The inhibition effect of caffeic acid on NOX/ROS-dependent macrophages M1-like polarization contributes to relieve the LPS-induced mice mastitis.

The mammary gland is an adipose tissue containing not only adipocytes but also epithelial, endothelial, and immune cells. Epithelial cells and macrophages, as the integral components of the immune system, are on the front line of defense against infection. Our preliminary work proved that caffeic acid (CA) can effectively inhibit the inflammatory cascade of bovine mammary epithelial cells (BMEC) induced by lipopolysaccharide (LPS) and maintain cellular integrity and viability. Here, we investigated the therapeutic effect of CA on LPS-induced mice mastitis and explored its regulatory mechanism on macrophage inflammatory response induced by LPS in vitro. Firstly, the mice mastitis model was established by intramammary injection with 10 µg LPS, after which different CA doses (5, 10, 15 mg/kg) were administered. Then, the pathological section, myeloperoxidase (MPO) activity, proinflammatory factors and chemokines releasement, and redox state of mammary tissues were assessed, confirming CA's effectiveness on mice mastitis. In vitro, we validated the therapeutic relevance of CA in relieving LPS-induced RAW264.7 inflammatory and oxidative stress responses. Moreover, we further provided evidence that CA significantly reduced LPS-induced reactive oxygen species (ROS) generation via NADPH oxidase (NOX), which improved the imbalance relationship between nuclear factor kappa-B (NF-κB) and NF-E2 p45-related factor 2 (Nrf2) and led to a marked weakening of M1 polarization. The NOX-ROS signal inhibited by CA weakened the oxidative burst and neutrophil chemotaxis of macrophages, thus alleviating the immune cascade in mammary gland tissue and reducing the LPS-induced inflammatory damage. Collectively, CA would be a potential candidate or antibacterial synergist for curbing mastitis.

Association of weight-adjusted waist index with all-cause mortality among non-Asian individuals: a national population-based cohort study.

INTRODUCTION: The Weight-Adjusted Waist Index (WWI) is a new indicator of obesity that is associated with all-cause mortality in Asian populations. Our study aimed to investigate the linear and non-linear associations between WWI and all-cause mortality in non-Asian populations in the United States, and whether WWI was superior to traditional obesity indicators as a predictor of all-cause mortality. METHODS: We conducted a cohort study using data from the 2011-2018 National Health and Nutrition Examination Survey (NHANES), involving 18,592 participants. We utilized Cox proportional hazard models to assess the association between WWI, BMI, WC, and the risk of all-cause mortality, and performed subgroup analyses and interaction tests. We also employed a receiver operating characteristics (ROC) curve study to evaluate the effectiveness of WWI, BMI, and WC in predicting all-cause mortality. RESULTS: After adjusting for confounders, WWI, BMI, and WC were positively associated with all-cause mortality. The performance of WWI, BMI, and WC in predicting all-cause mortality yielded AUCs of 0.697, 0.524, and 0.562, respectively. The data also revealed a U-shaped relationship between WWI and all-cause mortality. Race and cancer modified the relationship between WWI and all-cause mortality, with the relationship being negatively correlated in African Americans and cancer patients. CONCLUSIONS: In non-Asian populations in the United States, there is a U-shaped relationship between WWI and all-cause mortality, and WWI outperforms BMI and WC as a predictor of all-cause mortality. These findings may contribute to a better understanding and prediction of the relationship between obesity and mortality, and provide support for effective obesity management strategies.

Polysaccharide from Atractylodes macrocephala Koidz Binding with Zinc Oxide Nanoparticles as a Novel Mucosal Immune Adjuvant for H9N2 Inactivated Vaccine.

