Baker's yeast beta glucan supplementation was associated with an improved innate immune mRNA expression response after exercise.

Beta glucans are found in many natural sources, however, only Baker's Yeast Beta Glucan (BYBG) has been well documented to have structure-function effects that are associated with improved innate immune response to stressors (e.g., exercise, infection, etc.). The purpose was to identify a BYBG-associated mRNA expression pattern following exercise. Participants gave IRB-approved consent and were randomized to BYBG (Wellmune®; N=9) or Placebo (maltodextrin; N=10) for 6-weeks prior to performing 90 min of whole-body exercise. Paxgene blood samples were collected prior to exercise (PRE), after exercise (POST), two hours after exercise (2H), and four hours after exercise (4H). Total RNA was isolated and analyzed for the expression of 770 innate immune response mRNA (730 mRNA targets; 40 housekeepers/controls; Nanostring nCounter). The raw data were normalized against housekeeping controls and expressed as Log2 fold change from PRE for a given condition. Significance was set at p < 0.05 with adjustments for multiple comparisons and false discovery rate. We identified 47 mRNA whose expression was changed after exercise with BYBG and classified them to four functional pathways: 1) Immune Cell Maturation (8 mRNA), 2) Immune Response and Function (5 mRNA), 3) Pattern Recognition Receptors and DAMP or PAMP Detection (25 mRNA), and 4) Detection and Resolution of Tissue Damage (9 mRNA). The identified mRNA whose expression was altered after exercise with BYBG may represent an innate immune response pattern and supports previous conclusions that BYBG improves immune response to a future sterile inflammation or infection.

In silico optimization of analogs derived pro-adrenomedullin peptide to evaluate antimicrobial potential.

Diverse computational approaches have been widely used to assist in designing antimicrobial peptides with enhanced activities. This tactic has also been used to address the need for new treatment alternatives to combat resistant bacterial infections. Herein, we have designed eight variants from a natural peptide, pro-adrenomedullin N-terminal 20 peptide (PAMP), using an in silico pattern insertion approach, the Joker algorithm. All the variants show an α-helical conformation, but with differences in the helix percentages according to circular dichroism (CD) results. We found that the C-terminal portion of PAMP may be relevant for its antimicrobial activities, as revealed by the molecular dynamics, CD, and antibacterial results. The analogs showed variable antibacterial potential, but most were not cytotoxic. Nevertheless, PAMP2 exhibited the most potent activities against human and animal-isolated bacteria, showing cytotoxicity only at a substantially higher concentration than its minimal inhibitory concentration (MIC). Our results suggest that the enhanced activity in the profile of PAMP2 may be related to their particular physicochemical properties, along with the adoption of an amphipathic α-helical arrangement with the conserved C-terminus portion. Finally, the peptides designed in this study can constitute scaffolds for the design of improved sequences.

A Portable Automated Microfluidic Platform for Point-of-Care Testing for Multiple Mycotoxins in Wine.

Food safety requires point-of-care testing (POCT) for mycotoxins, since their presence in wine significantly impacts the wine industry and poses a severe threat to human life. Traditional detection methods are usually limited to detecting one mycotoxin and cannot achieve high-throughput, automated, and rapid quantitative analysis of multiple mycotoxins in real samples. Here, we propose a portable automated microfluidic platform (PAMP) integrating a chemiluminescence (CL) imaging system and a microfluidic chip to realize POCT for multiple mycotoxins in real samples, simplifying complex manual operations, shortening the detection time, and improving the detection sensitivity. Specially, silicone films were used as substrates on microfluidic chips to incubate mycotoxin conjugations, and the streptavidin-biotin (SA-B) system and an indirect immunoassay were implemented on silicone films to improve the sensitivity of reaction results. Interestingly, these methods significantly improved detection results, resulting in sensitive detection of mycotoxins, including zearalenone (ZEA) ranging from 1 to 32 ng/mL, aflatoxin B1 (AFB1) ranging from 0.2 to 6.4 ng/mL, and ochratoxin A (OTA) ranging from 2 to 64 ng/mL. The recovery of samples reached 91.39-109.14%, which verified the reliability and practicability of the PAMP. This PAMP enables sensitive and rapid detection of multiple mycotoxins in markets or wineries that lack advanced laboratory facilities. Therefore, it is essential to develop a portable microfluidic platform for POCT to detect mycotoxins in real samples.

NLRC5 senses NAD+ depletion, forming a PANoptosome and driving PANoptosis and inflammation.

