TNF-α Gene Expression Research Articles

Pro-inflammatory regulatory mechanism of C5a/C5aR pathway in GCRV-infected grass carp (Ctenopharyngodon idella)

Death of grass carp (Ctenopharyngodon idella) caused by grass carp reovirus (GCRV) infection severely restricts the development of the grass carp industry. The interaction of C5a and C5a receptor (C5aR) in the complement system promotes inflammation and aggravates bleeding in grass carps. To reveal the regulatory mechanism of the C5a/C5aR pro-inflammatory response after GCRV infection, the expression of C5 in the liver and muscle tissues of grass carp after GCRV infection was analyzed using immunofluorescence. The effects of over expression of C5a and the C5aR antagonist PMX205 in grass carp kidney cell line (CIK) on the phosphorylation of p38 and ERK and the gene and protein expression of IL-6 and TNF-α were also analyzed using RT-qPCR and western blot. The results showed that the expression of the C5a protein in the liver and muscle of grass carp infected with GCRV first increased and then decreased, and the C5a protein level peaked on the 5th day after infection. In CIK cells, over expression of C5a significantly increased C5aR expression, phosphorylation of p38 and ERK, and overexpression of IL-6 and TNF-α. Treatment of CIK cells with PMX205 significantly decreased the expression of C5a and C5aR, phosphorylation of p38 and ERK proteins, and expression of IL-6 and TNF-α after GCRV infection. These results implied that the interaction of C5a and C5aR in grass carp resulted in the phosphorylation of p38 and ERK proteins, a significant increase in the expression of IL-6 and TNF-α, and ultimately induction of an inflammatory response of grass carp infected with GCRV. These results provide a theoretical basis for relieving grass carp hemorrhagic diseases by regulating the interaction of C5a and C5aR.

Effect of Dayak Onion (Eleutherine bulbosa (Mill.) Urb.) on the Immune Response and Gene Expression of Nile Tilapia (Oreochromis niloticus) Infected with Aeromonas hydrophila

Abstract Dayak onion (Eleutherine bulbosa (Mill.) Urb.) has the potential to be an immunostimulant to benefit fish health. This study aimed to evaluate the effectiveness of dietary supplementation with the powder and crude ethanol extracts of Dayak onion on the hematology and immune response parameters of Nile tilapia challenged with Aeromonas hydrophila. The research used a completely randomized design with six treatments and three replications. Nile tilapias were fed with Dayak onion (w/v) consisting of powders of 5% (P5), 10% (P10), and 15% (P15), while crude extract was 0.5% (E05) and given during rearing for 30 days. Positive control (C+) and negative control (C-) were used without Dayak onion. Nile tilapia in treatment C+, P5, P10, and P15 were injected with A. hydrophila 10⁶ CFU mL⁻1, while that in treatment C- was injected with phosphate buffered saline (PBS) intramuscularly. The results showed that the dietary with 15% powder and 0.5% crude extract significantly improved the hematology and immune response parameters compared to the control after the challenge against A. hydrophila (P<0.05). Dayak onion supplementation effectively increased the health status based on hematology and immune response parameters of Nile tilapia against A. hydrophila infection. Highlight Research Dayak onion has significant potential to improve nonspecific immunity in fish. Dayak onion improves hematology and immune response of Nile tilapia. Dayak onion can significantly regulate the immune system against A. hydrophila infection. Dayak onion significantly increases the production of the gene expression of IL-1β and TNF-α.

Development and characterization of phage display-derived anti-toxin antibodies neutralizing TcdA and TcdB of Clostridioides difficile.

