Effects of moxibustion on intestinal barrier function and TLR4/NF-κB p65 signaling pathway in obese rats. / è‰¾ç¸å¯¹è‚¥èƒ–å¤§é¼ è‚ é“å±éšœåŠŸèƒ½åŠTLR4/NF-κB p65信号通路的影响.

OBJECTIVES: To observe the effects of moxibustion on intestinal barrier function and Toll-like receptor 4 (TLR4)/nuclear factor-κB p65 (NF-κB p65) signaling pathway in obese rats and explore the mechanism of moxibustion in the intervention of obesity. METHODS: Fifty-five Wistar rats of SPF grade were randomly divided into a normal group (10 rats) and a modeling group (45 rats). In the modeling group, the obesity model was established by feeding high-fat diet. Thirty successfully-modeled rats were randomized into a model group, a moxibustion group, and a placebo-control group, with 10 rats in each one. In the moxibustion group, moxibustion was applied at the site 3 cm to 5 cm far from the surface of "Zhongwan" (CV 12), with the temperature maintained at (46±1 ) ℃. In the placebo-control group, moxibustion was applied at the site 8 cm to 10 cm far from "Zhongwan" (CV 12), with the temperature maintained at (38±1) ℃. The intervention was delivered once daily for 8 weeks in the above two groups. The body mass and food intake of the rats were observed before and after intervention in each group. Using ELISA methool, the levels of serum triacylglycerol (TG), total cholesterol (TC) and lipopolysaccharide (LPS) were detected and the insulin resistance index (HOMA-IR) was calculated. HE staining was used to observe the morphology of colon tissue. The mRNA expression of zonula occludens-1 (ZO-1), Occludin, Claudin-1, TLR4 and NF-κB p65 in the colon tissue was detected by quantitative real-time PCR; and the protein expression of ZO-1, Occludin, Claudin-1, TLR4 and NF-κB p65 was detected by Western blot in the rats of each group. RESULTS: Compared with the normal group, the body mass, food intake, the level of HOMA-IR, and the serum levels of TC, TG and LPS were increased in the rats of the model group (P<0.01); those indexes in the moxibustion group were all reduced when compared with the model group and the placebo-control group respectively (P<0.01, P<0.05). Compared with the normal group, a large number of epithelial cells in the mucosa of colon tissue was damaged, shed, and the inflammatory cells were infiltrated obviously in the interstitium in the rats of the model group. When compared with the model group, in the moxibustion group, the damage of the colon tissue was recovered to various degrees and there were few infiltrated inflammatory cells in the interstitium, while, the epithelial injury of the colon tissue was slightly recovered and the infiltrated inflammatory cells in the interstitium were still seen in the placebo-control group. The mRNA and protein expressions of ZO-1, Occludin and Caudin-1 were decreased in the model group compared with those in the normal group (P<0.01). When compared with the model group and the placebo-control group, the mRNA and protein expressions of these indexes were increased in the moxibustion group (P<0.01, P<0.05). In the model group, the mRNA and protein expressions of TLR4 and NF-κB p65 were increased when compared with those in the normal group (P<0.01), and the mRNA and protein expressions of these indexes were reduced in the moxibustion group when compared with those in the model group and the placebo-control group (P<0.01). CONCLUSIONS: Moxibustion can reduce the body mass and food intake, regulate the blood lipid and improve insulin resistance in the rats of obesity. It may be related to alleviating inflammatory response through improving intestinal barrier function and modulating the intestinal TLR4/NF-κB p65 signaling pathway.

[Effect and mechanism of Maxing Shigan Decoction on reducing inflammatory response in rats with cough variant asthma via TLR4/MyD88/NF-κB and p38 MAPK signaling pathways].

This study aims to investigate the effect and mechanism of Maxingshigan Decoction on inflammation in the rat model of cough variant asthma(CVA). The SPF-grade SD rats of 6-8 weeks were randomized into normal, model, Montelukast sodium, and low-, medium-, and high-dose Maxing Shigan Decoction groups, with 8 rats in each group. The CVA rat model was induced by ovalbumin(OVA) and aluminum hydroxide sensitization and ovalbumin stimulation. The normal group and model group were administrated with equal volume of normal saline by gavage, and other groups with corresponding drugs by gavage. After the experiment, the number of white blood cells in blood and the levels of interleukin-6(IL-6), interleukin-10(IL-10), and tumor necrosis factor-α(TNF-α) in the serum were measured. The lung tissue was stained with hematoxylin-eosin(HE). Western blot was employed to determine the protein levels of nuclear factor-κB(NF-κB), Toll-like receptor 4(TLR4), myeloid differentiation protein(MyD88), and mitogen-activated protein kinase(MAPK) in the lung tissue. Real-time PCR was carried out to measure the mRNA levels of TLR4 and MyD88 in the lung tissue. Compared with the normal group, the model group showed increased white blood cells, elevated IL-6 and TNF-α levels(P<0.01), lowered IL-10 level(P<0.01), up-regulated protein levels of TLR4, MyD88, p-p65/NF-κB p65, and p-p38 MAPK/p38 MAPK(P<0.01) and mRNA levels of TLR4 and MyD88(P<0.01) in the lung tissue. HE staining showed obvious infiltration of inflammatory cells around the airway and cell disarrangement in the model group. Compared with the model group, Montelukast sodium and high-dose Maxing Shigan Decoction reduced the white blood cells, lowered the IL-6 and TNF-α levels(P<0.01), and elevated the IL-10 level(P<0.01). Moreover, they down-regulated the protein levels of TLR4, MyD88, p-p65/NF-κB p65, p-p38 MAPK/p38 MAPK in the lung tissue(P<0.01) and the mRNA levels of TLR4 and MyD88 in the lung tissue(P<0.01). HE staining showed that Montelukast sodium and high-dose Maxing Shigan Decoction reduced inflammatory cell infiltration and cell disarrangement. The number of white blood cells, the levels of IL-10 and TNF-α in the serum, the protein levels of TLR4, MyD88, p-p65/NF-κB p65, and p-p38 MAPK/p38 MAPK, and the mRNA levels of TLR4 and MyD88 in the lung tissue showed no significant differences between the Montelukast sodium group and high-dose Maxing Shigan Decoction group. Maxing Shigan Decoction can inhibit airway inflammation in CVA rats by inhibiting the activation of TLR4/MyD88/NF-κB and p38 MAPK signaling pathways.

