P38 MAPK Research Articles

A p38 MAPK-ROS axis fuels proliferation stress and DNA damage during CRISPR-Cas9 gene editing in hematopoietic stem and progenitor cells.

Exvivo activation is a prerequisite to reaching adequate levels of gene editing by homology-directed repair (HDR) for hematopoietic stem and progenitor cell (HSPC)-based clinical applications. Here, we show that shortening culture time mitigates the p53-mediated DNA damage response to CRISPR-Cas9-induced DNA double-strand breaks, enhancing the reconstitution capacity of edited HSPCs. However, this results in lower HDR efficiency, rendering exvivo culture necessary yet detrimental. Mechanistically, exvivo activation triggers a multi-step process initiated by p38 mitogen-activated protein kinase (MAPK) phosphorylation, which generates mitogenic reactive oxygen species (ROS), promoting fast cell-cycle progression and subsequent proliferation-induced DNA damage. Thus, p38 inhibition before gene editing delays G1/S transition and expands transcriptionally defined HSCs, ultimately endowing edited cells with superior multi-lineage differentiation, persistence throughout serial transplantation, enhanced polyclonal repertoire, and better-preserved genome integrity. Our data identify proliferative stress as a driver of HSPC dysfunction with fundamental implications for designing more effective and safer gene correction strategies for clinical applications.

The bta-miR-22-3p can alleviate LPS-induced inflammatory response in yak endometrial epithelial cells by targeting KSR2

Bovine endometritis is a common reproductive system disease in dairy cows that leads to decreased milk production and reproductive performance, causing significant economic losses for farmers. Research has shown that microRNAs (miRNAs) play a significant role in regulating the expression of biological genes and are closely related to the occurrence of inflammation, including bta-miR-22-3p. However, the specific molecular mechanisms by which miRNAs regulate bovine endometritis remain unclear. To investigate the regulatory mechanism of bta-miR-22-3p in yak endometritis, uterine tissues were collected from three healthy bos grunniens and three bos grunniens with endometritis, approximately 21 days postpartum. Various methods were employed, including real-time quantitative polymerase chain reaction (RT-qPCR), Western blot (WB), enzyme-linked immunosorbent assay (ELISA), and immunofluorescence (IF). The results demonstrated that overexpression of bta-miR-22-3p led to a significant decrease (P<0.05) in factors related to the mitogen-activated protein kinase (MAPK) signaling pathway and associated inflammatory factors, such as extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (P38), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β). Furthermore, dual-luciferase assays confirmed that the kinase suppressor of the Ras 2 (KSR2) gene is a downstream target of bta-miR-22-3p. Overexpression of bta-miR-22-3p inhibited the expression of KSR2. When KSR2 was inhibited, the levels of MAPK signaling pathway-related factors and inflammation also significantly decreased(P<0.05). Thus, bta-miR-22-3p suppresses the activation of the MAPK signaling pathway through the inhibition of KSR2, resulting in a reduction of inflammatory factors. In conclusion, this study demonstrated that bta-miR-22-3p targets the KSR2 gene to alleviate LPS (Lipopolysaccharide)-induced inflammatory damage.

Protective effect of 6'-Sialyllactose on LPS-induced macrophage inflammation via regulating Nrf2-mediated oxidative stress and inflammatory signaling pathways.

Macrophages play a central role in cardiovascular diseases, like atherosclerosis, by accumulating in vessel walls and inducing sustained local inflammation marked by the release of chemokines, cytokines, and matrix-degrading enzymes. Recent studies indicate that 6'-sialyllactose (6'-SL) may mitigate inflammation by modulating the immune system. Here, we examined the impact of 6'-SL on lipopolysaccharide (LPS)-induced acute inflammation using RAW 264.7 cells and a mouse model. In vivo, ICR mice received pretreatment with 100 mg/kg 6'-SL for 2 h, followed by intraperitoneal LPS injection (10 mg/kg) for 6 h. In vitro, RAW 264.7 cells were preincubated with 6'-SL before LPS stimulation. Mechanistic insights were gained though Western blotting, qRT-PCR, and immunofluorescence analysis, while reactive oxygen species (ROS) production was assessed via DHE assay. 6'-SL effectively attenuated LPS-induced p38 MAPK and Akt phosphorylation, as well as p65 nuclear translocation. Additionally, 6'-SL inhibited LPS-induced expression of tissue damage marker MMP9, IL-1β, and MCP-1 by modulating NF-κB activation. It also reduced ROS levels, mediated by p38 MAPK and Akt pathways. Moreover, 6'-SL restored LPS-suppressed Nrf2 and HO-1 akin to specific inhibitors SB203580 and LY294002. Consistent with in vitro results, 6'-SL decreased oxidative stress, MMP9, and MCP-1 expression in mouse endothelium following LPS-induced macrophage activation. In summary, our findings suggest that 6'-SL holds promise in mitigating atherosclerosis by dampening LPS-induced acute macrophage inflammation.

