Intervention effect of Tuina pressing and kneading the Huantiao (GB30) acupoint on NF-κB p65 protein at spinal cord dorsal horn in sciatica rats / 中国骨伤

OBJECTIVE@#To observe the analgesic effect of Tuina by pressing and kneading the Huantiao (GB30) acupoint on rats with chronic constriction injury (CCI) and to explore the analgesic mechanism of Tuina on sciatica rats.@*METHODS@#Thirty-two SPF male SD rats weighing 180 to 220 g were randomly divided into fore groups:blank group (without any treatment), sham group (only exposed without sciatic nerve ligating), model group (sciatic nerve ligating) and Tuina group (manual intervention after lsciatic nerve ligating). The CCI model was prepared by ligating the right sciatic nerve of the rats, on the third day of modeling, the rats in the Tuina group were given pressing and kneading the Huantiao (GB30) point for 14 days, and the changes of paw withdrawal threshold(PWT), paw withdrawal latency(PWL) were measured before and on the 1st, 3rd, 7th, 10th, 14th and 17th days after modeling. The changes of sciatic functional index(SFI) were measured before and on the 1st and 17th day after modeling. The morphological changes of the sciatic nerve were observed by hematoxylin-eosin(HE) staining;and the differences in NF-κB protein expression in the right dorsal horn of the spinal cord of rats were detected.@*RESULTS@#Following modeling, there was no significant difference in PWT, PWL and SFI between the blank group and the sham group (P>0.05), but the PWT, PWL and SFI of the model group and the Tuina group decreased significantly (P<0.01). After manual intervention, the pain threshold of rats in Tuina group increased. On the 8th day of manual intervention (the 10th day after modeling), PWT in Tuina group increased significantly compared with that in model group (P<0.01). On the 5th day of manual intervention (the 7th day after modeling), the PWL of the massage group was significantly higher than that of the model group (P<0.01). The pain threshold of rats in Tuina group continued to rise with the continuous manipulation intervention. After 14 days of manipulative intervention, the sciatic nerve function index of rats in the Tuina group increased significantly(P<0.01). Compared with the blank group and sham group, the myelinated nerve fibers of sciatic nerve in the model group were disordered and the density of axons and myelin sheath was uneven. Compared with the model group, the nerve fibers of rats in the Tuina group were gradually continuous and the axons and myelin sheath were more uniform than those in the model group. Compared with the blank group and sham group, the expression of NF-κB protein in the right spinal dorsal horn of the model group was significantly increased(P<0.01). Compared with the model group, the expression of NF-κB protein in the right spinal dorsal horn of rats in Tuina group decreased significantly(P<0.01).@*CONCLUSION@#Pressing and kneading the Huantiao (GB30) point restores nerve fiber alignment;and improves the PWT、PWL and SFI in the CCI model by decreasing NF-κB p65 protein expression in the spinal dorsal horn. There fore, Tuina demmstrates an analgesic effect and improves the gait of rats with sciatica.

Intervention of Osteoking in Rats with Myofascial Pain Syndrome / 中国实验方剂学杂志

ObjectiveTo clarify the intervention effect of Osteoking （OK） in rats with myofascial pain syndrome （MPS） and preliminarily explore the pharmacological mechanism of OK in relieving chronic pain from the perspective of anti-inflammatory disease. MethodThe 60 SD rats were divided into normal group， model group， low， medium， and high dose OK groups （0.66， 1.31， 2.63 mL·kg-1）， and positive celecoxib group （21 mg·kg-1）. The MPS rat model was established by beating combined with the centrifugal exercise method， and the OK and celecoxib were given at the same time. SMALGO paw pressure pain manometer detected the shock pain point tenderness threshold of rats， and the Von-Frey needle and acetone stimulation method detected the mechanical hyperalgesia threshold and cold hyperalgesia stimulation response respectively. Eight weeks and 10 weeks after modeling， the spontaneous discharge state and convulsion response of MPS rats were determined by electromyograph （EMG） instrument. The gait changes of MPS rats were detected using a CatWalk gait analyzer. The expression levels of interleukin-1 β （IL-1β）， tumor necrosis factor-α （TNF-α）， substance P （SP）， and bradykinin （BK） were measured by enzyme-linked immunosorbent assay （ELISA）. The protein expression levels of nuclear transcription factor-κB （NF-κB） inhibiting protein α （IκBα）， phosphorylates （p）- IκBα， NF-κB p65， and p-NF-κB p65 were detected in MPS rats by Western blot. The positive expression of p-NF-κB p65 was detected by immunofluorescence. ResultCompared with the normal group， the model group shows 100% positive rates for EMG signal and local convulsions response at both the 8th and 10th weeks. The tenderness threshold and mechanical hyperalgesia threshold are significantly reduced. Cold hyperalgesia score is significantly increased， and gait is abnormal. The expression levels of serum and trigger points IL-1β， TNF-α， SP， BK， p-IκBα， and p-NF-κB p65， as well as the positive expression intensity of p-NF-κB p65 are significantly increased （P<0.01）. Compared with the model group， the positive rate of EMG detection and local convulsion response is significantly reduced in the medium and high dose OK groups （P<0.05）. The tenderness threshold and mechanical hyperalgesia threshold increase significantly in the medium and high dose OK groups， and the cold hyperalgesia score is significantly reduced in the high dose OK group （P<0.01）. The standing time， swing time， and walking period are significantly increased. The swing speed， maximum contact area， and maximum contact intensity are significantly decreased in the high dose OK group （P<0.05）. Moreover， the protein expression levels of p-IκBα/IκBα and p-NF-κB p65/NF-κB p65 are significantly reduced in the medium and high dose OK groups （P<0.05，P<0.01）. The positive expression intensity of p-NF-κB p65 is significantly decreased in the high dose OK group （P<0.01）. ConclusionThe mechanism of OK in relieving the pain in trigger points of MPS and improving gait abnormalities is related to the downregulation of the NF-κB p65 inflammatory signaling pathway to reduce the expression of inflammatory factors and pain mediators in blood and trigger point tissue.