TcdA and TcdB are known as the major virulence attributes of Clostridioides difficile. Hence, neutralizing the TcdA and TcdB activities can be considered as an efficient therapeutic approach against C. difficile infection (CDI). In this work, we utilized phage display technique to select single-chain fragment variable (scFv) fragments as recombinant antibodies displayed on the surface of phages, which specifically target native TcdA, or TcdB (nTcdA and nTcdB), and their recombinant C-terminal combined repetitive oligopeptide (CROP) domains (rTcdA and rTcdB). After three rounds of biopanning, abundance of phage clones displaying high reactivity with TcdA or TcdB was quantified through enzyme-linked immunosorbent assay (ELISA). Furthermore, selected scFvs were characterized by cell viability and neutralization assays. The gene expression of immunological markers, IL-8 and TNF-α, was examined in treated Caco-2 cells by RT-qPCR. The epitopes of neutralizing scFvs were also identified by molecular docking. Totally, 18 scFv antibodies (seven for TcdA and 11 for TcdB) were identified by ELISA. Among selected scFvs, two clones for TcdA (rA-C2, A-C9) and three clones for TcdB (rB-B4, B-F5, B-F11) exhibited the highest neutralizing activity in Caco-2 and Vero cells. Moreover, the cocktail of anti-TcdA and anti-TcdB antibodies notably decreased the mRNA expression of TNF-α and IL-8 in Caco-2 cells. Molecular docking revealed that the interaction between scFv and toxin was mostly restricted to CROP domain of TcdA or TcdB. Our results collectively provided more insights for the development of neutralizing scFvs against C. difficile toxins using phage display. Further research is needed to meticulously evaluate the potential of scFvs as an alternative treatment for CDI using animal models and clinical trials.IMPORTANCETargeting the major toxins of Clostridioides difficile by neutralizing antibodies is a novel therapeutic approach for CDI. Here, we report a panel of new anti-TcdA (rA-C2, A-C9) and anti-TcdB (rB-B4, B-F5, and B-F11) recombinant antibody fragments (scFvs) isolated from Tomlinson I and J libraries using phage display technique. These scFv antibodies were capable of neutralizing their respective toxin and showed promise as potential therapeutics against TcdA and TcdB of C. difficile in different in vitro models. In addition, in silico analysis showed that at least two neutralization mechanisms, including inhibiting cell surface binding of toxins and inhibiting toxin internalization can be proposed for the isolated scFvs in this work. These findings provide more insights for the applicability of specific scFvs toward C. difficile toxins at in vitro level. However, further research is required to evaluate the potential application of these scFvs as therapeutic agents for CDI treatment in clinical setting.

High affinity of protocatechuic acid to human serum albumin and modulatory effects against oxidative stress and inflammation in alveolar epithelial cells: Modulation of pulmonary fibrosis

Protocatechuic acid (PCA), C7H6O4, has been shown to possess potential antioxidant properties. But its interaction with the main plasma carrier protein, human serum albumin (HSA), as well as its antioxidant mechanism remains largely unknown. It has been shown that induced pulmonary fibrosis can be modulated through mitigating epithelial apoptosis mediated by the prohibition of oxidative stress. Therefore, in this study, the interaction of PCA and HSA was explored by spectroscopy, calorimetry (DSC), as well as molecular docking studies. Also, the protective effects of PCA against lipopolysaccharide (LPS)-induced cytotoxicity and oxidative stress were evaluated by MTT, ROS, ELISA, real-time PCR, and western blot assays. It was shown that under the interaction of HSA with PCA a spontaneous interaction occurs with the contribution of hydrophobic forces, which results in the formation of a stable complex. Cellular assays showed that PCA reduced LPS-induced cytotoxicity in human type II alveolar epithelial cells (AECs) through mitigation of ROS production, release and gene expression of TNf-α and IL-1β as pro-inflammatory mediators, and caspase-3 gene and protein as an apoptotic factor. Also, it was shown that PCA can reduce the expression of NF-κB at the protein level, indicating a possible inhibition of pulmonary fibrosis via regulating the NF-κB signaling pathway. In conclusion, PCA could be a promising therapeutic agent for the control of oxidative stress in AECs which is an important factor in redox modulatory therapy, while its pharmacodynamics can be modulated by interaction with HSA.

Glucomannan enhanced the macrophage activity in exposure to methicillin-resistant Staphylococcus aureus (MRSA): in-vitro study.

The increasing number of methicillin-resistant Staphylococcus aureus persuade the need for preventive measures. Glucomannan is a polysaccharide choice for developing immunological strategies. This study aimed to investigate changes in gene expression and phagocytic activity of macrophage cells in the presence of glucomannan. The effect of different concentrations of glucomannan (25, 50, and 100 μg/mL) on the phagocytic activity of macrophage cells was measured using the colony count method. The expression of Tumor Necrosis Factor-alpha (TNF-α) and Inducible Nitric Oxide Synthase (iNOS) genes was evaluated by Real-Time PCR. The concentrations of glucomannan significantly reduced the bacterial Colony-Forming Unit (CFU) and increased the phagocytic activity of macrophage cells. The maximum effect of glucomannan on iNOS and TNF-A genes expression was 100 μg/mL. Glucomannan should be considered an adjuvant that stimulates the immune system. It may increase the expression of TNF-α and iNOS genes and the phagocytic activity of macrophage cells against methicillin-resistant Staphylococcus aureus.