Cortisol suppresses lipopolysaccharide-induced in vitro inflammatory response of large yellow croaker (Larimichthys crocea) via the glucocorticoid receptor and p38 mitogen-activated protein kinase pathways.

Glucocorticoids (GCs) are well-established anti-inflammatory agents, with cortisol, an endogenous GC, exerting pivotal regulatory effects on normal physiological processes. However, the immune regulatory role of cortisol in teleost fish, particularly in inflammation induced by pathogenic infection, remains largely unexplored. Here, we revealed that lipopolysaccharide (LPS) triggers a pro-inflammatory response in the large yellow croaker (Larimichthys crocea), as evidenced by increased expression of key pro-inflammatory cytokines and activation of the mitogen-activated protein kinase (MAPK) signaling pathway. We further explored the immunosuppressive capacity of cortisol in LPS-stimulated large yellow croaker kidney cells (PCK cells) and in vitro tissues of the large yellow croaker. Our findings indicated that cortisol effectively suppresses LPS-induced overexpression of pro-inflammatory cytokines and p38 MAPK pathway activation. Moreover, the immunosuppressive effects of cortisol were reversed by pretreatment with mifepristone, a glucocorticoid receptor (GR) antagonist. Collectively, this study delineated the inhibitory role of cortisol in the LPS-induced inflammatory cascade in large yellow croaker and underscores the significance of GR in mediating this response. These insights advance our comprehension of GCs-mediated immune modulation and provide a theoretical basis for the application of cortisol in disease prevention and the selective breeding of disease-resistant traits in aquaculture.

RIPK2 Is Crucial for the Microglial Inflammatory Response to Bacterial Muramyl Dipeptide but Not to Lipopolysaccharide.

Receptor-interacting serine/threonine protein kinase 2 (RIPK2) is a kinase that is essential in modulating innate and adaptive immune responses. As a downstream signaling molecule for nucleotide-binding oligomerization domain 1 (NOD1), NOD2, and Toll-like receptors (TLRs), it is implicated in the signaling triggered by recognition of microbe-associated molecular patterns by NOD1/2 and TLRs. Upon activation of these innate immune receptors, RIPK2 mediates the release of pro-inflammatory factors by activating mitogen-activated protein kinases (MAPKs) and nuclear factor-kappa B (NF-κB). However, whether RIPK2 is essential for downstream inflammatory signaling following the activation of NOD1/2, TLRs, or both remains controversial. In this study, we examined the role of RIPK2 in NOD2- and TLR4-dependent signaling cascades following stimulation of microglial cells with bacterial muramyl dipeptide (MDP), a NOD2 agonist, or lipopolysaccharide (LPS), a TLR4 agonist. We utilized a highly specific proteolysis targeting chimera (PROTAC) molecule, GSK3728857A, and found dramatic degradation of RIPK2 in a concentration- and time-dependent manner. Importantly, the PROTAC completely abolished MDP-induced increases in iNOS and COX-2 protein levels and pro-inflammatory gene transcription of Nos2, Ptgs2, Il-1β, Tnfα, Il6, Ccl2, and Mmp9. However, increases in iNOS and COX-2 proteins and pro-inflammatory gene transcription induced by the TLR4 agonist, LPS, were only slightly attenuated with the GSK3728857A pretreatment. Further findings revealed that the RIPK2 PROTAC completely blocked the phosphorylation and activation of p65 NF-κB and p38 MAPK induced by MDP, but it had no effects on the phosphorylation of these two mediators triggered by LPS. Collectively, our findings strongly suggest that RIPK2 plays an essential role in the inflammatory responses of microglia to bacterial MDP but not to LPS.

