ABSTRACT
ObjectiveTo investigate the effect of Chuanshanlong granule on Toll-like receptor 4 （TLR4）/myeloid differentiation protein 88 （MyD88）/nuclear transcription factor -κB （NF-κB） signaling pathway， and to explore the mechanism of its treatment of autoimmune thyroiditis （AIT） in rats. MethodForty AIT models were established following excess iodine and injection of porcine thyroglobulin and Freund's adjuvant into Lewis rats for six weeks. Then the rats were randomly divided into the model group， Chuanshanlong granule low-， medium- and high-dose group （0.52， 1.03， 2.06 g·kg-1·d-1）， with ten in each group. Rats in the Chuanshanlong granule low-， medium- and high-dose groups were separately given 0.01 mL·g-1·d-1 Chuanshanlong granule， and those in the normal group and the model group were given the same volume of deionized water for eight weeks. Serum of rats was taken to measure thyroglobulin antibody （TgAb） and thyroid peroxidase antibody （TPOAb） by enzyme-linked immunosorbent assay （ELISA）， and the concentrations of free triiodothyronine （FT3）， free thyroxine （FT4） and thyroid stimulating hormone （TSH） were detected. The rat thyroid lobes were stained with hematoxylin and eosin （HE）， and the pathological changes were observed under light microscope. In addition， the relative expression of TLR4， MyD88， NF-κB protein and mRNA was determined by immunohistochemistry and real-time polymerase chain reaction （Real-time PCR）. ResultCompared with the conditions in the normal group， the serum concentrations of TPOAb and TgAb （P<0.01） and FT3 and FT4 （P<0.01） increased and TSH decreased （P<0.01） in the model group. Compared with the conditions in the model group， the concentrations of TPOAb and TgAb in the Chuanshanlong granule treatment groups reduced （P<0.01）， and the concentrations of FT3 and FT4 were lowered （P<0.01） while TSH increased （P<0.01） in the Chuanshanlong granule high-dose group. HE staining showed that there was lymphocyte infiltration in the thyroid follicular space， a large number of destroyed or diminished follicular cavities， decreased colloid content， and thinned or destroyed follicular wall in the model group， while the thyroid lymphocyte infiltration in the Chuanshanlong granule treatment groups was significantly less and the structure of thyroid follicles was more complete than those in the model group. Compared with the normal group， the model group had up-regulated relative expression of TLR4， MyD88 and NF-κB protein （P<0.01） and mRNA （P<0.01）. Compared with the model group， the Chuanshanlong granule high-dose group had down-regulated relative expression of TLR4 protein and mRNA （P<0.05）， MyD88 protein （P<0.01） and mRNA （P<0.05）， and NF-κB protein and mRNA （P<0.01）. ConclusionChuanshanlong granule may play a therapeutic role in AIT by inhibiting the activation of TLR4/MyD88/NF-κB signaling pathway.
ABSTRACT
ObjectiveTo investigate the intervention effect of total glucosides of paeony （TGP） on the renal injury of MRL/lpr mice based on the Toll-like receptor 9 （TLR9）/myeloid differentiation factor 88 （MyD88）/nuclear transcription factor-κB （NF-κB） signaling pathway and explore the immunological mechanism of TGP in preventing and treating systemic lupus erythematosus （SLE）. MethodMRL/lpr female mice of SPF grade were randomly divided into a model group， a dexamethasone group （0.15 g·kg-1）， and high- （0.078 g·kg-1） and low-dose （0.039 g·kg-1） TGP groups， and female C57BL/6J mice were assigned to a blank group， with 7 mice in each group. Mice in each group were treated with corresponding drugs or normal saline by gavage at the same time every day. After 4 weeks， samples were collected. The kidney and spleen were weighed， and the organ index was calculated. Serum creatinine （SCr） and blood urea nitrogen （BUN） levels in each group were detected by biochemical assay. Hematoxylin-eosin （HE） staining was used to observe the histopathological changes in the kidney. The degree of renal fibrosis was evaluated by Masson staining. The serum levels of interleukin （IL）-2， interferon （IFN）-α， IL-4， and anti-nuclear antibody （ANA） were detected by enzyme-linked immunosorbent assay （ELISA）. The mRNA expression of TLR9， MyD88， and NF-κB p65 in renal tissues was detected by real-time quantitative polymerase chain reaction （Real-time PCR）. The protein expression of TLR9 and NF-κB p65 in renal tissues was detected by immunofluorescence. The protein expression of TLR9， MyD88， and NF-κB p65 in renal and spleen tissues was tested by Western blot. ResultCompared with the blank group， the model group showed increased SCr， BUN， spleen index， and kidney index （P<0.05）， deteriorated pathological injury and fibrosis in renal tissues， elevated serum levels of IFN-α， IL-4， and ANA， decreased level of IL-2 （P<0.05）， and up-regulated TLR9， MyD88， and NF-κB p65 mRNA and protein levels in the kidney and spleen （P<0.05）. Compared with the model group， the TGP groups displayed reduced SCr， BUN， spleen index， and kidney index （P<0.05）， relieved pathological damage and fibrosis in renal tissues， decreased serum levels of IFN-α， IL-4， and ANA （P<0.05）， increased level of IL-2， and declining mRNA and protein expression levels of TLR9， MyD88， and NF-κB p65 in the kidney and spleen （P<0.05）. ConclusionTGP may inhibit the expression of downstream inflammatory factors to regulate immunity and resist SLE-induced renal injury by regulating the TLR9/MyD88/NF-κB signaling pathway.
