Expression Levels Of iNOS Research Articles

The combined use of thymoquinone and metformin provides more effective neuroprotection in a mouse model of MPTP-induced Parkinson’s disease

Thymoquinone (TQ) is known for its antioxidant properties, and although metformin (MM) is known as an antidiabetic drug, it is suggested that it reduces neurodegeneration. The study aimed to investigate the neuroprotective effects of TQ and MM, particularly when used together, in relation to Parkinson’s disease (PD). In the study, sixty-eight male C57BL/6 mice weighing 25–30 g were divided into five groups as follows: control, MPTP, MPTP+TQ, MPTP+MM, and MPTP+TQ+MM. MM (500 mg/kg, orally) and TQ (5 mg/kg, i.p.) were administered for 21 days. Motor coordination and locomotor activities were evaluated by the pole test. TOS and TAS analyses were conducted to determine oxidative stress levels in the substantia nigra. Dopaminergic degeneration in the substantia nigra was evaluated by analyzing Tyrosine hydroxylase (TH). To evaluate the apoptotic pathway, the expression levels of iNOS, BDNF, Complex 1, Bax, Bcl-2, Cytochrome C, AIF, and Caspase-3 were examined immunohistochemically. Compared to the MPTP-treated group, TQ, MM and MM+TQ treatment provided significant improvement in locomotor activity in mice, significantly increased antioxidant activity, significantly reduced the expression levels of iNOS, Bax, Cytochrome C, Caspase-3, and AIF, significantly increased BDNF, Bcl-2, and Complex 1 expressions, and significantly increased the number of TH positive cells. The separate use of TQ and MM exhibits neuroprotective activity, however, we showed that using TQ and MM in combination may be more effective. This may provide preclinical evidence supporting the therapeutic potential of their combined use for treating PD.

MiR-214-3p inhibits LPS-induced macrophage inflammation and attenuates the progression of dry eye syndrome by regulating ferroptosis in cells.

Dry eye disease (DED) is an ocular illness caused by insufficient tear secretion or poor tear quality, and inflammation is a key factor in its pathogenesis. Previous studies have shown that miRNAs are important regulatory factors in DED. The purpose of this study was to explore the potential mechanism by which miR-214-3p influenced the DED process by regulating the macrophage inflammatory response. We induced THP-1 cells to differentiate into M0 macrophages with 100 ng/mL phorbol-12-myristate-13-acetate (PMA) and then added 15 ng/mL lipopolysaccharide (LPS) to induce inflammation. The expression of related genes and proteins was detected via RT‒qPCR, Western blotting, ELISA and immunofluorescence staining; cell viability was measured using the CCK-8 assay; and flow cytometry was used to detect ROS levels. In tear and serum samples from DED patients, the levels of miR-214-3p, IL-10, and Arg1 were decreased, and the levels of IL-6, TNF-α, IL-1β, and iNOS expression were increased. Moreover, the overexpression of miR-214-3p attenuated the effect of LPS and inhibited M1 polarization and inflammation in macrophages. Mechanistically, miR-214-3p inhibited macrophage ferroptosis by downregulating TFRC expression, thereby inhibiting macrophage M1 polarization and inflammation and alleviating the progression of DED. Our study indicated that the upregulation of miR-214-3p expression might be a new target for DED therapy.

Structural Characterization and Anti-inflammatory Activities of a Purified Polysaccharide from Veronica persica Poir.

In this manuscript, the polysaccharide (VPP-I) from Veronica persica Poir., was characterized in detail by high performance gel permeation chromatography (HPGPC), high performance liquid chromatography (HPLC), Fourier transform infrared spectroscopy (FT-IR) and nuclear magnetic resonance spectroscopy (NMR). In addition, lipopolysaccharide (LPS) induced inflammation model of RAW264.7 cells was used to evaluate the in vitro anti-inflammatory activity of VPP-I. The results showed that the relative molecular weight of VPP-I was 2.355 KDa, which was mainly composed of mannose (Man), glucose (Glc) and galactose (Gal) in a ratio of 1 : 32.46 : 28.76. Moreover, the VPP-I contained sugar alcohol derivatives of T-DGlcp(1→, →4)-D-Galp(1→, →3,6)-D-Manp(1→, →4)-D-Glcp(1→, →6)-D-Galp(1→ and →6)-D-Glcp(1→. In vitro anti-inflammatory results showed that VPP-I could inhibit the secretion of IL-β, IL-6 and TNF-α in RAW264.7 cells induced by LPS. Moreover, compared to the LPS group, the mRNA expression levels of iNOS, COX-2, IL-β, IL-6 and TNF-α produced by RAW264.7 were significantly decreased after treatment with VPP-I (P<0.05). In addition, VPP-I could increase the SOD and GSH-Px enzymes activity and decrease the content of MDA in LPS-induced RAW264.7 cells (P<0.05). In summary, this paper laid theoretical foundation for the application of Veronica persica Poir. in the field of medicine.

