This study aims to investigate the protective effect and potential mechanism of Jingfang Granules(JF) on the mouse model of chronic fatigue syndrome(CFS). Mice were randomized into normal, model, and low-, medium-, and high-dose(0.9, 1.8, and 3.6 g·kg~(-1)·d~(-1), respectively) JF groups according to the body weight. In addition to the normal group, other groups of mice received exhaustive swimming training and tail suspension training every day for the modeling of CFS. The mice in each administration group were administrated with JF at the corresponding dose by gavage, and those in the other groups were administrated with an equal amount of purified water. The exhaustive swimming and tail suspension tests were conducted in each group. The UV-glutamate dehydrogenase method was used to determine the serum level of urea nitrogen(UREA), and the lactate dehydrogenase(LDH) assay kit was used to determine the LDH level. Enzyme-linked immunosorbent assay was employed to measure the levels of interleukin-6(IL-6) and tumor necrosis factor-α(TNF-α) in the serum, muscle tissue, and brain tissue of mice in each group. Western blot was employed to determine the expression levels of Toll-like receptor 4(TLR4), myeloid differentiation factor 88(MyD88), nuclear factor-kappa B(NF-κB) and their phosphorylated proteins in the muscle tissue of mice. The 16S rDNA sequencing and ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) were adopted to detect the changes of intestinal flora and intestinal metabolites in mice. Compared with the model group, JF significantly prolonged the swimming exhaustion time and shortened the tail suspension time of the model mice, lowered the levels of LDH and UREA in the serum as well as the levels of IL-6 and TNF-α in the serum, muscle tissue, and brain tissue of CFS mice. In addition, JF down-regulated the expression of TLR4, MyD88, and p-NF-κB/NF-κB in the muscle tissue of CFS mice compared with the model group. The results of 16S rDNA sequencing demonstrated that JF ameliorated the intestinal flora disorder of CFS mice. The results of UPLC-MS/MS revealed that JF significantly affected the histidine metabolism pathway in the intestinal tract of CFS mice. Spearman analysis displayed that histamine, a metabolite involved in histidine metabolism, was negatively correlated with the abundance of Clostridia＿UCG-014, Dubosiella, and RF39 and positively correlated with the abundance of Coriobacteriaceae＿UCG-002. The metabolite imidazole-4-acetaldehyde was negatively correlated with the abundance of Clostridia＿UCG-014, Dubosiella, and RF39 and positively correlated with the abundance of Coriobacteriaceae＿UCG-002. In conclusion, JF can increase the swimming exhaustion time, reduce the immobility time of tail suspension, lower serum LDH and UREA levels, and alleviate inflammation response. It may exert the therapeutic effect by improving intestinal flora homeostasis and inhibiting histidine metabolism by down-regulating the expression of proteins in the TLR4/MyD88/NF-κB signaling pathway, thereby relieving the symptoms of CFS in mice.
Animals ; Mice ; Gastrointestinal Microbiome/drug effects* ; Drugs, Chinese Herbal/administration & dosage* ; Male ; Metabolomics ; Fatigue Syndrome, Chronic/genetics* ; NF-kappa B/genetics* ; Humans ; Interleukin-6/genetics* ; Myeloid Differentiation Factor 88/genetics* ; Toll-Like Receptor 4/genetics* ; Tumor Necrosis Factor-alpha/genetics* ; Disease Models, Animal

