Anti-inflammatory effects of amarogentin on 2,4-dinitrochlorobenzene-induced atopic dermatitis-like mice and in HaCat cells.

BACKGROUND: Amarogentin (AMA) is a secoiridoid glycoside extracted from Swertia and Gentiana roots and exhibits many biological effects such as antioxidative, anti-inflammatory, and antitumor activities. Atopic dermatitis (AD) is a chronic inflammatory skin disease caused by disorders in the regulation of multiple inflammatory cytokines. No effective cure has been found for AD now. METHODS: We constructed the HaCat and splenocyte model and tested the inhibitory effect of AMA on IL-4, IL-6, and IL-13 secretions using enzyme-linked immunosorbent assay (ELISA). The AD mouse model was constructed and treated with AMA, the severity of skin lesions was observed, epidermal tissue was collected, and epidermal thickness and mast cell infiltration were observed using hematoxylin and eosin and toluidine blue staining, respectively. The expression of kallikrein-related peptidase 7 (KLK7) and filaggrin (FLG) was detected using immunostaining and Western blot analysis. The mRNA expression of KLK7 and FLG was detected using quantitative polymerase chain reaction (qPCR). Blood immunoglobulin E (IgE) secretion was detected. RESULTS: AMA inhibited IL-6 secreted by tumor necrosis factor (TNF)-α-induced HaCaT cells and reduced IL-4 and IL-13 secreted by phytohemagglutinin (PHA)-induced primary cells in the mice spleen. It was found that the treatment of AMA with 2,4-dinitrochlorobenzene-induced AD-like mice could promote the recovery of dermatitis, reduce the score of dermatitis severity and the scratching frequency, treat the skin lesions, reduce the epidermal thickness, decrease the infiltration of mast cells, reduce the IgE level in serum, decrease the expression levels of AD-related cytokines, increase protein and mRNA expression of FLG, and reduce the protein and mRNA expression of KLK7 in the skin tissues of AD-like mice. CONCLUSION: In conclusion, AMA inhibits inflammatory response at the cellular level, and AMA reduces the validation response of specific dermatitis mice, relieves pruritus, and repairs the damaged skin barrier.

Link Between Senescence and Cell Fate: Senescence-Associated Secretory Phenotype and Its Effects on Stem Cell Fate Transition.

Senescence is a form of durable cell cycle arrest elicited in response to a wide range of stimuli. Senescent cells remain metabolically active and secrete a variety of factors collectively termed senescence-associated secretory phenotype (SASP). SASP is highly pleiotropic and can impact numerous biological processes in which it has both beneficial and deleterious roles. The underlying mechanisms by which SASP exerts its pleiotropic influence remain largely unknown. SASP serves as an environmental factor, which regulates stem cell differentiation and alters its routine. The latter can potentially be accomplished through dedifferentiation, transdifferentiation, or reprogramming. Behavioral changes that cells undergo when exposed to SASP are involved in several senescence-associated physiological and pathological phenomena. These findings provide clues for identifying possible interventions to reduce the deleterious effects without interfering in the beneficial outcomes. In this study, we discuss the multifaceted effects of SASP and the changes occurring in cellular states upon exposure to SASP factors.

Therapeutic effects of quinine in a mouse model of atopic dermatitis.

Atopic dermatitis (AD) is a chronic inflammatory skin disease that seriously affects quality of life. Quinine is a bitter taste receptor agonist that exhibits antimalarial effects. The aim of the present study was to examine the therapeutic effects of quinine in AD­like mice. AD was induced with 2,4­dinitrochlorobenzene, and the mice were treated with 10 mg/kg quinine for 1, 4 and 7 days. A total of 60 BALB/c mice were divided into the following groups: Healthy, AD­like, AD­like + quinine and healthy + quinine, with 1, 4 and 7 days groups for each treatment. Blood was extracted from all mice and ELISA was performed to detect immunoglobulin E (IgE) levels. H&E­stained tissue sections were prepared from skin lesions on the backs of the mice and pathological changes were observed. Cytokines were detected via ELISA, and the filaggrin (FLG) and kallikrein­7 (KLK7) proteins were detected via western blotting and immunohistochemistry. IKKα and NF­κB mRNA were analyzed via reverse transcription­quantitative PCR. Quinine ameliorated skin damage in the AD­like mice, reduced IgE expression in the blood, inhibited expression of IKKα and NF­κB, reduced cytokine secretion, reduced KLK7 expression, reduced scratching frequency, increased FLG expression and repaired the skin barrier. These results suggested that quinine exhibited therapeutic effects in AD­like mice.