Structure characteristics, protective effect and mechanisms of ethanol-fractional polysaccharides from Dendrobium officinale on acute ethanol-induced gastritis.

Gastritis is a common disease characterized by gastric ulcers and severe bleeding. Excessive daily alcohol consumption can cause acute gastritis, impacting individuals' quality of life. This study aims to explore the protective effects of different ethanol-fractional polysaccharides of Dendrobium officinale (EPDO) on acute alcohol-induced gastric injury in vivo. Results showed that EPDO-80, identified as a ß-glucan, exhibited significant anti-inflammatory properties in pathology. It could reduce the area of gastric mucosal injury and cell infiltration. EPDO-80 had a dose-effect relationship in reducing the levels of malondialdehyde and cyclooxygenase-2 and decreasing the levels of inflammation mediators such as tumor necrosis factor α. More extensively, EPDO-80 could inhibit the activation of the TNFR/IκB/NF-κB signaling pathway, reducing the production of TNF-α mRNA and cell apoptosis in organs. Conversely, EPDO-80 could promote changes in the gut microbiota structure. These findings suggest that EPDO-80 could have great potential in limiting oxidative stress and inflammation mediated by inhibiting the NF-κB signaling pathway, which is highly related to its ß-glucan structure and functions in gut microbiota.

L-AP Alleviates Liver Injury in Septic Mice by Inhibiting Macrophage Activation via Suppressing NF-κB and NLRP3 Inflammasome/Caspase-1 Signal Pathways.

Liver injury and progressive liver failure are severe life-threatening complications in sepsis, further worsening the disease and leading to death. Macrophages and their mediated inflammatory cytokine storm are critical regulators in the occurrence and progression of liver injury in sepsis, for which effective treatments are still lacking. l-Ascorbic acid 6-palmitate (L-AP), a food additive, can inhibit neuroinflammation by modulating the phenotype of the microglia, but its pharmacological action in septic liver damage has not been fully explored. We aimed to investigate L-AP's antisepticemia action and the possible pharmacological mechanisms in attenuating septic liver damage by modulating macrophage function. We observed that L-AP treatment significantly increased survival in cecal ligation and puncture-induced WT mice and attenuated hepatic inflammatory injury, including the histopathology of the liver tissues, hepatocyte apoptosis, and the liver enzyme levels in plasma, which were comparable to NLRP3-deficiency in septic mice. L-AP supplementation significantly attenuated the excessive inflammatory response in hepatic tissues of septic mice in vivo and in cultured macrophages challenged by both LPS and ATP in vitro, by reducing the levels of NLRP3, pro-IL-1ß, and pro-IL-18 mRNA expression, as well as the levels of proteins for p-I-κB-α, p-NF-κB-p65, NLRP3, cleaved-caspase-1, IL-1ß, and IL-18. Additionally, it impaired the inflammasome ASC spot activation and reduced the inflammatory factor contents, including IL-1ß and IL-18 in plasma/cultured superannuants. It also prevented the infiltration/migration of macrophages and their M1-like inflammatory polarization while improving their M2-like polarization. Overall, our findings revealed that L-AP protected against sepsis by reducing macrophage activation and inflammatory cytokine production by suppressing their activation in NF-κB and NLRP3 inflammasome signal pathways in septic liver.

[Ligusticum cycloprolactam inhibits IL-1ß-induced apoptosis and inflammation of rat chondrocytes via HMGB1/TLR4/NF-κB signaling pathway].

Chondrocytes are unique resident cells in the articular cartilage, and the pathological changes of them can lead to the occurrence of osteoarthritis(OA). Ligusticum cycloprolactam(LIGc) are derivatives of Z-ligustilide(LIG), a pharmacodynamic marker of Angelica sinensis, which has various biological functions such as anti-inflammation and inhibition of cell apoptosis. However, its protective effect on chondrocytes in the case of OA and the underlying mechanism remain unclear. This study conducted in vitro experiments to explore the molecular mechanism of LIGc in protecting chondrocytes from OA. The inflammation model of rat OA chondrocyte model was established by using interleukin-1ß(IL-1ß) to induce. LIGc alone and combined with glycyrrhizic acid(GA), a blocker of the high mobility group box-1 protein(HMGB1)/Toll-like receptor 4(TLR4)/nuclear factor-kappa B(NF-κB) signaling pathway, were used to intervene in the model, and the therapeutic effects were systematically evaluated. The viability of chondrocytes treated with different concentrations of LIGc was measured by the cell counting kit-8(CCK-8), and the optimal LIGc concentration was screened out. Annexin V-FITC/PI apoptosis detection kit was employed to examine the apoptosis of chondrocytes in each group. The enzyme-linked immunosorbent assay(ELISA) was employed to measure the expression of cyclooxygenase-2(COX-2), prostaglandin-2(PGE2), and tumor necrosis factor-alpha(TNF-α) in the supernatant of chondrocytes in each group. Western blot was employed to determine the protein levels of B-cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax), caspase-3, HMGB1, TLR4, and NF-κB p65. The mRNA levels of HMGB1, TLR4, NF-κB p65, and myeloid differentiation factor 88(MyD88) in chondrocytes were determined by real-time fluorescent quantitative PCR(RT-qPCR). The safe concentration range of LIGc on chondrocytes was determined by CCK-8, and then the optimal concentration of LIGc for exerting the effect was clarified. Under the intervention of IL-1ß, the rat chondrocyte model of OA was successfully established. The modeled chondrocytes showed increased apoptosis rate, promoted expression of COX-2, PGE2, and TNF-α, up-regulated protein levels of Bax, caspase-3, HMGB1, TLR4, and NF-κB p65 and mRNA levels of HMGB1, TLR4, NF-κB p65, and MyD88, and down-regulated protein level of Bcl-2. However, LIGc reversed the IL-1ß-induced changes of the above factors. Moreover, LIGc combined with GA showed more significant reversal effect than LIGc alone. These fin-dings indicate that LIGc extracted and derived from the traditional Chinese medicine A. sinensis can inhibit the inflammatory response of chondrocytes and reduce the apoptosis of chondrocytes, and this effect may be related to the HMGB1/TLR4/NF-κB signaling pathway. The pharmacological effect of LIGc on protecting chondrocytes has potential value in delaying the progression of OA and improving the clinical symptoms of patients, and deserves further study.