Mechanism of Tibetan medicine Ershiwuwei Songshi Pills against liver injury induced by acetaminophen in mice based on Keap1/Nrf2 and TLR4/NF-κB p65 signaling pathways / 中国中药杂志

The present study investigated the mechanism of the Tibetan medicine Ershiwuwei Songshi Pills(ESP) against the liver injury induced by acetaminophen(APAP) in mice based on the kelch-like ECH-associated protein 1(Keap1)/nuclear transcription factor E2 related factor 2(Nrf2) and Toll-like receptor 4(TLR4)/nuclear factor-kappa B(NF-κB) p65 signaling pathways. Kunming mice were randomly divided into a blank control group, a model group, an N-acetyl-L-cysteine(NAC) group, and high-(400 mg·kg~(-1)), medium-(200 mg·kg~(-1)), and low-dose(100 mg·kg~(-1)) ESP groups. After 14 days of continuous administration, except for those in the control group, the mice were intraperitoneally injected with 200 mg·kg~(-1) APAP. After 12 h, the serum and liver tissues of mice were collected. Hematoxylin-eosin(HE) staining was performed on pathological sections of the liver, and the levels of aspartate aminotransferase(AST) and alanine aminotransferase(ALT) in the serum and the levels of glutathione(GSH), malondialdehyde(MDA), superoxide dismutase(SOD), catalase(CAT), myeloperoxidase(MPO), and total antioxidant capacity(T-AOC) in liver tissue homogenate were detected to observe and analyze the protective effect of ESP on APAP-induced liver injury in mice. The serum levels of tumor necrosis factor-alpha(TNF-α), interleukin-1 beta(IL-1β), and interleukin-6(IL-6) were determined by enzyme-linked immunosorbent assay(ELISA). The protein expression of Nrf2, Keap1, TLR4, and NF-κB p65 in the liver was determined by Western blot. Quantitative real-time was used to determine the mRNA expression of glutamate-cysteine ligase catalytic subunit(GCLC), glutamate-cysteine ligase regulatory subunit(GCLM), heme oxygenase-1(HO-1), and NAD(P)H dehydrogenase quinone 1(NQO-1) in the liver to explore the mechanism of ESP in improving APAP-induced liver damage in mice. As revealed by results, compared with the model group, the ESP groups showed improved liver pathological damage, decreased ALT and AST levels in the serum and MDA and MPO content in the liver, increased GSH, SOD, CAT, and T-AOC in the liver, reduced TNF-α and IL-6 levels in the serum, down-regulated expression of Keap1 in the liver cytoplasm and NF-κB p65 in the liver nucleus, up-regulated expression of Nrf2 in the liver nucleus, insignificant change in TLR4 expression, and elevated relative mRNA expression levels of antioxidant genes GCLC, GCLM, HO-1, and NQO-1. ESP can reduce the oxidative damage and inflammation caused by APAP, and the mechanism may be related to the Keap1/Nrf2 signaling pathway and the signal transduction factors on the TLR4/NF-κB p65 pathway.

Effects of different extracts of Dendrobium officinale on rats with chronic pharyngitis induced by pepper water combined with ammonia / 中国中药杂志

Dendrobium officinale can serve as Chinese medicinal material effective in nourishing yin, clearing heat, and producing fluid, and is used to treat throat diseases, but its active substances and mechanism are not clear. To clarify the active fraction and underlying mechanism of D. officinale against chronic pharyngitis(CP), the present study induced a CP model in rats by pepper water combined with low-concentration ammonia, and crude polysaccharides of D. officinale(DOP), non-polysaccharides of D. officinale(DON), and total extract of D. officinale(DOT)(0.33 g·kg~(-1), calculated according to the crude drug) were administered by gavage for six weeks. The changes in oral secretions and pharyngeal conditions of rats with CP were observed and rated. The hematological indicators were determined by an automatic hematology analyzer. The serum levels of pro-inflammatory factors, such as tumor necrosis factor-alpha(TNF-α), interleukin 1β(IL-1β), and interleukin 6(IL-6), and T-lymphocyte cytokines, including interferon γ(IFN-γ), interleukin 4(IL-4), interleukin 17(IL-17), and transforming growth factor β1(TGF-β1) were detected by the enzyme-linked immunosorbent assay(ELISA). The proportions of CD3~+, CD4~+, and CD8~+cells in peripheral blood T lymphocyte subsets were determined by the flow cytometry. The histomorphological changes of the pharynx were observed by hematoxylin-eosin(HE) staining. The protein expression of nuclear factor-κB P65(NF-κB P65), cyclooxygenase-2(COX-2), F4/80, and monocyte chemoattractant protein-1(MCP-1) in the pharynx were detected by immunohistochemistry and Western blot. The results showed that DOP and DON could significantly relieve pharyngeal lesions, reduce white blood cells(WBC) and lymphocytes(LYMP), decrease the levels of pro-inflammatory factors TNF-α, IL-6, and IL-1β, and inhibit the protein expression of NF-κB P65, COX-2, F4/80, and MCP-1 in the pharynx. DOP was superior in reducing oral secretions and serum IL-17 level and inferior in increasing CD4~+/CD8~+ratio to DON. It is suggested that both polysaccharides and non-polysaccharides of D. officinale have anti-PC effects and the anti-inflammatory mechanism may be related to the regulation of T lymphocyte distribution and inhibition of the inflammatory signaling pathways mediated by NF-κB P65. The anti-inflammatory effect of DOP may be related to the regulation of Th17/Treg balance, while that of DON may be related to the regulation of the Th/Tc ratio.

