Phosphorylation Levels Of P38 Research Articles

Actoeside mitigated the renal proximal tubule cells damage triggered by high glucose through miR-766/VCAM1/NF-κB signalling pathway

Context Diabetic nephropathy (DN) triggered by diabetes mellitus is one of the primary causes of end-stage renal failure worldwide. Objective This study intends to explore the function and potential mechanism of actoeside on renal proximal tubule (HK-2) cells damage induced by high-glucose (HG). Methods The DN model was established in HK-2 cells with 30 mM HG treatment. The viability, apoptosis and inflammation of HK-2 cells were analysed severally via CCK-8, flow cytomery and ELISA. The key factors related to NF-κB were detected by western blotting. Results Actoeside attenuated the HG-induced HK-2 cells damage. The differentially expression of miR-766 and VCAM1 in DN patients was reversed by actoeside. Moreover, the increased phosphorylation levels of p65 NF-κB/IκBα induced by HG were attenuated by actoeside. Conclusions Actoeside promoted the growth and repressed the apoptosis and inflammation of HK-2 cells via miR-766/VCAM1/NF-κB signalling pathway, affording a promising idea for the treatment of DN.

CD14 Involvement in Third-degree Skin Burn-induced Myocardial Injury via the MAPK Signaling Pathway.

This study investigated the potential genes and related pathways in burn-induced myocardial injury. Rat myocardial injury induced by third-degree burn and the histopathological structures, apoptosis, and cardiac injury markers were then identified using hematoxylin & eosin staining, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining, and enzyme-linked immunosorbent assay. Next, differentially expressed mRNAs were screened through next-generation sequencing (NGS), followed by functional annotation and key gene validation through quantitative reverse transcription-polymerase chain reaction. Subsequently, CD14 was screened out, and small interfering RNAs against CD14 were transfected to H9C2 cells to further verify the role of CD14 in burn-induced injury. The results showed that third-degree burn could markedly damage the structure of myocardial tissue, induce the apoptosis of myocardial cells, and increase the levels of myocardial injury-related markers, suggesting that burns could induce myocardial injury in rats. Besides, NGS data discovered that third-degree burn could result in 416 differentially upregulated mRNAs and 285 differentially downregulated mRNAs in myocardial tissue. It was also disclosed that differentially expressed mRNAs were mainly enriched in the phosphatidylinositol 3-kinase/Akt, mitogen-activated protein kinase (MAPK), and tumor necrosis factor signaling pathways. Furthermore, cell viability was significantly decreased in H9C2 cells treated with 10% rat burn serum. CD14 was significantly differentially expressed and screened out for further studies. Treatment with burn serum can significantly upregulate the phosphorylation level of extracellular signal-regulated kinase, p38, and c-Jun N-terminal kinase and the expression of cleaved caspase-3 and downregulate the expression of Bcl2 when compared with those in negative control of small interfering RNA transfected H9C2 cells, whereas interfering with CD14 expression reversed the effects of burn serum. The study demonstrated that burn serum treatment could activate the MAPK signaling pathway to promote cell apoptosis, and it can be reversed by interfering with the expression of CD14.

Inhibitory Effects of Palmatine on P2X7 Receptor Expression in Trigeminal Ganglion and Facial Pain in Trigeminal Neuralgia Rats

Trigeminal Neuralgia (TN) refers to recurrent severe paroxysmal pain in the distribution area of the trigeminal nerve, which seriously affects the quality of life of patients. This research applied the chronic constriction injury of the infraorbital nerve (CCI—ION) approach to induce an animal model of TN in rats. The mechanical pain threshold of each group of rats was determined postoperatively; the expression of P2X7 receptor in trigeminal ganglion (TG) was assessed by qRT-PCR, immunofluorescence and Western blot; and the changes of the proinflammatory cytokines IL-1β and TNF-α in serum of rats were detected by ELISA. The results showed that the administration of palmatine in the TN rats could reduce the mechanical pain threshold, significantly decrease the expression of P2X7 receptor in TG, and lower the serum concentrations of IL-1β and TNF-α, compared to the sham group. In addition, the phosphorylation level of p38 in TG of TN rats was significantly decreased after treatment with palmatine. Likewise, inhibition of P2X7 expression by shRNA treatment could effectively counteract the adversary changes of pain sensitivity, IL-1β and TNF-α production, and p38 phosphorylation in TN rats. Our data suggest that palmatine may alleviate mechanical facial pain in TN rats possibly by reducing the expression of P2X7 receptor in TG of TN rats, which may be attributable to inhibiting p38 phosphorylation and reducing the release of IL-1β and TNF-α.