NLRs constitute a large, highly conserved family of cytosolic pattern recognition receptors that are central to health and disease, making them key therapeutic targets. NLRC5 is an enigmatic NLR with mutations associated with inflammatory and infectious diseases, but little is known about its function as an innate immune sensor and cell death regulator. Therefore, we screened for NLRC5's role in response to infections, PAMPs, DAMPs, and cytokines. We identified that NLRC5 acts as an innate immune sensor to drive inflammatory cell death, PANoptosis, in response to specific ligands, including PAMP/heme and heme/cytokine combinations. NLRC5 interacted with NLRP12 and PANoptosome components to form a cell death complex, suggesting an NLR network forms similar to those in plants. Mechanistically, TLR signaling and NAD+ levels regulated NLRC5 expression and ROS production to control cell death. Furthermore, NLRC5-deficient mice were protected in hemolytic and inflammatory models, suggesting that NLRC5 could be a potential therapeutic target.

TRPM8 affects relative "cooling and heating" of subcellular organelles in microglia in a context-dependent manner.

Thermoregulation and thermal homeostasis at the cellular and subcellular organelle level are poorly understood events. In this work, we used BV2, a microglial cell line, and a series of thermo-sensitive subcellular organelle-specific probes to analyze the relative changes in the spatio-temporal temperatures of different subcellular organelles, both qualitatively and quantitatively. These methodologies allowed us to understand the thermal relationship of different subcellular organelles also. We modulated BV2 cells by pharmacological application of activator or inhibitor of TRPM8 ion channel (a cold-sensitive ion channel) and/or by treating the cells with LPS, a molecule that induces pathogen-associated molecular patterns (PAMPs) signaling. We demonstrate that the temperatures of individual organelles remain variable within a physiological range, yet vary in different conditions. We also demonstrate that treating BV2 cells by TRPM8 modulators and/or LPS alters the organelle temperatures in a specific and context-dependent manner. We show that TRPM8 modulation and/or LPS can alter the relationship of mitochondrial membrane potential to mitochondrial temperature. Our work suggests that mitochondrial temperature positively influences ER temperature and negatively influences Golgi temperature. Golgi temperature positively influences membrane temperature. This understanding of thermal relationships may be crucial for dissecting cellular structures, function, and stress signaling and may be relevant for different diseases.

Pathological remodeling of reactive astrocytes: Involvement of DNA methylation and downregulation of homeostatic genes.

Astrocytes provide metabolic support to neurons, maintain ionic and water homeostasis, and uptake and recycle neurotransmitters. After exposure to the prototypical PAMP lipopolysaccharide (LPS), reactive astrocytes increase the expression of pro-inflammatory genes, facilitating neurodegeneration. In this study, we analyzed the expression of homeostatic genes in astrocytes exposed to LPS and identified the epigenetic factors contributing to the suppression of homeostatic genes in reactive astrocytes. Primary astrocytic cultures were acutely exposed to LPS and allowed to recover for 24, 72 h, and 7 days. As expected, LPS exposure induced reactive astrogliosis and increased the expression of pro-inflammatory IL-1B and IL-6. Interestingly, the acute exposure resulted in persistent hypermethylation of astroglial DNA. Similar hypermethylation was observed in highly reactive astrocytes from the traumatic brain injury (TBI) penumbra in vivo. Hypermethylation was accompanied by decreased expression of homeostatic genes including LDHA and Scl16a1 (MCT1) both involved in the lactate shuttle to neurons; glutamine synthase (GS) responsible for glutamate processing; Kcnj10 (Kir4.1) important for K+ homeostasis, and the water channel aquaporin-4 (Aqp4). Furthermore, the master regulator of DNA methylation, MAFG-1, as well as DNA methyl transferases DNMT1 and DNMT3a were overexpressed. The downregulation of homeostatic genes correlated with increased methylation of CpG islands in their promoters, as assessed by methylation-sensitive PCR and increased DNMT3a binding to the GS promoter. Treatment with decitabine, a DNMT inhibitor, prevented the LPS- and the HMGB-1-induced downregulation of homeostatic genes. Decitabine treatment also prevented the neurotoxic effects of these astrocytes in primary cortical cultures. In summary, our findings reveal that the pathological remodeling of reactive astrocytes encompasses not only the pro-inflammatory response but, significantly, also entails a long-term suppression of homeostatic gene expression with methylation of crucial CpG islands within their promoters.

Bivalent histone modifications: how phytopathogens evade plant immunity.