Toll-like receptor signaling in neurons modulates C. elegans feeding behavior in a hunger state-dependent manner

Animals face the risk of encountering pathogenic microbes while foraging for resources. Assessing the risk of nutrition vs. infection can result in the behavioral regulation of immune processes. Behavioral immunity in the nematode roundworm Caenorhabditis elegans (C. elegans) is regulated, in part, by the innate immune molecule TOL-1: a homolog of vertebrate Toll-like Receptor (TLR) proteins that influences C. elegans pathogen avoidance behaviors by promoting the development of CO2-detecting chemosensory neurons. While TOL-1′s role in pathogen avoidance is well established, its role in an opposing behavior – foraging – has not been examined. In addition to pathogenic bacteria, preferred food for C. elegans, such as Escherichia coli (E. coli), create significant and aversive environmental CO2 levels which may limit feeding behaviors in a tol-1 dependent manner. We have found that in addition to conferring antibacterial immunity, TOL-1 signals in neurons through the p38 MAPK PMK-1 to promote turning behavior and limit foraging when food is abundant and that the anorectic TOL-1/PMK-1 pathway is attenuated during starvation to promote foraging These data highlight the dynamic role of a conserved innate immune cascade in neurons during both high and low hunger states and identify mechanisms underlying the neuro-immune control of feeding strategies.

Reversing metabolic reprogramming by CPT1 inhibition with etomoxir promotes cardiomyocyte proliferation and heart regeneration via DUSP1 ADP-ribosylation-mediated p38 MAPK phosphorylation

The neonatal mammalian heart has a remarkable regenerative capacity, while the adult heart has difficulty to regenerate. A metabolic reprogramming from glycolysis to fatty acid oxidation occurs along with the loss of cardiomyocyte proliferative capacity shortly after birth. In this study, we sought to determine if and how metabolic reprogramming regulates cardiomyocyte proliferation. Reversing metabolic reprogramming by carnitine palmitoyltransferase 1 (CPT1) inhibition, using cardiac-specific Cpt1a and Cpt1b knockout mice promoted cardiomyocyte proliferation and improved cardiac function post-myocardial infarction. The inhibition of CPT1 is of pharmacological significance because those protective effects were replicated by etomoxir, a CPT1 inhibitor. CPT1 inhibition, by decreasing poly(ADP-ribose) polymerase 1 expression, reduced ADP-ribosylation of dual-specificity phosphatase 1 in cardiomyocytes, leading to decreased p38 MAPK phosphorylation, and stimulation of cardiomyocyte proliferation. Our present study indicates that reversing metabolic reprogramming is an effective strategy to stimulate adult cardiomyocyte proliferation. CPT1 is a potential therapeutic target for promoting heart regeneration and myocardial infarction treatment.

PARP15 is a Susceptibility Locus for Clarkson Disease (Monoclonal Gammopathy-Associated Systemic Capillary Leak Syndrome).

Vascular leakage is a deadly complication of severe infections, ranging from bacterial sepsis to malaria. Worldwide, septicemia is among the top 10 causes of lethality because of the shock and multiorgan dysfunction that arise from the host vascular response. In the monoclonal gammopathy-associated capillary leak syndrome (MG-CLS), even otherwise mundane infections induce recurrent septic-like episodes of profound microvascular hyperpermeability and shock. There are no defined genetic risk factors for MG-CLS or effective treatments for acute crises. We characterized predicted loss-of-function mutations in PARP15 (poly[ADP-ribose] polymerase 15), a protein of unknown function that is absent in mice, in patients with MG-CLS. We analyzed barrier function in PARP15-deficient vascular endothelial cells and vascular leakage in mice engineered to express WT or loss-of-function variant human PARP15. We discovered several loss-of-function PARP15 variants associated with MG-CLS. These mutations severely reduced PARP15 enzymatic function. The presence of the most frequently detected variant (G628R) correlated with clinical markers of severe vascular leakage. In human microvascular endothelial cells, PARP15 suppressed cytokine-induced barrier disruption by ADP-ribosylating the scaffold protein JIP3 (c-Jun N-terminal kinase-interacting protein 3) and inhibiting p38 MAP kinase activation. Mice expressing enzymatically inactive human PARP15(G628R) were significantly more prone to inflammation-associated vascular leakage than mice expressing WT PARP15 in a p38-dependent fashion. PARP15 represents a previously unrecognized genetic susceptibility factor for MG-CLS. PARP15-mediated ADP ribosylation is an essential and genetically determined mechanism of the human vascular response to inflammation.

