Inducible Nitric Oxide Synthase Expression Research Articles

Increasing the radiation-induced cytotoxicity by silver nanoparticles and docetaxel in prostate cancer cells.

Radiation therapy is utilized for treatment of localized prostate cancer. Nevertheless, cancerous cells frequently develop radiation resistance. While higher radiation doses have not always been effective, radiosensitizers have been extensively studied for their ability to enhance the cytotoxic effects of radiation.So, this study aims to evaluate the possible radiosensitization effects of docetaxel (DTX) and silver nanoparticles (SNP) in LNCaP cells. The cytotoxic effects of DTX, SNP and2Gy of X-Ray radiation treatments were assessed in human LNCaP cell line using the MTT test after 24 h. Moreover, the effects of DTX, SNP and radiation on Epidermal growth factor (EGF), Caspase 3, inducible nitric oxide synthase and E-cadherin gene expression were analyzed using the Real-time PCR method. The level of Hydrogen peroxide (H2O2), an oxidative stress marker, was also detected 24h after various single and combined treatments. The combinations of SNP (in low toxic concentration) and/or DTX (0.25× IC50 and 0.5 × IC50 concentrations for triple and double combinations respectively) with radiation induced significant cytotoxicity in LNCaP cells in comparison to monotherapies. These cytotoxic effects were associated with the downregulation of EGF mRNA. Additionally, H2O2 levels increased after Radiation + SNP + DTX triple combination and double combinations including Radiation + SNP and Radiation + DTX versus single treatments. The triple combination treatment also increased Caspase 3 and and E-cadherin mRNA levels in compared to single treatments in LNCaP cells. Our results indicate that the combination of SNP and DTX with radiation induces significant anti-cancer effects. Upregulation of Caspase 3 and E-cadherin gene expression, and decreased mRNA expression level of EGF may be exerted specifically by use of this combination versus single treatments.

Protective Effects of an Oligo-Fucoidan-Based Formula against Osteoarthritis Development via iNOS and COX-2 Suppression following Monosodium Iodoacetate Injection

Osteoarthritis (OA) is a debilitating joint disorder characterized by cartilage degradation and chronic inflammation, accompanied by high oxidative stress. In this study, we utilized the monosodium iodoacetate (MIA)-induced OA model to investigate the efficacy of oligo-fucoidan-based formula (FF) intervention in mitigating OA progression. Through its capacity to alleviate joint bearing function and inflammation, improvements in cartilage integrity following oligo-fucoidan-based formula intervention were observed, highlighting its protective effects against cartilage degeneration and structural damage. Furthermore, the oligo-fucoidan-based formula modulated the p38 signaling pathway, along with downregulating cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression, contributing to its beneficial effects. Our study provides valuable insights into targeted interventions for OA management and calls for further clinical investigations to validate these preclinical findings and to explore the translational potential of an oligo-fucoidan-based formula in human OA patients.

Tectorigenin inhibits inflammatory responses in murineinflammatory bowel disease and LPS-stimulated macrophages via inactivating MAPK signaling pathway.

Considering the antihepatitis effects of Tectorigenin (TEC), and the same adenosine mitogen-activated protein kinase (MAPK) pathway in both hepatitis and inflammatory bowel disease (IBD) models, exploring the role of TEC in IBD is contributive to develop a new treatment strategy against IBD. The IBD mouse model was constructed by feeding with dextran sodium sulfate (DSS) and injection of TEC. Afterward, the mouse body weight, colon length, and disease activity index (DAI) were tested to assess the enteritis level. Mouse intestine lesions were detected by hematoxylin and eosin staining. Murine macrophages underwent lipopolysaccharide (LPS) induction to establish an inflammation model. Cell viability was determined by cell counting kit-8 assay. Enzyme-linked immunosorbent assay was performed to measure interleukin 6(IL-6) and tumor necrosis factor-α (TNF-α) levels. Cyclooxygenase-2(COX-2) and inducible nitric oxide synthase(iNOS) expressions were quantified via quantitative reverse transcription polymerase chain reaction. Levels of MAPK pathway-related proteins (p-P38, P38, p-Jun N-terminal kinase(JNK), JNK, signal-regulated kinase(ERK), p-ERK), COX-2 and iNOS were quantitated by Western blot. TEC improved the inflammatory response through ameliorating weight loss, shortening colon, and increasing DAI score in IBD mouse. Expressions of intestinal inflammatory factors (IL-6, TNF-α, iNOS and COX-2) and MAPK pathway-related proteins (p-P38, p-JNK, and p-ERK) were increased both in DSS-induced mouse intestinal tissue, but TEC inhibited expressions of inflammatory factors. The same increased trend was identified in LPS-induced macrophages, but TEC improved macrophage inflammation, as evidenced by downregulation of inflammatory factors. TEC mitigates IBD and LPS-induced macrophage inflammation in mice via inhibiting MAPK signaling pathway.