[Mechanism of Fushen Granules treat peritoneal dialysis-related peritonitis by regulating TLR4/NF-κB pathway:based on network pharmacology and animal experiments].

Network pharmacology was employed to probe into the mechanism of Fushen Granules in treating peritoneal dialysis-rela-ted peritonitis(PDRP) in rats. The main active components of Fushen Granules were searched against the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform, and their targets were predicted. PDRP-related targets were retrieved from DisGeNET and other databases. The common targets shared by the drug and the disease were identified by the online tool, and protein-protein interaction(PPI) network of the common targets. The obtained 276 common targets were imported into DAVID for GO function enrichment and KEGG pathway enrichment. The main signaling pathway of Fushen Granules in the treatment of PDRP was predicted as Toll-like receptor 4(TLR4)/nuclear factor(NF)-κB. The rat model of uremia was induced by 5/6 nephrectomy. From two weeks after operation, the rat model of peritoneal dialysis(PD) was established by intraperitoneal injection of 20 mL dialysate with 1.25% glucose every day. The sham operation group and model group received 2 mL normal saline by gavage every day. The rats in Fushen Gra-nules groups were administrated with 2 mL solutions of low-(0.54 g·kg~(-1)), medium-(1.08 g·kg~(-1)) and high-dose(2.16 g·kg~(-1)) Fushen Granules every day. The bifico group received 2 mL(113.4 mg·kg~(-1)) of bifico solution every day. At the end of the 8th week, the levels of serum creatinine(Scr) and blood urea nitrogen(BUN) in each group were measured. The serum levels of hypersensitive C reactive protein(hs-CRP), tumor necrosis factor(TNF)-α, and interleukin(IL)-6 were measured, and the pathological changes in the colon tissue were observed by hematoxylin-eosin(HE) staining. The serum levels of lipopolysaccharide(LPS) and lipopolysaccharide-binding protein(LBP) of rats were measured, and the expression levels of LBP, TLR4, NF-κB p65, inhibitor of κB kinase α(IκBα), TNF-α, and IL-1ß in the colon tissue were determined. Compared with sham operation group, the model group had abnormal structure of all layers of colon tissue, sparse and shorter intestinal villi, visible edema in mucosal layer, wider gap, obvious local inflammatory cell infiltration, significantly decreased body weight(P<0.01), and significantly increased kidney function index(Scr, BUN) content(P<0.01). Serum levels of inflammatory cytokines(hs-CRP, TNF-α, IL-6), LPS and LBP were significantly increased(P<0.01), protein expressions of LBP, TLR4, NF-κB p65, TNF-α and IL-1ß were significantly increased(P<0.01), and protein expressions of IκBα were significantly decreased(P<0.01). Compared with model group, intestinal villi damage in colonic tissue of rats in low-, medium-and high-dose Fushen Granules groups and bifico group were alleviated to different degrees, edema in submucosa was alleviated, space was narrowed, and inflammatory cell infiltration in lamina propria was reduced. The contents of renal function index(Scr, BUN) and serum inflammatory factors(hs-CRP, TNF-α, IL-6) were significantly decreased(P<0.05 or P<0.01) in medium-and high-dose Fushen Granules groups and bifico group(P<0.05 or P<0.01). Serum LPS and LBP contents in Fushen Granules group and bifico group were significantly decreased(P<0.01), protein expressions of LBP, TLR4, NF-κB p65, TNF-α and IL-1ß in Fushen Granules group were significantly decreased(P<0.05 or P<0.01), and protein expressions of IκBα were significantly increased(P<0.01). The expression of LBP protein in bifico group was significantly decreased(P<0.01). The results suggest that Fushen Granules can protect the residual renal function of PD rats, reduce the inflammatory response, and protect the colon tissue. Based on network pharmacology, TLR4/NF-κB pathway may be the main signaling pathway of Fushen granule in the treatment of PDRP. The results showed that Fushen Granules could improve intestinal inflammation and protect intestinal barrier to prevent PDRP by regulating the expression of key factors in TLR4/NF-κB pathway in colon of PD rats.

[Mechanism of traditional Chinese medicine in treating ulcerative colitis-related colorectal cancer by regulating NF-κB-related signaling pathways: a review].