Reducing the Allergenicity of α-Lactalbumin after Lipid Peroxidation.

This study analyzed the effect of lipid peroxidation using 2,2'-azobis(2-amidinopropane)dihydrochloride (AAPH) and acrolein on the in vitro and in vivo allergenicity of α-lactalbumin (α-La). The structure of oxidized α-La was evaluated by sodium dodecyl sulfate polyacrylamide gel electrophoresis, fluorescence spectroscopy, and circular dichroism, whereas the changes in the allergenic properties were evaluated. Lipid peroxidation induced changes to the structural properties that might destroy and/or mask α-La epitopes. In comparison to native α-La, oxidation complexes caused a decrease in the immunoglobulin E (IgE) binding capacity, as observed via immunoblotting. Moreover, the capacity to release mediators and cytokines from KU812 cells was also greatly reduced. In vivo, oxidation with AAPH and acrolein caused a significant reduction in IgE, IgG, IgG1, mast cell protease 1, and plasma histamine, along with the reduction of mast surface c-Kit+ and FcεRI+ expression. Therefore, these results indicate that oxidation via AAPH and acrolein can potentially reduce the allergenicity of α-La, which can help with the better understanding of the changes in allergenicity of milk allergen by lipid peroxidation.

The effect of different dietary structure on gastrointestinal dysfunction in children with cerebral palsy and epilepsy based on gut microbiota.

BACKGROUND: Gastrointestinal (GI) difficulties are very common among children with cerebral palsy (CP) and comorbid epilepsy. GI function is influenced by dietary structure on gut microbiota. The aim of this study was to compare gut microbiota differences in two dietary groups of this population and examine whether such differences are related to GI dysfunction. METHODS: Forty children with CP and epilepsy were recruited from a social welfare center, including 23 consuming a fluid diet (liquid diet group) and 17 consuming a normal diet (general diet group). Bacterial DNA was extracted from feces, the V3-V4 region of the 16S rRNA gene was amplified from the DNA, and high-throughput sequencing of the amplified sequences was performed. Microbe prevalence levels were compared on multiple phylogenic levels. RESULTS: Gut microbial populations differed substantially between the liquid diet group and general diet group. The only two phyla that differed significantly between the two groups were Bacteroidetes (p = 0.034) and Actinobacteria (p = 0.013). Regarding representation of genera, Prevotella species were selectively predominant in the general diet group (25.849% vs. 3.612% in the liquid diet group, p < 0.001), while Bifidobacterium species were selectively predominant in the liquid diet group (24.929% vs. 12.947% in the general diet group, p = 0.013). The gut microbiota of children in the general diet group contained more butyric acid-producing microbiota which was also common in healthy people (e.g. Lachnoclostridium, Dorea, Ruminococcus, Faecalibacterium, Roseburia, and Coprococcus). The gut microbiota of children in liquid diet group however, were rich in symbiotic pathogenic bacteria (e.g. Collinsella, Alistipes, and Eggerthella). CONCLUSION: The gut microbiota of children with CP and epilepsy consuming a liquid diet had elevated levels of symbiotic pathogens and diminished intestinal barrier protection bacteria, relative to a general diet group. These differences in bacterial microbiota were associated with GI dysfunction symptoms.