Xianglian pill alleviates ulcerative colitis by inhibiting M1 macrophage polarization via modulation of energy metabolite itaconate

BackgroundXianglian pill (XLP) is a traditional Chinese medicine (TCM) that is widely used to treat ulcerative colitis (UC). However, its mechanism of action in UC is unclear. PurposeThis study aimed to investigate the mechanism of action of XLP in treating UC and role of M1 macrophage polarization in this process. Study designIn vivo experiments were performed using UC mice while in vitro experiments were conducted using RAW264.7 cells. MethodsMice were administered 3 % dextran sulfate to induce UC model and then treated with XLP. Changes in histopathology and pro-inflammatory cytokines were evaluated. The levels of M1 macrophages in mesenteric lymph nodes were detected by flow cytometry. Colon metabolite levels were analyzed using an energy metabolomic assay. To assess itaconate's impact, both in vivo (mice) and in vitro (RAW264.7 cells) models were employed. Immunofluorescence staining was used to measure the expression levels of TNF-α, IL-6, and iNOS, while qRT-PCR was utilized to quantify the mRNA levels of TET2, STAT1, and Nfkbiz. ResultsXLP alleviated ulcerative damage and reduced TNF-α and IL-6 levels in colon, and also downregulated the levels of M1 macrophages and modulated the state of energy metabolism. Specifically, XLP significantly increased ITA level in colonic tissue and this increase was significantly associated with decreased levels of M1 macrophages and alleviation of UC following XLP treatment. Moreover, ITA directly suppressed the polarization of macrophage from M0 to M1 phenotype, accompanied by the decrease of TNF-α, IL-6, and iNOS levels. Further, ITA decreased inflammatory responses in M1 macrophage by inhibiting the TET2/STAT1 and TET2/NF-κB signaling pathways. ConclusionXLP can treat UC by suppressing M1 macrophage polarization via increasing the level of energy metabolite ITA.

Amentoflavone maintaining extracellular matrix homeostasis and inhibiting subchondral bone loss in osteoarthritis by inhibiting ERK, JNK and NF-κB signaling pathways

Amentoflavone (AF), a plant biflavone isolated from Selaginella sinensis ethanol extract, is characterized by anti-inflammatory and anti-oxidant properties. According to previous studies, inflammation and oxidative stress are closely related to the pathophysiology of osteoarthritis (OA). However, the effects and mechanisms of AF on OA have not been elucidated.To investigate the inhibitory effects and its molecular mechanism of AF on extracellular matrix (ECM) degradation stimulated by IL-1β as well as subchondral bone loss induced by RANKL in mice chondrocytes. Quantitative PCR was used to detect the mRNA expression of genes related to inflammation, ECM, and osteoclast differentiation. Protein expression level of iNOS, COX-2, MMP13, ADAMTS5, COL2A1, SOX9, NFATc1, c-fos, JNK, ERK, P65, IκBα was measured by western blotting. The levels of TNF-α and IL-6 in the supernatants were measured by ELISA. The amount of ECM in chondrocytes was measured using toluidine blue staining. The levels of Aggrecan and Col2a1 in chondrocytes were measured using immunofluorescence. Tartrate-resistant acid phosphatase (TRAP) staining, F-actin staining and immunofluorescence were used to detect the effect of AF on osteoclast differentiation and bone resorption. The effect of AF on destabilization of the medial meniscus (DMM)-induced OA mice can be detected in hematoxylin–eosin (H&E) staining, Safranin O green staining and immunohistochemistry.AF might drastically attenuated IL-1β-stimulated inflammation and reduction of ECM formation by blocking ERK and NF-κB signaling pathways in chondrocytes. Meanwhile, AF suppressed the formation of osteoclasts and the resorption of bone function induced by RANKL. In vivo, AF played a protective role by stabilizing cartilage ECM and inhibiting subchondral bone loss in destabilization of the medial meniscus (DMM)-induced OA mice, further proving its protective effect in the development of OA. Our study show that AF alleviated OA by suppressing ERK, JNK and NF-κB signaling pathways in OA models in vitro and DMM-induced OA mice, suggesting that AF might be a potential therapeutic agent in the treatment of OA.

A Novel Trichinella spiralis Galectin Strengthens the Macrophage ADCC Killing of Larvae via Driving M1 Polarization.