The study aims to observe the effects and explore the mechanisms of Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination in the treatment of the inflammatory response of mice with atherosclerosis(AS) via the Toll-like receptor 4(TLR4)/myeloid differentiation primary response protein 88(MyD88)/nuclear factor-κB(NF-κB) signaling pathway. Male ApoE~(-/-) mice were randomly assigned into a model group, a Buyang Huanwu Decoction group, an Astragali Radix-Angelicae Sinensis Radix combination group, and an atorvastatin group, and male C57BL/6J mice of the same weeks old were used as the control group. Other groups except the control group were given high-fat diets for 12 weeks to establish the AS model, and drugs were administrated by gavage. Aortic intimal hyperplasia thickness, blood lipid level, plasma inflammatory cytokine levels, M1/M2 macrophage markers, and expression levels of proteins in TLR4/MyD88/NF-κB pathway in the vessel wall were measured to evaluate the effects of drugs on AS lesions and inflammatory responses. The results showed that the AS model was successfully established with the ApoE~(-/-) mice fed with high-fat diets. Compared with the control group, the model group showed elevated plasma total cholesterol(TC), triglyceride(TG), and low-density lipoprotein cholesterol(LDL-c) levels(P<0.05), thickened intima(P<0.01), and increased plasma tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) levels(P<0.01). Moreover, the model group showed increased expression of vascular cell adhesion molecule-1(VCAM-1) and inducible nitric oxide synthase(iNOS)(P<0.01), inhibited expression of endothelial nitric oxide synthase(eNOS) and cluster of differentiation 206(CD206)(P<0.01), and up-regulated mRNA and protein levels of TLR4, MyD88, NF-κB inhibitor alpha(IκBα), and NF-κB in the vessel wall(P<0.05). Compared with the model group, Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination lowered the plasma TC and LDL-c levels(P<0.01), alleviated the intimal hyperplasia(P<0.01), and reduced the plasma TNF-α and IL-6 levels(P<0.05). Moreover, the two interventions promoted the expression of eNOS and CD206(P<0.05), inhibited the expression of VCAM-1 and iNOS(P<0.01), and down-regulated the mRNA and protein levels of TLR4, MyD88, IκBα, and NF-κB(P<0.05) in the vessel wall. This study indicated that Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination could delay the progression of AS, inhibit the polarization of vascular wall macrophages toward M1 type, and attenuate vascular inflammatory response by inhibiting the activation of TLR4/MyD88/NF-κB signaling pathway in the vascular wall. Astragali Radix and Angelicae Sinensis Radix were the main pharmacological substances in Buyang Huanwu Decoction for alleviating the AS vascular inflammatory response.
Mice ; Male ; Animals ; NF-kappa B/metabolism* ; Toll-Like Receptor 4/metabolism* ; NF-KappaB Inhibitor alpha/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Interleukin-6/metabolism* ; Myeloid Differentiation Factor 88/metabolism* ; Vascular Cell Adhesion Molecule-1/metabolism* ; Cholesterol, LDL ; Hyperplasia ; Mice, Inbred C57BL ; Atherosclerosis/genetics* ; Apolipoproteins E/therapeutic use* ; RNA, Messenger

Objective To investigate the role of microRNA-18a (miR-18a) in the pathogenesis of allergic rhinitis in mice. Methods Twenty-two BALB/c mice were randomly divided into a blank group, a model group and a miR-18a group. Mice in the model group and the miR-18a group were injected intraperitoneally with obumin (OVA) suspension to prepare allergic rhinitis models, and mice in the miR-18a group were simultaneously given lentiviral vector plasmid for overexpression of miR-18a. Allergy symptoms were evaluated by the behavioral score and HE staining. The plasma levels of interleukin-1β (IL-1β), IL-6 and tumor necrosis factor α (TNF-α) were measured by ELISA. The distribution of CD45⁺ cells in nasal mucosa was measured by immunofluorescence histochemistry, and CD45⁺ cells in nasal lavage fluid were measured by flow cytometry. The mRNA expression levels of IL-1β, IL-6 and TNF-α in nasal mucosa tissues were measured by fluorescence quantitative PCR, and the protein expressions of Toll like receptor 4 (TLR4), nuclear factor κB p65 (NF-κB p65), inhibitor of NF-κB α (IκBα) and phosphorylated IκBα (p-IκBα) in nasal mucosa were measured by Western blot analysis. Results Compared with the blank group, the plasma levels of IL-1β, IL-6, and TNF-α in the model group increased significantly. The number of CD45⁺ cells in both nasal mucosa tissue and nasal irrigation fluid increased, and the mRNA levels of IL-1β, IL-6 and TNF-α and the protein expression levels of TLR4, NF-κB p65 and p-IκBα in nasal mucosa increased. Compared with the model group, the plasma levels of IL-1β, IL-6 and TNF-α in the miR-18a group decreased significantly. The number of CD45⁺ cells in both nasal mucosa tissue and nasal lavage fluid decreased, and the mRNA levels of IL-1β, IL-6 and TNF-α and the exprotein expression levels of TLR4, NF-κB p65 and p-IκBα in nasal mucosa decreased. Conclusion miR-18a can inhibit the occurrence and development of allergic rhinitis, and its molecular mechanism is related to the inhibition of TLR4/NF-κB pathway activation.
Animals ; Mice ; Disease Models, Animal ; Inflammation ; Interleukin-6/genetics* ; MicroRNAs/genetics* ; NF-kappa B/metabolism* ; NF-KappaB Inhibitor alpha ; Rhinitis, Allergic ; RNA, Messenger ; Toll-Like Receptor 4/metabolism* ; Tumor Necrosis Factor-alpha/genetics*