Disease activity in chronic inflammatory demyelinating polyneuropathy: association between circulating B-cell subsets, cytokine levels, and clinical outcomes.

Chronic inflammatory demyelinating polyneuropathy (CIDP), a common and treatable autoimmune neuropathy, is frequently misdiagnosed. The aim of this study is to evaluate the relationship between immunological markers and clinical outcome measures in a mixed cohort of patients with typical CIDP and CIDP variants at different disease stages. Twenty-three typical, 16 multifocal and five distal CIDP patients were included. Twenty-five sex and age-matched healthy controls and 12 patients with Charcot-Marie-Tooth type 1A (CMT1A) disease served as controls. Peripheral B-cell populations were analyzed by flow cytometry. IL6, IL10, TNFA mRNA and mir-21, mir-146a, and mir-155-5p expression levels were evaluated by real-time polymerase chain reaction in peripheral blood mononuclear cells (PBMC) and/or skin biopsy specimens. Results were then assessed for a possible association with clinical disability scores and intraepidermal nerve fiber densities (IENFD) in the distal leg. We detected a significant reduction in naive B cells (P ≤ 0.001), plasma cells (P ≤ 0.001) and regulatory B cells (P < 0.05), and an elevation in switched memory B cells (P ≤ 0.001) in CIDP compared to healthy controls. CMT1A and CIDP patients had comparable B-cell subset distribution. CIDP cases had significantly higher TNFA and IL10 gene expression levels in PBMC compared to healthy controls (P < 0.05 and P ≤ 0.01, respectively). IENFDs in the distal leg showed a moderate negative correlation with switched memory B-cell ratios (r = -0.51, P < 0.05) and a moderate positive correlation with plasma cell ratios (r = 0.46, P < 0.05). INCAT sum scores showed a moderate positive correlation with IL6 gene expression levels in PBMC (r = 0.54, P < 0.05). Altered B-cell homeostasis and IL10 and TNFA gene expression levels imply chronic antigen exposure and overactivity in the humoral immune system, and seem to be a common pathological pathway in both typical CIDP and CIDP variants.

Sleeve Gastrectomy Decrease Body Weight, Fasting Blood Glucose, and Gene Expression of TNF-α and IL-1 in The Abdominal Aorta of Rats with Obesity and Diabetes Mellitus

BACKGROUND: Sleeve gastrectomy (SG) is one option to significantly reduce body weight while also protect the cardiovascular system by controlling hyperglycemia and inflammatory markers. Secretion of tumor necrosis factor (TNF)-α and interleukin (IL)-1 could induce obesity- and diabetes mellitus (DM)-related inflammation, however its association with SG procedure has not been elucidated well. Therefore, TNF-α and IL-1 gene expression on the abdominal aorta of obese and DM rats that went through SG procedure were evaluated.METHODS: Fifteen rats were divided into 3 groups: lean-non-DM rats model (C1 group), obese-DM rats model (C2 group), and obese-DM rats model underwent SG (T group). Before and 10 days after the SG procedure, rats’ body weight and fasting blood glucose (FBG) were measured. Ten days after the procedure, TNF-α and IL-1 gene expression were also evaluated by PCR.RESULTS: In the end of study, mean body weight and FBG levels in C1 group (231.80±4.32 gram; 68.60±2.07 mg/dL) and T group (232.00±5.33 gram; 114.40±3.20 mg/dL) were significantly lower than in C2 group (264.60± 3.28 gram; 271.00±6.89 mg/dL). TNF-α and IL-1 gene expressions were also found to be significantly lower in the C1 group (1.01±0.01 rfu; 1.01±0.01 rfu) and T group (1.97±0.57 rfu; 1.21±0.78 rfu) compared to the C2 group (224.12±47.59 rfu; 1.85±0.73 rfu).CONCLUSION: SG could decrease body weight and FBG, as well as TNF-α and IL-1 gene expression in the abdominal aorta of rats with obesity and DM, hence SG could be a useful method in reducing body weight and controlling hyperglycemia and inflammatory markers.KEYWORDS: sleeve gastrectomy, TNF-α, interleukin-1, obesity, diabetes mellitus

Effects of heat stress abatement on systemic and mammary inflammation in lactating dairy cows