Galectin recognizes β-galactosides through its carbohydrate recognition domains (CRDs). This study aimed to determine the biological features of a novel Trichinella spiralis galectin (galactoside-binding lectin family protein, TsGLFP) and its role in driving macrophage M1 polarization and enhancing ADCC killing of larvae. TsGLFP belongs to the galectin family and has two CRDs. The complete TsGLFP cDNA sequence was cloned and then expressed in Escherichia coli BL21. The results of qPCR, Western blot, and indirect immunofluorescence tests (IIFTs) revealed that TsGLFP was expressed in various stages of T. spiralis worms and principally localized at the cuticle and around the female embryos of the nematode. rTsGLFP had the function of agglutinating mouse erythrocytes, and this agglutination activity could be inhibited by lactose. After the mouse macrophage RAW264.7 was incubated with rTsGLFP, the expression level of the M1 genes (iNOS, IL-6, and TNF-α) and NO production were obviously increased. After incubating macrophages with rTsGLFP, there was a noticeable rise in the expression levels of p-IκB-α and p-NF-κB p65. Additionally, rTsGLFP enhanced the macrophage's ability to kill newborn larvae by ADCC cytotoxicity. When the macrophages were pretreated with the specific p-NF-κB p65 inhibitor PDTC, and then stimulated with rTsGLFP, the expression levels of iNOS, NO, and p-NF-κB p65 and the macrophages' ADCC cytotoxicity were distinctly decreased. These findings indicated that rTsGLFP enhanced the macrophage ADCC killing of larvae by driving M1 polarization through activating the NF-κB pathway.

NIR stimulated epigallocatechin gallate loaded polydopamine with enhanced antibacterial and ROS scavenging abilities for improved infectious wound healing

Infectious wound healing is complicated with and limited by infection and oxidative stress at the wound site. In recent years, various evidences suggest that nanozymes with multiple enzymatic activities have enabled the development of novel strategies for infectious wound healing. In this study, epigallocatechin gallate loaded polydopamine (P@E) was developed to act as a potent reactive oxygen species (ROS) scavenger for scavenging ROS, alleviating inflammatory responses, and promoting infectious wound healing. Combining with near infrared (NIR) irradiation, P@E presented excellent antibacterial ability of Escherichia coli (E. coli, 93.6%) and methicillin-resistant Staphylococcus aureus (MRSA, 87.6%). Specifically, P@E+NIR exhibited the most potent antioxidant, anti-inflammatory and cell proliferation behaviors through down-regulating intracellular ROS levels (81.9% and 94.3% for NIH3T3 and RAW264.7 respectively) and inducible nitric oxide synthase (iNOS) expression level (55.7%), and up-regulating the expression levels of arginase-1 (Arg-1, 71.4%), heat shock protein 70 (HSP70, 48.6%) and platelet endothelial cell adhesion molecule (CD31, 35.3%) compared to control group. Meanwhile, it also efficiently induced M2 directional polarization of lipopolysaccharide induced murine macrophages to achieve anti-inflammation, indicated by the down-regulation of CD86 (86.2%), and up-regulation of CD206 (85.6%). Significantly, it was also observed that P@E+NIR presented the excellent behaviors of inhibiting wound infection, alleviating wound inflammation, as well as promoting skin tissue repairing. Altogether, it has developed the strategy of using P@E combining with NIR irradiation for the synergistic enhanced healing of infectious skin wound, which can serve as a promising therapeutic strategy for its clinical treatment.

Triggering receptor expressed on myeloid cells-1 aggravates obliterative bronchiolitis via enhancing the proinflammatory phenotype of macrophages

BackgroundTriggering Receptor Expressed on Myeloid cells-1 (TREM-1) plays an important role in innate immune system. However, whether and how TREM-1 contributes to obliterative bronchiolitis (OB) progression remains unclear. MethodsA murine orthotopic tracheal transplantation model was constructed to mimic the pathogenesis of OB. qPCR and immunoblotting were used to measure TREM-1 expression. RNA sequencing was used to investigate the impact of TREM-1 on proinflammatory phenotype of macrophages. Trem-1 knockout mice and Nlrp3 knockout mice were generated to investigate the role of the TREM-1/NLRP3 pathway in the proinflammatory phenotype of macrophages. The infiltration of immune cells within the grafts was quantified using immunofluorescence staining. Flow cytometry was used to detect the proportion of different immune cells in mice spleen and the expression levels of iNOS and co-stimulatory molecules in macrophages. ResultsThe expression of TREM-1 was upregulated in the mouse OB model. Genetic ablation or pharmacological inhibition of TREM-1 ameliorated OB, whereas the stimulation of TREM-1 using anti-TREM-1 agonistic antibody exacerbated OB. Moreover, Trem-1 ablation reduced the infiltration of iNOS+ macrophages and limited the T cell responses. In vitro studies revealed that Trem-1 deletion impaired the proinflammatory function and antigen presentation ability of macrophages. Additionally, Trem-1 knockout inhibited the activation of NLRP3 signaling pathway. NLRP3 overexpression restored the proinflammatory phenotype of Trem-1 knockout macrophages. ConclusionsThese findings indicated that TREM-1 could promote the proinflammatory phenotype of macrophages through NLRP3 inflammasome activation, thereby exacerbating OB progression. These findings indicated that TREM-1 may serve as a therapeutic target for OB treatment.