Antibacterial activity and mechanism of sanguinarine against Providencia rettgeri in vitro.

BACKGROUND: Sanguinarine (SAG), a benzophenanthridine alkaloid, occurs in Papaveraceas, Berberidaceae and Ranunculaceae families. Studies have found that SAG has antioxidant, anti-inflammatory, and antiproliferative activities in several malignancies and that it exhibits robust antibacterial activities. However, information reported on the action of SAG against Providencia rettgeri is limited in the literature. Therefore, the present study aimed to evaluate the antimicrobial and antibiofilm activities of SAG against P. rettgeri in vitro. METHODS: The agar dilution method was used to determine the minimum inhibitory concentration (MIC) of SAG against P. rettgeri. The intracellular ATP concentration, intracellular pH (pHin), and cell membrane integrity and potential were measured. Confocal laser scanning microscopy (CLSM), field emission scanning electron microscopy (FESEM), and crystal violet staining were used to measure the antibiofilm formation of SAG. RESULTS: The MIC of SAG against P. rettgeri was 7.8 µg/mL. SAG inhibited the growth of P. rettgeri and destroyed the integrity of P. rettgeri cell membrane, as reflected mainly through the decreases in the intracellular ATP concentration, pHin and cell membrane potential and significant changes in cellular morphology. The findings of CLSM, FESEM and crystal violet staining indicated that SAG exhibited strong inhibitory effects on the biofilm formation of P. rettgeri and led to the inactivity of biofilm-related P. rettgeri cells.

Successful management of seven cases of critical COVID-19 with early noninvasive-invasive sequential ventilation algorithm and bundle pharmacotherapy.

We report the clinical and laboratory findings and successful management of seven patients with critical coronavirus disease 2019 (COVID-19) requiring mechanical ventilation (MV). The patients were diagnosed based on epidemiological history, clinical manifestations, and nucleic acid testing. Upon diagnosis with COVID-19 of critical severity, the patients were admitted to the intensive care unit, where they received early noninvasive-invasive sequential ventilation, early prone positioning, and bundle pharmacotherapy regimen, which consists of antiviral, anti-inflammation, immune-enhancing, and complication-prophylaxis medicines. The patients presented fever (n = 7, 100%), dry cough (n = 3, 42.9%), weakness (n = 2, 28.6%), chest tightness (n = 1, 14.3%), and/or muscle pain (n = 1, 14.3%). All patients had normal or lower than normal white blood cell count/lymphocyte count, and chest computed tomography scans showed bilateral patchy shadows or ground glass opacity in the lungs. Nucleic acid testing confirmed COVID-19 in all seven patients. The median MV duration and intensive care unit stay were 9.9 days (interquartile range, 6.5-14.6 days; range, 5-17 days) and 12.9 days (interquartile range, 9.7-17.6 days; range, 7-19 days), respectively. All seven patients were extubated, weaned off MV, transferred to the common ward, and discharged as of the writing of this report. Thus, we concluded that good outcomes for patients with critical COVID-19 can be achieved with early noninvasive-invasive sequential ventilation and bundle pharmacotherapy.

MeCP2 inhibits proliferation and migration of breast cancer via suppression of epithelial-mesenchymal transition.

Methyl-CpG-binding protein 2 (MeCP2) is an important epigenetic regulator for normal neuronal maturation and brain glial cell function. Additionally, MeCP2 is also involved in a variety of cancers, such as breast, prostate, lung, liver and colorectal. However, whether MeCP2 contributes to the progression of breast cancer remains unknown. In the present study, we investigated the role of MeCP2 in cell proliferation, migration and invasion in vitro. We found that knockdown of MeCP2 inhibited expression of epithelial-mesenchymal transition (EMT)-related markers in breast cancer cell lines. In conclusion, our study suggests that MeCP2 inhibits proliferation and invasion through suppression of the EMT pathway in breast cancer.

Covalent conjugation with (-)-epigallo-catechin 3-gallate and chlorogenic acid changes allergenicity and functional properties of Ara h1 from peanut.