To examine the effects of evaporative cooling on systemic and mammary inflammation of lactating dairy cows, 30 multiparous Holstein cows (parity = 2.4, 156 d in milk) were randomly assigned to 1 of 2 treatments: cooling (CL) with fans and misters or not (NC). The experiment was divided into a 10-d baseline when all cows were cooled, followed by a 36-d environmental challenge when cooling was terminated for NC cows. The onset of environmental challenge was considered as d 1. Temperature-humidity index averaged 78.4 during the environmental challenge. Milk yield and dry matter intake (DMI) were recorded daily. Blood and milk samples were collected from a subset of cows (n = 9/treatment) on d -3, 1, 3, 7, 14, and 28 of the experiment to measure cortisol, interleukin 10 (IL10), tumor necrosis factor-α (TNF-α), haptoglobin, and lipopolysaccharide binding protein (LBP). Mammary biopsies were collected from a second subset of cows (n = 6/treatment) on d -9, 2, 10, and 36 to analyze gene expression of cytokines and haptoglobin. A subset of cows (n = 7/treatment) who were not subjected to mammary biopsy collection received a bolus of lipopolysaccharides (LPS) in the left rear quarter on d 30 of the experiment. Blood was sampled from cows and milk samples from the LPS-infused quarter were collected at -4, 0, 3, 6, 12, 24, 48, and 96 h relative to infusion, for analyses of inflammatory products. Deprivation of cooling decreased milk yield and DMI. Compared with CL cows, plasma cortisol concentration of NC cows was higher on d 1 but lower on d 28 of the experiment (cooling × time). Deprivation of cooling did not affect circulating TNF-α, IL10, haptoglobin, or LBP. Compared with CL cows, NC cows tended to have higher milk IL10 concentrations but did not show effects in TNF-α, haptoglobin, or LBP. No differences were observed in mammary tissue gene expression of TNF-α, IL10, and haptoglobin. Milk yield declined after LPS infusion but was not affected by treatment. Compared with CL cows, NC cows had greater milk somatic cell count following intramammary LPS infusion. Non-cooled cows had lower circulating TNF-α and IL10 concentrations and tended to have lower circulating haptoglobin concentrations than CL cows. Milk IL10 and TNF-⍺ concentrations were higher 3 h after LPS infusion for NC cows compared with CL cows. Additionally, NC cows tended to have higher milk haptoglobin concentration after LPS infusion than CL cows. In conclusion, deprivation of evaporative cooling had minimal effects on lactating cows' basal inflammatory status, but upregulated mammary inflammatory responses after intramammary LPS infusion.

Inhibition of pro-inflammatory signaling in human primary macrophages by enhancing arginase-2 via target site blockers

The modulation of macrophage phenotype from a pro-inflammatory to an anti-inflammatory state holds therapeutic potential in the treatment of inflammatory disease. We have previously shown that arginase-2 (Arg2), a mitochondrial enzyme, is a key regulator of the macrophage anti-inflammatory response. Here, we investigate the therapeutic potential of Arg2 enhancement via target site blockers (TSBs) in human macrophages. TSBs are locked nucleic acid antisense oligonucleotides that were specifically designed to protect specific microRNA recognition elements (MREs) in human ARG2 3' UTR mRNA. TSBs targeting miR-155 (TSB-155) and miR-3202 (TSB-3202) MREs increased ARG2 expression in human monocyte-derived macrophages. This resulted in decreased gene expression and cytokine production of TNF-α and CCL2 and, for TSB-3202, in an increase in the anti-inflammatory macrophage marker, CD206. Proteomic analysis demonstrated that a network of pro-inflammatory responsive proteins was modulated by TSBs. In silico bioinformatic analysis predicted that TSB-3202 suppressed upstream pro-inflammatory regulators including STAT-1 while enhancing anti-inflammatory associated proteins. Proteomic data were validated by confirming increased levels of sequestosome-1 and decreased levels of phosphorylated STAT-1 and STAT-1 upon TSB treatment. In conclusion, upregulation of Arg2 by TSBs inhibits pro-inflammatory signaling and is a promising novel therapeutic strategy to modulate inflammatory signaling in human macrophages.