Effect of electroacupuncture on pulmonary function and M1 polarization of alveolar macrophages in rats with chronic obstructive pulmonary disease / 针刺研究

OBJECTIVE: To observe the effect of electroacupuncture (EA) at "Zusanli" (ST36) and "Feishu" (BL13) on M1 polarization of alveolar macrophages (AM) in rats with chronic obstructive pulmonary disease(COPD), so as to explore its anti-inflammatory mechanism underlying improvement of COPD. METHODS: Forty SD rats were randomly divided into normal and normal＋EA, COPD model and COPD＋EA groups (n＝10 in each group). The COPD model was established by simple fumigation. EA (4 Hz/20 Hz, 1 to 2 mA) was applied to bilateral ST36 and BL13 for 30 min, once every other day for 2 weeks. The pulmonary function including the forced vital capacity (FVC), forced expiratory volume in 0.1 and 0.3 s (FEV0.1, FEV0.3, FEV0.1/FVC, and FEV0.3/FVC) was detected by using a small animal respiratory function detector. Histopathological changes of the lung were displayed by H．E. staining. The contents of tumor necrosis factor-α (TNF-α) and induced nitric oxide synthase (iNOS) in the broncho alveolar lavage fluid (BALF) were assayed by ELISA. The expression of M1 polarization markers (CD86，iNOS), myeloid differentiation factor 88(MyD88) and nuclear factor-κB p65(NF-κB p65) in AM were detected by Western blot and quantitative real time-PCR, separately. The distribution and expression of CD86 in the lung were detected by immunohistochemistry. RESULTS: Following modeling, the levels of FVC, FEV0.1, FEV0.3, ratios of FVE0.1/FVC and FEV0.3/FVC were significantly decreased (P<0.01), while the contents of TNF-α and iNOS in the BALF, expression of CD86, iNOS, MyD88 and NF-κB p65 mRNAs and proteins in the AM, and CD86 immunoactivity in the lung were significantly increased in the model group relevant to the normal group (P<0.01). After the intervention, the decrease of the lung function and increase of the above-mentioned genes and proteins were all reversed in the COPD＋EA group (P<0.05, P<0.01). CONCLUSION: EA at ST36 and BL13 can reduce pulmonary inflammation in COPD rats, which may be related to its function in inhibiting M1 polarization of AM via down-regulating MyD88/NF-κB p65 signaling pathway.

Effect of Shaoyaotang on Expressions of TLR4, NF-κB p65 and IL-6 in Rats with Damp-heat Ulcerative Colitis / 中国实验方剂学杂志

Objective:To explore the mechanism of Shaoyaotang in the treatment of ulcerative colitis (UC) based on toll-like receptor 4 (TLR4)/nuclear factor kappaB (NF-κB) signaling pathway. Method:A total of 50 Wistar rats were selected, including half male and half female. The damp-heat UC rat model was replicated by the methods of the combination of diseases and syndromes and the combination of 2, 4, 6-nitrobenzene sulfonic acid (TNBS) and ethanol. After the successful modeling, the model rats were randomly divided into model group, salazulesulfonate group, and low, medium and high-dose Shaoyaotang groups, and 10 rats (half male and half female) were selected as the blank control group. Low, medium and high-dose Shaoyaotang groups were given 6, 12, 24 g·kg-1 by gavage, and salazonyl arsenic group was given 1 g·kg-1 by gavage. Blank control group was given the equal volume of normal saline for 21 consecutive days. Colon samples were collected after the last administration, and the expressions of TLR4, NF-κB p65 and IL-6 mRNA in colon tissues were detected by fluorescent quantitative polymerase chain reaction (Real-time PCR）, and the expressions of TLR4, NF-κB p65 and IL-6 protein in colon tissues were detected by Western blot. Result:Compared with the blank control group, the relative expressions of TLR4, NF-κB p65, IL-6 mRNA and protein in the model group were significantly increased (P<0.05). Compared with the model group, the expression levels of TLR4, NF-κB p65 and IL-6 mRNA and protein in the salazopyridine group and Shaoyaotang groups were significantly decreased (P<0.05). Conclusion:Shaoyaotang can inhibit the development of UC by regulating the expressions of TLR4, NF-κB p65 and IL-6 mRNA and proteins in the TLR4/NF-κB pathway.

Effect of Huangqi Guizhi Wuwutang on AGEs/RAGE/NF-κB Signaling Pathway in Diabetic Peripheral Neuropathy Rats / 中国实验方剂学杂志

Objective:To study the effect of Huangqi Guizhi Wuwutang on receptor of advanced glycation end products(AGEs)/advanced glycation endproducts (RAGE)/nuclear transcription factor-kappa B p65 (NF-κB p65) signaling pathway in the diabetic peripheral neuropathy rats through an animal modeling experiment, and discuss the mechanism of Huangqi Guizhi Wuwutang in alleviating diabetic peripheral neuropathy. Method:Rat model of diabetic peripheral neuropathy was established by high-fat diet and intraperitoneal injection with streptozotocin (STZ). After successful modeling, Huangqi Guizhi Wuwutang intervention began in the fifth week. The patients in high-dose group (19.40 g∙kg-1∙d-1), middle-dose group (4.85 g∙kg-1∙d-1) and low-dose group (2.43 g∙kg-1∙d-1) were given by gavage continuously for 12 weeks. The western medicine control group was given 25 mg∙kg-1∙d-1 by gavage. After the experiment, serum interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) were detected by enzyme-linked immunosorbent assay (ELISA). Real-time fluorescent quantitative polymerase chain reaction (Real-time PCR) was used to detect RAGE and NF-κB p65 mRNA expressions in sciatic nerve tissue. The expressions of RAGE, NF-κB and phosphorylation(p)-NF-κB p65 proteins in sciatic nerve tissues were detected by Western blot (WB). Result:Compared with the normal group, serum IL-1β and TNF-α protein levels, RAGE mRNA and NF-κB p65 mRNA levels, RAGE protein, NF-κB p65 protein and p-NF-κB p65 protein levels were significantly increased in the model group (P<0.01), the ratio of p-NF-κB p65 to NF-κB p65 was increased, and the phosphorylation of NF-κB p65 was enhanced (P<0.01). After the intervention of Huangqi Guizhi Wuwutang, compared with the model group, serum IL-1β and TNF-α protein levels, RAGE and NF-κB p65 mRNA levels, RAGE protein, NF-κB p65 protein and p-NF-κB p65 protein levels were all decreased (P<0.01), the ratio of p-NF-κB p65 to NF-κB p65 was decreased in high-dose group (P<0.01). The effect was obvious with the increase of dose of astragalus cassia twig. Conclusion:Huangqi Guizhi Wuwutang can alleviate diabetic peripheral neuropathy, and its mechanism may be related to blocking the expression of RAGE on tissue cell surface in AGEs/RAGE/NF-κB signaling pathway, inhibiting the activation of NF-κB and inducing TNF-α triggered oxidative stress and excessive inflammatory response, so as to avoid cell damage and dysfunction.

