present work suggests that the use of drugs such as corticosteroids or NF-kB signaling inhibitors may serve as neuroprotectors from EHEC infection.

ROS and ERK Pathway Mechanistic Approach on Hepatic Insulin Resistance After Chronic Oral Exposure to Cadmium NOAEL Dose.

Cadmium is a critical toxic agent in occupational and non-occupational settings and acute and chronic environmental exposure situations that have recently been associated with metabolic disease development. Until now, the no observed adverse effect level (NOAEL) of cadmium has not been studied regarding insulin resistance development. Therefore, we aimed to monitor whether chronic oral exposure to cadmium NOAEL dose induces insulin resistance in Wistar rats and investigate if oxidative stress and/or inflammation are related. Male Wistar rats were separated into control (standard normocalorie diet + water free of cadmium) and cadmium groups (standard normocalorie diet + drinking water with 15 ppm CdCl2). At 15, 30, and 60 days, oral glucose tolerance, insulin response, and insulin resistance were analyzed using mathematical models. In the liver glycogen, triglyceride, pro- and anti-inflammatory cytokines, cadmium, zinc, metallothioneins, and redox balance were quantified. Immunoreactivity analysis of proteins involved in metabolic and mitogenic insulin signaling was performed. The results showed that a cadmium NOAEL dose after 15 days of exposure causes ROS and mitogenic arm of insulin signaling to increase while hepatic glycogen diminishes. At 30 days, Cd accumulation accentuated ROS production, hepatic triglyceride overaccumulation, and mitogenic signals that develop insulin resistance. Finally, inflammation and lipid peroxidation appear after 60 days of Cd exposure, while lipids and carbohydrate homeostasis deteriorate. In conclusion, environmental exposure to cadmium NAOEL dose causes hepatic Cd accumulation and ROS overproduction that chronically declines the antioxidant defense, deteriorates metabolic homeostasis associated with the mitogenic pathway of insulin signaling, and induces insulin resistance.

BRAF/MEK inhibition in NSCLC: mechanisms of resistance and how to overcome it.

Targeted therapy for oncogenic genetic alterations has changed the treatment paradigm of advanced non-small cell lung cancer (NSCLC). Mutations in the BRAF gene are detected in approximately 4% of patients and result in hyper-activation of the MAPK pathway, leading to uncontrolled cellular proliferation. Inhibition of BRAF and its downstream effector MEK constitutes a therapeutic strategy for a subset of patients with NSCLC and is associated with clinical benefit. Unfortunately, the majority of patients will develop disease progression within 1 year. Preclinical and clinical evidence suggests that resistance mechanisms involve the restoration of MAPK signaling which becomes inhibition-independent due to upstream or downstream alterations, and the activation of bypass pathways, such as the PI3/AKT/mTOR pathway. Future research should be directed to deciphering the mechanisms of cancer cells' oncogenic dependence, understanding the tissue-specific mechanisms of BRAF-mutant tumors, and optimizing treatment strategies after progression on BRAF and MEK inhibition.