Ulcerative colitis is a chronic, recurrent, and nonspecific intestinal inflammatory disease, which is difficult to cure and has the risk of deterioration into related tumors. Long-term chronic inflammatory stimulation can increase the risk of cancerization. With the signaling pathway as a key link in the regulation of tumor microenvironments, nuclear factor-kappa B(NF-κB) is an important regulator of intestinal inflammation. It can also be co-regulated as downstream factors of other signaling pathways, such as TLR4, MAPK, STAT, PI3K, and so on. At present, a large number of animal experiments have proved that traditional Chinese medicine(TCM) can reduce inflammation by interfering with NF-κB-related signaling pathways, improve intestinal inflammation, and inhibit the progression of inflammation to tumors. This article reviewed the relationship between NF-κB-related signaling pathways and the intervention mechanism of TCM, so as to provide a reference for the clinical treatment of ulcerative colitis and the optimization of related cancer prevention strategies.

Yanggan Jiangmei Formula alleviates hepatic inflammation and lipid accumulation in non-alcoholic steatohepatitis by inhibiting the NF-κB/NLRP3 signaling pathway.

The role of NF-κB and the NLRP3 inflammasome in the chronic inflammatory microenvironment of non-alcoholic steatohepatitis (NASH) has been posited as crucial. The Yanggan Jiangmei Formula (YGJMF) has shown promise in ameliorating hepatic steatosis in NASH patients, yet its pharmacological mechanisms remain largely unexplored. This study was conducted to investigate the efficacy of YGJMF in NASH and to elucidate its pharmacological underpinnings. To simulate NASH both in vivo and in vitro, high-fat-diet (HFD) rats and HepG2 cells stimulated with free fatty acids (FFAs) were utilized. The severity of liver injury and lipid deposition was assessed using serum indicators, histopathological staining, micro-magnetic resonance imaging (MRI), and the liver-to-muscle signal intensity ratio (SIRL/M). Furthermore, a combination of enzyme-linked immunosorbent assay (ELISA), immunohistochemistry (IHC), immunofluorescence, real-time quantitative polymerase chain reaction (RT-qPCR), and Western blotting analyses was employed to investigate the NF-κB/NLRP3 signaling pathway and associated cytokine levels. The results from liver pathology, MRI assessments, and biochemical tests in rat models demonstrated YGJMF's significant effectiveness in reducing liver damage and lipid accumulation. Additionally, YGJMF markedly reduced hepatocyte inflammation by downregulating inflammatory cytokines in both liver tissue and serum. Furthermore, YGJMF was found to disrupt NF-κB activation, consequently inhibiting the assembly of the NLRP3 inflammasome in both the in vitro and in vivo models. The preliminary findings of this study suggest that YGJMF may alleviate hepatic steatosis and inhibit the NF-κB/NLRP3 signaling pathway, thereby exerting anti-inflammatory effects in NASH.

Ethanol extract of Cyathulae Radix inhibits osteoclast differentiation and bone loss.

Cyathulae Radix, a traditional Chinese medicine and a common vegetable, boasts a history spanning millennia. It enhances bone density, boosts metabolism, and effectively alleviates osteoporosis-induced pain. Despite its historical use, the molecular mechanisms behind Cyathulae Radix's impact on osteoporosis remain unexplored. In this study, we investigated the effects and mechanisms of Cyathulae Radix ethanol extract (CEE) in inhibiting osteoporosis and osteoclastogenesis. Eight-week-old female mice underwent ovariectomy and were treated with CEE for eight weeks. Micro-computed tomography (micro-CT) assessed histomorphometric parameters, bone tissue staining observed distal femur histomorphology, and three-point bending tests evaluated tibia mechanical properties. Enzyme-linked immunosorbent assay (ELISA) measured serum estradiol (E2), receptor activator for nuclear factor B ligand (RANKL), and osteoprotegerin (OPG) levels. Osteoclastogenesis-related markers were analyzed via Western blotting (WB) and quantitative real-time polymerase chain reaction (qRT-PCR). Additionally, CEE effects on RANKL-induced osteoclast formation and bone resorption were investigated in vitro using tartrate-resistant acid phosphatase (TRAP) staining, qRT-PCR, and WB assay. Compared with the ovariectomy (OVX) group, CEE treatment enhanced trabecular bone density, maximal load-bearing capacity, and various histomorphometric parameters. Serum E2 and OPG levels significantly increased, while Receptor activator of nuclear factor-κB (RANK) decreased in the CEE group. CEE downregulated matrix metallopeptidase 9 (MMP-9), Cathepsin K (CTSK), and TRAP gene and protein expression. In bone marrow macrophages (BMMs), CEE reduced mature osteoclasts, bone resorption pit areas, and MMP-9, CTSK, and TRAP expression during osteoclast differentiation. Compared with DMSO treatment, CEE markedly inhibited RANK, TNF receptor associated factor 6 (TRAF6), Proto-oncogene c-Fos (c-Fos), Nuclear factor of activated T-cells cytoplasmic 1 (NFATc1) expressions, and Extracellular regulated protein kinases (ERK), c-Jun N-terminal kinase (JNK), NF-kappa B-p65 (p65) phosphorylation in osteoclasts. In conclusion, CEE significantly inhibits OVX-induced osteoporosis and RANKL-induced osteoclastogenesis, potentially through modulating the Estrogen Receptor (ER)/RANK/NFATc1 signaling pathway.

Yemazhui () ameliorates lipopolysaccharide-induced acute lung injury modulation of the toll-like receptor 4/nuclear factor kappa-B/nod-like receptor family pyrin domain-containing 3 protein signaling pathway and intestinal flora in rats.

