Protective effect of breviscapine against brain injury induced by intrauterine inflammation in preterm rats and its mechanism / 中国当代儿科杂志

OBJECTIVES@#To study the protective effect of breviscapine against brain injury induced by intrauterine inflammation in preterm rats and its mechanism.@*METHODS@#A preterm rat model of brain injury caused by intrauterine inflammation was prepared by intraperitoneal injections of lipopolysaccharide in pregnant rats. The pregnant rats and preterm rats were respectively randomly divided into 5 groups: control, model, low-dose breviscapine (45 mg/kg), high-dose breviscapine (90 mg/kg), and high-dose breviscapine (90 mg/kg)+ML385 [a nuclear factor erythroid 2-related factor 2 (Nrf2) inhibitor, 30 mg/kg] (n=10 each). The number and body weight of the live offspring rats were measured for each group. Hematoxylin-eosin staining was used to observe the pathological morphology of the uterus and placenta of pregnant rats and the pathological morphology of the brain tissue of offspring rats. Immunofluorescent staining was used to measure the co-expression of ionized calcium binding adaptor molecule-1 (IBA-1) and nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) in the cerebral cortex of offspring rats. ELISA was used to measure the levels of interleukin-6 (IL-6), interleukin-8 (IL-8), and interleukin-1β (IL-1β) in the brain tissue of offspring rats. Western blotting was used to measure the expression of Nrf2 pathway-related proteins in the brain tissue of offspring rats.@*RESULTS@#Pathological injury was found in the uterus, and placenta tissue of the pregnant rats and the brain tissue of the offspring rats, and severe microglia pyroptosis occurred in the cerebral cortex of the offspring rats in the model group. Compared with the control group, the model group had significant reductions in the number and body weight of the live offspring rats and the protein expression levels of Nrf2 and heme oxygenase-1 (HO-1) in the brain tissue of the offspring rats (P<0.05), but significant increases in the relative fluorescence intensity of the co-expression of IBA-1 and NLRP3, the levels of the inflammatory factors IL-6, IL-8, and IL-1β, and the protein expression levels of NLRP3 and caspase-1 in the brain tissue of the offspring rats (P<0.05). Compared with the model group, the breviscapine administration groups showed alleviated pathological injury of the uterus and placenta tissue of the pregnant rats and the brain tissue of the offspring rats, significant increases in the number and body weight of the live offspring rats and the protein expression levels of Nrf2 and HO-1 in the brain tissue of the offspring rats (P<0.05), and significant reductions in the relative fluorescence intensity of the co-expression of IBA-1 and NLRP3, the levels of the inflammatory factors IL-6, IL-8, and IL-1β, and the protein expression levels of NLRP3 and caspase-1 in the brain tissue of the offspring rats (P<0.05). The high-dose breviscapine group had a significantly better effect than the low-dose breviscapine (P<0.05). ML385 significantly inhibited the intervention effect of high-dose breviscapine (P<0.05).@*CONCLUSIONS@#Breviscapine can inhibit inflammatory response in brain tissue of preterm rats caused by intrauterine inflammation by activating the Nrf2 pathway, and it can also inhibit microglial pyroptosis and alleviate brain injury.

Effect of Electroacupuncture on Pain Transition and Content of Protein Kinase Cε in Dorsal Root Ganglia in Hyperalgesia Rats / 针刺研究

OBJECTIVE: To observe the effect of electroacupuncture (EA)on mechanical pain transition and content of protein kinase C epsilon(PKCε)in dorsal root ganglia (DRG) in inflammatory articular pain rats，so as to explore its peripheral mechanism underlying relieving transition from acute to chronic pain. METHODS: 1)In the first part of the present study，male SD rats were equally randomized into blank control，sham hyperalgesic priming(HP), and real HP groups(n＝6 in each). The HP model was established by subcutaneous injection of 1% carrageenan (100 µL) into the left hind paw (the first injection)，followed by injection of PGE 2 (100 ng/25 µL, the second injection) into the dorsum pedis of the same hind paw 7 days after the first injection. The mechanical withdrawal threshold (MWT) of the ipsilateral paw was detected before and 4, 24, 48, 72 h, and 7 d after the first injection，and 1, 4, 24 and 48 h after the second injection. 2) In the second part，SD rats were randomly divided into sham-HP，real HP，sham-EA and EA groups(n＝6 in each). The sham-HP and HP models were made in the same way as those in the first part. Bilateral "Zusanli"(ST 36)and "Kunlun"(BL 60)were punctured with filiform needles and also stimulated with electrical current：2 Hz/100 Hz，0.5－1.5 mA(0.5 mA increase per 10 min)for 30 min，1 time/d from the 1st carrageenan injection on till the end of the experiments. PKCε protein expression in the L 4－L 6 DRGs was assayed by Western blot 48 h after the second injection. RESULTS: 1)In the first part of the study，compared with the sham-HP group，the MWT at 4, 24、48 h after carrageenan injection and 4, 24 and 48 h after PGE 2 injection were significantly decreased in the HP model group(P<0.01). 2)In the second part，compared with the HP group，the MWT at 24、48 and 72 h after carrageenan injection, and 24 and 48 h after PGE 2 injection were significantly up-regulated in the EA group(P<0.05，P<0.01). 3)The relative content of PKCε in the DRGs(L 4－L 6)was significantly higher in the HP group than in the sham-HP group(P<0.01)，but considerably lower in the EA group than in the HP group (P<0.01)．. CONCLUSION: EA has a good effect on pain conversion in inflammatory joint pain rats，which may be related to its effect in down-regulating the PKCε level in the ipsilateral lumbar DRGs.

