Middle-dose Group Research Articles

Effects of lactational exposure to low-dose BaP on allergic and non-allergic immune responses in mice offspring

Benzo[a]pyrene (BaP) can induce developmental and reproductive toxicity; however, the full scope of its immunotoxic effects remains unknown. This study aimed to assess effects of lactational exposure to low-dose BaP (comparable to human exposure) on potential allergic\ on-allergic immune responses in murine offspring. Lactating C3H/HeJ dams were orally dosed with BaP at 0, 0.25, 5.0, or 100 pmol/animal/week) at post-natal days [PND] 1, 8, and 15. Five-weeks-old pups then received intratracheally ovalbumin (OVA) every 2 weeks for 6 weeks. Following the final exposure, mice were processed to permit analyses of bronchoalveolar lavage (BAL) fluid cell profiles as well as levels of lung inflammatory cytokines and chemokines, serum OVA-specific immunoglobulin, and mediastinal lymph node (MLN) cell activation/proliferation. In OVA-sensitized male offspring, lactational low-dose BaP exposure led to enhanced (albeit not significantly) macrophage, neutrophil, and eosinophil infiltration to, and increased T-helper (TH)-2 cytokine production in, the lungs. In females, BaP exposure, regardless of dose, led to slightly enhanced lung levels of macrophages and eosinophils, and of inflammatory molecules. Protein levels of interleukin (IL)-33 in the OVA + BaP (middle dose) group, and interferon (IFN)-γ in the OVA + BaP (low dose) group, were higher than that of the OVA (no BaP) group. Ex vivo studies showed lactational exposure to BaP partially induced activation of T-cells and antigen-presenting cells (APCs) in the MLN cells of both male and female offspring, with or without OVA sensitization. Further, IL-4 and IFNγ levels in MLN culture supernatants were elevated even without OVA-re-stimulation in OVA + BaP groups. In conclusion, lactational exposure to low-dose BaP appeared to exert slight effects on later allergic and non-allergic immune responses in offspring by facilitating development of modest TH2 responses and activating MLN cells. In addition, lactational exposures to BaP might give rise to gender differences in allergic/non-allergic immune responses of offspring.

Scutellaria baicalensis Attenuates Airway Remodeling via PI3K/Akt/NF-κB Pathway in Cigarette Smoke Mediated-COPD Rats Model.

Background. Scutellaria baicalensis (SB) is commonly used in traditional Chinese medicine for chronic inflammatory diseases. This study aims to investigate the effects of the early intervention with SB on airway remodeling in a well-established rat model of COPD induced by cigarette smoking. Methods. COPD model in Sprague Dawley (SD) rats were established by exposing them to smoke for 6 days/week, for 12 weeks, 24 weeks, or 36 weeks. Meanwhile, rats were randomly divided into normal control group, model group, Budesonide (BUD) group, and the SB (low, middle, and high) dose groups with 8 rats in each group and 3 stages (12 weeks, 24 weeks, and 36 weeks). After treatment, the pulmonary function was evaluated by BUXCO system and the morphology changes of the lungs were observed with HE and Masson staining. The serum IL-6, IL-8, and IL-10 and TNF-α, TGF-beta (TGF-β1), MMP-2, MMP-9, and TIMP-1 levels in BALF were detected by ELISA-kit assay. The protein expression levels of AKT and NF-κB (p65) were determined by western blot (WB). Results. The oral of SB significantly improved pulmonary function (PF) and ameliorated the pathological damage and attenuated inflammatory cytokines infiltration into the lungs. Meanwhile, the levels of TGF-β, MMP-2, MMP-9, and TIMP-1 were partially significantly decreased. The levels of PI3K/AKT/NF-κB pathway were also markedly suppressed by SB. Conclusions. SB could significantly improve the condition of airway remodeling by inhibiting airway inflammation and partially quenching TGF-β and MMPs via PI3K/AKT/NF-κB pathway.