TRPV1 activation and internalization is part of the LPS-induced inflammation in human iPSC-derived cardiomyocytes

The non-selective cation channel transient receptor potential vanilloid 1 (TRPV1) is expressed throughout the cardiovascular system. Recent evidence shows a role for TRPV1 in inflammatory processes. The role of TRPV1 for myocardial inflammation has not been established yet. Human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (hiPSC-CM) from 4 healthy donors were incubated with lipopolysaccharides (LPS, 6 h), TRPV1 agonist capsaicin (CAP, 20 min) or the antagonist capsazepine (CPZ, 20 min). TRPV1 expression was studied by PCR and western blotting. TRPV1 internalization was analyzed by immunofluorescence. Interleukin-6 (IL-6) secretion and phosphorylation of JNK, p38 and ERK were determined by ELISA. TRPV1-associated ion channel current was measured by patch clamp. TRPV1-mRNA and -protein were expressed in hiPSC-CM. TRPV1 was localized in the plasma membrane. LPS significantly increased secretion of IL-6 by 2.3-fold, which was prevented by pre-incubation with CPZ. LPS induced TRPV1 internalization. Phosphorylation levels of ERK, p38 or JNK were not altered by TRPV1 stimulation or inhibition. LPS and IL-6 significantly lowered TRPV1-mediated ion channel current. TRPV1 mediates the LPS-induced inflammation in cardiomyocytes, associated with changes of cellular electrophysiology. LPS-induced inflammation results in TRPV1 internalization. Further studies have to examine the underlying pathways and the clinical relevance of these findings.

Benzoylaconine Modulates LPS-Induced Responses Through Inhibition of Toll-Like Receptor-Mediated NF-κB and MAPK Signaling in RAW264.7 Cells.

Previous studies have shown that benzoylaconine (BAC), a representative monoester alkaloid, has a potential anti-inflammatory effect. This study investigated the underlying molecular mechanisms using the mode of LPS-activated RAW264.7 macrophage cells. Our findings showed that BAC significantly suppressed the release of pro-inflammatory cytokines and mediators, including IL-6, TNF-α, IL-1β, ROS, NO, and PGE2. BAC treatment also effectively downregulated the elevated protein levels of iNOS and COX-2 induced by LPS in a dose-dependent manner. In this study, we found that BAC inhibited LPS-induced NF-κB activation by reducing the phosphorylation and degradation of IκBα by western blotting and blocking the nuclear translocation of p65 using an immunofluorescence assay. The elevated protein levels of JNK, p38, and ERK phosphorylation after LPS stimulation were restored effectively by BAC treatment. The protein expression of Toll-like receptor 4 (TLR4) and LPS-induced phosphorylation of TAK1, which is a crucial upstream regulatory factor of TLR-induced MAPK and NF-κB signaling, were inhibited by BAC in activated RAW264.7 macrophages. Moreover, BAC decreased the levels of TAK1 phosphorylation and pro-inflammatory cytokines and mediators associated with MAPK and NF-κB activation, similar to TLR4 inhibitor TAK-242. These findings demonstrated that BAC exhibited an anti-inflammatory effect by the inhibition of TLR-induced MAPK and NF-κB pathways, indicating that it could potentially be used for treating inflammatory diseases.

Iron Chelator Induces Apoptosis in Osteosarcoma Cells by Disrupting Intracellular Iron Homeostasis and Activating the MAPK Pathway.

Osteosarcoma is a common malignant bone tumor in clinical orthopedics. Iron chelators have inhibitory effects on many cancers, but their effects and mechanisms in osteosarcoma are still uncertain. Our in vitro results show that deferoxamine (DFO) and deferasirox (DFX), two iron chelators, significantly inhibited the proliferation of osteosarcoma cells (MG-63, MNNG/HOS and K7M2). The viability of osteosarcoma cells was decreased by DFO and DFX in a concentration-dependent manner. DFO and DFX generated reactive oxygen species (ROS), altered iron metabolism and triggered apoptosis in osteosarcoma cells. Iron chelator-induced apoptosis was due to the activation of the MAPK signaling pathway, with increased phosphorylation levels of JNK, p38 and ERK, and ROS generation; in this process, the expression of C-caspase-3 and C-PARP increased. In an orthotopic osteosarcoma transplantation model, iron chelators (20 mg/kg every day, Ip, for 14 days) significantly inhibited the growth of the tumor. Immunohistochemical analysis showed that iron metabolism was altered, apoptosis was promoted, and malignant proliferation was reduced with iron chelators in the tumor tissues. In conclusion, we observed that iron chelators induced apoptosis in osteosarcoma by activating the ROS-related MAPK signaling pathway. Because iron is crucial for cell proliferation, iron chelators may provide a novel therapeutic strategy for osteosarcoma.