Assessing the antiproliferative properties of various teas against the DU-145 prostate cancer cell line: A combined in vitro and in silico investigation

BackgroundPhytotherapy emerges as a promising solution to the challenges posed by cancer treatment. Black, green, and purple teas, renowned for their antioxidant properties, exhibit efficacy against various cancers, including prostate cancer. Study DesignThis was an experimental study conducted from the Kenya Medical Research Institute (KEMRI). Aim of the Studythe present study aimed to evaluate the Cytotoxic effect of black, green and purple teas against prostate cancer cell line DU-145 and in silico prediction of Tea catechins interaction with prostate cancer proteins such as 3D-crystal structures of Androgen-receptor(5T8E) and Human p38 MAPK(1KV2). MethodsThe study started by quantifying tea catechins (Epigallocatechin gallate (EGCG), Epigallocatechin EGC, Epicatechin gallate (ECG) and Epicatechin (EC) with high-liquid chromatography (HPLC), cytotoxicity analysis of black, green and purple tea crude extracts against DU-145 cell line at 24hr and 48 hrs by Resazurin assay and molecular docking was performed with Autodock Vina 1.1.2 software. ResultsEGCG appeared the most predominant catechin with 552.2 ± 10.61 (mg g-1 DW) while EC was the lowest detected with 118.08 ± 9.45 (mg g-1 DW). Purple tea extract exhibited high antiproliferative activity with IC50 of 98.69±1.99 µg/ml at 24hr and 91.21±6.91 µg/ml at 48hr, green tea followed with 121 ± 5.97µg/ml at 24hr and 107.1 ± 2.03 µg/ml at 48hr IC50; black tea was the least effective tea at both 24hr and 48hr with IC50 of 290.8 ± 2.46µg/ml and 237.8 ± 5.59µg/ml respectively. Prediction of catechin target genes and Prostate cancer (Pca)-related genes discovered 121 catechin target genes and 34,926 Pca-related genes. EGC exhibited high binding interaction to 5T8E with the lowest affinity of -9.1Kcal/mol followed by EGCG at -8.4 (Kcal/mol then EC with -8.2 (Kcal/mol) and EGC at -8.1 (Kcal/mol). Catechins interaction with 1KV2 was in the following order EGCG (-9.4 Kcal/mol) ˃ECG (-8.2 Kcal/mol) ˃EC (-8.2 Kcal/mol) ˃EGC (-8.1 Kcal/mol). ConclusionIn vitro experiment detected a very significant antiproliferative activity of black, green and purple tea crude extracts against the DU-145 cell line; furthermore, in silico analysis revealed a strong binding interaction with proteins associated with prostate cancer, indicating that black, green, and purple teas, along with catechins, hold promise as potential herbal remedies for combating prostate cancer.

DLK1 and DLK2, two non-canonical ligands of NOTCH receptors, differentially modulate the osteogenic differentiation of mesenchymal C3H10T1/2 cells

BackgroundC3H10T1/2 is a mesenchymal cell line capable of differentiating into osteoblasts, adipocytes and chondrocytes. The differentiation of these cells into osteoblasts is modulated by various transcription factors, such as RUNX2. Additionally, several interconnected signaling pathways, including the NOTCH pathway, play a crucial role in modulating their differentiation into mature bone cells. We have investigated the roles of DLK1 and DLK2, two non-canonical inhibitory ligands of NOTCH receptors, in the osteogenic differentiation of C3H10T1/2 cells.ResultsOur results corroborate existing evidence that DLK1 acts as an inhibitor of osteogenesis. In contrast, we demonstrate for the first time that DLK2 enhances this differentiation process. Additionally, our data suggest that NOTCH2, 3 and 4 receptors may promote osteogenesis, as indicated by their increased expression during this process, whereas NOTCH1 expression, which decreases during cell differentiation, might inhibit osteogenesis. Moreover, treatment with DAPT, a NOTCH signaling inhibitor, impeded osteogenic differentiation. We have confirmed the increase in ERK1/2 MAPK and p38 MAPK phosphorylation in C3H10T1/2 cells induced to differentiate to osteoblasts. Our new findings reveal increased ERK1/2 MAPK phosphorylation in differentiated C3H10T1/2 cells with a decrease in DLK1 expression or an overexpression of DLK2, which is coincident with the behavior of those transfectants where we have detected an increase in osteogenic differentiation. Additionally, p38 MAPK phosphorylation increases in differentiated C3H10T1/2 cells with reduced DLK1 levels.ConclusionsOur results suggest that DLK1 may inhibit osteogenesis, while DLK2 may promote it, by modulating NOTCH signaling and the phosphorylation of ERK1/2 and p38 MAPK pathways. Given the established inhibitory effect of DLK proteins on NOTCH signaling, these new insights could pave the way for developing future therapeutic strategies aimed at treating bone diseases.