This study aims to investigate the effect of modified Danggui Shaoyao Powder on the suppressor of cytokine signaling 3(SOCS3)/Toll-like receptor 4(TLR4) signaling pathway in gastric tissue of rats with chronic atrophic gastritis(CAG).Sixty SPF-grade SD rats were randomly assigned into the normal group, model group, Moluo Pills group, and high-, medium-, and low-dose groups of modified Danggui Shaoyao Powder.The rats in other groups except the normal group were treated with N-methyl-N'-nitro-N-nitrosoguanidine(MNNG) to establish the CAG model.After 12 weeks of modeling, the rats in each group were administrated with corresponding drugs by gavage for 8 weeks.After the last administration, the histopathological changes of rat gastric mucosa were observed via hematoxylin-eosin(HE) staining.The serum levels of IL-6, TNF-α, and CRP were determined by enzyme-linked immunosorbent assay(ELISA).The mRNA levels of SOCS3 and TLR4 were determined by real-time PCR.The protein levels of SOCS3, TLR4, JAK2, p-JAK2, STAT3, and p-STAT3 in rat gastric tissue were measured by Western blot.Immunohistochemical method was employed to determine the protein levels of NF-κB, MyD88, NLRP3, Bcl-2, Bax, and Bad in rat gastric tissue.The results showed that modified Danggui Shaoyao Powder alleviated gastric mucosal atrophy of rats, significantly lowered the levels of IL-6, TNF-α, and CRP in rat serum, up-regulated the mRNA level of SOCS3, and down-regulated the mRNA level of TLR4 in rat gastric tissue.Furthermore, modified Danggui Shaoyao Powder up-regulated the protein level of SOCS3, down-regulated the protein levels of TLR4, p-JAK2, p-STAT3, NF-κB, MyD88, NLRP3, Bax, and Bad, and promoted the expression of Bcl-2 protein.Therefore, modified Danggui Shaoyao Powder may mitigate the gastric mucosal atrophy of rats by regulating the SOCS3/TLR4 signaling pathway.
Animals ; Atrophy ; Gastritis, Atrophic/genetics* ; Interleukin-6/metabolism* ; Myeloid Differentiation Factor 88/metabolism* ; NF-kappa B/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Powders ; RNA, Messenger ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Suppressor of Cytokine Signaling 3 Protein/metabolism* ; Toll-Like Receptor 4/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; bcl-2-Associated X Protein/metabolism*