In vitro assessment of Dracocephalum lindbergii for growth inhibition, apoptosis induction, and cytokine modulation against Leishmaniamajor

Leishmaniasis is a vector-borne disease caused by several species of flagellate protozoan parasites belonging to the genus Leishmania, which are part of the Trypanosomatidae family. This study aims to evaluate the in vitro anti-Leishmania activity of Dracocephalum lindbergii Rech.f (D. lindbergii) on the growth, apoptosis induction, and proliferation of Leishmania major. To conduct this study, the aerial parts of D. lindbergii were collected during the flowering stage in May 2022 from North Khorasan province, Iran. The plant material was extracted using various solvents, starting with those of lower polarity and progressing to those of higher polarity. To assess the impact of the D. lindbergii fraction on L. major promastigotes, amastigotes, and macrophages (THP-1), cell viability was determined using the MTT assay. Additionally, flow cytometry with the Annexin V-PE apoptosis detection kit was employed to distinguish between viable, necrotic, and apoptotic promastigotes in response to treatment with the 100 % methanolic fraction of D. lindbergii (D). The expression levels of TNF-α, IFN-γ, IL-12, IL-10, TGF-β, IL4, iNOS, and GAPDH were quantified using real-time PCR (qPCR) on the macrophage cell line. Each treatment approach exhibited marked anti-leishmanial effects across different concentrations over 24, 48, and 72 h of incubation, showing statistically significant differences compared to the untreated control group (P < 0.001). Different concentrations of D, MAT, and AmpB, both individually and in combination, significantly reduced the total number of intramacrophage amastigotes compared to the untreated control groups at 24, 48, and 72 h. The results also showed time-dependent variations in the anti-Leishmania activity of the fraction. In terms of cellular morphology, treated cells exhibited changes such as shrinkage, cytoplasmic condensation, and reduced mobility, particularly noticeable after 24 h of treatment. Additionally, fraction D demonstrated significant antioxidant properties. This study highlights the potential of D. lindbergii as an anti-Leishmania agent, with the 100 % methanolic fraction emerging as a promising candidate for the development of novel treatments for leishmaniasis.

Anti-inflammatory Effect of Symplocos prunifolia Extract in an In Vitro Model of Acute Pneumonia.

Acute pneumonia is a respiratory disease characterized by inflammation within the lung tissue, exhibiting higher morbidity rates and mortality rates among immunocompromised children and older adults. Symplocos species have been traditionally used as herbal remedies for conditions like dysentery, skin ulcers, diarrhea, and dyspepsia. Contemporary research has employed various Symplocos species in the study of diverse diseases. However, the exact efficacy and mechanisms of action of Symplocos Prunifolia remain unknown. Therefore, this study investigated the anti-inflammatory mechanism of S. prunifolia extract (SPE) in A549 and RAW264.7 cells stimulated by lipopolysaccharide (LPS). SPE significantly reduced nitric oxide (NO) production and the protein expression levels of like inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-stimulated RAW 264.7 cells. Furthermore, it reduced the protein expression levels of iNOS, COX-2 and the levels of pro-inflammatory cytokines in LPS-stimulated A549 cells. The mechanism underlying the anti-inflammatory effect of SPE was associated with the inhibition of LPS stimulated the phosphoinositide-3-kinase/protein kinase B (PI3K/Akt) and Mitogen-activated protein kinase (MAPK) phosphorylation. Moreover, we confirmed that SPE decreased the nuclear translocation of nuclear factor-κB (NF-κB)/p65 stimulated by LPS. In conclusion, these results demonstrate that SPE alleviates inflammatory responses by deactivating the PI3K/Akt, MAPK, and NF-κB signaling pathways. Our findings suggest that SPE is a potential candidate for acute pneumonia prevention.

Effect of goose-derived adiponectin peptide gADP3 on LPS-induced inflammatory injury in goose liver