Inhibition of TNF-α/IFN-γ induced RANTES expression in HaCaT cell by naringin.

CONTEXT: Naringin is a bioflavonoid derivative and is predominantly found in Citrus paradisi Macf., Citrus sinensis (Linn.) Osbeck, Citrus unshiu Marc., Citrus reticulata Blanco cv. Nobilis, Citrus tachibana (Makino) Tanaka, Citrus junos Sieb. ex Tanaka (Rutaceae), and related citrus species. It has anti-inflammatory effects that have been well-documented, but the mechanism is poorly characterized. OBJECTIVE: The effect of naringin on production of RANTES (regulated upon activation normal T-cell expressed and secreted) in human HaCaT cells was investigated here for the first time. MATERIALS AND METHODS: The HaCaT cells were cultured in Dulbecco's modified Eagle's medium (DMEM) and the proliferation of cell was determined by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT). The cells were divided into three groups including control group, tumor necrosis factor alpha (TNF-α)/interferon gamma (IFN-γ)-stimulated group, and naringin pretreatment group (first incubated in the presence of naringin and then exposed to TNF-α/IFN-γ). The concentration of RANTES in the supernatants was determined by enzyme-linked immunosorbent assay (ELISA). The expression of RANTES mRNA was analyzed by reverse transcription-polymerase chain reaction (RT-PCR). The expression of nuclear factor kappa B (NF-κB) P65 protein was detected with immunocytochemical method and western blot method. RESULTS: Naringin hardly inhibits HaCaT cells growth at concentrations rising from 0.25 to 1 mmol/L. However, RANTES expression detected in supernatant stimulated with TNF-α/IFN-γ reduced 15 and 16%, respectively, when cultured with 0.25, 0.5 mmol/L naringin. Furthermore, 1 mmol/L naringin significantly decreased RANTES mRNA level. Finally, naringin decreased the expression of NF-κB P65 protein in nuclei. DISCUSSION AND CONCLUSION: Naringin can inhibit the increased production of RANTES, which is partially via NF-κB-dependent signal pathway.

Mechanisms of gross saponins of Tribulus terrestris via activating PKCepsilon against myocardial apoptosis induced by oxidative stress / 药学学报

This study is to observe the effect of gross saponins of Tribulus terrestris (GSTT) on protein kinase Cepsilon (PKCepsilon) and apoptosis-associated protein in the apoptosis of cultured cardiocyte apoptosis induced by hydrogen peroxide (H2O2), and to explore the mechanisms of GSTT against myocardial apoptosis. Primary cardiocytes were isolated and cultured. Myocardial apoptosis was induced by H2O2 and analyzed with flow cytometry. Protein content of phospho-PKCepsilon, Bcl-2, and Bax were detected with Western blotting analysis. Cleaved caspase-3 protein content was determined with immunocytochemical technique. After the pretreatment of 100 mg x L(-1) GSTT, compared with H2O2 group, GSTT could not only decrease the apoptotic percentage in cardiocytes damaged by H2O2 (P < 0.01), but also reduce protein contents of Bax and cleaved caspase-3 (P < 0.01), and increase protein content of phospho-PKCepsilon and Bcl-2 significantly (P < 0.01). PKC inhibitor chelerythrine (Che) could prevent partly the effect of GSTT against myocardial apoptosis (P < 0.05 and P < 0.01). Mechanisms of GSTT against myocardial apoptosis might be associated with inhibition of mitochondrial apoptosis pathway after PKCepsilon activation.