Based on network pharmacology and in vivo experiment, this study explored the therapeutic effect of Tetrastigma hemsle-yanum(SYQ) on sepsis and the underlying mechanism. The common targets of SYQ and sepsis were screened out by network pharmacology, and the "SYQ-component-target-sepsis" network was constructed. The protein-protein interaction(PPI) network was established by STRING. Gene Ontology(GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment were performed based on DAVID to predict the anti-sepsis mechanism of SYQ. The prediction results of network pharmacology were verified by animal experiment. The network pharmacology results showed that the key anti-sepsis targets of SYQ were tumor necrosis factor(TNF), interleukin(IL)-6, IL-1β, IL-10, and cysteinyl asparate specific proteinase 3(caspase-3), which were mainly involved in Toll-like receptor 4(TLR4)/myeloid differentiation factor 88(MyD88)/nuclear factor kappaB(NF-κB) signaling pathway. The results of animal experiment showed that SYQ can decrease the content of C-reactive protein(CRP), procalcitonin(PCT), lactate dehydrogenase(LDH), IL-6, TNF-α, and IL-1β, increase the content of IL-10, and down-regulate the protein levels of Bcl-2-associa-ted X(Bax)/B-cell lymphoma 2(Bcl2), cleaved caspase-3, TLR4, MyD88, and p-NF-κB p65/NF-κB p65. In summary, SYQ plays an anti-inflammatory role in the treatment of sepsis by acting on the key genes related to inflammation and apoptosis, such as TNF-α, IL-6, IL-lβ, IL-10, Bax, Bcl2, and cleaved caspase-3. The mechanism is the likelihood that it suppresses the TLR4/MyD88/NF-κB signaling pathway, which verifies relative prediction results of network pharmacology.
Animals ; Anti-Inflammatory Agents/therapeutic use* ; C-Reactive Protein ; Caspase 3/metabolism* ; Interleukin-10 ; Interleukin-6/metabolism* ; Lactate Dehydrogenases/metabolism* ; Myeloblastin/metabolism* ; Myeloid Differentiation Factor 88/metabolism* ; NF-kappa B/metabolism* ; Network Pharmacology ; Procalcitonin/therapeutic use* ; Sepsis/genetics* ; Toll-Like Receptor 4/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; bcl-2-Associated X Protein/metabolism*

This paper aims to explore the activity of Codonopsis canescens extract against rheumatoid arthritis(RA) based on the Toll-like receptors(TLRs)/mitogen-activated protein kinases(MAPKs)/nuclear factor kappa B(NF-κB) signaling pathways and its mechanism. The ultra-performance liquid chromatography-quadrupole time-of-flight/mass spectrometry(UPLC-Q-TOF-MS) was used to identify the components of C. canescens extract. Forty-eight male SD rats were randomly divided into six groups, namely the normal group, the model group, the methotrexate(MTX) tablet group, and the low, medium, and high-dose C. canescens extract(ZDS-L, ZDS-M, and ZDS-H) groups, with 8 rats in each group. The model of collagen-induced arthritis in rats was induced by injection of bovine type Ⅱ collagen emulsion. MTX(2.5 mg·kg~(-1)), ZDS-L, ZDS-M, and ZDS-H(0.3 g·kg~(-1), 0.6 g·kg~(-1), and 1.2 g·kg~(-1)) were administrated by gavage. Rats in the normal group and the model group received distilled water. MTX was given once every three days for 28 days, and the rest medicines were given once daily for 28 days. Body weight, degree of foot swelling, arthritis index, immune organ index, synovial histopathological changes, and serum levels of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), and interleukin-6(IL-6) were observed. Protein expressions of TLR2, TLR4, NF-κB p65, p38 MAPK, and p-p38 MAPK in rats were determined by Western blot. Thirty-four main components were identified by UPLC-Q-TOF-MS, including 15 flavonoids, 7 phenylpropanoids, 4 terpenoids, 4 organic acids, 2 esters, and 2 polyalkynes. As compared with the normal group, the body weight of the model group was significantly decreased(P<0.01), and foot swelling(P<0.05, P<0.01), arthritis index(P<0.01), and the immune organ index(P<0.01) were significantly increased. The synovial histopathological injury was obviously observed in the model group. The serum levels of inflammatory factors TNF-α, IL-1β, and IL-6 were significantly increased(P<0.01), and the protein expression levels of TLR2, TLR4, NF-κB p65, p-p38 MAPK/p38 MAPK in the synovial tissue were significantly increased(P<0.01) in the model group. As compared with the model group, the body weights of the ZDS dose groups were increased(P<0.01), and the degree of foot swelling(P<0.01) and the arthritis index were decreased(P<0.05, P<0.01). The immune organ index was decreased(P<0.01) in the ZDS dose groups, and the synovial tissue hyperplasia and inflammatory cell infiltration were alleviated. The serum levels of TNF-α, IL-1β, and IL-6 were significantly decreased(P<0.05, P<0.01), and the protein expression levels of TLR2, TLR4, NF-κB p65, p-p38 MAPK/p38 MAPK were decreased(P<0.05, P<0.01) in the ZDS dose groups. C. canescens extract containing apigenin, tricin, chlorogenic acid, aesculin, ferulic acid, caffeic acid, and oleanolic acid has a good anti-RA effect, and the mechanism may be related to the inhibition of TLRs/MAPKs/NF-κB signaling pathways.
Animals ; Cattle ; Male ; Rats ; Arthritis, Experimental/drug therapy* ; Arthritis, Rheumatoid/drug therapy* ; Body Weight ; Codonopsis/chemistry* ; Interleukin-6/blood* ; NF-kappa B/genetics* ; p38 Mitogen-Activated Protein Kinases/metabolism* ; Plant Extracts/therapeutic use* ; Rats, Sprague-Dawley ; Signal Transduction ; Toll-Like Receptor 2/metabolism* ; Toll-Like Receptor 4/metabolism* ; Tumor Necrosis Factor-alpha/pharmacology*