Bivalent histone modifications regulate gene expression during development, but little is known about their function in plant-microbe interactions. In a recent report, Zhao et al. showed that expression of bivalent chromatin-marked gene 1 (BCG1), containing a pathogen-associated molecular pattern (PAMP) motif, is epigenetically regulated by trimethylation of lysine 4 (H3K4me3) and lysine 27 (H3K27me3) of histone H3 to evade plant immunity.

The NLR family of innate immune and cell death sensors.

Nucleotide-binding oligomerization domain (NOD)-like receptors, also known as nucleotide-binding leucine-rich repeat receptors (NLRs), are a family of cytosolic pattern recognition receptors that detect a wide variety of pathogenic and sterile triggers. Activation of specific NLRs initiates pro- or anti-inflammatory signaling cascades and the formation of inflammasomes-multi-protein complexes that induce caspase-1 activation to drive inflammatory cytokine maturation and lytic cell death, pyroptosis. Certain NLRs and inflammasomes act as integral components of larger cell death complexes-PANoptosomes-driving another form of lytic cell death, PANoptosis. Here, we review the current understanding of the evolution, structure, and function of NLRs in health and disease. We discuss the concept of NLR networks and their roles in driving cell death and immunity. An improved mechanistic understanding of NLRs may provide therapeutic strategies applicable across infectious and inflammatory diseases and in cancer.

An Evaluation of Type 1 Interferon Related Genes in Male and Female-Matched, SARS-CoV-2 Infected Individuals Early in the COVID-19 Pandemic.

SARS-CoV-2 infection has claimed just over 1.1 million lives in the US since 2020. Globally, the SARS-CoV-2 respiratory infection spread to 771 million people and caused mortality in 6.9 million individuals to date. Much of the early literature showed that SARS-CoV-2 immunity was defective in the early stages of the pandemic, leading to heightened and, sometimes, chronic inflammatory responses in the lungs. This lung-associated 'cytokine storm' or 'cytokine release syndrome' led to the need for oxygen supplementation, respiratory distress syndrome, and mechanical ventilation in a relatively high number of people. In this study, we evaluated circulating PBMC from non-hospitalized, male and female, COVID-19+ individuals over the course of infection, from the day of diagnosis (day 0) to one-week post diagnosis (day 7), and finally 4 weeks after diagnosis (day 28). In our early studies, we included hospitalized and critically care patient PBMC; however, most of these individuals were lymphopenic, which limited our assessments of their immune integrity. We chose a panel of 30 interferon-stimulated genes (ISG) to evaluate by PCR and completed flow analysis for immune populations present in those PBMC. Lastly, we assessed immune activation by stimulating PBMC with common TLR ligands. We identified changes in innate cells, primarily the innate lymphoid cells (ILC, NK cells) and adaptive immune cells (CD4+ and CD8+ T cells) over this time course of infection. We found that the TLR-7 agonist, Resiquimod, and the TLR-4 ligand, LPS, induced significantly better IFNα and IFNγ responses in the later phase (day 28) of SARS-CoV-2 infection in those non-hospitalized COVID-19+ individuals as compared to early infection (day 0 and day 7). We concluded that TLR-7 and TLR-4 agonists may be effective adjuvants in COVID-19 vaccines for mounting immunity that is long-lasting against SARS-CoV-2 infection.

Unique pathways downstream of TLR-4 and TLR-7 activation: sex-dependent behavioural, cytokine, and metabolic consequences.

Introduction: Post-infection syndromes are characterised by fatigue, muscle pain, anhedonia, and cognitive impairment; mechanistic studies exploring these syndromes have focussed on pathways downstream of Toll-like receptor (TLR) 4 activation. Here, we investigated the mechanistic interplay between behaviour, metabolism, and inflammation downstream of TLR-7 activation compared to TLR-4 activation in male and female CD1 mice. Methods: Animals received either a TLR-4 (LPS; 0.83 mg/kg) or TLR-7 (R848, 5 mg/kg) agonist, or saline, and behaviour was analysed in an Open Field (OF) at 24 h (n = 20/group). Plasma, liver, and prefrontal cortex (PFC) were collected for gene expression analysis at 24 h and 1H-NMR metabolomics. Results: TLR-4 and TLR-7 activation decreased distance travelled and rearing in the OF, but activation of each receptor induced distinct cytokine responses and metabolome profiles. LPS increased IL-1ß expression and CXCL1 in the PFC, but TLR7 activation did not and strongly induced PFC CXCL10 expression. Thus, TLR7 induced sickness behaviour is independent of IL-1ß expression. In both cases, the behavioural response to TLR activation was sexually dimorphic: females were more resilient. However, dissociation was observed between the resilient female mice behaviour and the levels of gene cytokine expression, which was, in general, higher in the female mice. However, the metabolic shifts induced by immune activation were better correlated with the sex-dependent behavioural dimorphisms; increased levels of antioxidant potential in the female brain are intrinsic male/female metabolome differences. A common feature of both TLR4 and TLR7 activation was an increase in N-acetyl aspartate (NAA) in the PFC, which is likely be an allostatic response to the challenges as sickness behaviour is inversely correlated with NAA levels. Discussion: The results highlight how the cytokine profile induced by one PAMP cannot be extrapolated to another, but they do reveal how the manipulation of the conserved metabolome response might afford a more generic approach to the treatment of post-infection syndromes.