Ara h1 is a major allergen from peanut. We investigated the effect of covalent conjugation of Ara h1 and dietary polyphenols on allergenicity and functional properties of Ara h1. Enzyme-linked immunosorbent assay revealed that the covalent conjugation of dietary polyphenols significantly reduced the IgE binding capacity of Ara h1. Covalent binding of dietary polyphenols with Ara h1 reduced histamine release by 40% in basophils. The decreased IgE binding capacity of Ara h1 could be ascribed to changes in protein conformation. The IgE epitope of Ara h1 might be blocked by polyphenols at the binding site. Analysis of pepsin digestion of Ara h1-polyphenol conjugates indicated that the covalent binding increased pepsin digestibility and reduced IgE binding capacity. Furthermore, covalent conjugation of Ara h1 with polyphenols decreased denaturation temperature and increased antioxidant activity. Ara h1 conjugated with polyphenols may be a promising approach for reducing the allergenicity of Ara h1.

Ribonuclease T2 from Aspergillus fumigatus promotes T helper type 2 responses through M2 polarization of macrophages.

Allergic bronchopulmonary aspergillosis (ABPA) is an allergic immunological response to Aspergillus fumigatus (Af) exposure, which induces a strong T helper 2 (Th2) response via mechanisms that have yet to be elucidated. The aim of the present study was to investigate the hypothesis that T2 ribonuclease from Af (Af RNASET2) induces M2­type macrophage polarization to produce a T helper 2 (Th2) immune response. Recombinant Af RNASET2 (rAf RNASET2) was expressed and purified in a prokaryotic pET system and BALB/c mice were immunized with rAf RNASET2 for in vivo analyses. Expression levels of M2 polarization factors were evaluated in RAW264.7 macrophages treated with rAf RNASET2 in vitro using flow cytometry, reverse transcription­quantitative PCR, and western blot analysis. The results predicted that the mature Af RNASET2 protein (382 amino acids; GenBank no. MN593022) contained two conserved amino acid sequence (CAS) domains, termed CAS­1 and CAS­2, which are also characteristic of the RNASET2 family proteins. The protein expression levels of the Th2­related cytokines interleukin (IL)­4, IL­10, and IL­13 were upregulated in mice immunized with rAf RNASET2. RAW264.7 macrophages treated with rAf RNASET2 showed increased mRNA expression levels of M2 factors [arginase 1, Il­10, and Il­13]; however, there was no difference in cells treated with rAf RNASET2 that had been inactivated with a ribonuclease inhibitor (RNasin). The protein expression levels of IL­10 in macrophage culture supernatant were also increased following stimulation with rAf RNASET2. In addition, rAf RNASET2 upregulated the expression of phosphorylated mitogen activated protein kinases (MAPKs) in RAW264.7 cells, whereas MAPK inhibitors attenuated rAf RNASET2­induced IL­10 expression in RAW264.7 cells. In conclusion, the present study reveals that high rAf RNASET2 activity is required for rAf RNASET2­induced M2 polarization of macrophages and suggests an important immune regulatory role for Af RNASET2 in ABPA pathogenesis.

COVID-19 managed with early non-invasive ventilation and a bundle pharmacotherapy: A case report.

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic has become an immense public health burden, first in China and subsequently worldwide. Developing effective control measures for COVID-19, especially measures that can halt the worsening of severe cases to a critical status is of urgent importance. CASE SUMMARY: A 52-year-old woman presented with a high fever (38.8 °C), chills, dizziness, and weakness. Epidemiologically, she had not been to Wuhan where COVID-19 emerged and did not have a family history of a disease cluster. A blood test yielded a white blood cell count of 4.41 × 109/L (60.6 ± 2.67% neutrophils and 30.4 ± 1.34% lymphocytes). Chest imaging revealed bilateral ground-glass lung changes. Based on a positive nasopharyngeal swab nucleic acid test result and clinical characteristics, the patient was diagnosed with COVID-19. Following treatment with early non-invasive ventilation and a bundle pharmacotherapy, she recovered with a good outcome. CONCLUSION: Early non-invasive ventilation with a bundle pharmacotherapy may be an effective treatment regimen for the broader population of patients with COVID-19.