Effect of Xiao Xumingtang Combined with Ultra-acupuncture Along Governor Meridian on Autophagy-related Protein NF-κB p65 in Cerebral Ischemia Reperfusion Model / 中国实验方剂学杂志

Objective:To investigate the effect of Xiao Xumingtang combined with super-acupuncture along governor meridian on autophagy-related protein nuclear factor-kappa B p65 (NF-κB p65) in cerebral ischemia-reperfusion model rats, so as to study the relationship between autophagy-related protein NF-κB p65 and brain protection mechanism, and look for the best intervention time point of acupuncture. Method:A total of 152 adult SD male rats were randomly divided into sham operation group, model group, high-dose Xiao Xumingtang group (high-dose drug group), low-dose Xiao Xumingtang group (low-dose drug group) and acupuncture group. There were seven groups including high-dose Xiao Xumingtang + acupuncture group (high acupuncture group) and low-dose Xiao Xumingtang + acupuncture (low acupuncture group). Model group, high-dose drug group, low-dose drug group, and acupuncture group were divided into 4 subgroups according to 30 minutes, 2, 4， 6 h of ischemia-reperfusion, with 6 animals in each group. After successful modeling, according to Zea Longa's neural function score, eligible rats were included into the corresponding groups. The sham operation group only received carotid artery dissection, the model group was only modeled without any treatment, high and low-dose Xiao Xumingtang groups were calculated based on the body surface area of the animal and given 60 g·kg-1·d-1 and 15 g·kg-1·d-1 drug by gavage for treatment, acupuncture was performed to smooth governor meridian and regulate the mind. After 14 days of consecutive treatment, neurological function was scored. Western blot was used to detect the expression of autophagy-related protein NF-κB p65 in rat brain tissue. Result:Compared with the sham operation group, the neurological impairment scores of the model group, the high-dose drug group, the low-dose drug group, the acupuncture group, the high acupuncture group, and the low acupuncture group were significantly increased at each time point (P<0.01). The neurological impairment scores were significantly lower at each time point than those of the high-dose drug group, low-dose drug group, acupuncture group, high acupuncture group, and low acupuncture group (P<0.01), compared with the sham operation group, NF-κB p65 in model group, high-dose drug group, low-dose group, acupuncture group, high acupuncture group and low acupuncture group was significantly increased in the brain tissue at each time point (P<0.01), compared with the model group, the expression of NF-κB p65 protein in the brain tissue of model group, high-dose drug group, low-dose group, acupuncture group, high acupuncture group and low acupuncture group was decreased at each time point (P<0.05), particular in the high acupuncture group (P<0.01). Conclusion:Xiao Xumingtang combined with ultra-early acupuncture along governor meridian can significantly alleviate neurological impairment in rats with cerebral ischemia-reperfusion model. Xiao Xumingtang combined with ultra-early acupuncture along governor meridian can inhibit cerebral ischemia-reperfusion model rats. The activity of autophagy-related protein NF-κB P65 protects the brain function. There is no significant difference in the brain protective effect of Xiao Xumingtang combined with ultra-early acupuncture along governor meridian within 6 hours.

Effect of Shenling Baizhusan on Protein and mRNA Expression of NF-κB p65，IκBα，IκKβ in Ulcerative Colitis Rats with Syndrome of Dampness Stagnancy Due to Spleen Deficiency / 中国实验方剂学杂志

Objective:To observe the effect of Shenling Baizhusan on the protein and mRNA expression of inhibitor of nuclear factor kappa B kinase (IκK)/inhibitor of nuclear factor kappa B(IκB)/nuclear factor kappa B(NF-κB) signaling pathway in the colon of rats with ulcerative colitis (UC) of spleen deficiency and dampness stagnation type, and to explore the mechanism of Shenling Baizhusan in the treatment of UC. Method:The 48 Wistar rats were randomly divided into normal group, model group, Shenling Baizhusan group (15.6 g·kg-1) and osalazine sodium group (0.68 g·kg-1), 12 rats in each group. The model of UC with spleen deficiency and dampness stagnation was reproduced by trinitrobenzene sulfonic acid (TNBS)/ethanol enema combined with environment and diet intervention.Serum levels of tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β(IL-1β) were determined by enzyme-linked immunosorbent assay（ELISA）. The expression of NF-κB p65, IκBα, IκKβ protein in colon tissue was measured by Western blot and immunohistochemical method, and the mRNA expression of NF-κB p65, IκBα and IκKβ in colon tissue of rats in each group was detected and compared by real time polymerase chain reaction (Real-time PCR). Result:Compared with normal group,the levels of TNF-α,IL-6 and IL-1β in the serum, the protein and mRNA expression of NF-κB p65, IκKβ in colon tissue of the model group was significantly higher than that of normal group (P<0.01), and the protein and mRNA expression of IκBα was significantly lower than that of normal group (P<0.01). Compared with model group,the protein and mRNA expression of NF-κB p65, IκKβ in colon tissue of the Shenling Baizhusan group and osalazine sodium group were significantly decreased (P<0.01), and the protein and mRNA expression of IκBα was significantly increased (P<0.01). Conclusion:Shenling Baizhusan can obviously down regulate the protein and mRNA expression of NF-κB p65, IκKβ,up regulate the expression of IκBα in colon tissue of UC rats with spleen deficiency and dampness stagnation. The inhibition of IκK/IκB/NF-κB signal pathway activation by Shenling Baizhusan is an important mechanism of its role in protecting intestinal mucosa.