Guanosine-specific single-stranded ribonuclease effectors of a phytopathogenic fungus potentiate host immune responses.

Plants activate immunity upon recognition of pathogen-associated molecular patterns. Although phytopathogens have evolved a set of effector proteins to counteract plant immunity, some effectors are perceived by hosts and induce immune responses. Here, we show that two secreted ribonuclease effectors, SRN1 and SRN2, encoded in a phytopathogenic fungus, Colletotrichum orbiculare, induce cell death in a signal peptide- and catalytic residue-dependent manner, when transiently expressed in Nicotiana benthamiana. The pervasive presence of SRN genes across Colletotrichum species suggested the conserved roles. Using a transient gene expression system in cucumber (Cucumis sativus), an original host of C. orbiculare, we show that SRN1 and SRN2 potentiate host pattern-triggered immunity responses. Consistent with this, C. orbiculare SRN1 and SRN2 deletion mutants exhibited increased virulence on the host. In vitro analysis revealed that SRN1 specifically cleaves single-stranded RNAs at guanosine, leaving a 3'-end phosphate. Importantly, the potentiation of C. sativus responses by SRN1 and SRN2, present in the apoplast, depends on ribonuclease catalytic residues. We propose that the pathogen-derived apoplastic guanosine-specific single-stranded endoribonucleases lead to immunity potentiation in plants.

Eudesmane sesquiterpenoids with inhibitory effects on NO production from Artemisia princeps.

Five undescribed eudesmane methyl esters (1-5), three undescribed eudesmane-12,6-olides (6-8), and 21 known analogues (9-29) were isolated from the aerial part of Artemisia princeps Pamp. Their structures were established by detailed analysis of the NMR and HRESIMS data. The absolute configurations of 1-8 were determined based on single-crystal X-ray diffraction analysis and ECD calculations. Moreover, the inhibitory effects on LPS-induced NO production in BV-2 microglial cells of all the isolated compounds were assessed. Except for compounds 2, 4, 10, and 11, the others showed significant inhibitory activities, with IC50 values of 0.73-18.66 µM, wherein the potential structure-activity relationship was also discussed.

Exploring Pseudomonas syringae pv. tomato biofilm-like aggregate formation in susceptible and PTI-responding Arabidopsis thaliana.

Bacterial biofilm-like aggregates have been observed in plants, but their role in pathogenicity is underinvestigated. In the present study, we observed that extracellular DNA and polysaccharides colocalized with green fluorescent protein (GFP)-expressing Pseudomonas syringae pv. tomato (Pst) aggregates in Arabidopsis leaves, suggesting that Pst aggregates are biofilms. GFP-expressing Pst, Pst ΔalgU ΔmucAB (Pst algU mutant), and Pst ΔalgD ΔalgU ΔmucAB (Pst algU algD mutant) were examined to explore the roles of (1) alginate, a potential biofilm component; (2) Pst AlgU, thought to regulate alginate biosynthesis and some type III secretion system effector genes; and (3) intercellular salicylic acid (SA) accumulation during pathogen-associated molecular pattern-triggered immunity (PTI). Pst formed extensive aggregates in susceptible plants, whereas aggregate numbers and size were reduced in Pst algU and Pst algD algU mutants, and both multiplied poorly in planta, suggesting that aggregate formation contributes to Pst success in planta. However, in SA-deficient sid2-2 plants, Pst algD algU mutant multiplication and aggregate formation were partially restored, suggesting plant-produced SA contributes to suppression of Pst aggregate formation. Pst algD algU mutants formed fewer and smaller aggregates than Pst algU mutants, suggesting both AlgU and AlgD contribute to Pst aggregate formation. Col-0 plants accumulated low levels of SA in response to Pst and both mutants (Pst algU and Pst algD algU), suggesting the regulatory functions of AlgU are not involved in suppressing SA-mediated plant defence. Plant PTI was associated with highly reduced Pst aggregate formation and accumulation of intercellular SA in flg22-induced PTI-responding wild-type Col-0, but not in PTI-incompetent fls2, suggesting intercellular SA accumulation by Arabidopsis contributes to suppression of Pst biofilm-like aggregate formation during PTI.