H9N2 avian influenza poses a significant public health risk, necessitating effective vaccines for mass immunization. Oral inactivated vaccines offer advantages like the ease of administration, but their efficacy often requires enhancement through mucosal adjuvants. In a previous study, we established a novel complex of polysaccharide from Atractylodes macrocephala Koidz binding with zinc oxide nanoparticles (AMP-ZnONPs) and preliminarily demonstrated its immune-enhancing function. This work aimed to evaluate the efficacy of AMP-ZnONPs as adjuvants in an oral H9N2-inactivated vaccine and the vaccine's impact on intestinal mucosal immunity. In this study, mice were orally vaccinated on days 0 and 14 after adapting to the environment. AMP-ZnONPs significantly improved HI titers, the levels of specific IgG, IgG1 and IgG2a in serum and sIgA in intestinal lavage fluid; increased the number of B-1 and B-2 cells and dendritic cell populations; and enhanced the mRNA expression of intestinal homing factors and immune-related cytokines. Interestingly, AMP-ZnONPs were more likely to affect B-1 cells than B-2 cells. AMP-ZnONPs showed mucosal immune enhancement that was comparable to positive control (cholera toxin, CT), but not to the side effect of weight loss caused by CT. Compared to the whole-inactivated H9N2 virus (WIV) group, the WIV + AMP-ZnONP and WIV + CT groups exhibited opposite shifts in gut microbial abundance. AMP-ZnONPs serve as an effective and safe mucosal adjuvant for oral WIV, improving cellular, humoral and mucosal immunity and microbiota in the gastrointestinal tract, avoiding the related undesired effects of CT.

Laponite/lactoferrin hydrogel loaded with eugenol for methicillin-resistant Staphylococcus aureus-infected chronic skin wound healing.

Severe bacterial infections can give rise to protracted wound healing processes, thereby posing a significant risk to a patient's well-being. Consequently, the development of a versatile hydrogel dressing possessing robust bioactivity becomes imperative, as it holds the potential to expedite wound healing and yield enhanced clinical therapeutic outcomes. In this context, the present study involves the formulation of an injectable multifunctional hydrogel utilizing laponite (LAP) and lactoferrin (LF) as foundational components and loaded with eugenol (EG). This hydrogel is fabricated employing a straightforward one-pot mixing approach that leverages the principle of electrostatic interaction. The resulting LAP/LF/EG2% composite hydrogel can be conveniently injected to address irregular wound geometries effectively. Once administered, the hydrogel continually releases lactoferrin and eugenol, mitigating unwarranted oxidative stress and eradicating bacterial infections. This orchestrated action culminates in the acceleration of wound healing specifically in the context of MRSA-infected wounds. Importantly, the LAP/LF/EG2% hydrogel exhibits commendable qualities including exceptional injectability, potent antioxidant attributes, and proficient hemostatic functionality. Furthermore, the hydrogel composition notably encourages cellular migration while maintaining favorable cytocompatibility. Additionally, the hydrogel manifests noteworthy bactericidal efficacy against the formidable multidrug-resistant MRSA bacterium. Most significantly, this hydrogel formulation distinctly expedites the healing of MRSA-infected wounds by promptly inducing hemostasis, curbing bacterial proliferation, and fostering angiogenesis, collagen deposition, and re-epithelialization processes. As such, the innovative hydrogel material introduced in this investigation emerges as a promising dressing for the facilitation of bacterial-infected wound healing and consequent tissue regeneration.

Eugenol alleviates Salmonella Typhimurium-infected cecal injury by modulating cecal flora and tight junctions accompanied by suppressing inflammation.

BACKGROUND: Salmonella enterica serovar Typhimurium (ST) mainly exists in poultry and poultry related products, which are common sources of human salmonellosis. So, ST is an important zoonotic pathogen that threatens public health and safety. Eugenol has been noted for its antibacterial and anti-inflammatory properties, and it is expected to develop into an antibacterial therapy in vivo. METHODS: Herein, the effects of eugenol against ST infection in terms of intestinal flora, cecal tight junction, and cecal inflammation in broilers was evaluated in this study. RESULTS: The results showed that oral eugenol pretreatment obviously relieved the histopathological changes and ultrastructural injury of cecum caused by ST infection. Further analysis found that eugenol lessened the number of ST in the cecum, restrained Proteobacteria and Ruminococcus, and kept the ratio of Firmicutes to Bacteroidetes (F/B), improved the relative abundance of Alistipes compared to the infection control. Moreover, eugenol sustained the expression of ZO-1, claudin-1, and occluding tight junction proteins, reduced the mRNA levels of myeloid differentiation factor 88 (MyD88), toll-like receptor-4 (TLR4) and inducible nitric oxide synthesis (iNOS), suppressed the phosphorylation of p65 and IκBα in the NF-κB signaling pathway, as well as the mRNA levels of TNF-α, IL-1ß, IL-2, and IL-18 in the cecum tissue. CONCLUSION: Taken together, eugenol could protect broilers against ST infection via maintaining intestinal flora, tight junctions involved in mucosal barrier function, then accordingly limiting the excessive inflammation, finally contributed to improving relative weight gains and survival rates of broilers.