ABSTRACT 1. This study evaluated the effects and mechanisms of action of the peptide gADP3 on hepatic inflammatory injury induced by lipopolysaccharide (LPS). 2. Hepatic inflammatory injury was induced in geese by intraperitoneal injection of LPS and gADP3, and the adiponectin receptor agonist AdipoRon (positive control) was used for potential amelioration. Serum inflammatory factor levels, liver function-related biochemical indicators and oxidative stress-related biochemical parameters in the liver tissues were determined. The expression levels of adiponectin and its receptors, inflammation and oxidative stress-related genes and key signalling molecules involved in adiponectin, inflammation and oxidative stress signalling pathways in liver tissues were detected. 3. The peptide gADP3 alleviated inflammatory cell infiltration and hepatic inflammatory changes, reversed the decrease in serum albumin (ALB), total protein (TP), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) content or activity induced by LPS and increased the activity of the antioxidant enzymes CAT (catalase), SOD (superoxide dismutase) and GSH-Px (glutathione peroxidase). 4. The peptide gADP3 upregulated the expression of antioxidant enzyme-related genes GCLC, HO-1 and NQO1 in liver tissues, decreased the levels of inflammatory factors like TNF-α, IL-1β, IL-6, IFN-γ and TGF-β and reduced mRNA expression levels of inflammatory-related genes TNF-α, IL-1β, iNOS and TGF-β. Additionally, it increased the mRNA and protein expression levels of adiponectin and its receptors, as well as key molecules in the adiponectin signalling pathway like AMPK and PPARα. In addition, gADP3 reversed the changes in mRNA or protein expression of inflammatory and oxidative stress signalling pathway-related genes P38MAPK, NF-κBP65, TLR4 and Nrf2 in liver tissues caused by LPS treatment. 5. In conclusion, goose-derived adiponectin peptide gADP3, similar to the adiponectin receptor agonist AdipoRon, attenuated LPS-induced hepatic inflammatory injury in geese.

Inhibitory Effects of Naringenin on LPS-Induced Skin Inflammation by NF-κB Regulation in Human Dermal Fibroblasts.

Flavonoids are important natural compounds characterized by their extensive biological activities. Citrus flavonoids represent a significant segment of the broader flavonoid category. Naringenin, an integral part of this series, is recognized for its powerful anti-inflammatory and antioxidant properties. In addition, considering the lack of existing research on naringenin's potential effectiveness and intracellular mechanisms of action in skin-related applications, especially as a cosmetic ingredient, this study aimed to explore naringenin's role in reducing the fundamental generation of reactive oxygen species. This was achieved by examining its inhibitory effects on the expression levels of NADPH oxidase and iNOS, ultimately leading to a reduction in NO production. This research examined the anti-inflammatory and antioxidant capacities of naringenin by employing a cellular senescence model of LPS-induced HDFs. The evaluation of naringenin's efficacy was validated through several investigative procedures, including the NF-κB luciferase assay, ELISA assay, and qRT-PCR. To verify the anti-inflammatory effectiveness of naringenin, we measured the responsive elements of NF-κB using a luciferase reporter assay. This assessment revealed that naringenin could decrease the concentration of genes activated by NF-κB. Moreover, we found that naringenin inhibited the transcriptional expression of known NF-κB-regulated inflammatory cytokines, including IL-1β, IL-6, and IL-8. In addition, results from the qRT-PCR analysis indicated that naringenin facilitated a reduction in iNOS expression. Based on the data gathered and analyzed in this study, it can be conclusively inferred that naringenin possesses promising potential as a cosmetic ingredient, offering both anti-inflammatory and antioxidant benefits.

Exploring the synergistic effects of chuanxiong rhizoma and Cyperi rhizoma in eliciting a rapid anti-migraine action based on pharmacodynamics and pharmacokinetics

Ethnopharmacological relevanceHerb-herb combination has been used to maximize the therapeutic efficacy in the theory of traditional Chinese medicine. Chuanxiong rhizoma (called Chuanxiong in Chinese, CX) and Cyperi rhizoma (called Xiangfu in Chinese, XF) have been used alone or in combination (CRCR) to treat migraine dating back to Eastern Jin Dynasty (AD317) of China. But no data demonstrate the possible necessities or advantages of combining CX and XF for migraine. Aim of the studyThis study explores the combination mechanism based on pharmacodynamics and pharmacokinetics. Materials and methodsA nitroglycerin-induced acute migraine model in rats was used to evaluate the anti-migraine effects of CRCR and the individual herbs using behavior, real time polymerase chain reaction and Western blot experiments. The absorption characteristics of active components involved in the anti-migraine action were analyzed by UPLC-MS/MS. ResultsCX and CRCR significantly reversed the abnormal levels of vasoactive substances (5-HT, CGRP, MMP-2 and MMP-9) to normal levels, but XF did not. XF and CRCR significantly decreased the pro-inflammatory cytokines (IL-1β, IL-6, and TNF-a), and increased the anti-inflammatory cytokines (IL-4 and IL-10). CRCR significantly decreased the mRNA expression levels of c-fos, iNos and nNos, and the corresponding protein expression levels of c-Fos, iNOS, and nNOS. CRCR inhibited NOS/NO pathway by downregulating the expression levels of NOS and NO. Furthermore, CRCR significantly increased the intestinal absorption rate and amount, and changed the pharmacokinetic parameters of active components in comparison with the individual herbs. ConclusionsCX had an advantage in regulating vasoactive substances, and XF focused on regulating inflammatory cytokines. CRCR is more effective in treating migraine than the individual herbs by depending on the synergistic action of CX and XF. This research provided some critical evidences on synergistic action between herb-herb interactions, and revealed the potential advantages of herb-herb combination in traditional Chinese medicine.