Il12a Deletion Aggravates Sepsis-Induced Cardiac Dysfunction by Regulating Macrophage Polarization

Cardiac dysfunction is a well-recognized complication of sepsis and is associated with the outcome and prognosis of septic patients. Evidence suggests that Il12a participates in the regulation of various cardiovascular diseases, including heart failure, hypertension and acute myocardial infarction. However, the effects of Il12a in sepsis-induced cardiac dysfunction remain unknown. In our study, lipopolysaccharide (LPS) and cecal ligation and puncture (CLP) model were used to mimic sepsis, and cardiac Il12a expression was assessed. In addition, Il12a knockout mice were used to detect the role of Il12a in sepsis-related cardiac dysfunction. We observed for the first time that Il12a expression is upregulated in mice after LPS treatment and macrophages were the main sources of Il12a. In addition, our findings demonstrated that Il12a deletion aggravates LPS-induced cardiac dysfunction and injury, as evidenced by the increased serum and cardiac levels of lactate dehydrogenase (LDH) and cardiac creatine kinase-myocardial band (CK-MB). Moreover, Il12a deletion enhances LPS-induced macrophage accumulation and drives macrophages toward the M1 phenotype in LPS-treated mice. Il12a deletion also downregulated the activity of AMP-activated protein kinase (AMPK) but increased the phosphorylation levels of p65 (p-p65) and NF-κB inhibitor alpha (p-IκBα). In addition, Il12a deletion aggravates CLP-induced cardiac dysfunction and injury. Treatment with the AMPK activator AICAR abolishes the deterioration effect of Il12a deletion on LPS-induced cardiac dysfunction. In conclusion, Il12a deletion aggravated LPS-induced cardiac dysfunction and injury by exacerbating the imbalance of M1 and M2 macrophages. Our data provide evidence that Il12a may represent an attractive target for sepsis-induced cardiac dysfunction.

Progranulin attenuated asthma inflammation by inhibiting the expression of IL-6

Objective: To investigate the role and mechanism of progranulin (PGRN) in asthma. Methods: Control group and model group were set up in wild and IL-6 knockout (IL-6 ko) mice, respectively. For asthma model, mice were intraperitoneally sensitized with 100 μg OVA on days 0 and 7, followed by aerosol challenges with 5% OVA for 30 min per day from day 14 to 21, and mice were sacrificed 24 h after the last challenge. The mice in control group were treated in the same way with PBS. Bronchoalveolar lavage fluid (BALF) was collected for leukocytes count and differential count. The pathological changes of lung tissues were observed by H&E staining. The cytokines in lung homogenate, serum and BALF were detected by Q-PCR and ELISA. The in vitro model of asthma was induced by stimulating A549 or BEAS-2B cells with IL-13. Each group was replicated in three wells and four groups were designed: PBS group, IL-13 treatment group, IL-13 + rhPGRN treatment group, inhibitors of p38 phosphorylation (SB203508) treatment group. The cells or supernatant were collected after 0~48 h. PGRN and IL-6 levels were determined by Q-PCR and ELISA, the level of p38 phosphorylation was tested by Western blot (WB). Results: Compared with control group, PGRN levels were decreased in lung homogenate and BALF (P＜0.05), and PGRN presented a downtrend in serum, however, the level of IL-6 in BALF was increased in asthma mice (P＜0.01). In IL-6 ko asthma mice, compared with the wild asthma mice, leukocytes, especially neutrophils in BALF were decreased (P＜0.05), but PGRN was increased (P＜0.05), lung pathological damage was significantly alleviated. In vitro experiments, compared with PBS group, PGRN level was decreased (P＜0.05), IL-6 level was increased (P＜0.01), phosphorylation of p38 was activated in IL-13 treatment group. Compared with IL-13 treatment group, in IL-13 + PGRN treatment group, IL-6 level was decreased (P＜0.05); phosphorylation of p38 was inhibited (P＜0.05); and the production of IL-6 (P＜0.05) was decreased after treatment with inhibitor of p38 phosphorylation. Conclusion: PGRN inhibited the production of IL-6 by suppressing the p38 phosphorylation to alleviate asthmatic airway inflammation.