Associations between weight-adjusted waist index and bone mineral density: results of a nationwide survey.

BACKGROUND: The weight-adjusted waist circumference index (WWI) is a novel obesity indicator that offers improved accuracy in assessing both muscle and fat mass compared to traditional measures. This study aimed to investigate the association between WWI and bone mineral density (BMD) in adults. METHODS: Weighted multivariate logistic regression, subgroup analysis, interaction tests and restricted cubic spline (RCS) curves were used to explore the relationship between WWI and BMD based on data from the National Health and Nutrition Examination Survey (NHANES). RESULTS: This study had 40,568 individuals in total. At all four measurement sites, we detected a negative linear correlation between WWI and BMD. Even when quartile factors for WWI were created, this unfavorable connection maintained. In comparison to those in the lowest quartile, those in the highest percentile of WWI showed declines in lumbar BMD of 0.08 g/cm2 and femoral neck BMD of 0.03 g/cm2, respectively. This adverse correlation, nevertheless, differed among several categories. CONCLUSIONS: Our findings suggest an adverse correlation between WWI and BMD among US adults. Employing WWI as a tool for osteoporosis prevention in the general population may enhance interventions.

The synergy of tea tree oil nano-emulsion and antibiotics against multidrug-resistant bacteria.

AIMS: We determined the synergistic effects of tea tree essential oil nano-emulsion (nanoTTO) and antibiotics against multidrug-resistant (MDR) bacteria in vitro and in vivo. Then, the underlying mechanism of action of nanoTTO was investigated. METHODS AND RESULTS: Minimum inhibitory concentrations and fractional inhibitory concentration index (FICI) were determined. The transepithelial electrical resistance (TEER) and the expression of tight junction (TJ) protein of IPEC-J2 cells were measured to determine the in vitro efficacy of nanoTTO in combination with antibiotics. A mouse intestinal infection model evaluated the in vivo synergistic efficacy. Proteome, adhesion assays, quantitative real-time PCR, and scanning electron microscopy were used to explore the underlying mechanisms. Results showed that nanoTTO was synergistic (FICI ≤ 0.5) or partial synergistic (0.5 < FICI < 1) with antibiotics against MDR Gram-positive and Gram-negative bacteria strains. Moreover, combinations increased the TEER values and the TJ protein expression of IPEC-J2 cells infected with MDR Escherichia coli. The in vivo study showed that the combination of nanoTTO and amoxicillin improved the relative weight gain and maintained the structural integrity of intestinal barriers. Proteome showed that type 1 fimbriae d-mannose specific adhesin of E. coli was downregulated by nanoTTO. Then, nanoTTO reduced bacterial adhesion and invasion and inhibited the mRNA expression of fimC, fimG, and fliC, and disrupted bacterial membranes.

Associations between systemic immune-inflammation index and abdominal aortic calcification: Results of a nationwide survey.

BACKGROUND AND AIM: The Systemic Immune-Inflammation Index (SII) is a novel index of inflammation assessment that appears to be superior to the common single blood index in the assessment of cardiovascular disease. The purpose of this study was to investigate the association between SII and abdominal aortic calcification (AAC) in adults. METHODS AND RESULTS: Multivariate logistic regression, sensitivity analysis, and smoothing curve fitting were used to investigate the relationship between SII and AAC based on data from the National Health and Nutrition Examination Survey (NHANES) 2013-2014. Subgroup analysis and interaction tests were used to investigate whether this association was stable across populations. There was a positive association between SII and ACC in 3036 participants >40 years of age. In the fully adjusted model, each 100-unit increase in SII was associated with a 4% increase in the risk of developing severe AAC [1.04 (1.02, 1.07)]. Participants in the highest quartile of SII had a 47% higher risk of developing severe AAC than those in the lowest quartile [1.47 (1.10, 1.99)]. This positive association was more pronounced in older adults >60 years of age. CONCLUSIONS: SII is positively associated with AAC in US adults. Our findings imply that SII has the potential to improve AAC prevention in the general population.