Study on Preventive Effects and Mechanism of Ginsenoside Rg 1 on Focal Cerebral Ischemia-reperfusion Injury Model Rats / 中国药房

OBJECTIVE：To st udy preventive effect and mec hanism of ginsenoside Rg 1 on focal cerebral ischemia-reperfusion injury（CIRI）model rats. METHODS ：Totally 78 SD rats were randomly divided into sham operation group ，model group ， butylphthalide control group （positive control ，10 mL/kg），ginsenoside Rg 1 low-dose，medium-dose and high-dose groups （10， 20，40 mg/kg），with 13 rats in each group. Administration groups were give relevant medicine intraperitoneally ，sham operation group and model group were given constant volume of normal saline intraperitoneally ，once a day ，for consecutive 7 d. After medication，except for the sham operation group ，focal CIRI model was induced by middle cerebral artery occlusion （MCAO） method in other groups. After modeling ，neurological deficit scoring was performed according to the modified neurological difict scoring standard ； TTC staining was used to detected the percentage of cerebral infarction of rats ；the cerebral water content was measured by dry/wet weight method ；serum contents of IL- 1β and IL-6 were detected by ELISA ；the protein expressions of p-p 38 MAPK and p-NF-κB p65 in cerebral tissue were determined by immunohistochemistry and Western blotting assay. RESULTS ： Compared with sham operation ，neurological deficits score ，percentage of cerebral infarction and cerebral water content ，serum contents of IL- 1β and IL-6，positive expression numbers of cells and protein expressions of p-p 38 MAPK and p-NF-κB p65 in cerebral tissue were increased significantly in model group （P＜0.05 or P＜0.01）. Compared with model group ，above index levels of administration groups were all decreased significantly （P＜0.05 or P＜0.01），and the effect of ginsenoside Rg 1 had a dose-dependent trend ；there was no significant difference of all above indexes between ginsenoside Rg 1 middle-dose，high-dose groups and butylphthalide control group （P＞0.05）. CONCLUSIONS ：Ginsenoside Rg 1 has a certain preventive effect on focal CIRI model rats ，the mechanism of which may be associated with down-regulating the protein expression of p-p 38 MAPK and p-NF-κB p65，inhibiting the release of inflammatory factors such as IL- 1β and IL-6.

Effect of Xiao Chaihutang on NF-κB Signaling Pathway in Synovial Tissue of CFA Rats / 中国实验方剂学杂志

Objective:To observe the expression of tumor necrosis factor receptor-associated death domain (TARDD), nuclear transcription factor-κB inhibiting protein α(IκBα)IκB kinase-α (IKKα) and nuclear transcription factor (NF)-κB p65 protein in the NF-κB signaling pathway of synovial tissues of complete Freund's adjuvant (CFA) rats after treatment with Xiao Chaihutang (XCHT). Method:In animal experiments, SPF health adult female Wistar rats were used to prepare the CFA animal model of rats with rheumatoid arthritis with Freund's complete adjuvant and cattle Ⅱ collagen type. According to the random number table, the rats were randomly divided into the normal group, the model group, the low-dose XCHT group, the medium-dose XCHT group, the high-dose XCHT group, and the Tripterygium glucosides group. The drugs were given at 7 d after the model was built. Both normal group and model group were given water for injection,and low-dose XCHT group(5.94 g·kg-1),medium-dose XCHT group(11.88 g·kg-1),high-dose XCHT group(23.76 g·kg-1),Tripterygium glucosides group(0.006 3 g·kg-1) were given corresponding drugs by gavage for three times a day, 2 mL/time. The histopathology of rat ankle joint was observed, and the protein expressions of TARDD,IKKα,IκBα,NF-κB p65 in the NF-κB signaling pathway in synovial tissue of CFA rats were detected by Western blot. Result:With the increase of the dosage of XCHT, the histopathological score of the right posterior ankle joint of the experimental rats was increased. And in the protein expressions of TARDD,IKKα,IκBα,NF-κB p65 in NF-κB signaling pathway in Synovial Tissue of CFA rats, compared with the model group, the statistical results of the low-dose XCHT group showed decreased protein expressions (PPPα, IκB α, NF-κB p65 in the NF-κB signaling pathway were significantly increased (PPα, IκBα, NF-κB p65 key protein expressions in the NF-κB signaling pathway and protein expressions in low-dose XCHT group were obviously lower (PPConclusion:This study shows that as the dose of Xiao Chaihutang increases, it could effectively improve synovitis, and suppress the expressions of key proteins in the inflammatory signaling pathway of NF-κB, thereby preventing inflammation and suppressing bone erosion.

Effects of Taoren Chengqitang on Intestinal Mucosal Barrier and TLR9 Signaling Pathway in Septic Rats / 中国实验方剂学杂志

Objective: To study the protective effect of Taoren Chengqitang on intestinal mucosal barrier in septic rats and its possible mechanism. Method: Rats were divided into sham operation group, model group (replication of septic rats with cecal ligation and perforation), low, middle and high-dose Taoren Chengqitang groups (2.85,5.70,8.55 g·kg-1), and dexamethasone group (0.01 g·kg-1),with 12 rats in each group. After last administration, rats were put to death, the morphological changes of intestinal mucosa were observed under electron microscope, the bacterial translocation rates in lymphoglandulae mesentericae, liver, kidney and spleen tissues were detected; the levels of serum tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and diamine oxidase (DAO) and intestinal fatty acid binding protein (i-FABP) levels in small intestine tissues were detected by enzyme-linked immunosorbent assay (ELISA).Expressions of Toll like receptor 9 (TLR9), myeloid differentiation factor 88 (MyD88) and nuclear factor-kappa B subunit p65 in small intestine tissues were detected by Western blot and immunohistochemistry. Result: Compared with sham operated group, the bacterial translocation rate, TNF-α and IL-1β levels in model group increased significantly, DAO, i-FABP, mucosal thickness and villus height decreased significantly, and protein expressions of TLR9, MyD88 and nucleus NF-κB p65 increased significantly (PPκB p65 protein decreased significantly. Compared with model group, the bacterial translocation rate, TNF-α and IL-1β levels in organs of low, medium and high-dose Taoren Chengqitang groups and dexamethasone group decreased significantly, DAO, i-FABP, mucosal thickness and villus height increased significantly, while the protein expressions of TLR9, MyD88 and nucleus NF-κB p65 decreased significantly (PκB p65 protein increased significantly. Conclusion: Taoren Chengqitang has a certain protective effect on intestinal mucosal barrier in septic rats, which may be related to the inhibition of TLR9 signaling pathway.