Absence of PD-L1 signaling hinders macrophage defense against Mycobacterium tuberculosis via upregulating STAT3/IL-6 pathway

The blockade of programmed death-ligand 1 (PD-L1) pathway has been clinically used in cancer immunotherapy, while its effects on infectious diseases remain elusive. Roles of PD-L1 signaling in the macrophage-mediated innate immune defense against M.tb is unclear. In this study, the outcomes of tuberculosis (TB) in wild-type (WT) mice treated with anti-PD-1/PD-L1 therapy and macrophage-specific Pdl1-knockout (Pdl1ΔΜΦ) mice were compared. Treatment with anti-PD-L1 or anti-PD-1 benefited protection against M.tb infection in WT mice, while Pdl1ΔΜΦ mice exhibited the increased susceptibility to M.tb infection. Mechanistically, the absence of PD-L1 signaling impaired M.tb killing by macrophages. Furthermore, elevated STAT3 activation was found in PD-L1-deficient macrophages, leading to increased interleukin (IL)-6 production and reduced inducible nitric oxide synthase (iNOS) expression. Inhibiting STAT3 phosphorylation partially impeded the increase in IL-6 production and restored iNOS expression in these PD-L1-deficient cells. These findings provide valuable insights into the complexity and mechanisms underlying anti-PD-L1 therapy in the context of tuberculosis.

Salvimulticanol from Salvia multicaulis suppresses LPS-induced inflammation in RAW264.7 macrophages: in vitro and in silico studies.

Sustained inflammatory responses can badly affect several vital organs and lead to chronic inflammation-related disorders, such as atherosclerosis, pneumonia, rheumatoid arthritis, obesity, diabetes, Alzheimer's disease, and cancers. Salvia multicaulis is one of the widely distributed plants that contains several biologically active phytochemicals and diterpenoids with anti-inflammatory effects. Therefore, finding alternative and safer natural plant-extracted compounds with good curative anti-inflammatory efficiencies is an urgent need for the clinical treatment of inflammation-related diseases. In the current study, S. multicaulis Vahl was used to extract and isolate two compounds identified as salvimulticanol and candesalvone B methyl ester to examine their effects against inflammation in murine macrophage RAW264.7 cells that were induced by lipopolysaccharide (LPS). Accordingly, after culturing RAW264.7 cells and induction of inflammation by LPS (100ng/ml), cells were exposed to different concentrations (9, 18, 37.5, 75, and 150µM) of each compound. Then, Griess assay for detection of nitric oxide (NO) levels and western blotting for the determination of inducible nitric oxide synthase (iNOS) expression were performed. Molecular docking and molecular dynamics (MD) simulation studies were employed to investigate the anti-inflammatory mechanism. Our obtained results validated that the level of NO was significantly decreased in the macrophage cell suspensions as a response to salvimulticanol treatment in a dose-dependent manner (IC50: 25.1 ± 1.2µM) as compared to the methyl ester of candesalvone B which exerted a weaker inhibition (IC50: 69.2 ± 3.0µM). This decline in NO percentage was comparable with a down-regulation of iNOS expression by western blotting. Salvimulticanol strongly interacted with both the Toll-like receptor 4 (TLR4)/myeloid differentiation factor 2 (MD-2) complex and the inhibitor of nuclear factor kappa-B (NF-κB) kinase subunit beta (IKKβ) to disrupt their inflammatory activation due to the significant hydrogen bonds and effective interactions with amino acid residues present in the target proteins' active sites. S.multicaulis is a rich natural source of the aromatic abietane diterpenoid, salvimulticanol, which exerted a strong anti-inflammatory effect through targeting iNOS and diminishing NO production in LPS-induced RAW264.7 cells in a mechanism that is dependent on the inhibition of TLR4-MD-2 and IKKβ as activators of the classical NF-κB-mediated inflammatory pathway.