Protective effect of betaine supplementation in low-met diet against Aeromonas hydrophila-induced intestinal injury in grass carp (Ctenopharyngodon idella)

This study evaluated the effect of supplemental betaine in low-methionine (Met) diet concerning the regulatory role of bacteria-induced intestinal injury in fish. 540 grass carp (Ctenopharyngodon Idella) (210 ± 0.68 g) were selected and randomly divided into 6 groups, which contained a reference group fed with the diet supplemented with DL-Met (3.1 g kg−1 diet) and 5 groups fed on the diets added with increasing levels of betaine (0–6.4 g kg−1 diet) for 60 days. Then, a 14-day Aeromonas hydrophila challenge was conducted. Compared with BET0 group, results indicated that appropriate betaine supplementation (1) enhanced antioxidant enzymes activities (except glutathione S-transferase pi-1 (GSTP1) and copper zinc superoxide dismutase (CuZnSOD) and glutathione (GSH) contents, and NF-E2-related-factor 2 (Nrf2) protein nuclear translocation, while reduced reactive oxygen species (ROS), and representative oxidative damage markers; (2) up-regulated the transcription abundance of anti-apoptosis-related molecules while reduced DNA-ladder occurrence and the transcription levels of pro-apoptosis-related genes; (3) down-regulated the transcription of tight junction-related proteins (such as zonula occludens-1 (ZO-1) etc.) in fish intestines. Furthermore, correlation analysis revealed that feeding betaine ameliorating the oxidative damage, apoptosis and tight junction breakdown was possibly related to intestinal Nrf2, p38 mitogen-activated protein kinase (p38 MAPK) and c-Jun N-terminal kinases (JNK), and myosin light chain kinase (MLCK) signaling, respectively. According to the regression analysis for intestinal ROS abundance, claudin-b and caspase-2 transcription, the appropriate betaine supplementations were estimated to be 3.82, 3.98, and 3.62 g kg−1 diet, respectively. Notably, betaine has the potential to spare part Met and the efficacy of betaine relative to Met was 206.67 % based on intestinal ROS contents in grass carp (210–776 g).

GSK3179106 ameliorates lipopolysaccharide-induced inflammation and acute lung injury by targeting P38 MAPK

Acute lung injury (ALI) is a serious acute respiratory disease that can cause alveolar-capillary barrier disruption and pulmonary edema, respiratory failure and multiple organ dysfunction syndrome. However, there is no effective drugs in clinic until now. GSK3179106 has been reported can alleviate intestinal stress syndrome, but the protective effect of GSK3179106 on ALI has not been elucidated. The present study will evaluate the pharmacological activity of GSK3179106 on lipopolysaccharide (LPS)-induced inflammation and lung injury and clarify its underlying mechanism. We found that GSK3179106 significantly attenuated LPS-induced lung injury in vivo, accompanied by inhibited infiltration of inflammatory cells and reduced expression of inflammatory cytokines. Meanwhile, GSK3179106 dose-dependently reduced the LPS-induced IL-6 expression both in protein and gene levels in macrophages. Mechanistically, GSK3179106 could inhibited the phosphorylation of P38 MAPK induced by LPS. Importantly, results showed that there is a direct combination between GSK3179106 and P38 MAPK. Together, our findings not only clarified the anti-inflammatory activity of GSK3179106 but also discovered its new clinical indications. Therefore, compound GSK3179106 may be a potential candidate for the treatment of acute inflammatory diseases.

Effect of extracellular vesicles derived from inflammatory H9C2 cardiomyocytes on the polarization of macrophages and its mechanism

Abstract Objective The aim of this study was to investigate the effect of Extracellular vesicles (EVs) derived from inflammatory cardiomyocytes on the polarization of macrophages and its related mechanism. Methods EVs derived from the inflammatory and normal cardiomyocyte cell line H9C2 were extracted by differential centrifugation respectively. The extracted EVs were characterized by nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM) and Western blot analysis (WB). The RAW264.7 macrophages in the inflammatory environment were treated with EVs, the proliferative activities were detected by CCK8, the expression of inflammatory factors were detected by qRT-PCR, and the differentiation of RAW264.7 macrophages into M1 and M2 was analyzed by flow cytometry. In vivo experiments, the animal model of viral myocarditis was established in Balb/c mice with intraperitoneal injection of CVB3. EVs derived from inflammatory H9C2 cardiomyocytes and medium were injected into mice by tail vein injection, respectively. The mice myocardium was stained by immunohistochemistry, and the total RNA of myocardial tissue was extracted for the detection of inflammatory factors by RT-PCR. In terms of mechanism research, mRNA expression profiles of RAW264.7 cells after EVs treatment were analyzed by RNA sequencing for functional annotation and mechanism study. At the same time, the extracted EVs derived from H9C2 normal group and the inflammatory group were subjected to 4D-Lable Free technology for quantitative proteomic analysis, and the results were analyzed bioinformatics, and then by WB analysis.Results We found that EVs derived from inflammatory H9C2 cardiomyocytes decreased the proliferative activity of RAW264.7 macrophages, the expression of pro-inflammatory cytokines IL-6, IL-1β, TNF-ɑ, and the proportion of M1 macrophages in the inflammatory state, while the proportion of M2 macrophages, which mainly inhibit inflammation, increased, and the expression levels of anti-inflammatory cytokines IL-10 and CD-206 increased. Results of immunohistochemical staining and RT-PCR showed that the EVs of inflammatory H9C2 regulated the heterogeneity of infiltrating macrophages and the expression of several inflammatory cytokines in mouse myocardium inflammatory tissue. Meanwhile, in vitro studies have found that the regulatory effect of EVs derived from inflammatory H9C2 cardiomyocytes on macrophage heterogeneity is mediated by MAPK signaling pathway. Conclusion EVs derived from inflammatory H9C2 cardiomyocytes can regulate the polarization of macrophages, change the expression of inflammatory factors, and regulate the myocardial inflammatory microenvironment, which may be achieved by mediating the p38 MAPK pathway through the delivery of PP2A.