Mechanism of Modified Erchentang on Signaling Pathway of TLR4/MyD88/NF-κB in Lung Tissue of Rats with Chronic Obstructive Pulmonary Disease / 中国实验方剂学杂志

Objective: To study the effect of modified Erchentang on expressions of Toll-like receptor 4 (TLR4), myeloid differentiation factor (MyD88) and nuclear factor-κB (NF-κB) genes in the lung tissue homogenate of rats with chronic obstructive pulmonary disease (COPD). Method: Forty SD rats were randomly divided into normal group, model group, modified Erchentang group and EVP4593 (NF-κB inhibitor) group. Rat COPD models were prepared through cigarette smoke and tracheal dripping with lipopolysaccharide (LPS). After the modeling, normal and model groups were intragastrically given normal saline solution, EVP4593 group was given EVP4593(1 mg · kg-1) through subcutaneous injection, and modified Erchentang group was given corresponding herbal drugs intragastrically (10 g · kg-1) for 14 days. The levels of high mobility group box 1(HMGB1), chemokines CXCL-2, CXCL-3 and monocyte chemoattractant protein-1 (MCP-1) in rats serum were detected by enzyme-linked immunosorbent assay in rats serum. The expressions of Toll-like receptors 4(TLR4), myeloid differentiation factor (MyD88) and nuclear factor-κB p65 (NF-κB p65) mRNA were detected by Real-time fluorescence quantitative PCR (Real-time PCR) method. Western blot were used to detect the levels of TLR4, MyD88, NF-κB p65 and p-NF-κB p65 protein. Immunohistochemistry (IHC) method was used to detect the localization and expressions of TLR4, MyD88 and p-NF-κB p65 protein in the lung tissue. Result: The mRNA and protein expressions of TLR4, MyD88 and NF-κB p65 were increased significantly (PPκB p65 mRNA and protein were decreased significantly (PConclusion: Modified Erchentang may inhibit the inflammatory response of COPD effectively. The mechanism may be related to the inhibition of the expressions of the signal molecule genes involved in the TLR4/MyD88/NF-κB pathway and the reduction of the release of HMGB1, CXCL-2, CXCL-3 and MCP-1.

Ligustrazine regulates LPS-induced apoptosis and inflammatory response of osteoarthritis chondrocytes via inhibiting phosphorylation of NF-κB P65 / 中国免疫学杂志

Objective: To explore the effect of ligustrazine on the LPS-induced apoptosis and inflammatory response of osteoarthritis chondrocytes. Methods: Osteoarthritis model was induced by LPS. Chondrocytes were divided into four group: control group, ligustrazine ( 20 μmol/L) group, LPS ( 100 ng/ml) group and ligustrazine ( 20 μmol/L) +LPS ( 100 ng/ml) group. Apoptosis was measured by Hoechst33258 staining. The levels of nitric oxide ( NO), tumor necrosis factor α ( TNF-α) and interleukin ( IL) -6 were detected by ELISA. The protein levels of collagenⅡ, aggrecan, matrix metalloproteinase 13 ( MMP-13), NF-κB P65 and p-NF-κB P65 were tested by Western blot. Results: The LPS-induced abnormal cell morphology and decreased number of cells were ameliorated by ligustrazine ( 20 μmol/L). The apoptosis in LPS group was higher than control group ( P＜0. 05). The LPS-induced enhancive apoptosis was reduced by ligustrazine ( P＜0. 05). Compared with control group, the expression of collagenⅡ and aggrecan was alleviated with increased expression of MMP-13 ( P＜0. 05). The LPS-induced declined expression of collagenⅡ and aggrecan and elevated expression of MMP-13 was inhibited by ligustrazine ( P＜0. 05). The levels of NO, TNF-α and IL-6 in LPS group were higher than control group ( P＜0. 05). The levels of NO, TNF-α and IL-6 in LPS+ligustrazine group were lower than LPS group ( P＜0. 05). Compared with control group, the rate of pP65/P65 in LPPS group was enhanced ( P＜ 0. 05). The LPS-indiced increased rate of p-P65/P65 was decreased by ligustrazine ( P ＜0. 05). Conclusion: Ligustrazine alleviates the LPS-induced apoptosis and inflammatory response of osteoarthritis chondrocytes via inhibiting phosphorylation of NF-κB P65.