The alleviation of difenoconazole-induced kidney injury in common carp (Cyprinus carpio) by silybin: Involvement of inflammation, oxidative stress, and apoptosis

Triazole fungicides, such as difenoconazole (DFZ), are frequently used to control fungus in crops that pollute water. The common carp (Cyprinus carpio) (hereafter referred to as “carp”) is an excellent bio-indicator of water quality. The seeds of the silymarin plant contain a flavonolignan called silybin (SYB), which is used to treat liver disease. To explore SYB's involvement in DFZ-triggered kidney damage in carps, an H&E assay was conducted, and ROS level was also examined. The results demonstrated that SYB alleviated DFZ-induced destruction of kidney tissue structure in carps, as well as alleviating the elevation of kidney ROS level in carps. RT-qPCR and Western blot were used to detect inflammation-, oxidative stress- and apoptosis-related factors at mRNA level and protein level. The experimental findings indicated that relative to the DFZ group, SYB + DFZ co-treatment reduced inflammation-related mRNA level of il-6, il-1β and tnf-α, elevated mRNA level of il-10. It also reduced protein expression levels of NF-κB and iNOS. In addition, SYB + DFZ co-treatment reduced DFZ-induced increase in the oxidative stress-related mRNA indicators sod and cat, and decreased the protein expression levels of Nrf2 and NQO1. SYB reduced the DFZ-induced increase in pro-apoptotic gene Bax mRNA and protein expression levels and the DFZ-induced decrease in anti-apoptotic gene Bcl-2 mRNA and protein expression levels. In summary, SYB potentially mitigates DFZ-induced kidney damage in carp by addressing inflammation, oxidative stress, and apoptosis. Our results establish a theoretical foundation for the clinical advancement of freshwater carp feeds.

Haperforatones A-M, thirteen undescribed limonoids from Harrisonia perforata with anti-inflammatory activity

UPLC-Q-TOF-MS combined with mass defect filtering strategies were applied for the phytochemical investigation of Harrisonia perforata, leading to the isolation of thirteen undescribed limonoids named haperforatones A-M (1–13) and seventeen known compounds (14–30). Particularly, haperforatones D-E (4–5) have an unprecedented A, B, C, D-seco-6, 7-nor-C-24-limonoid skeleton, structurally stripped of the five-membered lactone ring B and formed a double bond at the C-5 and C-10 positions. Their 2D structures and relative configurations were identified using spectroscopic data. The absolute configurations of 1, 4, and 6 were established via X-ray diffraction crystallography. All 30 compounds were evaluated for anti-inflammatory potential in LPS-induced Raw 264.7 cell lines. Among those tested compounds, the most potent activity against LPS-induced NO generation was demonstrated by haperforatone F (6), with the IC50 value of inhibition NO production of 7.2 µM. Additionally, 6 could significantly inhibit IL-1β and IL-6 release and markedly downregulate the protein expression level of iNOS in the LPS-stimulated RAW264.7 cells at 10 µM. The possible mechanism of NO inhibition of 6 was also investigated using molecular docking, which revealed the interaction of compound 6 with the iNOS protein.

Induction of Nitric Oxide and Its Role in Facial Nerve Regeneration According to the Method of Facial Nerve Injury.

Nitric oxide (NO) is an important molecule in cell communication that also plays an important role in many biological processes. Given the dual role of NO in nerve degeneration and regeneration after facial nerve injury, we sought to delve deeper into its role through a systematic literature review. A comprehensive review of the literature employing SCOPUS, PubMed, Cochrane Library, EMBASE, and Google Scholar databases was conducted to evaluate the induction and role of NO in neurodegeneration and regeneration after facial nerve injury. From the 20 papers ultimately reviewed, the central findings were that neuronal nitric oxide synthase(nNOS), endothelial nitric oxide synthase (eNOS), and induced nitric oxide synthase (iNOS) increased or decreased depending on the method of facial nerve damage, damaged area, harvested area, and animal age, and were correlated with degeneration and regeneration of the facial nerve. Research conducted on rats and mice demonstrated that NO, nNOS, eNOS, and iNOS play significant roles in nerve regeneration and degeneration. However, the relationship between nerve damage and NO could not be defined by a simple causal relationship. Instead, the involvement of NOS depends on the type of nerve cell, source of NO, timing, and location of expression, age of the target animal, and proximity of the damage location to the brainstem. Consequently, nNOS, eNOS, and iNOS expression levels and functions may vary significantly.