Morpho-physiological and phytohormonal changes during the induction of adventitious root development stimulated by exogenous IBA application in Magnolia biondii Pamp / Alterações morfofisiológicas e fito-hormonais durante a indução do desenvolvimento radicular adventício estimulado pela aplicação exógena do AIB em Magnolia biondii Pamp

Magnolia biondii Pamp is an important ornamental tree species widely grown and used as a rootstock in the propagation of different Magnolia varieties. In the current studies, anatomical, physiological and endogenous hormones were studied to check the effect of IBA 750 mg/L on the adventitious rooting and to provide theoretical and technical support for the propagation of Magnolia biondii Pamp through stem cuttings. Two thousand stem cuttings were prepared and divided into two groups i.e., IBA treated cuttings and water control. For the evaluation of antioxidant enzyme activities, and endogenous hormones levels, samples were collected on the day of planting and each 5th day and further steps were carried out in the laboratory according to the protocols and proper precautions. For the anatomical observations, samples were collected on the 13th, 15th, and 17th day for IBA treated cuttings while 21st, 23rd, and 25th day for control. Collected samples were preserved in the FAA solution and further observations were carried out in the laboratory. Anatomical observations showed that it took 13 days for the differentiation of root primordia to the appearance of young adventitious roots in IBA treated cuttings, while it took 21 days to develop primordia in the control. Antioxidant enzyme activities involved in ROS were significantly higher in the IBA treated cuttings compared to control. POD showed a peak on the 13th day before the emergence of roots in IBA treated cuttings while it showed a peak on the 21st day in the control. PPO showed a peak on the 21st day in the IBA treated cuttings while it showed a peak on the 29th day in the control. SOD showed a peak on the 17th day in IBA treated cuttings, while it showed a peak on the 25th day in the control. Exogenous application of IBA enhanced the endogenous IAA and GA3 levels compared to CK, while it reduced the levels of ABA continuously at the time of rooting and then increased gradually. Inclusively, our study suggests that IBA 750 mg/L is efficient for the rooting of Magnolia biondii Pamp cuttings, as it enhanced the process of antioxidant enzyme activities, endogenous hormones levels and reduced the time of root formation which is evident from the anatomical observations.

Magnolia biondii Pamp é uma importante espécie de árvore ornamental muito cultivada e utilizada como porta-enxerto na propagação de diferentes variedades de Magnolia. Nos estudos atuais, hormônios anatômicos, fisiológicos e endógenos foram estudados para verificar o efeito do AIB na dose de 750 mg / L no enraizamento adventício e fornecer suporte teórico e técnico para a propagação de M. biondii Pamp por meio de estacas. Duas mil estacas foram preparadas e divididas em dois grupos, ou seja, tratadas com AIB e controle de água. Para a avaliação das atividades das enzimas antioxidantes e dos níveis de hormônios endógenos, as amostras foram coletadas no dia do plantio e a cada 5 dias, enquanto as demais etapas foram realizadas em laboratório de acordo com os protocolos e os devidos cuidados. Para as observações anatômicas, as amostras foram coletadas no 13º, 15º e 17º dias para estacas tratadas com AIB e no 21º, 23º e 25º dias para o controle. As amostras coletadas foram preservadas em solução FAA, e outras observações foram realizadas em laboratório. Observações anatômicas mostraram a necessidade de 13 dias para a diferenciação dos primórdios radiculares até o aparecimento de raízes adventícias jovens em estacas tratadas com AIB e de 21 dias para o desenvolvimento dos primórdios no controle. As atividades das enzimas antioxidantes envolvidas nas ROS foram significativamente maiores nas estacas tratadas com AIB em comparação com o controle. A POD apresentou pico no 13º dia antes da emergência das raízes nas estacas tratadas com AIB, enquanto no 21º dia apresentou pico no controle. A PPO teve pico no 21º dia nas estacas tratadas com AIB e no 29º dia no controle. A SOD apresentou pico no 17º dia nas estacas tratadas com AIB e no 25º dia no controle. A aplicação exógena de AIB aumentou os níveis endógenos de IAA e GA3 em relação ao controle, enquanto reduziu os níveis de ABA continuamente no momento do enraizamento e, em seguida, aumentou gradativamente. Inclusive, nosso estudo sugere que o AIB na dose de 750 mg / L é eficiente para o enraizamento de estacas de M. biondii Pamp, visto que potencializou o processo de atividades de enzimas antioxidantes e os níveis de hormônios endógenos, além de reduzir o tempo de formação de raízes, o que fica evidente nas observações anatômicas.