ABSTRACT
ObjectiveTo study the effect of Xianlian Jiedu prescription （XLJDP） on the activation of nuclear transcription factor-κB （NF-κB） signaling pathway induced by bromodomain-containing protein 4 （Brd4） in hypoxic microenvironment and to explore its mechanism in inhibiting the proliferation of colorectal cancer HT-29 cells. MethodThe human colorectal cancer HT-29 cells were cultured in a hypoxic incubator or normoxia incubator and treated with XLJDP at 0.8，1，1.2，1.6，3.2，6.4，and 12.8 g·L-1 for 48 h， respectively. Following the detection of cell vitality using methyl thiazolyl tetrazolium （MTT） colorimetry， the effects of XLJDP （1.25，2.5，and 5 g·L-1） on the cell mitochondrial membrane potential were determined using a fluorescent probe （JC-1）， and the apoptosis of colorectal cancer HT-29 cells was detected by flow cytometry. The cell colony formation assay and 5-ethynyl-2'-deoxyuridine （EDU） staining were conducted to test the proliferation of colorectal cancer HT-29 cells. The Western blot was carried out to measure the expression levels of Brd4 and its downstream relevant proteins such as c-Myc and hexamethylene bisacetamide-inducible protein 1 （HEXIM1）， as well as the effects of XLJDP on related proteins in the NF-κB signaling pathway. ResultCompared with the blank control group， XLJDP at 0.8，1，1.2，1.6，3.2，6.4，and 12.8 g·L-1 inhibited the vitality of colorectal cancer HT-29 cells （P<0.05 ， P<0.01）， with the median inhibitory concentration （IC50） under the hypoxic condition higher than that under the normoxia condition. Compared with the blank control group， XLJDP at 1.25，2.5，and 5 g·L-1 significantly decreased the mitochondria membrane potential， enhanced the apoptosis （P<0.05，P<0.01）， and lowered the number of cell colonies and also the EDU-positive cells （P<0.05， P<0.01）. The results of Western blot showed that compared with the blank control group， XLJDP at 1.25，2.5，and 5 g·L-1 down-regulated Brd4， c-Myc， p-NF-κB p65， and p-IκBα protein expression to varying degrees and up-regulated the expression of HEXIM1 （P<0.05， P<0.01）. ConclusionIn the hypoxic microenvironment， XLJDP inhibits the proliferation of colorectal cancer HT-29 cells regulated by Brd4， which may be related to its inhibition of the activation of NF-κB signaling pathway.
ABSTRACT
Sepsis， a common critical disease in the intensive care unit（ICU）， features high morbidity and mortality. At present， it is mainly tackled with western medicine， which may trigger a series of problems like antibiotic resistance， adverse hormonal reactions， and high cost after a long-term use. Therefore， exploring new efficient， safe， and cheap drugs and treatment modes has become the focus of our research at this stage. By virtue of unique advantages including "the concept of holism and individualized treatment based on syndrome differentiation"， Chinese medicine has accumulated quite rich experience in the prevention and treatment of sepsis. In recent years， research on the regulation of Chinese medicine on nuclear transcription factor-κB（NF-κB） signaling pathway in sepsis has kept emerging. On this basis， this paper reviewed the etiology and pathogenesis of sepsis， syndrome differentiation and treatment， NF-κB signaling pathway， and its intervention with Chinese medicine. It has been found that some single Chinese herbs and their extracts， Chinese herbal compounds， and Chinese herbal injections effectively inhibit the expression of such inflammatory factors as NF-κB-mediated tumor necrosis factor-α（TNF-α）， interleukin-1（IL-1）， and IL-6 as well as the related proteins， reduce the systemic inflammatory response and organ injury， and improve the prognosis by regulating the activation of NF-κB signaling pathway and the immune function of macrophages. However， due to the limitations of objective conditions， some studies also have the problems of fuzzy pro-inflammatory anti-inflammatory balance mechanism， unclear pharmacokinetics and low drug safety evaluation， which need to be further studied and explored in order to provide a new theoretical basis and diagnosis and treatment thinking for the treatment of sepsis with traditional Chinese medicine.