Aflatoxin B1 exposure deteriorates immune abnormalities in a BTBR T+ Itpr3tf/J mouse model of autism by increasing inflammatory mediators' production in CD19-expressing cells

Autism spectrum disorder (ASD) is a neurodevelopmental condition characterized by deficiencies in communication, repetitive and stereotyped behavioral patterns, and difficulties in reciprocal social engagement. The presence of immunological dysfunction in ASD has been well established. Aflatoxin B1 (AFB1) is a prevalent mycotoxin found in food and feed, causing immune toxicity and hepatotoxicity. AFB1 is significantly elevated in several regions around the globe. Existing research indicates that prolonged exposure to AFB1 results in neurological problems. The BTBR T+ Itpr3tf/J (BTBR) mice, which were used as an autism model, exhibit the primary behavioral traits that define ASD, such as repeated, stereotyped behaviors and impaired social interactions. The main objective of this work was to assess the toxic impact of AFB1 in BTBR mice. This work aimed to examine the effects of AFB1 on the expression of Notch-1, IL-6, MCP-1, iNOS, GM-CSF, and NF-κB p65 by CD19+ B cells in the spleen of the BTBR using flow cytometry. We also verified the impact of AFB1 exposure on the mRNA expression levels of Notch-1, IL-6, MCP-1, iNOS, GM-CSF, and NF-κB p65 in the brain of BTBR mice using real-time PCR. The findings of our study showed that the mice treated with AFB1 in the BTBR group exhibited a substantial increase in the presence of CD19+Notch-1+, CD19+IL-6+, CD19+MCP-1+, CD19+iNOS+, CD19+GM-CSF+, and CD19+NF-κB p65+ compared to the mice in the BTBR group that were treated with saline. Our findings also confirmed that administering AFB1 to BTBR mice leads to elevated mRNA expression levels of Notch-1, IL-6, MCP-1, iNOS, GM-CSF, and NF-κB p65 in the brain, in comparison to BTBR mice treated with saline. The data highlight that exposure to AFB1 worsens immunological abnormalities by increasing the expression of inflammatory mediators in BTBR mice.

Collagen sponge scaffolds loaded with Trichostatin A pretreated BMSCs-derived exosomes regulate macrophage polarization to promote skin wound healing

The process of wound healing includes the inflammatory stage, which plays an important role. Macrophages can promote inflammatory response and also promote angiogenesis, wound contraction and tissue remodeling required for wound healing. It is crucial to promote macrophages to polarize from M1 pro-inflammatory phenotype to M2 anti-inflammatory phenotype at a critical time for the quality of wound healing. Because mesenchymal stem cell-derived exosomes have broad therapeutic prospects in the field of tissue repair and regeneration, in this study, we explored whether trichostatin A pretreated bone marrow mesenchymal stem cells (BMSCs)-derived exosomes (T-Exo) could promote wound healing by binding to biomaterial scaffolds through certain anti-inflammatory effects. In the cell experiment, we established macrophage inflammation model and then treated with T-Exo, and finally detected the expression levels of macrophage polarization proteins CD206, CD86 and TNF-α, iNOS, and Arg-1 by Western Blot and immunofluorescence staining; detected the expression levels of inflammation-related genes TNF-α, iNOS, IL-1β, IL-10 and anti-inflammatory genes CD206 and Arg-1 by qRT-PCR; explored the promoting ability of T-Exo to promote cell migration and tube formation by cell scratch experiment and angiogenesis experiment. The results showed that T-Exo could promote the polarization of M1 macrophages to M2 macrophages, and promote the migration and angiogenesis of HUVECs. Because TSA pretreatment may bring about changes in the content and function of BMSCs-derived exosomes, proteomic analysis was performed on T-Exo and unpretreated BMSCs-derived exosomes (Exo). The results showed that the differentially expressed proteins in T-Exo were related to some pathways that promote angiogenesis, cell migration, proliferation, and re-epithelialization. Then, exosome/collagen sponge (T-Exo/Col) biological scaffolds were prepared, and the physicochemical properties and biocompatibility of the scaffolds were investigated. Animal skin wound models were established, and the therapeutic effect and anti-inflammatory effect of T-Exo/Col in wound repair were evaluated by small animal in vivo imaging, H&E staining, Masson trichrome staining, immunohistochemical staining, Western Blot, and qRT-PCR. The results showed that T-Exo significantly promoted wound healing by inhibiting inflammation, thereby further promoting angiogenesis and collagen formation in vivo. Moreover, the existence of Col scaffold in T-Exo/Col enabled T-Exo to achieve a certain sustained release effect. Finally, we further explored whether TSA exerts beneficial effects by inhibiting HDAC6 gene of BMSCs, but the results showed that knockdown of HDAC6 gene would cause oxidative stress damage to BMSCs, which means that TSA does not produce these beneficial effects by inhibiting HDAC6 gene. What molecular mechanisms TSA exerts beneficial effects through needs to be further elucidated in the future.