EPO Deficiency Upregulates GADD45b/p38 MAPK Axis, Mediating Schizophrenia-Related Synaptic and Cognitive Impairments.

Schizophrenia (SZ) is a chronic and severe mental illness associated with psychiatric symptoms, cognitive deficits, and social dysfunction. Current clinical interventions only limit relief of psychiatric symptoms and have minimal impact on cognitive impairments. Erythropoietin (EPO), known for its role in neurogenesis and synaptic plasticity, is significantly low in SZ patients. However, the role of EPO deficiency in SZ-associated cognitive deficits remains unclear. In this study, we used the MK801-induced SZ rat model to show that EPO levels were significantly decreased, correlating with cognitive impairments. EPO supplementation mitigated apoptosis, synaptic damage, and cognitive impairments caused by MK801. RNA-sequencing and Western blot analysis revealed increased expression of growth arrest and DNA damage 45b (GADD45b) in MK801-treated rats, reversed by EPO supplementation. Moreover, overexpression of GADD45b exacerbated neuronal loss and cognitive impairments in male Sprague-Dawley rats, while downregulation of GADD45b rescued these SZ-related pathologies. Notably, the benefits of EPO supplementation on SZ pathology were blocked by GADD45b overexpression. Inhibition of p38 MAPK, a GADD45b target, reduced MK801-induced apoptosis and synaptic damage. These findings uncover a novel etiopathogenic mechanism of SZ-related cognitive impairments, driven by EPO deficiency and the activation of the GADD45b/p38 MAPK axis.

The pro-tumor activity of INTS7 on lung adenocarcinoma via inhibiting immune infiltration and activating p38MAPK pathway

Lung adenocarcinoma (LUAD) is the most common lung cancer, accounting for 19.4% of all cancer deaths. Our previous study discovered that INTS7 expression was upregulated in LUAD, while the precise mechanism by which INTS7 exerts pro-cancer effects remains unknown. In our study, shRNA was used to knockdown the expression of INTS7 in A549 cells. Cancer behaviors in vitro were determined by CCK8 and transwell assays. Xenograft mice models were constructed to detect the tumorigenesis in vivo. Immunofluorescence and toluidine blue staining were used to test the immune infiltration. Bioinformatics analysis was adopted to predict the potential signaling pathways and construct INTS7-derived genomic prognostic model. Western blot was utilized to confirm the molecular pathways. In total, downregulation of INTS7 suppressed proliferation, invasion and migration of A549 cells, as well as tumor growth. Bioinformatics and western blot analysis indicated that p38MAPK pathway participated in the regulatory mechanism of INTS7. Moreover, INTS7 expression was negatively correlated with infiltration of memory B cells and mast cells, while positively correlated with infiltration of macrophages M2. A nomogram, including INTS7-derived risk score, was used to estimate individual’s survival probability. Generally, our findings provided comprehensive understanding of the molecular mechanisms about INTS7, and targeting INTS7 may represent a potential therapy for LUAD.