OBJECTIVES: Because intracerebral hemorrhage (ICH) has high morbidity, disability and mortality, it is significant to find new and effective treatments for ICH. This study aims to explore the effect of butyphthalide (NBP) on neuroinflammation secondary to ICH and microglia polarization. METHODS: A total of 48 healthy male SD rats were randomly divided into 6 groups: a sham 24 h group, a sham 72 h group, an ICH 24 h group, an ICH 72 h group, an ICH+NBP 24 h group, and an ICH+NBP 72 h group (8 rats per group). After operation, the neurological deficiencies were assessed based on improved Garcia scores and corner test. The expressions of Toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB), nuclear factor erythroid 2-related factor 2 (Nrf2), aquaporin-4 (AQP4), zonula occludens-1 (ZO-1), occludin, CD68, CD86, and CD206 were observed by Western blotting. Inflammatory cytokines were detected by ELISA. The immunofluorescence was to detect the polarization of microglia. RESULTS: 1) Compared with the sham groups, the expression of TLR4 (24 h: P<0.05; 72 h: P<0.01), NF-κB (both P<0.01) and Nrf2 (both P<0.01) in the perihematoma of the ICH group was increased, leading to microglia activation (P<0.01). The expressions of IL-6 (24 h: P<0.05; 72 h: P<0.01) and TNF-α (both P<0.01), the pro-inflammatory cytokines were up-regulated, and the expression of anti-inflammatory cytokine IL-4 was down-regulated (both P<0.01). Besides, the expression of AQP4 was enhanced (both P<0.01). The protein level of tightly connected proteins (including ZO-1, occludin) was decreased (all P<0.01). The neurological function of the rats in the ICH group was impaired in the 2 time points (both P<0.01). 2) Compared with the sham group at 24 h and 72 h after the intervention of NBP, the expressions of TLR4 (both P<0.05) and NF-κB (both P<0.01) were significantly declined, and the expression of Nrf2 was further enhanced (both P<0.05) in the perihematoma of the ICH+NBP group. Furthermore, the expression of M1 microglia marker was inhibited (P<0.05), and the polarization of microglia to the M2 phenotype was promoted (P<0.01). 3) In terms of inflammation after ICH, the IL-4 expression in the ICH+NBP group was increased compared with the ICH group (24 h: P<0.05; 72 h: P<0.01); the expression of IL-6 was decreased significantly in the ICH+NBP 72 h group (P<0.01); the level of AQP4 was declined significantly in the ICH+NBP 24 h group (P<0.05), there was a downward trend in the 72-hour intervention group but without significant statistical difference. 4) Compared with the ICH group, the ZO-1 protein levels were increased (24 h: P<0.05; 72 h: P<0.01), and the symptoms of nerve defect were improved eventually (both P<0.05) in the ICH+NBP groups. CONCLUSIONS After ICH, the TLR4/NF-κB pathway is activated. The M1 microglia is up-regulated along with the release of detrimental cytokines, while the anti-inflammatory cytokines are down-regulated. The expression of AQP4 is increased, the tight junction proteins from the blood-brain barrier (BBB) is damaged, and the neurological function of rats is impaired. On the contrary, NBP may regulate microglia polarization to M2 phenotype and play a role in the neuroprotective effect mediated via inhibiting TLR4/NF-κB and enhancing Nrf2 pathways, which relieves the neuroinflammation, inhibits the expression of AQP4, repairs BBB, and improves neurological functional defects.
Animals ; Anti-Inflammatory Agents/therapeutic use* ; Cerebral Hemorrhage ; Cytokines/metabolism* ; Interleukin-4/therapeutic use* ; Interleukin-6/metabolism* ; Male ; Microglia/metabolism* ; NF-E2-Related Factor 2/metabolism* ; NF-kappa B/metabolism* ; Occludin/pharmacology* ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Toll-Like Receptor 4/genetics*