Chlorogenic acid regulates lipopolysaccharide-induced apoptosis and inflammatory response of human retinal vascular endothelial cells / 中国免疫学杂志

Objective: Diabetic retinopathy is one of the most common complications of diabetes. This study aims to explore the effect of chlorogenic acid ( CA) on lipopolysaccharide ( LPS) -induced apoptosis and inflammatory response of human retinal vascular endothelial cells ( HRECs). Methods: Cell proliferation was tested by CCK-8. Cell apoptosis was detected by flow cytometry. The levels of IL-6, TNF-α and IL-10 were measured by ELISA. The protein levels of Ki67, Bcl-2, Caspase-3, Bax, NF-κB P65 and P-NF-κB P65 were detected by Western blot. Results: The low concentration ( ＜50 μmol/L) of CA had no effect on cell viability of HRECs. The cell viability of HRECs was decreased by high concentration (＞ 50 μmol/L) of CA. Compared with control group, the proliferation in LPS group was reduced with enhancive apoptosis ( P ＜ 0. 05). Compared with LPS group, the proliferation in CA ( 20, 50 μmol/L) group was increased with attenuated apoptosis ( P ＜ 0. 05). The protein levels of Ki67 and Bcl-2 in LPS group were lower than control group ( P ＜0. 05). And the expression of Caspase-3 and Bax in LPS group was higher than control group ( P＜0. 05). Compared with LPS group, the protein levels of Ki67 and Bcl-2 in CA ( 20, 50 μmol/L) group were elevated with decreased expression of Caspase-3 and Bax ( P ＜0. 05). Moreover, the levels of IL-6 and TNF-α in LPS group were higher than control group ( P＜0. 05). And the levels of IL-10 in LPS group were lower than control group ( P＜0. 05). Compared with LPS group, the levels of IL-6 and TNF-α in CA ( 20, 50 μmol/L) group were reduced with enhanced levels of IL-10 ( P＜0. 05). In addition, the rate of p-P65/P65 in LPS group was higher than control group ( P＜0. 05). The rate of p-P65/P65 in CA ( 10, 20, 50 μmol/L) group was lower than LPS group ( P＜0. 05). Conclusion: Chlorogenic acid alleviates LPS-induced enhancive apoptosis and inflammatory response of HRECs via inhibiting activation of NF-κB P65.

Scalp-acupuncture Improves Neurological Function by Regulating Expression of NF-κB p 65 mRNA, IκB mRNA and IL-1 β and TNF-α in Parahippocampal Gyrus of Cerebral Ischemia Rats / 针刺研究

OBJECTIVE: To observe the effect of scalp-acupuncture intervention on the expression of parahippocampal factor-κB p 65 mRNA (NF-κB p 65 mRNA), IκB mRNA, interleukin-1 β (IL-1 β) and tumor necrosis factor-α (TNF-α) in rats with cerebral ischemia (CI), so as to investigate its molecular mechanisms underlying improving CI by reducing inflammatory response. METHODS: A total of 64 SD rats were randomized into normal control, model, medication and scalp-acupuncture groups, with 16 rats in each group. The focal CI model was established by middle cerebral artery occlusion (MCAO). Intraperitoneal injection of Pyrrolidine Dithiocarbamate (100 mg•kg－1•d－1) was administrated for rats in the medication group, once a day for 7 days. For rats of the scalp-acupuncture group, the acupuncture needles were rapidly inserted into bilateral Dingnieqianxiexian (MS 6) and Dingniehouxiexian (MS 7), followed by twirling the needles at 200 cycles/min for 1 min, once again every 10 min during 30 min's needle retention. The treatment was conducted once a day for 7 days. The neurologic deficit score (0－5 points, impaired consciousness, death, etc.) and neurological function score (motor, sensory and sensory tests, 0－10 points) were assessed according to Longa's (1989) and Schäbitz's (2004) methods, respectively. The expression levels of NF-κB p 65 mRNA and IκB mRNA in the parahippocampus gyrus tissue were detected by fluorescence quantitative reverse transcription-PCR, and IL-1 β and TNF-α proteins in the parahippocampus gyrus tissue were detected by immunohistochemistry. RESULTS: After modeling, the neurologic deficit and neurological function scores and the expression levels of NF-κB p 65 mRNA, IL-1 β and TNF-α in the parahip-pocampus were significantly increased in the model group than in the normal group (P<0.01), while the expression of IκB mRNA was considerably down-regulated (P<0.01). Following treatment intervention, the neurologic deficit and neurological function scores as well as NF-κB p 65 mRNA, and IL-1 β and TNF-α protein expression were significantly decreased in both scalp-acupuncture and medication groups compared with the model group (P<0.05, P<0.01), and the decreased expression of IκB mRNA was obviously increased (P<0.05)．. CONCLUSION: Scalp-acupuncture can improve neurologic function in cerebral ischemic rats, which is related with its effects in up-regulating the expression of IκB to inhibit the dissociation of NF-κB, then decreasing the expression of IL-1 β and TNF-α (reducing inflammatory response) in the parahippocampal gyrus tissue.

Toll-like Receptor 4/Nuclear Factor-κB Signaling in Synovial Tissue Is Involved in the Anti-inflammatory Effect of Moxibustion in Rats with Rheumatoid Arthritis / 针刺研究

OBJECTIVE: To observe the effect of moxibustion on Toll-like receptor 4/nuclear factor-κB (TLR 4/NF-κB) signaling in the synovial tissue of the ankle joint in rats with rheumatoid arthritis (RA), so as to analyze its biological mechanism underlying improvement of RA. METHODS: Fifty male SD rats were randomly divided into normal control, model, moxibustion, moxibustion ＋ TLR 4 agonist, and moxibustion ＋ TLR 4 antagonist groups (n＝10 rats in each). The RA model was established by subcutaneous injection of Freund's complete adjuvant (FCA, 0.1 mL/rat) at the right hind-paw and by being raised in a wind (air fan blowing), cold (about 10 ℃) and wet (purling) environment for 20 days. After 3 days of modeling, mild moxibustion was applied to bilateral "Shenshu" (BL 23) and "Zusanli" (ST 36) for 20 minutes, once daily for successive 10 days. The TLR 4 agonist (lipopolysaccharide) or TLR 4 antagonist (TAK-242) (1 mg/mL) was separately administered via the tail vein 30 min before performing moxibustion every time in the agonist group and the antagonist group. The expression of NF-κB inhibitory factor ɑ (IκBɑ), IκB kinase complex β(IκKβ), myeloid differentiation factor 88 (MyD 88), TLR 4, and NF-κB p 65 proteins in the synovial tissue of the ankle joint was detected by using Western blot. RESULTS: Following modeling, the girth of the swollen ankle joint was obviously bigger (P0.05), suggesting that activation of TLR 4 reduced the anti-inflammatory effect of moxibustion intervention. CONCLUSION: Moxibustion can reduce the ankle joint swelling in RA rats, which may be closely associated with its effect in down-regulating the expression of IκBɑ， IκKβ， TLR 4, MyD 88, and NF-κB p 65 proteins and in inhibiting TLR 4/NF-κB signaling in the synovial tissue of the ankle joint.