Morpho-physiological and phytohormonal changes during the induction of adventitious root development stimulated by exogenous IBA application in Magnolia biondii Pamp

Abstract Magnolia biondii Pamp is an important ornamental tree species widely grown and used as a rootstock in the propagation of different Magnolia varieties. In the current studies, anatomical, physiological and endogenous hormones were studied to check the effect of IBA 750 mg/L on the adventitious rooting and to provide theoretical and technical support for the propagation of Magnolia biondii Pamp through stem cuttings. Two thousand stem cuttings were prepared and divided into two groups i.e., IBA treated cuttings and water control. For the evaluation of antioxidant enzyme activities, and endogenous hormones levels, samples were collected on the day of planting and each 5th day and further steps were carried out in the laboratory according to the protocols and proper precautions. For the anatomical observations, samples were collected on the 13th, 15th, and 17th day for IBA treated cuttings while 21st, 23rd, and 25th day for control. Collected samples were preserved in the FAA solution and further observations were carried out in the laboratory. Anatomical observations showed that it took 13 days for the differentiation of root primordia to the appearance of young adventitious roots in IBA treated cuttings, while it took 21 days to develop primordia in the control. Antioxidant enzyme activities involved in ROS were significantly higher in the IBA treated cuttings compared to control. POD showed a peak on the 13th day before the emergence of roots in IBA treated cuttings while it showed a peak on the 21st day in the control. PPO showed a peak on the 21st day in the IBA treated cuttings while it showed a peak on the 29th day in the control. SOD showed a peak on the 17th day in IBA treated cuttings, while it showed a peak on the 25th day in the control. Exogenous application of IBA enhanced the endogenous IAA and GA3 levels compared to CK, while it reduced the levels of ABA continuously at the time of rooting and then increased gradually. Inclusively, our study suggests that IBA 750 mg/L is efficient for the rooting of Magnolia biondii Pamp cuttings, as it enhanced the process of antioxidant enzyme activities, endogenous hormones levels and reduced the time of root formation which is evident from the anatomical observations.

Resumo Magnolia biondii Pamp é uma importante espécie de árvore ornamental muito cultivada e utilizada como porta-enxerto na propagação de diferentes variedades de Magnolia. Nos estudos atuais, hormônios anatômicos, fisiológicos e endógenos foram estudados para verificar o efeito do AIB na dose de 750 mg / L no enraizamento adventício e fornecer suporte teórico e técnico para a propagação de M. biondii Pamp por meio de estacas. Duas mil estacas foram preparadas e divididas em dois grupos, ou seja, tratadas com AIB e controle de água. Para a avaliação das atividades das enzimas antioxidantes e dos níveis de hormônios endógenos, as amostras foram coletadas no dia do plantio e a cada 5 dias, enquanto as demais etapas foram realizadas em laboratório de acordo com os protocolos e os devidos cuidados. Para as observações anatômicas, as amostras foram coletadas no 13º, 15º e 17º dias para estacas tratadas com AIB e no 21º, 23º e 25º dias para o controle. As amostras coletadas foram preservadas em solução FAA, e outras observações foram realizadas em laboratório. Observações anatômicas mostraram a necessidade de 13 dias para a diferenciação dos primórdios radiculares até o aparecimento de raízes adventícias jovens em estacas tratadas com AIB e de 21 dias para o desenvolvimento dos primórdios no controle. As atividades das enzimas antioxidantes envolvidas nas ROS foram significativamente maiores nas estacas tratadas com AIB em comparação com o controle. A POD apresentou pico no 13º dia antes da emergência das raízes nas estacas tratadas com AIB, enquanto no 21º dia apresentou pico no controle. A PPO teve pico no 21º dia nas estacas tratadas com AIB e no 29º dia no controle. A SOD apresentou pico no 17º dia nas estacas tratadas com AIB e no 25º dia no controle. A aplicação exógena de AIB aumentou os níveis endógenos de IAA e GA3 em relação ao controle, enquanto reduziu os níveis de ABA continuamente no momento do enraizamento e, em seguida, aumentou gradativamente. Inclusive, nosso estudo sugere que o AIB na dose de 750 mg / L é eficiente para o enraizamento de estacas de M. biondii Pamp, visto que potencializou o processo de atividades de enzimas antioxidantes e os níveis de hormônios endógenos, além de reduzir o tempo de formação de raízes, o que fica evidente nas observações anatômicas.