CDDO-Me Elicits Anti-Breast Cancer Activity by Targeting LRP6 and FZD7 Receptor Complex.

Aberrant activation of the Wnt/ß-catenin pathway leads to the development of multiple cancers, including breast cancer. Development of therapeutic agents against this signaling pathway is an urgent need. In this study, we found that 2-cyano-3, 12-dioxooleana-1, 9(11)-dien-28-oic acid-methyl ester (CDDO-Me) could inhibit Wnt/ß-catenin signaling mainly through targeting the low-density lipoprotein receptor-related protein (LRP) 6 and Frizzled (FZD) 7 receptor complex. This compound induced the degradation and ubiquitination of LRP6 and Fzd7 via the lysosomal pathway. We further showed that CDDO-Me mediated the degradation of FZD7 in an LRP6 ectodomain-dependent manner. In breast cancer cells, treatment with CDDO-Me increased the degradation of LRP6 and FZD7 and reduced the levels of phosphorylated Disheveled (DVL) 2 and active ß-catenin, resulting in the downregulation of Wnt target genes and several cancer stem cell (CSC) marker genes. In a murine xenograft bearing mouse mammary tumor virus (MMTV)-Wnt1-driven mammary tumor, administration of CDDO-Me significantly inhibited tumor growth and was accompanied by reduced expression of phosphorylated and total LRP6, phosphorylated and unphosphorylated DVL2, active ß-catenin, several Wnt target genes, and CSC marker genes. Collectively, the results of our study present that CDDO-Me is a potent Wnt/ß-catenin signaling inhibitor that may be a promising therapeutic agent against breast cancer. SIGNIFICANCE STATEMENT: Blocking the membrane receptor complex consisting of low-density lipoprotein receptor-related protein (LRP) 6 and Frizzled (FZD) 7 may help developing therapeutic approaches for cancers, including breast cancers. Our study indicates that 2-cyano-3, 12-dioxooleana-1, 9(11)-dien-28-oic acid-methyl ester (CDDO-Me) can inhibit Wnt/ß-catenin signaling by inducing the ubiquitination and degradation of LRP6/FZD7 membrane receptor complex via a lysosomal pathway. We also found that the ectodomain of LRP6 is essential for CDDO-Me-induced FZD7 degradation. Defining CDDO-Me as a novel inhibitor of Wnt/ß-catenin signaling, our results provide insight into the mechanism of its anticancer activity.

γ-Mangostin Ameliorates Free Fatty Acid-Induced Lipid Accumulation via the SIRT1/LKB1/AMPK Pathway in HepG2 and L02 Cells.

Lipid accumulation is a typical characteristic of nonalcoholic fatty liver disease (NAFLD). The inhibition of lipid accumulation is regarded as a potential treatment for NAFLD. In this study, we investigated the effects of γ-mangostin or α-mangostin on lipid accumulation in a cell model. Analysis of the inhibitory effects of γ-mangostin on lipid accumulation revealed that it downregulated NAFLD-related biochemical parameters and stimulated the SIRT1/LKB1/AMPK pathway. Consequently, it suppressed lipid synthesis and enhanced fatty acid oxidation. Moreover, we demonstrated that the blockage of AMP-activated protein kinase (AMPK) by the pharmacological inhibitor Compound C abrogated the promoting effect of AMPK. Similar results were also observed for α-mangostin. The effects of α-mangostin on lipid accumulation were inferior to those of γ-mangostin. The differences in CPT1A activity might be originated from their different chemical structures. Our results suggested that γ-mangostin and α-mangostin can be exploited as potential candidates for NAFLD treatment.