Protective Effect of Valsartan on Podocyte Injury in Rats with Diabetic Nephropathy

To investigate the mechanism of valsartan protecting podocytes and inhibiting renal injury in diabetic rats. The rat model of diabetic nephropathy was induced by combination of valsartan and high-sugar and high-fat diet. The urinary protein content, renal index, inflammatory and antioxidant indexes in the kidney, renal pathological changes and podocyte holes were investigated. Membrane WT1 and P-Cadherin protein expression levels. Compared with the model group, the 24h urine protein content of valsartan was significantly decreased (P<0.05). Valsartan significantly inhibited the body weight of the model group (P<0.01), and significantly inhibited The increase of renal index (P<0.05); the high and middle doses of valsartan could significantly reduce the levels of IL-β, TNF-α and IL-6 in rat kidney (P<0.05-0.01). The valsartan high and middle dose groups significantly reduced MDA content in rat kidney (P<0.05), and significantly increased SOD activity (P<0.05). HE and PAS staining showed that valsartan was used in model group rats. The pathological changes were alleviated, and the glomerular morphology returned to normal. The protein expression of WT1 and P-Cadherin in the kidney of DN rats by Western blot showed that P- in the kidney tissue of valsartan rats. The expression levels of Cadherin and WT1 protein were significantly increased (P<0.05). Valsartan can regulate the expression of podocyte membrane proteins WT1 and P-Cadherin to protect podocytes, and then repair renal function and anti-diabetic nephropathy.

The optimal decoction time of Yinqiao Powder on inhibit influenza A virus FM1 in mice

Background: Yinqiao Powder (YQP) is administered in Traditional Chinese Medicine (TCM) for febrile infectious diseases and has been demonstrated to fight the prevalence of influenza. Objective: The present study aimed to identify the optimal decoction time of YQP on inhibit influenza A virus FM1 and investigate the molecular mechanisms of different decocting time YQP. Methods: The male BALB/c mice were made into pneumonia model of influenza A virus FM1. They were randomly assigned to twelve groups: blank control group, model control group, tamiflu (27.5 mg/Kg) group, 3 minutes YQP (High-dose group 30 g/Kg·d, Middle-dose group 15 g/Kg·d, and Low-dose group 7.5 g/Kg·d) groups, 6 minutes YQP (High-dose group 30 g/Kg·d, Middle-dose group 15 g/Kg·d, and Low-dose group 7.5 g/Kg·d) groups, and 12 minutes YQP (High-dose group 30 g/Kg·d, Middle-dose group 15 g/Kg·d, and Low-dose group 7.5 g/Kg·d) groups. All groups were administrated for 7 days. The mice were killed after the third and last days. Viral load of lung tissues were observed by Real-time PCR. Pulmonary index and inhibition rate were calculated by formula. Furthermore, the mice macrophage Ana-1 was infected by FM1, treated with different decocting time YQP drug-containing serum for 24 h. TLR3/4, MyD88, TRAF-6, TRAM and TRIF mRNA were detected by Real-time PCR. TLR3/4 proteins were observed by western blot. Results: All YQP intervention groups induced a reduction in the viral load and pulmonary index on the third and last days of infection, especially in 3 minutes YQP H/M/L groups and 6 minutes High-dose group. Meanwhile, YQP intervention groups led to significant decrease in TLR3/4, MyD88, TRAF-6, TRAM and TRIF mRNA levels and TLR3/4 proteins, in particular with 3 and 6 minutes YQP groups. Conclusion: YQP could protect the lung tissue and alleviate the inflammation caused by influenza A virus FM1. The optimal decoction time of YQP was 3to 6minutes on inhibit influenza A virus FM1 in mice.

Protective Effect of Norcantharidin on Collagen-Induced Arthritis Rats.