Advanced oxidation protein products trigger apoptosis and block epithelial-to-mesenchymal transition in crypt epithelial cells.

Advanced oxidation protein products (AOPPs) are uremic toxins. The present study aimed to investigate the effects of AOPPs on the epithelial mesenchymal transition (EMT) and apoptosis of rat crypt epithelial cells, and to assess the signaling pathways involved. The oxidized rat serum albumin was obtained by sodium hypochlorite modification as AOPPs, and the rat serum albumin (RSA) without sodium hypochlorite modification was set as the control. Different concentrations of AOPPs or RSA were incubated with rat crypt epithelial cells (IEC-6 cells). After culturing for 48 and 72 h, apoptosis was detected by flow cytometry. IEC-6 cells were divided into three groups: A normal group, an AOPPs group and an RSA group. Three groups of cells were collected following treatment for 2 h, and the phosphorylation levels of Akt and p65 NF-κB were detected by western blotting. After 72 h of treatment, the cells were collected and the apoptotic rate was detected by flow cytometry. The expression of EMT-related proteins was detected by reverse transcription-quantitative polymerase chain reaction and western blotting. The apoptotic rate of IEC-6 cells increased with the concentration of AOPPs, and the apoptotic rate of the AOPPs group was higher than that of the RSA group. The expression of fibronectin, snail, slug and collagen I in the AOPPs group was lower than that in the RSA group, while the expression of E-cadherin was not significantly different between the two groups. In addition, the expression of fibronectin, snail, slug and collagen I genes in the AOPPs-treated group was equal to or lower than that in the normal group. Compared with the normal group, the Akt phosphorylation level was decreased and the p65 phosphorylation level was increased in the AOPPs- or RSA-treated groups. Compared with the AOPPs-treated group, Akt and p65 phosphorylation levels in RSA-treated group were slightly higher. In conclusion, AOPPs trigger apoptosis and inhibit the EMT of rat crypt epithelial cells, which may be associated with the inhibition of Akt phosphorylation and the promotion of p65 phosphorylation.

Tetrahydrocurcumin Ameliorates Skin Inflammation by Modulating Autophagy in High-Fat Diet-Induced Obese Mice.

Obesity can induce chronic low-grade inflammation via oxidative stress. Tetrahydrocurcumin (THC) is a major curcumin metabolite with anti-inflammatory and antioxidant effects, but little is known about its effects on the skin of obese individuals. Thus, the aim of this study was to investigate the effects of THC on inflammatory cytokine production, oxidative stress, and autophagy in the skin of mice with high-fat diet- (HFD-) induced obesity. Eight-week-old C57BL/6J mice were fed a regular diet, HFD (60% of total calories from fat), or HFD supplemented with THC (100 mg/kg/day orally) for 12 weeks. We measured their body weights during the experimental period. After 12-week treatments, we performed western blotting and real-time polymerase chain reaction analyses on skin samples to evaluate the expression of inflammatory cytokines, oxidative stress markers, and autophagy markers. We observed higher tumor necrosis factor-α (TNF-α), NADPH oxidase 2 (Nox2), Nox4, and phosphorylated p65 levels; lower nuclear factor erythroid 2-related factor 2 (Nrf2) expression; and higher light chain 3 (LC3), autophagy-related 5 (Atg5), and Beclin 1 expression in the skin of HFD mice compared to the corresponding levels in the skin of mice fed with regular diet. THC administration decreased TNF-α, Nox2, Nox4, and phosphorylated p65 levels and activated the Nrf2 pathway. Interestingly, THC administration suppressed the expression of the autophagy markers LC3, Atg5, and Beclin 1. Overall, HFD-fed mice exhibited an elevation in inflammation, oxidative stress, and autophagy in their skin. THC ameliorated obesity-related skin pathology, and therefore, it is a potential therapeutic agent for obesity-related inflammatory skin diseases.

Gastrodin Attenuates Lipopolysaccharide-Induced Inflammatory Response and Migration via the Notch-1 Signaling Pathway in Activated Microglia.