The effects of the differentiated macrophages by dexamethasone on the immune responses

The characteristics of M2 macrophages suggest immunotherapeutic approaches for inducing immunological tolerance. The current study aimed to evaluate the effect of Dexamethasone (Dex) treatment on monocytes polarization and its impact on immune responses. The monocytes were extracted from the rat's blood samples. The effects of Dex concentration and treatment duration on monocyte viability, phagocytosis of rabbit red blood cell (RRBC) antigens, and cytokine gene expression were evaluated using MTT, ELISA, and Real-Time PCR analysis, respectively. The monocytes treated with Dex were injected into the rats as an autograft. The effects of the grafted cells were assessed on immune responses, monocyte differentiation, and pathological lesions, in comparison to the control groups.Treatment of monocytes with 10-5 M of Dex for 48 h increased the expression of IL-10 and TGF-β genes, while reducing the expression of TNF-α and IL-1 genes. The monocytes treated with antigen and Dex showed higher CD206 gene expression compared to CD80. The cells that were treated with Dex had the highest concentration of antigens after five days. Administration of the grafted cells to the animals has some significant effects on innate immune responses and no impact on pathological lesions. The group that received cells treated with Dex and antigen experienced a significant decrease in anti-RBC antibody titers. Additionally, there was a significant difference in the expression of cytokine genes and M2 differentiation markers among the groups that were evaluated. The effects of Dex on the viability and differentiation of monocytes depend on the dosage, timing, and duration of the treatment.

Effects of Supplementation with the Standardized Extract of Saffron (affron®) on the Kynurenine Pathway and Melatonin Synthesis in Rats.

Melatonin is a hormone that regulates sleep-wake cycles and is mainly synthesized in the pineal gland from tryptophan after its conversion into serotonin. Under normal conditions, less than 5% of tryptophan is reserved for the synthesis of serotonin and melatonin. The remaining 95% is metabolized in the liver through the kynurenine pathway. Increased levels of proinflammatory cytokines and cortisol increase the metabolism of tryptophan through the kynurenine pathway and reduce its availability for the synthesis of melatonin and serotonin, which may cause alterations in mood and sleep. The standardized saffron extract (affron®) has shown beneficial effects on mood and sleep disorders in humans, but the underlying mechanisms are not well understood. Thus, the aim of this work was to study the effects of affron® supplementation on the kynurenine pathway and the synthesis of melatonin in rats. For this purpose, adult male Wistar rats were supplemented for 7 days with 150 mg/kg of affron® or vehicle (2 mL/kg water) administered by gavage one hour before sleep. Affron® supplementation reduced body weight gain and increased the circulating levels of melatonin, testosterone, and c-HDL. Moreover, animals supplemented with affron® showed decreased serum levels of kynurenine, ET-1, and c-LDL. In the pineal gland, affron® reduced Il-6 expression and increased the expression of Aanat, the key enzyme for melatonin synthesis. In the liver, affron® administration decreased the mRNA levels of the enzymes of the kynurenine pathway Ido-2, Tod-2, and Aadat, as well as the gene expression of Il-1β and Tnf-α. Finally, rats treated with affron® showed increased mRNA levels of the antioxidant enzymes Ho-1, Sod-1, Gsr, and Gpx-3, both in the liver and in the pineal gland. In conclusion, affron® supplementation reduces kynurenine levels and promotes melatonin synthesis in rats, possibly through its antioxidant and anti-inflammatory effects, making this extract a possible alternative for the treatment and/or prevention of mood and sleep disorders.

TRPA1 and TPRV1 Ion Channels Are Required for Contact Lens-Induced Corneal Parainflammation and Can Modulate Levels of Resident Corneal Immune Cells.

Contact lens wear can induce corneal parainflammation involving CD11c+ cell responses (24 hours), γδ T cell responses (24 hours and 6 days), and IL-17-dependent Ly6G+ cell responses (6 days). Topical antibiotics blocked these CD11c+ responses. Because corneal CD11c+ responses to bacteria require transient receptor potential (TRP) ion-channels (TRPA1/TRPV1), we determined if these channels mediate lens-induced corneal parainflammation. Wild-type mice were fitted with contact lenses for 24 hours or 6 days and compared to lens wearing TRPA1 (-/-) or TRPV1 (-/-) mice or resiniferatoxin (RTX)-treated mice. Contralateral eyes were not fitted with lenses. Corneas were examined for major histocompatibility complex (MHC) class II+, CD45+, γδ T, or TNF-α+ cell responses (24 hours) or Ly6G+ responses (6 days) by quantitative imaging. The quantitative PCR (qPCR) determined cytokine gene expression. Lens-induced increases in MHC class II+ cells after 24 hours were abrogated in TRPV1 (-/-) but not TRPA1 (-/-) mice. Increases in CD45+ cells were unaffected. Increases in γδ T cells after 24 hours of wear were abrogated in TRPA1 (-/-) and TRPV1 (-/-) mice, as were 6 day Ly6G+ cell responses. Contralateral corneas of TRPA1 (-/-) and TRPV1 (-/-) mice showed reduced MHC class II+ and γδ T cells at 24 hours. RTX inhibited lens-induced parainflammatory phenotypes (24 hours and 6 days), blocked lens-induced TNF-α and IL-18 gene expression, TNF-α+ cell infiltration (24 hours), and reduced baseline MHC class II+ cells. TRPA1 and TRPV1 mediate contact lens-induced corneal parainflammation after 24 hours and 6 days of wear and can modulate baseline levels of resident corneal immune cells.