Anti-inflammatory properties of Bacillus pumilus TS1 in lipopolysaccharide-induced inflammatory damage in broilers

This study investigates whether Bacillus pumilus TS1 improves growth performance and alleviates inflammatory damage in broilers and explored its feasibility as an antibiotic alternative. We divided 240 one-day-old AA308 white-finned broilers into five groups (con, LPS, TS1L + LPS, TS1M + LPS and TS1H + LPS). The TS1L + LPS, TS1M + LPS and TS1H + LPS groups were fed TS1 for 15 days by gavage. The LPS, TS1L + LPS, TS1M + LPS and TS1H + LPS groups were injected intraperitoneally with 1 mg/kg LPS for three days. We investigated the probiotic and anti-inflammatory activities by measuring body weight, sequencing the intestinal flora and examining the structure of tissues by using pathological stain, real-time PCR, Western blotting and immunohistochemical detection. TS1 could improve growth performance and intestinal flora composition, also reduced different organ damage and inflammatory cytokine expression in serum and organs. The mechanism may involve upregulating HSP60 and HSP70 expression, targeting and regulating Nrf2 and P38 MAPK and modulating NF-κB and HO-1 expression at the transcriptional level in different organs. B. pumilus TS1 alleviated Inflammatory injury caused by LPS and attenuated the inflammatory response in broilers, and these effects were achieved through MAPK and Nrf2 regulation of HSPs/HO-1 in different organs. The above results suggested broilers fed with TS1 could release the LPS caused organ damage, and the most suggested dosage was 1.4 × 108 CFU/mL.

Erianin inhibits the progression of pancreatic cancer by directly targeting AKT and ASK1

BackgroundPancreatic cancer is a malignant tumor of the digestive tract with a high mortality rate. Erianin has antitumor activity, but the regulatory targets and mechanism of action in pancreatic cancer are unclear. The objective of this study was to evaluate the anti-pancreatic cancer activity of Erianin and explore its underlying mechanisms.MethodsA network pharmacology approach was used to investigate the mechanism of action of Erianin in pancreatic cancer cells. Cell proliferation was analyzed using CCK8, colony-formation, and EdU proliferation assays. Cell migration was evaluated through wound healing and transwell assays, as well as determination of the protein expression levels of EMT markers and β-catenin. Apoptosis and the cell cycle were measured using flow cytometry and JC-1 staining, respectively. The protein expression levels of p-Rb, CyclinB1, P21, Cleaved-PARP, and Cleaved-Caspase3 were assessed using western blotting. RNA sequencing (RNA-seq) and bioinformatics analyses were performed to elucidate the mechanism underlying the action of Erianin in pancreatic cancer. Western blotting was used to examine the expression levels of key proteins in the AKT, JNK, and p38 MAPK signaling pathways. Molecular docking and CETSA were used to test hypotheses. The tumor-suppressive ability of Erianin in vivo was assessed using a tumor-bearing assay in nude mice.ResultsNetwork pharmacology revealed that Erianin inhibited pancreatic cancer through multiple pathways. Erianin significantly inhibited pancreatic cancer cell proliferation and migration while promoting intracellular ROS and inducing apoptosis. Mechanistically, Erianin inhibited pancreatic cancer cell proliferation by regulating the AKT/FOXO1 and ASK1/JNK/p38 MAPK signaling pathways. In vivo experiments showed that Erianin inhibited subcutaneous tumor growth and promoted tumor tissue apoptosis in nude mice.ConclusionsThe component-target-pathway network revealed that Erianin exerted anti-cancer effects through multiple components, targets, and pathways. Erianin inhibited the proliferation and migration of pancreatic cancer cells and induced apoptosis through the AKT/FOXO1 and ASK1/JNK/p38 MAPK signaling pathways. These results indicate that Erianin is a promising agent for pancreatic cancer treatment.

APEX1 Knockdown Alleviates Inflammation and Fibrosis in Myocardial Infarction Through Promoting ZCCHC9 Expression and Blocking the p38 MAPK Signaling.