OBJECTIVE: To investigate the impact of Yemazhui (Herba Eupatorii Lindleyani, HEL) against lipopolysaccharide (LPS)-induced acute lung injury (ALI) and explore its underlying mechanism in vivo. METHODS: The chemical constituents of HEL were analyzed by ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry method. Then, HEL was found to suppress LPS-induced ALI in vivo. Six-week-old male Sprague-Dawley rats were randomly divided into 6 groups: control, LPS, Dexamethasone (Dex), HEL low dose 6 g/kg (HEL-L), HEL medium dose 18 g/kg (HEL-M) and HEL high dose 54 g/kg (HEL-H) groups. The model rats were intratracheally injected with 3 mg/kg LPS to establish an ALI model. Leukocyte counts, lung wet/dry weight ratio, as well as myeloperoxidase (MPO) activity were determined followed by the detection with hematoxylin and eosin staining, enzyme linked immunosorbent assay, quantitative real time polymerase chain reaction, western blotting, immunohistochemistry, and immunofluorescence. Besides, to explore the effect of HEL on ALI-mediated intestinal flora, we performed 16s rRNA sequencing analysis of intestinal contents. RESULTS: HEL attenuated LPS-induced inflammation in lung tissue and intestinal flora disturbance. Mechanism study indicated that HEL suppressed the lung coefficient and wet/dry weight ratio of LPS-induced ALI in rats, inhibited leukocytes exudation and MPO activity, and improved the pathological injury of lung tissue. In addition, HEL reduced the expression of tumor necrosis factor-alpha, interleukin-1beta (IL-1ß) and interleukin-6 (IL-6) in bronchoalveolar lavage fluid and serum, and inhibited nuclear displacement of nuclear factor kappa-B p65 (NF-κBp65). And 18 g/kg HEL also reduced the expression levels of toll-like receptor 4 (TLR4), myeloid differentiation factor 88, NF-κBp65, phosphorylated inhibitor kappa B alpha (phospho-IκBα), nod-like receptor family pyrin domain-containing 3 protein (NLRP3), IL-1ß, and interleukin-18 (IL-18) in lung tissue, and regulated intestinal flora disturbance. CONCLUSIONS: In summary, our findings revealed that HEL has a protective effect on LPS-induced ALI in rats, and its mechanism may be related to inhibiting TLR4/ NF-κB/NLRP3 signaling pathway and improving intestinal flora disturbance.

Protective effect of modified Huangqi Chifeng decoction on immunoglobulin A nephropathy through toll-like receptor 4/myeloid differentiation factor 88/nuclear factor-kappa B signaling pathway.

OBJECTIVE: To examine the nephroprotective mechanism of modified Huangqi Chifeng decoction (, MHCD) in immunoglobulin A nephropathy (IgAN) rats. METHODS: To establish the IgAN rat model, the bovine serum albumin, lipopolysaccharide, and carbon tetrachloride 4 method was employed. The rats were then randomly assigned to the control, model, telmisartan, and high-, medium-, and low-dose MHCD groups, and were administered the respective treatments via intragastric administration for 8 weeks. The levels of 24-h urinary protein, serum creatinine (CRE), and blood urea nitrogen (BUN) were measured in each group. Pathological alterations were detected. IgA deposition was visualized through the use of immunofluorescence staining. The ultrastructure of the kidney was observed using a transmission electron microscope. The expression levels of interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1), and transforming growth factor-ß1 (TGF-ß1) were examined by immunohistochemistry and quantitative polymerase chain reaction. Levels of toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88), and nuclear factor-kappa B (NF-κB) P65, were examined by immunohistochemistry, Western blotting, and quantitative polymerase chain reaction. RESULTS: The 24-h urine protein level in each group increased significantly at week 6, and worsen from then on. But this process can be reversed by treatments of telmisartan, and high-, medium-, and low-dose of MHCD, and these treatments did not affect renal function. Telmisartan, and high-, and medium-dose of MHCD reduced IgA deposition. Renal histopathology demonstrated the protective effect of high-, medium-, and low-dose of MHCD against kidney injury. The expression levels of MCP-1, IL-6, and TGF-ß1 in kidney tissues were downregulated by low, medium and high doses of MHCD treatment. Additionally, treatment of low, medium and high doses of MHCD decreased the protein and mRNA levels of TLR4, MyD88, and NF-κB. CONCLUSIONS: MHCD exerted nephroprotective effects on IgAN rats, and MHCD regulated the expressions of key targets in TLR4/MyD88/NF-κB signaling pathway, thereby alleviating renal inflammation by inhibiting MCP-1, IL-6 expressions, and ameliorating renal fibrosis by inhibiting TGF-ß1 expression.

Green tea polyphenols inhibit TBBPA-induced lung injury via enhancing antioxidant capacity and modulating the NF-κB pathway in mice.

Tetrabromobisphenol A (TBBPA) is a global pollutant. When TBBPA is absorbed by the body through various routes, it can have a wide range of harmful effects on the body. Green tea polyphenols (GTPs) can act as antioxidants, resisting the toxic effects of TBBPA on animals. The effects and mechanisms of GTP and TBBPA on oxidative stress, inflammation and apoptosis in the mouse lung are unknown. Therefore, we established in vivo and in vitro models of TBBPA exposure and GTP antagonism using C57 mice and A549 cells and examined the expression of factors related to oxidative stress, autophagy, inflammation and apoptosis. The results of the study showed that the increase in reactive oxygen species (ROS) levels after TBBPA exposure decreased the expression of autophagy-related factors Beclin1, LC3-II, ATG3, ATG5, ATG7 and ATG12 and increased the expression of p62; oxidative stress inhibits autophagy levels. The increased expression of the pro-inflammatory factors IL-1ß, IL-6 and TNF-α decreased the expression of the anti-inflammatory factor IL-10 and activation of the NF-κB p65/TNF-α pathway. The increased expression of Bax, caspase-3, caspase-7 and caspase-9 and the decreased expression of Bcl-2 activate apoptosis-related pathways. The addition of GTP attenuated oxidative stress levels, restored autophagy inhibition and reduced the inflammation and apoptosis levels. Our results suggest that GTP can attenuate the toxic effects of TBBPA by modulating ROS, reducing oxidative stress levels, increasing autophagy and attenuating inflammation and apoptosis in mouse lung and A549 cells. These results provide fundamental information for exploring the antioxidant mechanism of GTP and further for studying the toxic effects of TBBPA.