New insight into arginine and tryptophan metabolism in macrophage activation during tuberculosis.

Arginine and tryptophan are pivotal in orchestrating cytokine-driven macrophage polarization and immune activation. Specifically, interferon-gamma (IFN-γ) stimulates inducible nitric oxide synthase (iNOS) expression), leading to the conversion of arginine into citrulline and nitric oxide (NO), while Interleukin-4 (IL4) promotes arginase activation, shifting arginine metabolism toward ornithine. Concomitantly, IFN-γ triggers indoleamine 2,3-dioxygenase 1 (IDO1) and Interleukin-4 induced 1 (IL4i1), resulting in the conversion of tryptophan into kynurenine and indole-3-pyruvic acid. These metabolic pathways are tightly regulated by NAD+-dependent sirtuin proteins, with Sirt2 and Sirt5 playing integral roles. In this review, we present novel insights that augment our understanding of the metabolic pathways of arginine and tryptophan following Mycobacterium tuberculosis infection, particularly their relevance in macrophage responses. Additionally, we discuss arginine methylation and demethylation and the role of Sirt2 and Sirt5 in regulating tryptophan metabolism and arginine metabolism, potentially driving macrophage polarization.

Anti-inflammatory potential via the MAPK signaling pathway of Lactobacillus spp. isolated from canine feces.

Two probiotic candidates, Lactobacillus reuteri C1 (C1) and Lactobacillus acidophilus C5 (C5), which were previously isolated from canines, were evaluated in the present study. L. reuteri and L. acidophilus have anti-oxidant, anti-inflammatory, immune-enhancing, and anti-cancer properties and exhibit various probiotic effects in humans and animals. The strains C1 and C5 demonstrated good tolerance to acid and bile salt exposure, exhibited effective adhesion to HT-29 cell monolayer, and displayed sensitivity to antibiotics, thus affirming their probiotic characteristics. Moreover, C1 and C5 exhibited the ability to downregulate the expression of inducible NO synthase (iNOS), an immunomodulatory factor, leading to a reduction in NO production in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. These strains also demonstrated potent anti-inflammatory effects in LPS-stimulated RAW 264.7 cells, achieved through the augmentation of anti-inflammatory cytokine IL-10 expression and the inhibition of pro-inflammatory cytokine IL-1β expression. These anti-inflammatory effects of C1 and C5 were closely associated with the mitogen-activated protein kinase (MAPK) signaling pathway. The results of the present study suggest that the C1 and C5 probiotic candidates attenuate LPS-induced inflammation via the MAPK signaling pathway and the strains can be used as probiotics considering their anti-inflammatory potential.

Nitric oxide and tumor necrosis factor-⍺ levels are negatively correlated in endotoxin tolerance recovery in vitro

Endotoxin tolerance (ET) is the hyporesponsiveness to lipopolysaccharide (LPS) after prior exposure. It is characterized by the downregulation of pro-inflammatory cytokine levels. Although ET protects against inflammation, its abolishment or recovery is critical for immunity. Nitric oxide (NO) plays various roles in the development of ET; however, its specific role in ET recovery remains unknown. To induce ET, RAW264.7 cells (a murine macrophage cell line) were pre-exposed to LPS (LPS1, 100 ng/mL for 24 h) and subsequently re-stimulated with LPS (LPS2, 100 ng/mL for 24 h). Expression of cytokines, NO, nitrite and inducible NO synthase (iNOS) were measured after 0, 12, 24, and 36 h of resting after LPS1 treatment with or without the iNOS-specific inhibitor, 1400W. LPS2-induced tumor necrosis factor-⍺ (TNF-⍺) and interleukin-6 (IL-6) were downregulated after LPS1 treatment, confirming the development of ET. Notably, TNF-⍺ and IL-6 levels spontaneously rebounded after 12–24 h of resting following LPS1 treatment. In contrast, levles of NO, nitrite and iNOS increased during ET development and decreased during ET recovery. Moreover, 1400W inhibited ET development and blocked the early production of NO (<12 h) during ET recovery. Our findings suggest a negative correlation between iNOS-induced NO and cytokine levels in the abolishment of ET.

Anti-Inflammatory Activity of Cyanobacteria Pigment Extracts: Physiological Free Radical Scavenging and Modulation of iNOS and LOX Activity.