Microglia-mediated neuroinflammation is known to play a pivotal role in the pathogenesis of different neurological diseases. Gastrodin, a phenolic glucoside, has been reported to exert anti-inflammatory effects in activated microglia challenged with lipopolysaccharide (LPS); however, the underlying mechanism has remained obscure. The present study aimed to ascertain if Gastrodin would regulate the Notch signaling pathway involved in microglia activation. We show here that LPS increased the expression of various members of the Notch-1 pathway, including intracellular Notch receptor domain (NICD), recombining binding protein suppressor of hairless (RBP-Jκ) and transcription factor hairy and enhancer of split-1 (Hes-1) in microglia in postnatal rat brain and in BV-2 microglia. Remarkably, Gastrodin was found to markedly attenuate the expression of the above various biomarkers both in vivo and in vitro. Moreover, increased phosphorylation level of ERK, JNK and P38 induced by LPS was attenuated with pretreatment of Notch-1 signaling inhibitor, N-[N-(3,5-difluorophenacetyl)-1-alany1-Sphenyglycinet-butylester (DAPT) as well as Gastrodin. Gastrodin mimicked the effects of DAPT by inhibiting the LPS-induced expression of IL-1β, IL-6, IL-23, TNF-α and NO. Moreover, lentivirus transfection mediated NICD overexpression inhibited the anti-inflammatory effects of Gastrodin. Furthermore, the activation of Notch-1 signaling promoted microglia migration and Gastrodin could inhibit the migration of activated BV-2 microglia by regulating the Notch-1 signaling pathway. In light of the above, our results indicate that Notch-1 signaling pathway is involved in the anti-inflammatory effects of Gastrodin against LPS-induced microglia activation. These findings provide a new biological target of Gastrodin for the treatment of neuroinflammatory disorders.

Protective effect of myricetin on LPS-induced mastitis in mice through ERK1/2 and p38 protein author.

The inflammatory reaction of mammary gland tissue in dairy cattle leads to the occurrence of mastitis disease and causes huge economic loss. Myricetin (Myr), a flavonoid natural product, is extracted from the root, stem, and leaves of Myrica rubra. It has a wide range of biological activities, such as anti-oxidant, anti-inflammatory, and anti-tumor. The purpose of this experiment is to further explore the effect of Myr on mastitis and further explore its potential mechanism in LPS-induced mice mastitis model and LPS-induced mice mammary epithelial cells (mMECs). The results showed that Myr could significantly inhibit the expression of TNF-α, IL-6, and IL-1β in the mammary gland of mice. Furthermore, the results of mechanism studies show that Myr can significantly inhibit P38 and ERK1/2 protein phosphorylation levels in mice mammary tissue, and this result has been further verified at the cellular level. These results confirm that Myr can significantly inhibit mammary inflammation, and its potential mechanism is to play a protective role by inhibiting the phosphorylation level of P38 and ERK1/2 protein.

LPS stimulates gingival fibroblasts to express PD-L1 via the p38 pathway under periodontal inflammatory conditions

ObjectiveThe overall aim of this research was to investigate the differences in the expression of programmed death ligand 1 (PD-L1) in human gingival fibroblasts (HGFs) between a periodontal healthy group and a periodontal inflammatory group. and explore the possible mechanism involved. MethodsDifferences in PD-L1 mRNA and protein expression in HGFs from a periodontal healthy group and a periodontal inflammatory group were examined by qPCR and western blotting, respectively, and were further tested after lipopolysaccharide (LPS) stimulation in both groups. The effects of a p38 pathway inhibitor on the changes in p38 phosphorylation levels and PD-L1 expression after LPS stimulation were investigated in both groups. ResultsPD-L1 mRNA and protein levels in HGFs in the periodontal inflammatory group were significantly higher than those in the periodontal healthy group (p < 0.05). After 10 μg/mL LPS stimulation, PD-L1 mRNA levels in HGFs from both groups increased significantly (p < 0.05), peaking at 4 h, and the peak was significantly higher in the periodontal inflammatory group than in the periodontal healthy group (p < 0.05). However, PD-L1 protein expression was upregulated only in the inflammatory group (p < 0.05). Inhibition of the p38 pathway in HGFs decreased p38 phosphorylation in both groups (p < 0.05) but this treatment reversed the LPS-induced increase in PD-L1 mRNA and protein levels only in the inflammatory group (p < 0.05). ConclusionIn the periodontal inflammatory state, the expression of PD-L1 in HGFs is more easily activated, and may be influenced by the p38 pathway.

Rosiglitazone alleviates lipopolysaccharide-induced inflammation in RAW264.7 cells via inhibition of NF-κB and in a PPARγ-dependent manner.