Corynoline Suppresses Osteoclastogenesis and Attenuates ROS Activities by Regulating NF-κB/MAPKs and Nrf2 Signaling Pathways.

Declining estrogen production in postmenopausal females causes osteoporosis in which the resorption of bone exceeds the increase in bone formation. Although clinical drugs are currently available for the treatment of osteoporosis, sustained medication use is accompanied by serious side effects. Corydalis bungeana Herba, a famous traditional Chinese herb listed in the Chinese Pharmacopoeia Commission, constitutes various traditional Chinese Medicine prescriptions, which date back to thousands of years. One of the primary active components of C. bungeana Turcz. is Corynoline (Cor), a plant isoquinoline alkaloid derived from the Corydalis species, which possesses bone metabolism disease therapeutic potential. The study aimed at exploring the effects as well as mechanisms of Cor on osteoclast formation and bone resorption. TRAcP staining, F-actin belt formation, and pit formation were employed for assessing the osteoclast function. Western blot, qPCR, network pharmacology, and docking analyses were used for analyzing the expression of osteoclast-associated genes and related signaling pathways. The study focused on investigating how Cor affected OVX-induced trabecular bone loss by using a mouse model. Cor could weaken osteoclast formation and function by affecting the biological receptor activators of NF-κB and its ligand at various concentrations. Mechanistically, Cor inhibited the NF-κB activation, and the MAPKs pathway stimulated by RANKL. Besides, Cor enhanced the protein stability of the Nrf2, which effectively abolished the RANKL-stimulated ROS generation. According to an OVX mouse model, Cor functions in restoring bone mass, improving microarchitecture, and reducing the ROS levels in the distal femurs, which corroborated with its in vitro antiosteoclastogenic effect. The present study indicates that Cor may restrain osteoclast formation and bone loss by modulating NF-κB/MAPKs and Nrf2 signaling pathways. Cor was shown to be a potential drug candidate that can be utilized for the treatment of osteoporosis.

Ameliorative Effect of Macadamia Nut Protein Peptides on Acetaminophen-Induced Acute Liver Injury in Mice.

This study aims to examine the ameliorative effect of macadamia nut protein peptides (MPP) on acetaminophen (APAP)-induced liver injury (AILI) in mice, and develop a new strategy for identifying hepatoprotective functional foods. The molecular weight distribution and amino acid composition of MPP were first studied. Forty mice were then randomized into four groups: control group (CON), APAP model group, APAP+MPP low-dose group (APAP+L-MPP), and APAP+MPP high-dose group (APAP+H-MPP). The APAP+L-MPP (320 mg/kg per day) and APAP+H-MPP (640 mg/kg per day) groups received continuous MPP gavage for 2 weeks. A 12 h of APAP (200 mg/kg) gavage resulted in liver damage. Pathological alterations, antioxidant index levels, expression of toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB), and associated inflammatory factors were determined for each treatment group. The results revealed that the total amino acid content of MPP was 39.58 g/100 g, with Glu, Arg, Asp, Leu, Tyr, and Gly being the major amino acids. The molecular weight range of 0-1000 Da accounted for 73.54%, and 0-500 Da accounted for 62.84% of MPP. MPP ameliorated the pathological morphology and reduced the serum levels of alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase of AILI in mice. MPP significantly increased the activities of superoxide dismutase and glutathione peroxidase in the liver compared with the APAP group. MPP inhibited the expression of TLR4, NF-κB, interleukin-1ß (IL-1ß), and tumor necrosis factor-α (TNF-α) genes in AILI mice. MPP also inhibited the expression levels of inflammatory factors (TNF-α and IL-6). Our study concludes that MPP alleviates AILI in mice by enhancing antioxidant capacity and inhibiting TLR4/NF-κB pathway-related gene activation.

Zinc glycinate alleviates LPS-induced inflammation and intestinal barrier disruption in chicken embryos by regulating zinc homeostasis and TLR4/NF-κB pathway.