Associations between cadmium exposure and whole-body aging: mediation analysis in the NHANES.

INTRODUCTION: Even though cadmium (Cd) exposure and cellular senescence (telomere length) have been linked in previous studies, composite molecular aging biomarkers are more significant and reliable factors to consider when examining the connection between metal exposure and health outcomes. The purpose of this research was to assess the association between urinary cadmium (U-Cd) and whole-body aging (phenotypic age). METHODS: Phenotypic age was calculated from chronological age and 9 molecular biomarkers. Multivariate linear regression models, subgroup analysis, and smoothing curve fitting were used to explore the linear and nonlinear relationship between U-Cd and phenotypic age. Mediation analysis was performed to explore the mediating effect of U-Cd on the association between smoking and phenotypic age. RESULTS: This study included 10,083 participants with a mean chronological age and a mean phenotypic age of 42.24 years and 42.34 years, respectively. In the fully adjusted model, there was a positive relationship between U-Cd and phenotypic age [2.13 years per 1 ng/g U-Cd, (1.67, 2.58)]. This association differed by sex, age, and smoking subgroups (P for interaction < 0.05). U-Cd mediated a positive association between serum cotinine and phenotypic age, mediating a proportion of 23.2%. CONCLUSIONS: Our results suggest that high levels of Cd exposure are associated with whole-body aging.

Dietary inflammatory potential and biological aging among US adults: a population-based study.

OBJECTIVES: The rate of biological aging is influenced by various factors such as genetics, environment, and diet. The dietary inflammatory index (DII) is strongly associated with various chronic diseases. The aim of this study was to investigate the association between DII and biological aging in US adults using quantitative indicators. METHODS: Based on data from the National Health and Nutrition Examination Survey (NHANES) 1999-2018, weighted multiple linear regression models, generalized weighted models, and smoothed fitted curves were used to investigate the linear and nonlinear relationships of DII with four biological markers of aging (biological age, phenotypic age, telomere length, and serum klotho concentration). RESULTS: A total of 35,575 adult participants with complete data were included in the study. After adjusting for all confounders, significant positive correlations were found between DII with biological age [0.070 (0.045, 0.095)] and phenotypic age [0.421 (0.371, 0.471)], with an increase of 0.07 and 0.42 years in biological age and phenotypic age, respectively, for each increase in DII score. The negative correlations between DII with telomere length [ - 0.005 ( - 0.008, - 0.002)] and klotho [ - 3.874 ( - 7.409, - 0.338)] were significant only in partially adjusted models and differed across subgroups. CONCLUSIONS: In the current study, higher DII scores (greater pro-inflammatory dietary potential) were associated with biological aging. These findings may contribute to the development of aging prevention strategies through dietary interventions.

Associations between apolipoprotein B and bone mineral density: a population-based study.

BACKGROUND: Lipids are critical in bone metabolism, and several studies have highlighted their importance. This study aimed to investigate the relationship between apolipoprotein B (apo B) and bone mineral density (BMD) at different skeletal sites (lumbar spine, femoral neck, and total femur) and to compare the influence of apo B with other traditional lipid markers. METHODS: The study included participants from the National Health and Nutrition Examination Survey (NHANES) between 2011 and 2016 who had complete data for apo B and BMD at the three skeletal sites. We used weighted multivariate regression analysis, subgroup analysis, and interaction tests to examine associations. Restricted cubic spline (RCS) was used to examine the non-linear relationship. RESULTS: A total of 4,258 adults were included in the study. Multivariate linear regression analysis showed that the relationship between apo B and BMD varied by skeletal site: a negative association was found with lumbar spine BMD [ß = -0.054, 95%CI: (-0.073, -0.035)]. In contrast, a positive association was found with femoral neck BMD [ß = 0.031, 95%CI: (0.011, 0.051)] and no significant association between apo B and total femur BMD. CONCLUSIONS: Our findings suggest that apo B is associated with BMD in a site-specific manner.

Candidate Marker Genes for Diagnosis of Osteoarthritis and Prediction of Their Regulatory Mechanisms.