Cyanobacteria are among the oldest organisms colonizing Earth. Their great biodiversity and ability to biosynthesize secondary metabolites through a variety of routes makes them attractive resources for biotechnological applications and drug discovery. In this pioneer study, four filamentous cyanobacteria (Cephalothrix lacustris LEGE 15493, Leptolyngbya boryana LEGE 15486, Nodosilinea nodulosa LEGE 06104 and Leptothoe sp. LEGE 11479) were explored for their anti-inflammatory potential in cell and cell-free in vitro bioassays, involving different inflammatory mediators and enzymes. Extracts of different polarities were sequentially prepared and chemically characterized for their content of phycobiliproteins (PBPs) and carotenoids. HPLC-PDA analysis of the acetone extracts revealed β-carotene to be the dominant carotenoid (18.4-44.3 mg/g) and zeaxanthin as the dominant xanthophyll (52.7-192.9 mg/g), with Leptothoe sp. LEGE 11479 and Nodosilinea nodulosa LEGE 06104, respectively, being the richest strains. The PBP profile was in accordance with the color presented by the aqueous extracts, with Leptolyngbya boryana LEGE 15486 being the richest in phycocyanin (204.5 μg/mg) and Leptothoe sp. LEGE 11479 the richest in phycoerythrin (78.5 μg/mg). Aqueous extracts were more effective in superoxide anion radical scavenging, while acetone ones were more effective in scavenging nitric oxide radical (●NO) and in inhibiting lipoxygenase. Acetone extracts also reduced ●NO production in lipopolysaccharide-stimulated RAW 264.7 macrophages, with the mechanistic study suggesting a downregulation of the inducible nitric oxide synthase expression. Nodosilinea nodulosa LEGE 06104 and Leptothoe sp. LEGE 11479 acetone extracts presented the lowest IC50 values for the mentioned assays, pointing them out as promising resources for the development of new multi-target anti-inflammatory therapies.

Ionizing radiation‑induced modification of nialamide as an anti‑inflammatory agent against lipopolysaccharide‑induced RAW 264.7 and DH82 cells.

Nialamide is a non-selective monoamine oxidase inhibitor that was widely used as an antidepressant. However, it has been prohibited for decades in the depressive medicine market due to the adverse hepatotoxic side effects. The re-use of drugs that have been withdrawn from the market represents a promising approach for the development of novel incrementally modified drugs and, in this context, ionizing radiation can serve as a powerful tool for producing new drug candidates. The present study exposed nialamide to γ radiation at 50 kGy to obtain the novel cyclized benzylamide, nialaminosin (compound 2), along with five known compounds, 3-amino-N-benzylpropanamide (compound 3), 3-methoxy-N-benzylpropanamide (compound 4), 3-hydroxy-N-benzylpropanamide (HBPA; compound 5), N-benzylpropanamide (compound 6) and isonicotinamide (compound 7). Among the isolated compounds, HBPA was established to inhibit the lipopolysaccharide-induced overproduction of pro-inflammatory mediators, including nitric oxide (NO) and prostaglandin E2 and cytokines including TNF-α, IL-6 and IL-10, without causing cytotoxicity to both RAW 264.7 and DH82 cells. Furthermore, HBPA was found to reduce the protein expression of inducible NO synthase and cyclooxygenase-2 in macrophages and compared with nialamide, it was established to have more potent radical scavenging activity. The present study therefore suggested the application of HBPA for the improvement of anti-inflammatory properties using ionizing radiation technology on the withdrawn drug nialamide.

AMPK activation attenuates central sensitization in a recurrent nitroglycerin-induced chronic migraine mouse model by promoting microglial M2-type polarization