The effect of an immune challenge induced by a lipopolysaccharide (LPS) exposure on systemic zinc homeostasis and the modulation of zinc glycinate (Zn-Gly) was investigated using a chicken embryo model. 160 Arbor Acres broiler fertilized eggs were randomly divided into 4 groups: CON (control group, injected with saline), LPS (LPS group, injected with 32 µg of LPS saline solution), Zn-Gly (zinc glycinate group, injected with 80 µg of zinc glycinate saline solution) and Zn-Gly+LPS (zinc glycinate and LPS group, injected with the same content of zinc glycinate and LPS saline solution). Each treatment consisted of eight replicates of five eggs each. An in ovo feeding procedure was performed at 17.5 embryonic day and samples were collected after 12 hours. The results showed that Zn-Gly attenuated the effects of LPS challenge-induced upregulation of pro-inflammatory factor interleukin 1ß (IL-1ß) level (P =0.003). The LPS challenge mediated zinc transporter proteins and metallothionein (MT) to regulate systemic zinc homeostasis, with increased expression of the jejunum zinc export gene zinc transporter protein 1 (ZnT-1) and elevated expression of the import genes divalent metal transporter 1 (DMT1), Zrt- and Irt-like protein 3 (Zip3), Zip8 and Zip14 (P < 0.05). A similar trend could be observed for the zinc transporter genes in the liver, which for ZnT-1 mitigated by Zn-Gly supplementation (P =0.01). Liver MT gene expression was downregulated in response to the LPS challenge (P =0.004). These alterations caused by LPS resulted in decreased serum and liver zinc levels and increased small intestinal, muscle and tibial zinc levels. Zn-Gly reversed the elevated expression of the liver zinc finger protein A20 induced by the LPS challenge (P =0.025), while Zn-Gly reduced the gene expression of the pro-inflammatory factors IL-1ß and IL-6, decreased toll-like receptor 4 (TLR4) and nuclear factor kappa-B p65 (NF-κB p65) (P < 0.05). Zn-Gly also alleviated the LPS-induced downregulation of the intestinal barrier gene Claudin-1. Thus, LPS exposure prompted the mobilization of zinc transporter proteins and MT to perform the remodeling of systemic zinc homeostasis, Zn-Gly participated in the regulation of zinc homeostasis and inhibited the production of pro-inflammatory factors through the TLR4/NF-κB pathway, attenuating the inflammatory response and intestinal barrier damage caused by an immune challenge.

Warm Acupuncture Reduces Pain and Inflammation in Rats with Lumbar Disc Herniation Induced by Autologous Nucleus Pulposus Transplantation via Regulating p38MAPK/NF-κB Pathway.

Background: : Warm acupuncture (WA) has analgesic and anti-inflammatory effects. However, the underlying mechanism of these effects remain unclear. Objectives: : To explore the analgesic and anti-inflammatory effects of WA and the potential underlying mechanism in male Sprague-Dawley rats with non-compressive lumbar disk herniation (LDH) caused by autologous nucleus pulposus (NP) transplantation. Methods: : We used low-frequency (2 Hz) electrical stimulation and WA (40℃) to treat GB30 and BL54 acupoints in rats for 30 mins per day. We monitored the paw withdrawal threshold of rats during the experiment and measured serum cytokine levels using commercial kits. Dorsal root ganglion (DRG) tissue pathology was analyzed via H&E staining. We used qRT-PCR to measure the mRNA expression levels of IL-1ß, IL-6, and TNF-α genes in DRG. Western blot was used to analyze the expression levels of IL-1ß, IL-6, TNFα, P-p38MAPK, p38MAPK, P-IκBα, IκB α, and NF-κB p65 proteins. Results: : WA treatment significantly increased the pain threshold of rats, reduced serum IL-6, PEG2, NO, SP, NP-Y, and MMP-3 levels, and effected histopathological improvements in the DRG in rats. Moreover, WA treatment significantly downregulated the expression levels of inflammation-associated genes (Il-1ß, Il-6, and Tnf-α) and proteins (IL-1ß, IL-6, TNF-α, P-p38MAPK, P-IκBα, and NF-κB p65) in the DRG of non-compressive LDH rats. Conclusion: : WA can alleviate pain and inhibit inflammatory response in rats with non-compressive LDH caused by autologous NP transplantation, and these effects are likely associated with the inhibition of the p38MAPK/NF-κB pathway.

[Effect and mechanism of Yueju Pills on breast cancer induced by DMBA in rats].

This paper aims to explore the inhibitory effect of Yueju Pills on breast cancer and decipher the underlying mechanism. A total of 92 SPF-grade SD female rats were involved in this study, and 14 of them were randomly selected into control group. The remaining 78 rats were administrated with 7,12-dimethylbenzanthracene(DMBA) by gavage to establish the breast cancer model. The modeled rats were randomized into model, tamoxifen(1.9 mg·kg~(-1)·d~(-1)), and low-and high-dose(17, 34 g·kg~(-1)·d~(-1)) Yueju Pills groups. The mental state, food intake, and activities of the rats were observed daily, and the body weight was measured on alternate days. After 12 weeks of administration, the rats were sacrificed and the tumor weight was measured. The serum estrogen and progeste-rone levels were determined by enzyme-linked immunosorbent assay. The histopathological changes of the breast and tumor were observed by hematoxylin-eosin staining. Western blot was employed to measure the protein levels of glucose transporter 1(GLUT1), lactate dehydrogenase A(LDHA), phosphofructokinase muscle(PFKM), pyruvate kinase isozyme type M2(PKM2), hexokinase 2(HK2), nuclear factor-kappaB(NF-κB), and phosphorylated NF-κB. The intestinal microbiome was examined by 16S rRNA high-throughput sequencing. The results showed that compared with the model group, high and low-dose Yueju Pills showed the tumor inhibition rate of 15.8% and 64.5%, respectively, and the low dose group had stronger inhibitory effect. Compared with the control group, the model group presented elevated the levels of estrogen and progesterone in serum. The administration of Yueju Pills lowered such ele-vation, and the low-dose group showed stronger lowering effect(P<0.05). Compared with the model group, Yueju Pills reduced the glands with increased breast tissue, the degree of breast duct expansion, the number and area of acinar cavity, the secretions, and the layers of mammary epithelial cells. Furthermore, Yueju Pills down-regulated the expression of GLUT1, LDHA, PFKM, PKM2, HK2, and NF-κB(P<0.05) and altered the diversity, composition, structure, and abundance of intestinal flora. The results showed that Yueju Pills could inhibit breast cancer by regulating the secretion of estrogen and progesterone, glycolysis, inflammatory cytokines, and intestinal flora.

[Zhuyu Pills promote polarization of macrophages toward M2 phenotype to prevent atherosclerosis via PPARγ/NF-κB signaling pathway].