We have screened candidate marker genes for the diagnosis of osteoarthritis and predicted their regulatory mechanisms. Six expression chips of tissue samples and one expression chip of peripheral blood mononuclear cell (PMBC) samples were obtained from the GEO database. Differential analysis, GSEA, and WGCNA were performed on the integra-ted tissue sample data with batch correction. Can-didate genes were obtained from the intersection of the genes significantly related to osteoarthritis in the WGCNA and the differentially expressed genes. ROC analysis was performed on the candidate genes in the tissue and PMBC samples. Genes with AUC values greater than 0.6 were retained as final candidates, and their upstream regulatory miRNAs were predicted. A total of 106 genes with differential expression were found in osteoarthritis tissue samples, which were mainly enriched in cell cycle and p53 signalling pathways. WGCNA selected a gene module significantly correlated with the occurrence of osteoarthritis. Fourteen candidate genes were obtained from the intersection of the genes in the module and the differentially expressed genes. ROC analysis showed that among these 14 candidate genes, only ADM, CX3CR1 and GADD45A had AUC values greater than 0.6 in both tissue and PMBC samples. The AUC values of the gene set of these three genes were greater than 0.7. Multiple miRNAs were predicted to be regulators of these three genes. ADM, CX3CR1 and GADD45A have potential as diagnostic marker genes for osteoarthritis and may be regulated by multiple miRNAs.

A Wireless Infrared Thermometry Device for Postoperative Flap Monitoring: Proof of Concept in Patients.

BACKGROUND/NEED: Postoperative flap perfusion assessment methods still rely on the evaluation of traditional clinical indicators, which have the disadvantage of being subjective and burdensome. METHODOLOGY: This study describes a self-designed infrared wireless thermometer for flap blood supply monitoring and evaluates its efficacy in the postoperative monitoring of 40 free flaps. DEVICE DESCRIPTION: The device consists of multiple temperature and humidity modules as well as a wireless module, which has the advantages of low cost and continuous remote monitoring. PRELIMINARY RESULTS: The alarm time of the wireless infrared thermometer was 30.5 ± 3.1 hours, and the clinical observation reported 41.7 ± 13.6 hours. CURRENT STATUS: In future studies, the device will be tested on different types of flaps in a porcine model.

A wireless infrared thermometry device for postoperative flap monitoring: Proof of concept in a porcine flap model.

The application of flap surgery is becoming more and more widespread with the development of microsurgical techniques. Currently, postoperative blood flow monitoring of flaps is still mainly assessed by medical staff for traditional clinical parameters, which has the disadvantage of being subjective and unable to monitor in real-time. This study describes a self-contained infrared wireless infrared thermometry device for flap blood supply monitoring and evaluates its effectiveness on eight porcine flap models. A scapular muscle flap model was established using eight small pigs, and the vessels were ligated at irregular intervals using a lumir ligature to simulate arterial crisis and venous crisis. Laser Doppler flowmetry (LDF), the wireless infrared thermometry device, and traditional clinical observation methods were applied to monitor the blood supply of the flap and evaluate the effect. The time to the determination of blood supply disturbance by wireless infrared thermography (IRT) was 28.75 ± 3.30 min and 96.5 ± 27.09 min for the arterial and venous groups, respectively; by LDF was 6.00 ± 1.41 min and 52.75 ± 15.76 min; by clinical observation was 42.00 ± 8.60 min and 156.50 ± 40.91 min, respectively. Paired t-tests were performed between the wireless IRT device and clinical observations, and the statistical results were significantly different in the arterial group and not significantly different in the venous group. Paired t-testing of the wireless infrared thermometry device with the LDF also showed significant differences in the arterial group and non-significant differences in the venous group. This wireless infrared thermometry device outperforms traditional clinical observation methods in monitoring blood supply in a porcine skin flap model. Because of its low cost, real-time monitoring, simple operation, and non-invasive features, it has the potential to be used in clinical practice as a routine means of postoperative blood supply monitoring in flap surgery.

Paeonol accelerates skin wound healing by regulating macrophage polarization and inflammation in diabetic rats.