Effects of dietary lentinan on growth performance, nonspecific immunity and disease resistance to Aeromonas hydrophila in largemouth bass (Micropterus salmoides)

A study was conducted to evaluate the effects of dietary lentinan (LNT) on growth performance, nonspecific immunity and disease resistance to Aeromonas hydrophila in largemouth bass (Micropterus salmoides). Largemouth bass (initial weight: 10.31 ± 0.25 g) were divided into the CK, L1, L2, L3, L4 and L5 groups in triplicate and fed with diets containing LNT at 0%, 0.05%, 0.1%, 0.2%, 0.3% and 0.4% for eight weeks. Results showed that FBW, WGR, SGR and FE were significantly increased in the L3, L4 and L5 groups compared to the CK group. VSI, HSI, crude protein, crude fat and crude ash were similar among different treatment groups. Trypsin and lipase activities of intestinal tract were elevated in the L4 and L5 groups. The length of intestinal microvilli was increased in all experimental groups (P < 0.05). Lysozyme activity in serum and liver were elevated in the L3, L4 and L5 groups, SOD activity in serum and liver were elevated in the L1, L2, L3 and L4 groups, and CAT activity in serum and liver were elevated in the L4 group. Compared with the CK group, cytokine gene expressions of tnf-α and il-1β in foregut of the L3, L4 and L5 groups were significantly down-regulated (P < 0.05), and the expression of il-10 in the above three groups was significantly up-regulated (P < 0.05). After the Aeromonas hydrophila challenge test, survival rates were higher than 80% in the L4 group and lower than 40% in the CK group. In summary, our study suggested that LNT had a favorable effect on the overall health and growth of largemouth bass, and the optimal dietary concentration was 0.3%.

Effect of kelp powder on the resistance of Aeromonas hydrophila in the gut of hybrid snakeheads (Channa maculata ♀ × Channa argus ♂).

To assess the level of oxidative stress, expression of immune-related genes, histomorphology, and changes in the intestinal tract of hybrid snakeheads(Channa maculata ♀ × Channa argus ♂) under stress from kelp powder in place of flour against Aeromonas hydrophila. We set up experimental diets: a control (C) diet of 20% flour, an experimental (MR) diet of 10% kelp powder and 10% flour, and an experimental (FR) diet of 0% starch and 15% kelp powder. The experimental fish in each group were infected with Aeromonas hydrophila after 60 days of feeding. For this experiment, some of the experimental fish in group C were injected with PBS as a negative control group (PBS). The results showed that the C group had significantly higher SOD, CAT, and T-AOC activity and expression of TAK1, IKKβ, IL-1β, and TNF-α genes in the MyD88 pathway than the PBS group. CAT activity and the expression of TAK1, IL-1β and TNF-α genes in the MyD88 pathway were significantly lower in the MR group than in the C group. Furthermore, the number of goblet cells in the MR group was significantly higher than in the C group. Furthermore, microorganisms such as Bacteroidota and Actinobacteriota were significantly lower in the C group than in the PBS and FR groups, as were beneficial bacteria such as Clostridium_sensu_stricto_1 and Sphingomonas. Replacing flour with kelp powder increases hybrid snakehead gut resistance to Aeromonas hydrophila.

Proinflammatory and Immunomodulatory Gene and Protein Expression Patterns in Spinal Cord and Spleen Following Acute and Chronic High Thoracic Injury.