BackgroundEnergy metabolism disorders and neurogenic inflammation play important roles in the central sensitization to chronic migraine (CM). AMP-activated protein kinase (AMPK) is an intracellular energy sensor, and its activation regulates inflammation and reduces neuropathic pain. However, studies on the involvement of AMPK in the regulation of CM are currently lacking. Therefore, this study aimed to explore the mechanism underlying the involvement of AMPK in the central sensitization to CM.MethodsMice with recurrent nitroglycerin (NTG)-induced CM were used to detect the expression of AMPK protein in the trigeminal nucleus caudalis (TNC). Following intraperitoneal injection of the AMPK activator 5-aminoimidazole-4-carboxyamide ribonucleoside (AICAR) and inhibitor compound C, the mechanical pain threshold, activity level, and pain-like behaviors in the mice were measured. The expression of calcitonin gene-related peptide (CGRP) and cytokines, M1/M2 microglia, and NF-κB pathway activation were detected after the intervention.ResultsRepeated NTG injections resulted in a gradual decrease in AMPK protein expression, and the negative regulation of AMPK by increased ubiquitin-like plant homeodomain and RING finger domain 1 (UHRF1) expression may counteract AMPK activation by increasing ADP/ATP. AICAR can reduce the hyperalgesia and pain-like behaviors of CM mice, improve the activity of mice, reduce the expression of CGRP, IL-1β, IL-6, and TNF-α in the TNC region, and increase the expression of IL-4 and IL-10. Moreover, AMPK in TNC was mainly located in microglia. AICAR could reduce the expression of inducible NO synthase (iNOS) in M1 microglia and increase the expression of Arginase 1 (Arg1) in M2 microglia by inhibiting the activation of NF-κB pathway.ConclusionsAMPK was involved in the central sensitization of CM, and the activation of AMPK reduced neuroinflammation in NTG-induced CM mice. AMPK may provide new insights into interventions for energy metabolism disorders and neurogenic inflammation in migraine.

CircRNA_SLC8A1 promotes the survival of mycobacterium tuberculosis in macrophages by upregulating expression of autophagy-related protein SQSTM1/p62 to activate the NF-κB pathway

Macrophages act as the first immune defense line of the host against Mycobacterium tuberculosis (Mtb). A previous study showed that circRNA_SLC8A1 was significantly upregulated in Mtb-infected macrophages, but its regulatory mechanism in anti-tuberculosis infection is unclear. Therefore, this study aimed to investigate the role of circRNA_SLC8A1 in the anti-tuberculosis activity of macrophages. We showed that circRNA_SLC8A1 was upregulated in tuberculosis patients. Moreover, the binding sites of miR-20b-5p on circRNA_SLC8A1 and Sequestosome 1 (SQSTM1/p62) mRNA were predicted by StarBase and verified by the double luciferase reporter gene assay. Next, we found that miR-20b-5p expression was decreased, while SQSTM1 protein expression was increased in a time- and dose-dependent manner in the human macrophage U937 in response to Mtb infection. Furthermore, circRNA_SLC8A1 overexpression vector (circRNA_SLC8A1) or shRNA (sh-circRNA_SLC8A1) and/or miR-20b-5p mimic or inhibitor and/or SQSTM1 overexpression vector (SQSTM1) or small interfering RNA (si-SQSTM1) or its corresponding control were transfected into Mtb-infected macrophages. Results showed that overexpression of circRNA_SLC8A1 or miR-20b-5p inhibitor promoted the secretion of pro-inflammatory factors IL-1β, IL-6, and TNF-α, increased Nitric Oxide (NO) content and inducible nitric oxide synthase (iNOS) expression, inhibited Reactive oxygen species (ROS) production. Cleaved-caspase-3 protein expression, and cell apoptosis, and promoted Mtb survival. Silencing SQSTM1 inhibited secretion of pro-inflammatory factors and activation of the NF-κB pathway. Overexpression of miR‐20b‐5p blocked the promoting of circ‐SLC8A1 on SQSTM1 protein expression. In summary, circRNA_SLC8A1 sponged miR‐20b‐5p to upregulate SQSTM1/p62 expression and promoted Mtb survival in macrophages through the NF-κB signaling pathway.

Involvement of IL-17 A/IL-17 Receptor A with Neutrophil Recruitment and the Severity of Coronary Arteritis in Kawasaki Disease