Rosiglitazone is a synthetic peroxisome proliferator-activated receptor (PPAR)γ agonist widely used for the treatment of type 2 diabetes. Recent studies have demonstrated that rosiglitazone displays anti-inflammatory effects. The present study aimed to investigate whether rosiglitazone alleviates decreases in RAW264.7 cell viability resulting from lipopolysaccharide (LPS)-induced inflammation, as well as exploring the underlying mechanism. A macrophage inflammatory injury model was established by treating RAW264.7 cells with 100 ng/ml LPS. Cells were divided into LPS and rosiglitazone groups with different concentrations. Cell viability was assessed by performing an MTT assay. The expression of inflammatory cytokines was detected by conducting enzyme-linked immunosorbent assays and reverse transcription-quantitative PCR. Nitric oxidesecretion was assessed using the Griess reagent system. The expression levels of key nuclear factor-κB pathway-associated proteins were detected via western blotting. Rosiglitazone alleviated LPS-induced decrease in RAW264.7 cell viability and inhibited inflammatory cytokine expression in a concentration-dependent manner. Rosiglitazone significantly inhibited LPS-induced upregulation of p65 phosphorylation levels and downregulated IκBα expression levels. However, rosiglitazone-mediated inhibitory effects were reversed by PPARγ knockdown. The results of the present study demonstrated that rosiglitazone significantly inhibited LPS-induced inflammatory responses in RAW264.7 macrophage cells, which was dependent on PPARγ activation and NF-κB suppression.

Mangiferin ameliorates cardiac fibrosis in D-galactose-induced aging rats by inhibiting TGF-β/p38/MK2 signaling pathway.

Aging is the process spontaneously occurred in living organisms. Cardiac fibrosis is a pathophysiological process of cardiac aging. Mangiferin is a well-known C-glucoside xanthone in mango leaves with lots of beneficial properties. In this study, rat model of cardiac fibrosis was induced by injected with 150 mg/kg/d D-galactose for 8 weeks. The age-related cardiac decline was estimated by detecting the relative weight of heart, the serum levels of cardiac injury indicators and the expression of hypertrophic biomakers. Cardiac oxidative stress and local inflammation were measured by detecting the levels of malondialdehyde, enzymatic antioxidant status and proinflammatory cytokines. Cardiac fibrosis was evaluated by observing collagen deposition via masson and sirius red staining, as well as by examining the expression of extracellular matrix proteins via Western blot analysis. The cardiac activity of profibrotic TGF-β1/p38/MK2 signaling pathway was assessed by measuring the expression of TGF-β1 and the phosphorylation levels of p38 and MK2. It was observed that mangiferin ameliorated D-galactose-induced cardiac aging, attenuated cardiac oxidative stress, inflammation and fibrosis, as well as inhibited the activation of TGF-β1/p38/MK2 signaling pathway. These results showed that mangiferin could ameliorate cardiac fibrosis in D-galactose-induced aging rats possibly via inhibiting TGF-β/p38/MK2 signaling pathway.

Double‐Headed Nanosystem‐Curcumin Therapy Ameliorates Hepatic Stress and Ocular Complications of Diabetes