NLRP3 Inflammasome Involvement in Heart, Liver, and Lung Diseases-A Lesson from Cytokine Storm Syndrome.

Inflammation and inflammasomes have been proposed as important regulators of the host-microorganism interaction, playing a key role in morbidity and mortality due to the coronavirus disease 2019 (COVID-19) in subjects with chronic conditions and compromised immune system. The inflammasome consists of a multiprotein complex that finely regulates the activation of caspase-1 and the production and secretion of potent pro-inflammatory cytokines such as IL-1ß and IL-18. The pyrin containing NOD (nucleotide-binding oligomerization domain) like receptor (NLRP) is a family of intracellular receptors, sensing patterns associated to pathogens or danger signals and NLRP3 inflammasome is the most deeply analyzed for its involvement in the innate and adaptive immune system as well as its contribution to several autoinflammatory and autoimmune diseases. It is highly expressed in leukocytes and up-regulated in sentinel cells upon inflammatory stimuli. NLRP3 expression has also been reported in B and T lymphocytes, in epithelial cells of oral and genital mucosa, in specific parenchymal cells as cardiomyocytes, and keratinocytes, and chondrocytes. It is well known that a dysregulated activation of the inflammasome is involved in the pathogenesis of different disorders that share the common red line of inflammation in their pathogenetic fingerprint. Here, we review the potential roles of the NLRP3 inflammasome in cardiovascular events, liver damage, pulmonary diseases, and in that wide range of systemic inflammatory syndromes named as a cytokine storm.

Promoting intestinal antimicrobial defense and microbiome symbiosis contributes to IL-22-mediated protection against alcoholic hepatitis in mice.

Background: The hepatoprotective effect of interleukin 22 (IL-22) has been reported in several models of liver injuries, including alcohol-associated liver disease (ALD). However, the intestinal role of IL-22 in alcoholic hepatitis remains to be elucidated. Methods: Intestinal IL-22 levels were measured in mice fed with alcohol for 8 weeks. IL-22 was then administered to alcohol-fed mice to test its protective effects on alleviating alcoholic hepatitis, focusing on intestinal protection. Acute IL-22 treatment was conducted in mice to further explore the link between IL-22 and the induction of antimicrobial peptide (AMP). Intestinal epithelial cell-specific knockout of signal transducer and activator of transcription 3 (STAT3) mice were generated and used for organoid study to explore its role in IL-22-mediated AMP expression and gut barrier integrity. Results: After alcohol feeding for 8 weeks, the intestinal levels of IL-22 were significantly reduced in mice. IL-22 treatment to alcohol-fed mice mitigated liver injury as indicated by normalized serum transaminase levels, improved liver histology, reduced lipid accumulation, and attenuated inflammation. In the intestine, alcohol-reduced Reg3γ and α-defensins levels were reversed by IL-22 treatment. IL-22 also improved gut barrier integrity and decreased endotoxemia in alcohol-fed mice. While alcohol feeding significantly reduced Akkermansia, IL-22 administration dramatically expanded this commensal bacterium in mice. Regardless of alcohol, acute IL-22 treatment induced a fast and robust induction of intestinal AMPs and STAT3 activation. By using in vitro cultured intestinal organoids isolated from WT mice and mice deficient in intestinal epithelial-STAT3, we further demonstrated that STAT3 is required for IL-22-mediated AMP expression. In addition, IL-22 also regulates intestinal epithelium differentiation as indicated by direct regulation of sodium-hydrogen exchanger 3 via STAT3. Conclusion: Our study suggests that IL-22 not only targets the liver but also benefits the intestine in many aspects. The intestinal effects of IL-22 include regulating AMP expression, microbiota, and gut barrier function that is pivotal in ameliorating alcohol induced translocation of gut-derived bacterial pathogens and liver inflammation.

Exploring the heterogeneous transcriptional response of the CNS to systemic LPS and Poly(I:C).