It was reported that serum apurinic/apyrimidinic endodeoxyribonuclease 1 (APEX1) level was higher in acute myocardial infarction (AMI) patients than in angina. This study aimed to investigate the role and mechanism of APEX1 in AMI progression. The mRNA and protein levels of APEX1 and zinc finger CCHC domain containing 9 (ZCCHC9) in blood specimens of AMI patients and normal controls were determined by RT-qPCR and Western blot assays, respectively. H9c2 cardiomyocytes were treated with angiotensin II (Ang II) to induce cardiomyocyte injury and then transfected with small interfering RNA against APEX1 (si-APEX1) or overexpression plasmids of ZCCHC9 (pcDNA-ZCCHC9). The cell viability, apoptosis, inflammatory cytokine levels, and fibrosis-associated protein expression in H9c2 cells were evaluated. ZCCHC9 promoter methylation were detected with methylation-specific PCR (MSP) assay. Then, rescue experiments were performed to explore whether APEX1 mediated cardiomyocyte functions by regulating ZCCHC9 expression. Furthermore, we explored whether the APEX1/ZCCHC9 axis regulated cardiomyocyte injury in AMI via the p38 MAPK signaling pathway. Additionally, an AMI rat model was established using the left anterior descending artery (LAD) ligation method and multipoint intramyocardial injection (5 points, 2 µL/point) of lentivirus (1 × 109 TU/mL) carrying scramble or si-APEX1 was conducted before modeling. The rats were euthanized four weeks after AMI modeling, and blood samples and myocardial tissues were harvested. The infarct area, cell apoptosis, inflammation, and fibrosis in myocardial tissues were detected. APEX1 was upregulated and ZCCHC9 was downregulated in blood samples of AMI patients compared with normal controls. APEX1 knockdown or ZCCHC9 overexpression attenuated Ang II-induced viability reduction, apoptosis, inflammation, and fibrosis in cardiomyocytes. APEX1 inhibited ZCCHC9 expression by promoting DNA methyltransferase 1 (DNMT1)-mediated ZCCHC9 promoter methylation. ZCCHC9 knockdown abolished the protective effects of APEX1 knockdown on Ang II-induced cardiomyocyte injury. APEX1 knockdown inhibited the p38 MAPK signal signaling, and anisomycin reversed the effect of APEX1 knockdown on cardiomyocyte functions. Additionally, APEX1 knockdown alleviated apoptosis, inflammation, and fibrosis in myocardial tissues of AMI rats. APEX1 knockdown attenuated Ang II-induced apoptosis, inflammation, and fibrosis in cardiomyocytes although promoting ZCCHC9 expression and inhibiting the p38 MAPK signaling pathway, thus relieving myocardial infarction, inflammation, and fibrosis in AMI rats.

Hemozoin induces malaria via activation of DNA damage, p38 MAPK and neurodegenerative pathways in a human iPSC-derived neuronal model of cerebral malaria

Malaria caused by Plasmodium falciparum infection results in severe complications including cerebral malaria (CM), in which approximately 30% of patients end up with neurological sequelae. Sparse in vitro cell culture-based experimental models which recapitulate the molecular basis of CM in humans has impeded progress in our understanding of its etiology. This study employed healthy human induced pluripotent stem cells (iPSCs)-derived neuronal cultures stimulated with hemozoin (HMZ) - the malarial toxin as a model for CM. Secretome, qRT-PCR, Metascape, and KEGG pathway analyses were conducted to assess elevated proteins, genes, and pathways. Neuronal cultures treated with HMZ showed enhanced secretion of interferon-gamma (IFN-γ), interleukin (IL)1-beta (IL-1β), IL-8 and IL-16. Enrichment analysis revealed malaria, positive regulation of cytokine production and positive regulation of mitogen-activated protein kinase (MAPK) cascade which confirm inflammatory response to HMZ exposure. KEGG assessment revealed up-regulation of malaria, MAPK and neurodegenerative diseases-associated pathways which corroborates findings from previous studies. Additionally, HMZ induced DNA damage in neurons. This study has unveiled that exposure of neuronal cultures to HMZ, activates molecules and pathways similar to those observed in CM and neurodegenerative diseases. Furthermore, our model is an alternative to rodent experimental models of CM.

EQTL mapping in transgenic alpha-synuclein carrying Caenorhabditis elegans recombinant inbred lines.

Protein aggregation of α-synuclein (αS) is a genetic and neuropathological hallmark of Parkinson's disease (PD). Studies in the model nematode Caenorhabditis elegans suggested that variation of αS aggregation depends on the genetic background. However, which genes and genetic modifiers underlie individual differences in αS pathology remains unknown. To study the genotypic-phenotypic relationship of αS aggregation, we constructed a Recombinant Inbred Line (RIL) panel derived from a cross between genetically divergent strains C. elegans NL5901 and SCH4856, both harboring the human αS gene. As a first step to discover genetic modifiers 70 αS-RILs were measured for whole-genome gene expression and expression quantitative locus analysis (eQTL) were mapped. We detected multiple eQTL hot-spots, many of which were located on Chromosome V. To confirm a causal locus, we developed Introgression Lines (ILs) that contain SCH4856 introgressions on Chromosome V in an NL5901 background. We detected 74 genes with an interactive effect between αS and the genetic background, including the human p38 MAPK homologue pmk-1 that has previously been associated with PD. Together, we present a unique αS-RIL panel for defining effects of natural genetic variation on αS pathology, which contributes to finding genetic modifiers of PD.