Objective :Retinopathy and cataract formation are hallmarks of late-stage diabetes mellitus, and the primary contributors of vision loss. The etiologies of both ocular complications are multifaceted, but hyperglycemia-induced injuries stemming from systemic inflammation and oxidative stress are known to play major roles. Curcumin, a polyphenol with anti-inflammatory and anti-oxidant properties, may be suitable for treating diabetic complications, but its clinical translation has been hampered by poor bioavailability. Nanoparticles have been studied as means to improve the pharmacokinetic parameters of less tractable drugs. Here, we utilized a gambogic acid-conjugated nanoparticle system which had been shown to facilitate gastrointestinal uptake via the transferrin receptor, and enhance drug delivery to a variety of tissues. Using diabetic rats, we measured the pathological progression of this disease in the eye as well as the liver, an organ involved in insulin resistance and critical to blood glucose homeostasis. Method Curcumin was encapsulated in double-headed nanosystem by the emulsion-diffusion-evaporation method (Fig 1A). Streptozotocin-induced diabetic Sprague Dawley (SD) rats were divided into three groups: those given daily doses (by oral gavage) of plain curcumin (D+CUR-CMC, 40mg/kg), nanoparticle-curcumin (D+GA2-CUR, 20 mg/kg), or neither (untreated diabetic). Healthy SD rats formed the control group (C). Cataract scores, blood glucose, and body weight were recorded on a weekly basis through an 11-week trial. Tissues were harvested after euthanasia on the 12th week. Quantitation of protein and mRNA expression levels were carried out by Western blot and real-time PCR, respectively. Results Nanoparticle-curcumin treatment alone led to a significant delay in cataract formation by about one stage in the scoring metric. Plain curcumin, in spite of doubled dosage, had little or no effect on cataract formation (Figure 1, B and C). Interestingly, the nanoparticle-curcumin treatment did not reduce blood glucose (Fig 1D). Weight gain, which was inversely correlated with the severity of diabetes, appeared to be marginally improved by both curcumin treatments (Fig 1E). Analysis of liver tissues by Western blot showed marked increase in the levels of P38 phosphorylation (a stress signaling kinase) and P53 among the untreated and plain curcumin groups, whereas nanoparticle-curcumin treatment yielded levels comparable to the control group (Fig 2A). Similarly, real-time PCR detected significant down-regulation of anti-oxidant genes Txnrd1 and Mt1 in the untreated and plain curcumin groups that was attenuated in the nanoparticle-curcumin group. On the other hand, Atf3, a gene inducible by P53, was upregulated in the former groups (Fig 2B). Finally, analysis of GFAP (glial fibrillary acidic protein), a marker for gliosis and an early feature of diabetic retinopathy, revealed characteristic expression increases in the diabetic and plain curcumin groups. Meanwhile, nanoparticle-curcumin treatment resulted in greatly reduced glial cell expansion and penetration through the retinal layers (Fig 2D).

Sodium houttuyfonate relieves acute lung injury by nano micellar carrier and inhibits mitogen-activated protein kinase and nuclear factor kappa-B activation in mice

Acute lung injury (ALI) is characterized by increased pulmonary vascular permeability in response to the accumulation of inflammatory cells, release of inflammatory cytokines, and activated oxidative stress. The present study was performed to investigate the effect of sodium houttuyfonate (SH), an extract of Houttuynia cordata, on inflammatory response and oxidative stress in ALI induced by lipopolysaccharides (LPS). Male C57BL/6 mice were randomly allocated to control, LPS, dimethyl sulfoxide (DMSO), and SH groups. The ALI model was established by intratracheal LPS injection. Lung tissue was collected 6 h after LPS injection for histopathological analysis, measurement of wet-to-dry ratio, myeloperoxidase (MPO) and oxidative stress levels, and the p38, jun N-terminal kinase (JNK), extracellular regulated kinase (ERK), and p65 phosphorylation levels. Bronchoalveolar fluid (BALF) was collected for the detection of protein concentration, MPO and cytokine levels. The histopathological test showed that SH significantly alleviates damage to pulmonary tissue. Improved vascular permeability was indicated by reduced BALF protein level and lung wet-to-dry ratio in the SH group. MPO levels were decreased in lung tissue and BALF. Oxidative stress and inflammatory responses were inhibited by SH, as indicated by MDA, SOD and cytokine levels. The MAPK and NF-KB pathways were inhibited as shown by the attenuated phosphorylation of p38, JNK, ERK and p65. Sodium houttuyfonate exhibited a protective role against LPS-induced lung injury through anti-oxidative and anti-inflammatory effects. The MAPK and NF-K B pathways may be inhibited by sodium houttuyfonate. Sodium Houttuynin has a good effect on a variety of acute infectious diseases, but its solubility and stability are insufficient, which limits its efficacy. Nano delivery system can enhance the effective ingredients of traditional Chinese medicine, reduce the toxic and side effects of drugs, and improve their medicinal properties. Therefore, this paper adopts nano delivery system to assist drug use and improve research efficiency.

PLD1 promotes tumor invasion by regulation of MMP-13 expression via NF-κB signaling in bladder cancer

Invasion of bladder cancer (BC) cells from the mucosa into the muscle layer is canonical for BC progression while phospholipase D isoform 1 (PLD1) is known to mediate development of cancer through phosphatidic acid (PA) production. We therefore used in silico, in vitro and in vivo approaches to detail the effect of PLD1 on BC invasion. In BC patients, higher levels of PLD1 expression were associated with poor prognoses. PLD1 knockdown significantly suppressed cellular invasion by human BC cells and matrix metalloproteinase-13 (MMP-13) was observed to mediate this effect. In our mouse bladder carcinogenesis model, the development of invasive BCs was suppressed by PLD1 knockout and a global transcriptomic analysis in this model indicated MMP-13 as a potential tumor invasion gene with NF-κB (nuclear factor-kB) as its transcriptional regulator. Furthermore, PA administration increased MMP-13 expression in line with NF-κB p65 phosphorylation levels. Collectively, we demonstrate that PLD1 promotes tumor invasion of BC by regulation of MMP-13 expression through the NF-κB signaling pathway and that PLD1 might be a potential therapeutic target to prevent clinical progression in BC patients.