PurposeTo assess the role of the interleukin (IL)-17 A/IL-17 receptor A (IL-17RA) in Kawasaki disease (KD)-related coronary arteritis (CA).MethodsIn human study, the plasma levels of IL-17 A and coronary arteries were concurrently examined in acute KD patients. In vitro responses of human coronary endothelial cells to plasma stimulation were investigated with and without IL-17RA neutralization. A murine model of Lactobacillus casei cell-wall extract (LCWE)-induced CA using wild-type Balb/c and Il17ra-deficient mice were also inspected.ResultsThe plasma levels of IL-17 A were significantly higher in KD patients before intravenous immunoglobulin therapy, especially in those with coronary artery lesion. The pre-IVIG IL-17 A levels positively correlated with maximal z scores of coronary diameters and plasma-induced endothelial mRNA levels of chemokine (C-X-C motif) ligand-1, IL-8, and IL-17RA. IL-17RA blockade significantly reduced such endothelial upregulations of aforementioned three genes and inducible nitric oxide synthase, and neutrophil transmigration. IL-17RA expression was enhanced on peripheral blood mononuclear cells in pre-IVIG KD patients, and in the aortic rings and spleens of the LCWE-stimulated mice. LCWE-induced CA composed of dual-positive Ly6G- and IL-17 A-stained infiltrates. Il17ra-deficient mice showed reduced CA severity with the fewer number of neutrophils and lower early inducible nitric oxide synthase and chemokine (C-X-C motif) ligand-1 mRNA expressions than Il17ra+/+ littermates, and absent IL-17RA upregulation at aortic roots.ConclusionIL-17 A/IL-17RA axis may play a role in mediating aortic neutrophil chemoattraction, thus contributory to the severity of CA in both humans and mice. These findings may help to develop a new therapeutic strategy toward ameliorating KD-related CA.

Curcuma longa L. extract increased immune responses in RAW 264.7 cells and cyclophosphamide-induced BALB/c mice

Curcuma longa L. extract (CLE) exerts various biological functions including antioxidant, anti-inflammation, anticancer, and antiallergenic effects. However, its immune-enhancing capacity remains unclear. Therefore, the immune-enhancing effect of CLE was investigated in RAW 264.7 cells and cyclophosphamide (CPP)-induced immunosuppression model. CLE upregulated nitric oxide (NO) and reactive oxygen species production and increased inducible nitric oxide synthase and cyclooxygenase-2 expression without affecting the RAW 264.7 cells viability. The results of quantitative real-time reverse transcription polymerase chain reaction and sandwich enzyme-linked immunosorbent assay showed that CLE increased the gene expression and protein levels of tumor necrosis factor-α, interleukin-6, and interleukin-1β in RAW 264.7 cells. Moreover, CLE upregulated p65, I kappa B kinase α/β, and I kappa B α (IκBα) phosphorylation and downregulated IκBα expression in RAW 264.7 cells. CLE also increased p65 translocation from the cytoplasmic to the nucleus in RAW 264.7 cells. The oral administration of CLE increased organ indexes (including the spleen and thymus) and NO production in peritoneal macrophages and improved natural killer cell activity in CPP-induced immunosuppression BALB/c mice. Overall, CLE could be a useful health functional food material that can improve innate immunity via macrophage activation.

Libertellenone C attenuates oxidative stress and neuroinflammation with the capacity of NLRP3 inhibition

Neurodegenerative diseases (NDs) are common chronic diseases arising from progressive damage to the nervous system. Here, in-house natural product database screening revealed that libertellenone C (LC) obtained from the fermentation products of Arthrinium arundinis separated from the gut of a centipede collected in our Tongji campus, showed a remarkable neuroprotective effect. Further investigation was conducted to clarify the specific mechanism. LC dose-dependently reversed glutamate-induced decreased viability, accumulated reactive oxygen species, mitochondrial membrane potential loss, and apoptosis in SH-SY5Y cells. Network pharmacology analysis predicted that the targets of LC were most likely directly related to oxidative stress and the regulation of inflammatory factor-associated signaling pathways. Further study demonstrated that LC attenuated nitrite, TNF-α, and IL-1β production and decreased inducible nitric oxide synthase and cyclooxygenase expression in lipopolysaccharide-induced BV-2 cells. LC could directly inhibit NLRP3 inflammasome activation by decreasing the expression levels of NLRP3, ASC, cleaved Caspase-1, and NF-κB p65. Our results provide a new understanding of how LC inhibits the NLRP3 inflammasome in microglia, providing neuroprotection. These findings might guide the development of effective LC-based therapeutic strategies for NDs.