Diabetic ulcer is usually seen in people with uncontrolled blood sugar. Reportedly, many factors such as impaired glucose metabolism, and macrovascular and microvascular diseases caused angiogenesis disorders and delayed the healing of diabetic ulcers, thus affecting the body's metabolism, nutrition, and immune function. This study aimed to explore the effect of paeonol on skin wound healing in diabetic rats and the related mechanism. A rat model of diabetic ulcer was established. High glucose-treated mouse skin fibroblasts were co-cultured with M1 or M2-polarized macrophages treated with or without paeonol. H&E and Masson staining were used to reveal inflammatory cell infiltration and collagen deposition, respectively. Immunohistochemistry visualized the expression of Ki67, CD31, and vascular endothelial growth factor (VEGF). Western blot was used to detect interleukin (IL)-1ß, tumor necrosis factor (TNF)-α, IL-4, IL-10, CD31, VEGFA, and collagen I/III. The expression of iNOS and arginase 1 was revealed by immunofluorescence staining. Paeonol treatment augmented collagen deposition and the expression of Ki67, CD31, VEGF, and macrophage M2 polarization markers (IL-4 and IL-10) and reduced wound area, inflammatory cell infiltration, and macrophage M1 polarization markers (IL-1ß and TNF-α) in the ulcerated area. In vitro, paeonol treatment promoted M2-polarization and repressed M1-polarization in macrophages, thereby improving the repair of cell damage induced by high glucose. Paeonol accelerates the healing of diabetic ulcers by promoting M2 macrophage polarization and inhibiting M1 macrophage polarization.

Eugenol exposure in vitro inhibits the expressions of T3SS and TIF virulence genes in Salmonella Typhimurium and reduces its pathogenicity to chickens.

BACKGROUND: Salmonella enterica serovar Typhimurium (S. Typhimurium) is a common food-borne pathogen, which has the ability to infect a wide range of hosts. The increasing emergence of drug-resistant strains urgently requires new alternative therapies. Eugenol has been shown to be very effective against drug-resistant strains of Gram-negative and Gram-positive bacteria. The purpose of this study is to explore the effects of eugenol on the virulence factors and pathogenicity of S. Typhimurium. METHODS: The antibacterial activity of eugenol was investigated via the changes of cell morphology, fimbriae related-genes and virulence factors of S. Typhimurium, then the pathogenicity of S. Typhimurium pretreated by eugenol to chickens was evaluated. RESULTS: Susceptibility testing showed that eugenol possessed significant antimicrobial activity. Scanning electron microscope analysis showed eugenol treatment deformed the morphology with damaged fimbriae structure of S. Typhimurium. Real time PCR assay confirmed eugenol significantly down-regulated the expressions of virulence factors (hilA, hilD, sipA, sipC, spiC, misL) of Type III secretion system (T3SS) and adherence genes (fimA, fimH, fimD, fimY, fimZ, stm0551) of Type I fimbriae (TIF). Animal experiment proved that the pathogenicity of S. Typhimurium exposed by eugenol was reduced, which was evidenced by the higher survival rate, weight gains and organs indexes, the lower bacterial loads in organs. Meanwhile, the duodenal histopathological changes were mitigated, with a significantly decline in the expressions of TNF-α, IL-6 and IL-18. CONCLUSION: In summary, eugenol pretreatment may alleviate the pathogenicity of the S. Typhimurium to chickens via wrecking the fimbriae and inhibiting the mRNA expressions of virulence factors and adhesion molecules. These data dedicated the potential mechanisms of eugenol against S. Typhimurium in vitro.

Bactericidal activity of gallic acid against multi-drug resistance Escherichia coli.

The continuous emergence of multidrug-resistant (MDR) bacteria has posed an increasingly serious public health threat which urges people to develop some alternatives. Gallic acid (GA) is a natural ingredient in many traditional Chinese medicines, which has many biological activities, such as antibacterial, and antiseptic. Here, clinical isolates of MDR Escherichia coli (E. coli) were used to evaluate the antibacterial effect of GA and the underlying mechanism. The results revealed that GA exerted bactericidal activity and inhibited the formation of bacterial biofilm. GA enhanced the activities of ceftiofur sodium or tetracycline against E. coli, and facilitated antibiotic accumulation in bacteria. Further analysis of morphological alterations and efflux pump gene expressions confirmed that GA damaged outer and inner membranes, and suppressed the mRNA expressions of acrA, acrB, tolC, acrD and acrF involved in membrane permeability. In addition, GA showed protective effects against bacterial infection and improved the survival rates of Galleria mellonella and BALB/c mice. These data highlight a better understanding of GA against bacteria and provide an alternative strategy for MDR bacterial infection.