Herkinorin negatively regulates NLRP3 inflammasome to alleviate neuronal ischemic injury through activating Mu opioid receptor and inhibiting the NF-κB pathway.

Herkinorin is a novel opioid receptor agonist. Activation of opioid receptors, a member of G protein coupled receptors (GPCRs), may play an important role in Herkinorin neuroprotection. GPCRs may modulate NOD-like receptor protein 3 (NLRP3)-mediated inflammatory responses in the mechanisms of inflammation-associated disease and pathological processes. In this study, we investigated the effects of Herkinorin on NLRP3 and the underlying receptor and molecular mechanisms in oxygen-glucose deprivation/reperfusion (OGD/R)-treated rat cortex neurons. First, Western blot analysis showed that Herkinorin can inhibit the activation of NLRP3 and Caspase-1, decrease the expression of interleukin (IL)-1β, and decrease the secretion of IL-6 and tumour necrosis factor α detected by enzyme-linked immunosorbent assay in OGD/R-treated neurons. Then we found that Herkinorin downregulated NLRP3 levels by inhibiting the activation of nuclear factor kappa B (NF-κB) pathway, reducing the phosphorylation level of p65 and IκBα in OGD/R-treated neurons (p < .05 or .01, n = 3 per group). Instead, both the mu opioid receptor (MOR) inhibitor, β-funaltrexamine, and MOR knockdown reversed the effects of Herkinorin on NLRP3 (p < .05 or.01, n = 3 per group). Further, we found that the level of β-arrestin2 decreased in the cell membrane and increased in the cytoplasm after Herkinorin pretreatment in OGD/R-treated neurons. In co-immunoprecipitation experiments, Herkinorin increased the binding of IκBα with β-arrestin2, decreased the ubiquitination level of IκBα, and β-arrestin2 knockdown reversed the effects of Herkinorin on IκBα in OGD/R-treated neurons (p < .05 or .01, n = 3 per group). Our data demonstrated that Herkinorin negatively regulated NLRP3 inflammasome to alleviate neuronal ischemic injury through inhibiting NF-κB pathway mediated primarily by MOR activation. Inhibition of the NF-κB pathway by Herkinorin may be achieved by decreasing the ubiquitination level of IκBα, in which β-arrestin2 may play an important role.

Lentinula edodes extract inhibits matrix metalloproteinase expression and increases type I procollagen expression via the p38 MAPK/c-Fos signaling pathway in ultraviolet A and B-irradiated HaCaT keratinocytes

Objective: To determine the effect of Lentinula edodes extract on ultraviolet (UV) A and UVB-induced changes in matrix metalloproteinase (MMP) and type I procollagen expression using human immortalized HaCaT keratinocytes. Methods: Lentinula edodes ethanol extract (LEE) was obtained by extraction with 80% ethanol for 4 h at 80 °C. Effect of LEE on UV-induced alteration on the expression and production of MMPs and type I procollagen in keratinocytes was investigated using ELISA, RT-PCR, and Western blotting assay. To determine the underlying mechanism of LEE-mediated effects, mitogen-activated protein kinase (MAPK) and activator protein 1 signaling pathways were analysed by Western blotting assay. Results: LEE significantly inhibited the expression of MMP-1 and MMP-9 and increased the expression of type I procollagen in UVA and UVB-irradiated HaCaT keratinocytes. The phosphorylation levels of p38 were significantly inhibited by LEE whereas it did not affect c-Jun N-terminal kinase and extracellular signal-regulated kinase phosphorylation. Suppression of p38 phosphorylation was also accompanied by downregulation of UVA and UVB-induced increase in c-Fos. Conclusions: LEE effectively inhibits the expression of MMP-1 and MMP-9 and increases type I procollagen production through the p38 MAPK/c-Fos signaling pathway in UVA and UVB-irradiated HaCaT keratinocytes. This findings suggest that Lentinula edodes may be developed as a cosmetic material to suppress UV exposuremediated skin aging.