In addition to paralysis and loss of sensation, high-level spinal cord injury (SCI) causes sympathetic dysfunction that can lead to autonomic dysreflexia (AD) and chronic immune suppression involving splenic leukopenia. Evidence has shown that treatment with either gabapentin or blockade of TNFα mitigates maladaptive plasticity and the underlying hemodynamic dysfunction, spleen atrophy, and immune dysfunction associated with AD. Because significant improvements long term was noted following treatments only during acute stages of recovery, we sought to systematically examine changes in proinflammatory and immunomodulatory cytokines to ascertain the reason. Adult female Wistar rats underwent complete T4 spinal transection before euthanasia at systematic intervals from 3 days to 8 weeks after injury. Using qRT-PCR and meso scale discovery (MSD) assays, the gene and protein expression of TNFα and IFNγ in the spleen, upper thoracic (T4-9) and lumbosacral (L5-S6) spinal cords were analyzed. We found that spleen atrophy occurs in a biphasic manner compared to naïve controls, with significant decreases in the spleen mass noted at 3 days and 8 weeks after injury. Splenic TNFα mRNA and protein levels did not change significantly over time, while IFNγ gene expression dipped acutely with trends for increased protein levels at more chronic time points. TNFα protein increased significantly only in thoracic spinal cord segments from 3 to 14 days post-injury. IFNγ mRNA and protein levels remained unelevated in injured spinal cords over time, with trends for increased protein levels at 2 and 8 weeks in the lumbosacral segments. Novel temporal-spatial cytokine expression profiles reveal that TNFα protein levels are increased solely in upper thoracic segments after high thoracic SCI, while IFNγ remains unaltered. Splenic leukopenia and latent systemic immunosuppression are not associated with altered TNFα or IFNγ expression in the spleen or spinal cord.

The Effect of secretome-hypoxia mesenchymal-stem-cells and vitamin-d3 in type-2 diabetes-mellitus induced periodontitis rats

Background: A systemic metabolic disorder by insulin resistance, type-2 diabetes mellitus (T2DM), is characterised by an increased level of reactive oxygen species (ROS) and decreased superoxide dismutase (SOD), which is associated with the activation of proinflammatory cytokine pathways such as tumor necrosis (TNF)-α. This condition usually stimulates inflammation of the periodontal tissue (periodontitis). Vitamin-D3 can inhibit the release of inflammatory mediator and reduce the risk of chronic periodontitis. Purpose this study to determine the effect of the secretome-hypoxia mesenchymal-stem-cells (SH-MSCs)-Vitamin-D3 combination on the expression of SOD, IL-10, and TNF-α genes in T2DM periodontitis-induced rats.&#x0D; Method: 30 rats were randomly divided into five groups; normal sham condition, positive control, SH-MSCs group, vitamin-D3 group, and combination of SH-MSCs and vitamin D3 group. SH-MSCs were injected at doses of 150 uL in the gingival. In addition, vitamin D3 5000 IU 2.25 mcg were administrated orally. After 35 days, all rats were sacrificed, and qRT-PCR from gingival tissue was performed to identify the expression of SOD, IL-10 and TNF-α genes &#x0D; Results: The results a significant increase in SOD and IL-10 gene expression (p

Thymoquinone-loaded self-nano-emulsifying drug delivery system against ischemia/reperfusion injury.

In the present study, a self-nano-emulsifying drug delivery system (SNEDDS) was developed to evaluate the efficiency of thymoquinone (TQ) in hepatic ischemia/reperfusion. SNEDDS was pharmaceutically characterized to evaluate droplet size, morphology, zeta potential, thermodynamic stability, and dissolution/diffusion capacity. Animals were orally pre-treated during 10days with TQ-loaded SNEDDS. Biochemical analyses, hematoxylin-eosin staining, indirect immunofluorescence, and reverse transcription polymerase chain reaction (RT-PCR) were carried out to assess cell injury, oxidative stress, inflammation, and apoptosis. The TQ formulation showed good in vitro characteristics, including stable nanoparticle structure and size with high drug release rate. In vivo determinations revealed that TQ-loaded SNEDDS pre-treatment of rats maintained cellular integrity by decreasing transaminase (ALT and AST) release and preserving the histological characteristics of their liver. The antioxidant ability of the formulation was proven by increased SOD activity, reduced MDA concentration, and iNOS protein expression. In addition, this formulation exerted an anti-inflammatory effect evidenced by reduced plasma CRP concentration, MPO activity, and gene expressions of TLR-4, TNF-α, NF-κB, and IL-6. Finally, the TQ-loaded SNEDDS formulation promoted cell survival by enhancing the Bcl-2/Bax ratio. In conclusion, our results indicate that TQ encapsulated in SNEDDS significantly protects rat liver from I/R injury.