Effects of icariin on acute lung injury in mice induced by activation of the complement alternative pathway / 中国药理学通报

Aim To explore the protective effect and mechanism of icariin ( ICA ) on acute lung injury ( ALI) in mice induced by activation of the comple-ment alternative pathway. Methods 32 healthy KM mice were randomly divided into four groups: the nor-mal group, the model group, the PDTC group and the Icariin group, which received 7-day intragastric admin-istration respectively. Then cobra venom factor ( CVF) was used to activate specifically complement alternative pathway to induce acute lung injury in mice by intrave-nous injection. Myeloperoxidase ( MPO ) activity of lung homogenate, the cell count and the protein con- tent of bronchoalveolar lavage fluid ( BALF ) were measured. The concentration of IL-6, TNF-α, P-selec-tin and ICAM-1 in BALF and serum were determined by ELISA. The pathological change of lung tissue was observed by HE staining. The phosphorylation of NF-κB p65 in lung tissues was checked by immunohisto-chemistry. The effect of the transcriptional activity of NF-κB signal pathway in microvascular endothelial cells was measured by employing dual-luciferase re-porter assay system. Results ICA reduced MPO ac-tivity of lung homogenate, the cell count and the con-tent of IL-6, TNF-α, P-selectin in BALF obviously. The level of TNF-α, P-selectin and ICAM-1 in serum was decreased, the pulmonary inflammatory cell infil-tration was reduced, the phosphorylation of NF-κB p65 in lung was inhibited significantly and the transcrip-tional activity of NF-κB was also down-regulated. Con-clusion ICA can alleviate acute inflammatory re-sponse of ALI mice induced by activation of the com-plement alternative pathway. The mechanism may be highly related to the inhibition of inflammatory cell in-filtration in lung tissue, the down-regulation of phos-phorylation of NF-κB p65 and nuclear transcriptional activity.

Sal B attenuates rat hepatocytes injury induced by hypoxia/reoxygenation via modulation of SIRT1/NF-κB/p53 pathway / 中国药理学通报

Aim To investigate the effect of salvianolic acid B(Sal B) on the attenuation of rat hepatocyte in-jury induced by hypoxia/reoxygenation(H/R) and its possible molecular mechanism. Methods Rat hepato-cytes BRL-3A were cultured in vitro. H/R injury mod-el was established and then BRL-3A cells were pretrea-ted with Sal B. The viability of cells was measured by CCK-8 assay;the expression of ALT and AST was de-tected by microplate assay; the levels of TNF-α and IL-1β were determined by ELISA; the apoptosis was detected by flow cytometry;the protein and mRNA lev-els of SIRT1, NF-κB p65, p53, Bax and Bcl-2 were measured by Western blot and qPCR. Results H/R intervention decreased the viability and increased the apoptosis of cells;the production of ALT, AST, TNF-α and IL-1β was elevated;the protein and mRNA lev-els of SIRT1, Bcl-2 were reduced, but the levels of NF-κB p65, p53 and Bax increased. After pretreated with Sal B, the viability of cells increased while the apoptosis decreased; the expression of ALT, AST, TNF-α and IL-1β was inhibited;moreover,the protein and mRNA levels of SIRT1,Bcl-2 were enhanced,and the levels of NF-κB p65, p53 and Bax decreased sig-nificantly. Conclusion Sal B may attenuate rat hepa-tocyte injury induced by H/R via the SIRT1/NF-κB/p53 pathway.

Study on the Effects of Maggot Oil on the Healing of Rat with Acute Skin Trauma and Infection and Its Mechanism / 中国药房

OBJECTIVE:To study the effects of maggot oil on the healing of rat with acute skin trauma and infection and its mechanism,in order and to provide reference for further development of maggot oil. METHODS:SD rats were randomly divided into normal group,model group,maggot oil group and Jingwanhong treatment group (positive control),with 70 mice in each group. Except for normal group,acute skin trauma and infection model was induced in other groups by smearing Staphylococus aureus suspension at the wound. After modeling,normal group and model group were given normal saline,and maggot oil group and Jingwanhong treatment group were given relevant medicine 0.3 mL/100 g,at 9:00 and 17:00,for consecutive 15 d. Wound and wound healing time of rats were observed in each group. The content of hydroxyproline in wound was determined in 10 rats of each group after 1,2,4,6,8 d of administration. The content of water in wound was determined after 2,4,6,8 d after administration. 2 h after last administration,the content of lysozyme,the levels of inflammatory factors (TNF-α,IL-6),the expression of NF-κB p65(in cytoplasm and nucleus)and p-IκB-α(in cytoplasm)protein were determined in 10 rats of each group. RESULTS:Compared with normal group,wound edema of model group was obvious,and wound healing time was(14.3±2.1)d. After 4,6,8 d of medication,the content of hydroxyproline in wound of rats was decreased significantly in model group (P<0.05 or P<0.01). After 2,4,6,8 d of medication,the content of water in wound was increased significantly in model group(P<0.01). After 15 d of medication,the serum contents of lysozyme,TNF-α and IL-6 in rats were increased significantly in model group (P<0.01). The expression of NF-κB p65 (in cytoplasm) in wound was decreased significantly (P<0.01),while the expressions of NF-κB p65 (in nucleus) and p-IκB-α(in nucleus) protein were increased significantly (P<0.01). Compared with model group,above indicators of administration groups were improved significantly (P<0.05 or P<0.01). CONCLUSIONS:Maggot oil could protect tissue injury induced by acute skin wound infection,promote wound healing. The possible mechanism might play anti-inflammatory effect through promoting collagen production,increasing lysozyme content,regulating NF-κB signal pathway.