Enhanced Anti-Inflammatory and Non-Alcoholic Fatty Liver Disease (NAFLD) Improvement Effects of Bacillus subtilis-Fermented Fagopyrum tataricum Gaertner

In this study, we investigated the enhanced anti-inflammatory activity and the effects on non-alcoholic fatty liver disease (NAFLD) of fermented Fagopyrum tataricum (F. tataricum) Gaertner extract (FFT) through in vitro analysis. We utilized high-performance liquid chromatography (HPLC) to analyze the non-fermented F. tataricum Gaertner extract (NFT) and the marker components, rutin and quercetin in FFT, to confirm changes in composition due to fermentation. The anti-inflammatory activity of NFT and FFT was evaluated using a lipopolysaccharide (LPS)-induced RAW 264.7 cell inflammation model. Simultaneously, the NAFLD improvement effects were measured by evaluating lipid accumulation and the expression of lipid synthesis regulators in free fatty acid (FFA)-induced HepG2 cells. HPLC analysis confirmed an increase in rutin content after the fermentation of F. tataricum Gaertner. Upon treatment with NFT and FFT at a concentration of 400 μg/mL, LPS-induced nitric oxide (NO) production values in RAW 264.7 cells were reduced to 16.12 μM and 2.09 μM, respectively, indicating enhanced significant inhibition (p &lt; 0.05) of NO production through fermentation. FFT demonstrated the significant inhibition (p &lt; 0.05) of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) protein, and inflammatory cytokine mRNA expression through the nuclear factor kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways in LPS-induced RAW 264.7 cells. In FFA-induced HepG2 cells, FFT significant suppressed (p &lt; 0.05) lipid accumulation and the expression of sterol regulatory element binding protein (SREBP)-1c, CCAAT/enhancer binding protein (C/EBP)α proteins, and acetyl-CoA carboxylase (ACC) mRNA. The results of this study suggest the potential utilization of FFT as a material for improving NAFLD.

Inhibitory effects of bioactive compounds on UVB-induced photodamage in human keratinocytes: modulation of MMP1 and Wnt signaling pathways.

UVB radiation significantly threatens skin health, contributing to wrinkle formation and an elevated risk of skin cancer. This study aimed to explore bioactive compounds with potential UVB-protective properties. Using in silico analysis, we chose compounds to reduce binding energy with matrix metalloproteinase-1 (MMP1). Piperitoside, procyanidin C1, and mulberrofuran E emerged as promising candidates through this computational screening process. We investigated the UVB-protective efficacy of the selected compounds and underlying mechanisms in human immortalized keratinocytes (HaCaT). We also investigated the molecular pathways implicated in their action, focusing on the transforming growth factor (TGF)-β and wingless-related integration site (Wnt)/β-catenin signaling pathways. In UVB-exposed HaCaT cells (100mJ/cm2 for 30min), piperitoside, procyanidin C1, and mulberrofuran E significantly reduced reactive oxygen species (ROS) and lipid peroxides, coupled with an augmentation of collagen expression. These compounds suppressed MMP1, tumor necrosis factor-alpha (TNF-α), and inducible nitric oxide synthase (iNOS) expression, while they concurrently enhanced collagen-1 (COL1A1), β-catenin (CTNNB1), and superoxide dismutase type-1 (SOD1) expression. Furthermore, Wnt/β-catenin inhibitors, when administered subsequently, partially counteracted the reduction in MMP1 expression and alleviated inflammatory and oxidative stress markers induced by the bioactive compounds. In conclusion, piperitoside, procyanidin C1, and mulberrofuran E protected against UVB-induced damage in HaCaT cells by inhibiting MMP1 expression and elevating β-catenin expression. Consequently, these bioactive compounds emerge as promising preventive agents for UVB-induced skin damage, promoting skin health.

Abstract IA23: "NO Target for Cancer Stem Cells"

Abstract Developing novel therapies that target cancer metastasis, chemoresistant cancer stem cells, and augment the efficacy of systemic chemo/radiotherapies is crucial to reducing cancer-associated mortality. A protein within the tumor microenvironment that has gained interest as a promising therapeutic target is nitric oxide synthase (NOS). Our previous findings have shown that increased inducible nitric oxide synthase (iNOS) expression is a poor prognostic indicator and associated with worse overall survival in triple negative breast cancer (TNBC) patients. Nitric oxide (NO) is a unique molecule in its ability to target multiple oncogenic pathways in a spatial and temporal manner, such as PI3K, extracellular signal-regulated kinase (ERK), β-catenin pathway, transforming growth factor beta (TGFβ) signaling, and hypoxia-inducible factor (HIF). Growing evidence has also suggested that hyperactivation of the phosphoinositide-3-kinase (PI3K) survival signaling pathway may be one of the most common oncogenic aberrations in certain TNBC subsets, in particular metaplastic breast cancer (MpBC). We will summarize the role of NO in PI3K-Akt pathway activation in human TNBC, as well as data from The Cancer Genome Atlas of the role of iNOS and PI3K-Akt interaction. This review discusses the current knowledge of the roles of nitric oxide (NO) in tumor progression and cancer metastasis, and promising preclinical studies that evaluated NOS inhibitors as an anti-cancer therapy. Lastly, we will offer a perspective on utilizing NOS inhibitors in the clinical setting, highlighting the prospects and outstanding challenges. Citation Format: Jenny C. Chang. "NO Target for Cancer Stem Cells" [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr IA23.