Ferulic acid inhibits LPS-induced apoptosis in bovine mammary epithelial cells by regulating the NF-κB and Nrf2 signalling pathways to restore mitochondrial dynamics and ROS generation.

In bovine mammary epithelial cells (BMECs), a cascade of inflammatory reactions induced by lipopolysaccharide (LPS) has been shown to result in cell injury and apoptosis. The present study aims to reveal the protective effect of ferulic acid (FA) on LPS-induced BMEC apoptosis and explore its potential molecular mechanisms. First, we showed that FA had low cytotoxicity to BMECs and significantly decreased cell apoptosis and the proinflammatory response induced by LPS. Next, FA blocked LPS-induced oxidative stress by restoring the balance of the redox state and inhibiting mitochondrial dysfunction, the main contributor to LPS-induced apoptosis and ROS generation. Furthermore, the relief of inflammation and redox disturbance in the FA preconditioning group were accompanied by weaker NF-κB activation, enhanced Nrf2 activation and maintained cell viability compared to the LPS group. When BMECs were treated with FA alone, we observed that Nrf2 activation was induced before the inhibition of NF-κB activation and that the Keap1-Nrf2 relationship was disturbed. We concluded that FA prevented LPS-induced BMEC apoptosis by reversing the dominant relationship between NF-κB and Nrf2.

Characterization of tea tree oil nanoemulsion and its acute and subchronic toxicity.

Tea tree oil (TTO) is a popular topical use to treat skin infections. However, its poor aqueous solubility and stability have substantially limited its widespread application, including oral administration that might be therapeutic for enteric infections. In this study, mechanical ultrasonic methods were used to prepare TTO nanoemulsion (nanoTTO) with a mean droplet diameter of 161.80 nm ± 3.97, polydispersity index of 0.21 ± 0.01, and zeta potential of -12.33 ± 0.72 mV. The potential toxicity of nanoTTO was assessed by studying the oral median lethal dose (LD50) and repeated 28-day oral toxicity to provide a reference for in vivo application. Results showed that nanoTTO had no phase separation under a centrifugation test and displayed good stability during storage at -20, 4 and 25 °C over 60 days. Repeated-dose 28-day oral toxicity evaluation revealed no significant effects on growth and behavior. Assessments of hematology, clinical biochemistry, and histopathology indicated no obvious adverse effects in mice at 50, 100 and 200 mg/mL. These data suggest that nanoTTO can be considered a potential antimicrobial agent by oral administration due to its inhibitory effect on bacteria and relatively lower toxicity.

Caffeic Acid Prevented LPS-Induced Injury of Primary Bovine Mammary Epithelial Cells through Inhibiting NF-κB and MAPK Activation.

In our previous study, lipopolysaccharide (LPS) significantly reduced the cell viability of primary bovine mammary epithelial cells (bMEC) leading to cell apoptosis, which were prevented by caffeic acid (CA) through inhibiting NF-κB activation and reducing proinflammatory cytokine expression. While the underlying mechanism remains unclear, here, we determined that LPS induced the extensive microstructural damage of bMEC, especially the mitochondria and endoplasmic reticulum. Then, the obvious reduction of mitochondrial membrane potential and expression changes of apoptosis-associated proteins (Bcl-2, Bax, and casepase-3) indicated that apoptosis signaling through the mitochondria should be responsible for the cell viability decrease. Next, the high-throughput cDNA sequencing (RNA-Seq) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were employed to verify that the MAPK and JAK-STAT signaling pathways also were the principal targets of LPS. Following, the critical proteins (ERK, JNK, p38, and c-jun) of the MAPK signaling pathways were activated, and the release of proinflammatory cytokines (TNF-α, IL-1ß, IL-6, and IL-8) regulated by NF-κB and MAPKs was significantly increased, which can promote a cascade of inflammation that induces cell injury and apoptosis. Meanwhile, CA significantly inhibited the activation of MAPKs and the release of proinflammatory cytokines in a dose-dependent manner, which were similar to its effects on the NF-κB activation that we previously published. So we concluded that CA regulates the proteins located in the upstream of multiple cell signal pathways which can reduce the LPS-induced activation of NF-κB and MAPKs, thus weakening the inflammatory response and maintaining cell structure and function, which accordingly inhibit apoptosis.