Syringic acid attenuates acute lung injury by modulating macrophage polarization in LPS-induced mice

BackgroundAcute lung injury (ALI) is a continuum of lung changes caused by multiple lung injuries, characterized by a syndrome of uncontrolled systemic inflammation that often leads to significant morbidity and death. Anti-inflammatory is one of its treatment methods, but there is no safe and available drug therapy. Syringic acid (SA) is a natural organic compound commonly found in a variety of plants, especially in certain woody plants and fruits. In modern pharmacological studies, SA has anti-inflammatory effects and therefore may be a potentially safe and available compound for the treatment of acute lung injury. PurposeThis study attempts to reveal the protective mechanism of SA against ALI by affecting the polarization of macrophages and the activation of NF-κB signaling pathway. Trying to find a safer and more effective drug therapy for clinical use. MethodsWe constructed the ALI model using C57BL/6 mice by intratracheal instillation of LPS (10 mg/kg). Histological analysis was performed with hematoxylin and eosin (H&E). The wet-dry ratio of the whole lung was measured to evaluate pulmonary edema. The effect of SA on macrophage M1-type was detected by flow cytometry. BCA protein quantification method was used to determine the total protein concentration in bronchoalveolar lavage fluid (BALF). The levels of Interleukin (IL)-6, IL-1β, and tumor necrosis factor (TNF)-α in BALF were determined by the ELISA kits, and RT-qPCR was used to detect the expression levels of IL-6, IL-1β and TNF-α mRNA of lung tissue. Western blot was used to detect the expression levels of iNOS and COX-2 and the phosphorylation of p65 and IκBα in the NF-κB pathway in lung tissue. In vitro experiments were conducted with RAW267.4 cell inflammation model induced by 100 ng/ml LPS and A549 cell inflammation model induced by 10 μg/ml LPS. The effects of SA on M1-type and M2-type macrophages of RAW267.4 macrophages induced by LPS were detected by flow cytometry. The toxicity of compound SA to A549 cells was detected by MTT method which to determine the safe dose of SA. The expressions of COX-2 and the phosphorylation of p65 and IκBα protein in NF-κB pathway were detected by Western blot. ResultsWe found that the pre-treatment of SA significantly reduced the degree of lung injury, and the infiltration of neutrophils in the lung interstitium and alveolar space of the lung. The formation of transparent membrane in lung tissue and thickening of alveolar septum were significantly reduced compared with the model group, and the wet-dry ratio of the lung was also reduced. ELISA and RT-qPCR results showed that SA could significantly inhibit the production of IL-6, IL-1β, TNF-α. At the same time, SA could significantly inhibit the expression of iNOS and COX-2 proteins, and could inhibit the phosphorylation of p65 and IκBα proteins. in a dose-dependent manner. In vitro experiments, we found that flow cytometry showed that SA could significantly inhibit the polarization of macrophages from M0 type macrophages to M1-type macrophages, while SA could promote the polarization of M1-type macrophages to M2-type macrophages. The results of MTT assay showed that SA had no obvious cytotoxicity to A549 cells when the concentration was not higher than 80 μM, while LPS could promote the proliferation of A549 cells. In the study of anti-inflammatory effect, SA can significantly inhibit the expression of COX-2 and the phosphorylation of p65 and IκBα proteins in LPS-induced A549 cells. ConclusionSA has possessed a crucial anti-ALI role in LPS-induced mice. The mechanism was elucidated, suggesting that the inhibition of macrophage polarization to M1-type and the promotion of macrophage polarization to M2-type, as well as the inhibition of NF-κB pathway by SA may be the reasons for its anti-ALI. This finding provides important molecular evidence for the further application of SA in the clinical treatment of ALI.

Effects of oleogels (alpha-linolenic acid plus beeswax) extracted supplementation for approaching the therapeutic food ingredient: in vitro model

The dietary fatty acid intake for vascular disease has been highlighted for a decade. This study aimed to determine the effect of palmitic acid and fatty acid-incorporated beeswax (oleogels: OG) supplementation on anti-atherosclerotic effects through vascular smooth muscle cell (VSMC) activity. The A7r5 cells were cultured, cells were treated with three treatments, including 1) control, 2) palmitic acid (PA), and 3) alpha-linolenic acid (ALA) + beeswax, ratio 1:1 (OG) at different concentrations of 0, 25, 50 and 100 μM for 48 hrs. Cell proliferation, cell apoptosis, wound repair, and relative quantity of mRNA level of inflammatory cytokines and angiogenetic transcription factors were determined. The results showed there was a significantly reduced VSMCs proliferation with concentrationdependent (p &lt; 0.05). In the presence of OG at 100 μM, the wound area had healed after 24 hrs (59.2%) compared to PA (28.0%) group treatments. OG and PA supplementation significantly up-regulated eNOS and VEGF but down-regulated the TNF-α, CRP, CD36, iNOS, and NF-κB mRNA expression levels, while PA was a contrasting pattern (p &lt; 0.05). Therefore, supplementation of OG reduced the pro-inflammatory effects of inflammatory cytokines in macrophages and induced VEGF expression in VSMCs, contributing to anti-atherosclerotic effect.