Exploring the Synergistic Effect of Sildenafil and Green Tea Polyphenols on Breast Cancer Stem Cell-like Cells and their Parental Cells: A Potential Novel Therapeutic Approach.

Many cancer studies have intensely focused on the role of diet, among other factors involved in cancer establishment. The positive effect of green tea polyphenols (GTP) on controlling breast cancer cells has been reported in several studies. Cancer stem cell-like cells (CSC-LCs) possessing self-renewal, metastatic, and drug-resistant capacities are considered prominent therapeutic targets. In many tumors, inducible nitric oxide synthase (iNOS) expression levels are high; however, they have a dual effect on breast cancer pathogenesis. This study aimed to investigate the cytotoxicity of the iNOS agonist (Sildenafil) and antagonist (LNAME), both alone and in combination with GTP, on MDA-MB-231, CD44+/CD24- CSC-LCs, and their parental cells (MCF-7). The cell viability assay has been studied using the MTT assay. To analyze drug-drug combinations, CompuSyn and Combenefit software were used. The cytotoxicity mechanism was determined using flow cytometric analysis. L-NAME and GTP showed a synergistic effect on MDA-MB-231 and CSC-LCs. Such an effect was not observed on MCF-7. Sildenafil and GTP, on the other hand, showed synergistic cytotoxicity in all the cells mentioned above. Flow cytometric tests resulted in more than 70% apoptosis in MDA-MB-231 and MCF-7. Also, sub-G1 arrest among MCF-7 cells and a considerable decrease in ROS production by MDA-MB-231 cells following treatment with Sildenafil and GTP were observed. Sildenafil, in combination with flavonoids, may be considered a novel strategy for cancer treatment.

Novel insights on the probable mechanism associated with histamine oral model-inducing neuropathological and behavioral toxicity in rats.

Histamine (HIS) is an important chemical mediator that causes vasodilation and contributes to anaphylactic reactions. Recently, HIS is an understudied neurotransmitter in the central nervous system, and its potential role in neuroinflammation and neurodegeneration is a critical area of research. So, the study's goal is to investigate the consequences of repeated oral intake of HISon the rat's brain and explore the mechanistic way of its neurotoxicity. Thirty male rats were divided into three groups (n = 10). The following treatments were administered orally to all rats every day for 14 days. Group (1) was given distilled water, whereas groups (2 & 3) were given HIS at dosage levels 250and 500 mg/kg body weight (BWT), respectively. Brain tissue samples were collected at 7- and 14-days from the beginning of the experiment. Our results revealed that continuous oral administration of HIS at both doses for 14 days significantly reduced the BWT and induced severe neurobehavioral changes, including depression, dullness, lethargy, tremors, abnormal walking, and loss of spatial learning and memory in rats. In all HIS receiving groups, HPLC data showed a considerable raise in the HIS contents of the brain. Additionally, the daily consumption of HIS causes oxidative stress that is dose- and time-dependent which is characterized by elevation of malondialdehyde levels along with reduction of catalase activity and reduced glutathione levels. The neuropathological lesions were commonly observed in the cerebrum, striatum, and cerebellum and confirmed by the immunohistochemistry staining that demonstrating moderate to strong caspase-3 and inducible nitric oxide synthaseexpressions in all HIS receiving groups, mainly those receiving 500 mg/kg HIS. NF-κB, TNF-α, and IL-1β gene levels were also upregulated at 7- and 14-days in all HIS groups, particularly in those getting 500 mg/kg. We concluded that ROS-induced apoptosis and inflammation was the essential mechanism involved in HIS-mediated neurobehavioral toxicity and histopathology.