OBJECTIVETo study the protective effect of lipoxin A4 (LXA4) against sepsis induced by lipopolysaccharide (LPS) in rats with obesity and its effect on the expression of Toll-like receptor 4 (TLR4) and TNF receptor-associated factor 6 (TRAF6) in the liver.

METHODSA total of 60 male Sprague-Dawley rats aged three weeks were randomly divided into a normal group and an obesity group, with 30 rats in each group. A rat model of obesity was established by high-fat diet. Each of the two groups was further randomly divided into control group, sepsis group, and LXA4 group, and 8 rats were selected from each group. The rats in the control, sepsis, and LXA4 groups were treated with intraperitoneal injection of normal saline, LPS, and LXA4+LPS respectively. Twelve hours later, blood samples were collected from the heart and liver tissue samples were also collected. ELISA was used to measure the serum levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). Western blot was used to measure the protein expression of TLR4 and TRAF6 in liver tissue. Quantitative real-time PCR was used to measure the mRNA expression of TLR4 and TRAF6.

RESULTSAfter being fed with high-fat diet for 6 weeks, the obesity group had significantly higher average weight and Lee's index than the normal group (P<0.05). Compared with the normal group, the obesity group had significant increases in the serum levels of IL-6 and TNF-α (P<0.05). In the normal group or the obesity group, the sepsis subgroup had significant increases in the serum levels of IL-6 and TNF-α compared with the control subgroup (P<0.05), while the LXA4 subgroup had significant reductions in the two indices compared with the sepsis subgroup (P<0.05). Compared with the normal group, the obesity group had significant increases in the protein and mRNA expression of TLR4 and TRAF6 (P<0.05). In the normal group or the obesity group, the sepsis subgroup had significant increases in the protein and mRNA expression of TLR4 and TRAF6 compared with the control subgroup (P<0.05). Compared with the sepsis subgroup, the LXA4 subgroup had significant reductions in the protein and mRNA expression of TLR4 and TRAF6 (P<0.05).

CONCLUSIONSLXA4 can reduce the serum levels of IL-6 and TNF-α and alleviate inflammatory response. LXA4 can inhibit the expression of TLR4 and TRAF6 in the liver of septic rats, possibly by inhibiting the TLR4 signaling pathway.

Animals ; Humans ; Interleukin-6 ; genetics ; metabolism ; Lipoxins ; administration & dosage ; Liver ; drug effects ; metabolism ; Male ; Obesity ; complications ; drug therapy ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Sepsis ; complications ; drug therapy ; genetics ; metabolism ; Signal Transduction ; drug effects ; TNF Receptor-Associated Factor 6 ; genetics ; metabolism ; Toll-Like Receptor 4 ; genetics ; metabolism ; Tumor Necrosis Factor-alpha ; genetics ; metabolism