Screening for Chemical Characterization and Pharmacological Properties of Different Extracts from Nepeta italica.

Plants from the Nepeta genus have been proved to possess different pharmacological properties, among which are antimicrobial, antioxidant, anti-inflammatory, analgesic, and cytotoxic effects. Nepeta italica is a medicinal plant traditionally used for its analgesic effects, and in the present study, the phytochemical composition and biological effects of hexane, dichloromethane (DCM), ethyl acetate (EA), ethanol, ethanol-water, and water extracts of the aerial parts were investigated for determining phenolic composition, antioxidant effects, and anti-inflammatory effects in isolated mouse colon specimens exposed to lipopolysaccharide (LPS). Polar extracts were the richest in terms of phenolic compounds, especially rosmarinic acid. In parallel, ethanol, ethanol-water, and water extracts were also the most effective as scavenging/reducing and enzyme inhibition agents, especially towards cholinesterases and α-glucosidase, and in inhibiting the LPS-induced cyclooxygenase-2 (COX-2) and tumor necrosis factor α (TNFα) gene expression in mouse colon. This poses the basis for future in vivo investigations for confirming the protective effects of polar extracts of N. italica against inflammatory bowel diseases.

Shifts in immune responses of an invasive alien species: A test of the evolution of increased competitive ability hypothesis using American Eastern gray squirrels in Italy

Based on the Evolution of Increased Competitive Ability (EICA) hypothesis, a reduced investment in immunity, consequent to parasite loss, could partly explain the success of invasive alien species. We investigated variation in parasite load and immune responses of alien Eastern gray squirrels (Sciurus carolinensis) along the invasion wave of an expanding population. We first verified by fecal analyses that 1) parasite abundance decreased moving from the core towards the invasion front. Next, we used multiple measures of immunity to investigate whether, in response to the lower parasite pressure, individuals at the invasion front 2) dampened their costly inflammatory response, and 3) increased their investment in less expensive acquired immunity. We first explored variation in hematological variables related either to the inflammatory or the acquired response. On a subset of individuals, we carried out ex vivo cell cultures to analyse the basal expression of MHC class II genes and the expression of TNF-α genes in response to an immune challenge. Platelet counts and TNF-α expression suggested higher inflammation in individuals living at the invasion core, whereas parameters associated with an acquired response (lymphocyte counts and MHC II expression by spleen cells), conversely, were higher in squirrels at the front. Overall, our results suggest a shift between different immune strategies along the invasion wave, supporting a reduced investment in costly inflammatory responses and an increased investment in acquired immunity in individuals at the expanding edge of the range, which are subjected to high selective pressures for dispersal and reproduction.

Chemical Characterization of Different Extracts from Artemisia annua and Their Antioxidant, Enzyme Inhibitory and Anti-Inflammatory Properties.

Artemisia annua L. (Asteraceae Family) is an important plant in Asia that has been used for treating different diseases, including fever due to malaria, wounds, tubercolisis, scabues, pain, convulsions, diabetes, and inflammation. In this study we aimed to evaluate the effects of different polarity extracts (hexane, dichloromethane, ethyl acetate, ethanol, ethanol/water (70 %) and water) from A. annua against the burden of inflammation and oxidative stress occurring in colon tissue exposed to LPS. In parallel, chemical composition, antiradical, and enzyme inhibition effects against α-amylase, α-glucosidase, tyrosinase, and cholinesterases were evaluated. The water extract contained the highest content of the total phenolic with 34.59 mg gallic acid equivalent (GAE)/g extract, while the hexane had the highest content of the total flavonoid (20.06 mg rutin equivalent (RE)/g extract). In antioxidant assays, the polar extracts (ethanol, ethanol/water and water) exhibited stronger radical scavenging and reducing power abilities when compared to non-polar extracts. The hexane extract showed the best AChE, tyrosinase and glucosidase inhibitory effects. All extracts revealed effective anti-inflammatory agents, as demonstrated by the blunting effects on COX-2 and TNFα gene expression. These effects seemed to be not related to the only phenolic content. However, it is worthy of interest to highlight how the higher potency against LPS-induced gene expression was shown by the water extract ; thus suggesting a potential phytotherapy application in the management of clinical symptoms related to inflammatory colon diseases, although future in vivo studies are needed to confirm such in vitro and ex vivo observations.