Peripheral contact to pathogen-associated molecular patterns (PAMPs) evokes a systemic innate immune response which is rapidly relayed to the central nervous system (CNS). The remarkable cellular heterogeneity of the CNS poses a significant challenge to the study of cell type and stimulus dependent responses of neural cells during acute inflammation. Here we utilized single nuclei RNA sequencing (snRNAseq), serum proteome profiling and primary cell culture methods to systematically compare the acute response of the mammalian brain to the bacterial PAMP lipopolysaccharide (LPS) and the viral PAMP polyinosinic:polycytidylic acid (Poly(I:C)), at single cell resolution. Our study unveiled convergent transcriptional cytokine and cellular stress responses in brain vascular and ependymal cells and a downregulation of several key mediators of directed blood brain barrier (BBB) transport. In contrast the neuronal response to PAMPs was limited in acute neuroinflammation. Moreover, our study highlighted the dominant role of IFN signalling upon Poly(I:C) challenge, particularly in cells of the oligodendrocyte lineage. Collectively our study unveils heterogeneous, shared and distinct cell type and stimulus dependent acute responses of the CNS to bacterial and viral PAMP challenges. Our findings highlight inflammation induced dysregulations of BBB-transporter gene expression, suggesting potential translational implications on drug pharmacokinetics variability during acute neuroinflammation. The pronounced dependency of oligodendrocytes on IFN stimulation during viral PAMP challenges, emphasizes their limited molecular viral response repertoire.

Composition of Polygonatum zanlanscianense Pamp. Steam and Leaf Phenolic Extract and Its Protective Mechanism on t-BHP-Induced Oxidative Damage of HepG2 Cells.

Plant phenolic compounds have attracted considerable attention because of their health benefits. This study aimed to investigate the composition and antioxidant activity of phenol extracts from Polygonatum zanlanscianense Pamp. steam and leaf (PPP). The FTIR, UPLC-Q-Obtrip-MS, and HPLC-DAD methods were used to analyze the composition of PPP, and 20 phenolic compounds were preliminarily identified. Among them, the contents of hyperin, astragalin, and diosmetin levels were the highest. Treatment with PPP can significantly reduce t-BHP-induced cell damage in HepG2 cells, reactive oxygen species (ROS) accumulation, and malondialdehyde (MDA) content. Meanwhile, the superoxide dismutase (SOD), catalase (CAT), glutathione (GSH), and glutathione peroxidase (GSH-Px) activities can be increased. Moreover, PPP enhanced Nrf2 expression, which was consistent with that of heme oxygenase-1 (HO-1), glutamate-cysteine ligase catalytic subunit (GCLC), and NAD(P)H quinone oxidoreductase-1 (NQO1), whereas the expression of Keap1, the Nrf2 inhibitor, was decreased. All findings indicate that PPP can serve as a natural bioactive substance for preventing oxidative stress.

Superior Efficacy of Apathogenic Genotype I (V4) over Lentogenic Genotype II (LaSota) Live Vaccines against Newcastle Disease Virus Genotype VII.1.1 in Pathogen-Associated Molecular Pattern-H9N2 Vaccinated Broiler Chickens.

A comparison of the efficacy of apathogenic genotype I (V4) and lentogenic genotype II (LaSota) strains of live Newcastle disease virus (NDV) vaccines was performed following vaccination with pathogen-associated molecular pattern (PAMP) H9N2 avian influenza vaccine and challenge with velogenic NDV genotype VII.1.1 (vNDV-VII.1.1). Eight groups (Gs) of day-old chicks were used (n = 25). Groups 1-4 received a single dose of PAMP-H9N2 subcutaneously, while Gs (1, 5) and (2, 6) received eye drops of V4 and LaSota, respectively, as two doses. All Gs, except for 4 and 8, were intramuscularly challenged with vNDV-VII.1.1 at 28 days of age. No signs were detected in Gs 1, 5, 4, and 8. The mortality rates were 0% in Gs 1, 4, 5, and 8; 40% in G2; 46.66% in G6; and 100% in Gs 3 and 7. Lesions were recorded as minimal in Gs 1 and 5, but mild to moderate in Gs 2 and 6. The lowest significant viral shedding was detected in Gs 1, 2, and 5. In conclusion, two successive vaccinations of broilers with a live V4 NDV vaccine provided higher protection against vNDV-VII.1.1 challenge than LaSota. PAMP-H9N2 with live NDV vaccines induced more protection than the live vaccine alone.