This article aims to investigate the effect of Zhuyu Pills on atherosclerosis and decipher the underlying mechanism. The mouse model of atherosclerosis was induced by a high-fat diet, and the total modeling period was 12 weeks. A total of 47 ApoE~(-/-) mice successfully modeled were randomized into 5 groups, including 10 in the model group, 9 in each of low-, medium-, and high-dose(130.54, 261.08 and 522.16 mg·kg~(-1)·d~(-1), respectively) Zhuyu Pills groups, and 10 in the atorvastatin calcium(10.40 mg·kg~(-1)·d~(-1)) group. In addition, 10 C57BL/6J mice were included as the normal group. The mice in the normal group and model group were administrated with an equal volume of sterile distilled water, and those in other groups with corresponding agents by gavage once a day for 12 weeks. At the end of drug intervention, the levels of total cholesterol(TC), triglyceride(TG), high-density lipoprotein cholesterol(HDL-C), and low-density lipoprotein cholesterol(LDL-C) were measured by the biochemical method. Hematoxylin-eosin(HE) staining was employed to observe the plaque distribution in the aortic region. The serum levels of pro-inflammatory cytokines tumor necrosis factor-α(TNF-α) and interleukin(IL)-6 in M1 macrophages and anti-inflammatory cytokines IL-13 and IL-4 in M2 macrophages were determined by enzyme-linked immunosorbent assay(ELISA). The expression levels of inducible nitric oxide synthase(iNOS) and arginase-1(Arg-1) were examined by immunofluorescence. Real-time fluorescence quantitative polymerase chain reaction(real-time PCR) was employed to measure the mRNA levels of peroxisome proliferator-activated receptor γ(PPARγ), nuclear factor-κB(NF-κB), Arg-1, and iNOS in the aorta. Western blot was employed to determine the protein levels of PPARγ and NF-κB in the aorta. The results showed that compared with the normal group, the modeling elevated the TC, TG, and LDL-C levels, lowered the HDL-C level, caused large area thickening of the aortic intima, elevated the TNF-α and IL-6 levels, lowered the IL-4 and IL-13 levels, down-regulated the mRNA and protein levels of PPARγ and Arg-1, and up-regulated the mRNA and protein levels of iNOS and NF-κB in the aorta(P<0.01). Compared with the model group, low-, medium-, and high-dose Zhuyu Pills and atorvastatin calcium lowered the TC, TG, and LDL-C levels, elevated the HDL-C level, reduced the plaque area in a concentration-dependent manner, lowered the TNF-α and IL-6 levels, elevated the IL-4 and IL-13 levels, up-regulated the mRNA and protein levels of PPARγ and Arg-1, and down-regulated the mRNA and protein levels of NF-κB and iNOS in the aorta(P<0.05 or P<0.01). In conclusion, Zhuyu Pills may play an anti-atherosclerosis role by regulating PPARγ/NF-κB signaling pathway, inhibiting the polarization of macrophages toward the M1 phenotype, promoting the polarization of macrophages toward the M2 phenotype, and improving the inflammatory microenvironment of macrophages.

Ginsenoside Rb2 exhibits therapeutic value for male osteoporosis in orchiectomy mice by suppressing osteoclastogenesis and modulating NF-κB/MAPK signaling pathways.

Osteoporosis (OP) is a systemic disorder characterized by decreased bone mass as well as deteriorated microarchitecture. Although OP in men is common, it has received much less attention than that in women. Ginseng, a famous traditional herb in Asia, is used to strengthen and repair bones by invigorating vital bioenergy and maintaining body homeostasis in dietary intake and clinical applications. However, there is currently no study investigating the impact of ginseng and its active compounds on male osteoporosis. In this study, RNA sequencing and bioinformatic analysis were conducted to reveal the influence of Ginsenoside-Rb2 on RAW264.7 cells and its underlying signaling pathways. The potential anti-osteoporosis effects of Rb2 as well as its molecular mechanisms were elucidated in RAW264.7 cells and BMMs by TRAP staining, F-actin belt staining, qRT-PCR and WB. Moreover, orchiectomy (ORX) was utilized to demonstrate the influence of Rb2 on bone mass loss in vivo by micro-CT scanning, and H&E, TRAP, and IHC staining. The results suggested that Rb2 suppressed osteoclastogenesis and mitigated bone loss in orchiectomy mice through NF-κB/MAPK signaling pathways. These findings indicate that ginseng as well as its active component Rb2 have potential therapeutic value in the management of osteoporosis in men.

Tea polyphenols alleviate TBBPA-induced inflammation, ferroptosis and apoptosis via TLR4/NF-κB pathway in carp gills.

The extensive application of Tetrabromobisphenol A (TBBPA) leads to the pollution of part of the water environment and brings great safety risks to aquatic animals. As a natural extract, tea polyphenols (TPs) have antioxidant and anti-inflammatory effects. Gills are one of the immune organs of fish and constitute the first line of defense of the immune system. However, it was unclear whether TPs could mitigate TBBPA-induced gills injury. Therefore, an animal model was established to investigate the effect of TPs on TBBPA-induced gills. The results indicated that TBBPA changed the coefficient and tissue morphology of carp gills. In addition, TBBPA induced oxidative stress and inflammation, leading to ferroptosis and apoptosis in carp gills. Dietary addition of TPs significantly improved the antioxidant capacity of carp, effectively inhibited the overexpression of TLR4/NF-κB and its mediated inflammatory response. Moreover, TPs restored iron metabolism, reduced the expression of pro-apoptotic factors thereby alleviating ferroptosis and apoptosis in carp gills. This study enriched the protective effect of TPs and provided a new way to improve the innate immunity of carp.