BACKGROUND/AIMS: To investigate the expression of Toll-like receptor 4 (TLR4) in the pancreases of rats with acute necrotizing pancreatitis (ANP) and any changes upon treatment with pyrrolidine dithiocarbamate (PDTC), an inhibitor of nuclear factor kappaB (NF-kappaB), as well as to determine the relationship between TLR4 and NF-kappaB in ANP pathogenesis. METHODS: A total of 72 SD rats were randomly divided into three groups, namely, the control (sham-operation), ANP, and ANP with PDTC pretreatment groups. The PDTC-pretreated group was intraperitoneally injected with PDTC at a dose of 100 mg/kg 1 hour before the induction of ANP. The expressions of TLR4 and NF-kappaB in pancreatic tissue were evaluated by immunohistochemistry and Western blot analysis. The mRNA levels of cytokines tumor necrosis factor alpha, interleukin (IL)-1beta, and IL-6 were measured by reverse transcription polymerase chain reaction. RESULTS: The expressions of TLR4, NF-kappaB, and cytokine (NF-kappaB target) genes in the pancreatic tissue increased more significantly in the ANP groups than in the sham-operation group at 3, 6, and 12 hours. Pretreatment with PDTC alleviated the inflammatory activation in the pancreas with ANP, causing a significant decrease in the expressions of TLR4, NF-kappaB, and cytokine genes in the pancreatic tissue. CONCLUSIONS: The expressions of TLR4 and NF-kappaB were increased in the pancreases of rats with ANP. PDTC not only inhibits NF-kappaB but also suppresses the expression of TLR4 and downregulates the expression of the related cytokine genes.
Animals ; Antioxidants/*pharmacology ; Interleukin-1beta/genetics/metabolism ; Interleukin-6/genetics/metabolism ; Male ; NF-kappa B/*drug effects/metabolism ; Pancreas/metabolism/pathology ; Pancreatitis, Acute Necrotizing/chemically induced/*drug therapy ; Pyrrolidines/*pharmacology ; RNA, Messenger/metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Thiocarbamates/*pharmacology ; Toll-Like Receptor 4/*drug effects/metabolism ; Tumor Necrosis Factor-alpha/genetics/metabolism

BACKGROUND/AIMS: To investigate the expression of Toll-like receptor 4 (TLR4) in the pancreases of rats with acute necrotizing pancreatitis (ANP) and any changes upon treatment with pyrrolidine dithiocarbamate (PDTC), an inhibitor of nuclear factor kappaB (NF-kappaB), as well as to determine the relationship between TLR4 and NF-kappaB in ANP pathogenesis. METHODS: A total of 72 SD rats were randomly divided into three groups, namely, the control (sham-operation), ANP, and ANP with PDTC pretreatment groups. The PDTC-pretreated group was intraperitoneally injected with PDTC at a dose of 100 mg/kg 1 hour before the induction of ANP. The expressions of TLR4 and NF-kappaB in pancreatic tissue were evaluated by immunohistochemistry and Western blot analysis. The mRNA levels of cytokines tumor necrosis factor alpha, interleukin (IL)-1beta, and IL-6 were measured by reverse transcription polymerase chain reaction. RESULTS: The expressions of TLR4, NF-kappaB, and cytokine (NF-kappaB target) genes in the pancreatic tissue increased more significantly in the ANP groups than in the sham-operation group at 3, 6, and 12 hours. Pretreatment with PDTC alleviated the inflammatory activation in the pancreas with ANP, causing a significant decrease in the expressions of TLR4, NF-kappaB, and cytokine genes in the pancreatic tissue. CONCLUSIONS: The expressions of TLR4 and NF-kappaB were increased in the pancreases of rats with ANP. PDTC not only inhibits NF-kappaB but also suppresses the expression of TLR4 and downregulates the expression of the related cytokine genes.
Animals ; Antioxidants/*pharmacology ; Interleukin-1beta/genetics/metabolism ; Interleukin-6/genetics/metabolism ; Male ; NF-kappa B/*drug effects/metabolism ; Pancreas/metabolism/pathology ; Pancreatitis, Acute Necrotizing/chemically induced/*drug therapy ; Pyrrolidines/*pharmacology ; RNA, Messenger/metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Thiocarbamates/*pharmacology ; Toll-Like Receptor 4/*drug effects/metabolism ; Tumor Necrosis Factor-alpha/genetics/metabolism