To observe the effect of norcantharidin (NCTD) on collagen-induced arthritis (CIA) rats. Sixty Sprague-Dawley(SD) rats were randomly divided into 6 groups (n=10): normal group, CIA model group(model group), NCTD low-dose group [1.35 mg/(kg•d)], NCTD middle-dose group [2.7 mg/(kg•d)], NCTD high-dose group [5.4 mg/(kg•d)] and methotrexate (MTX) group [1.8 mg/(kg/w)]. Anesthetized rats were sacrificed by luxation of cervical vertebra after 4 weeks of administration. The arthritis scores were evaluated twice a week. The pathological changes in the ankle joints of rats were observed by hematoxylin-eosin (H&E) staining. The serum levels of interleukin (IL) 1β, IL-6, tumor necrosis factor (TNF)-α, vascular endothelial growth factor (VEGF), IL-17 and transform growth factor (TGF) β were detected by enzyme linked immunosorbent assay (ELISA). The mRNA expression of retinoid-related orphan nuclear receptorγt (RORγt) and forkhead box P3 (Foxp3) in peripheral blood lymphocytes were confirmed by real-time polymerase chain reaction. MTX and high-dose NCTD not only decreased the arthritis scores but also alleviated the pathological changes in CIA rats' ankle joints compared with the model group (P<0.05 or P<0.01). All doses of NCTD significantly inhibited the serum levels of IL-6, IL-17 and TNF-α in CIA rats (P<0.05). Only middle- and high-dose of NCTD prominently decreased serum IL-1β and TGF-β levels of CIA rats (P<0.05). However, NCTD has no effect on vascular endothelial growth factor (VEGF) level in CIA rats. The Foxp3 mRNA expression in all NCTD groups were increased significantly than in the model group (P<0.05). The mRNA expression of RORγt in NCTD high-dose group was decreased apparently in comparison with the model group (P<0.05). NCTD showed therapeutic effect on CIA rats by inhibition of cytokines and regulation of Th17/Treg cells.

Tangshen formula improves inflammation in renal tissue of diabetic nephropathy through SIRT1/NF-κB pathway.

The present study investigated the mechanism underlying the anti-inflammatory effects of Tangshen formula (TS) in Sprague Dawley (SD) rats with diabetic nephropathy (DN). A rat model of DN was established by intraperitoneal injection of 1% (40 mg/kg) streptozotocin and administration of a high fat and glucose diet. Subsequently, SD rats were randomly divided into six groups (n=8): A DN group, a valsartan group, a high-dose TS group, a middle-dose TS group, a low-dose TS group and a control group with normal SD rats. Once rats received their allocated treatment for 12 weeks, body weight and kidney weight were recorded, and fasting blood glucose, ratio of urinary protein, β2-MG and creatinine clearance rate were determined. Furthermore, hemodynamic indices, including plasma viscosity and whole blood reduction viscosity were detected. Immunohistochemistry was used to detect the infiltration of macrophages in the kidneys of rats. Reverse transcription-quantitative polymerase chain reaction and western blotting were performed to investigate the activation; mRNA and protein expression levels of monocyte chemoattractant protein-1 (MCP-1), macrophage migration inhibitory factor (MIF), nuclear factor-κB (NF-κB) and sirtuin-1 (SIRT1) in each group. In comparison with the DN group, each biochemical indicator of rats in the high-dose TS group was significantly decreased (P<0.05). Blood viscosity in each treatment group was significantly decreased when compared with the DN group (P<0.01). Hematoxylin and eosin staining indicated that the infiltration of macrophages was significantly decreased in the high-dose TS group when compared with the DN group (P<0.01). mRNA and protein expression levels of MCP-1 and MIF in the high-dose TS group were significantly decreased when compared with the DN group (P<0.05). In the treatment groups, SITR1 mRNA expression levels were significantly increased, whereas the mRNA expression levels of NF-κB were significantly decreased (P<0.01). Western blotting results indicated a significant decrease in the protein expression levels of acetylated NF-κB in the treatment groups when compared with the DN group (P<0.01) and the propensity of protein expression of the other inflammatory factors were consistent with the mRNA findings. The results of the high-dose TS group were similar to those of the valsartan group. The present study indicates that TS was able to activate SITR1, which lead to NF-κB deacetylation, thus reducing the release of inflammatory factors and decreasing the severity of diabetic nephropathy.

Phase I clinical trial of a novel autologous modified-DC vaccine in patients with resected NSCLC

BackgroundThe primary aim of this study was to evaluate the safety of a novel dendritic cell (DC) vaccine pulsed with survivin and MUC1, silenced with suppressor of cytokine signaling 1 (SOCS1), and immune stimulated with flagellin for patients with stage I to IIIA non-small cell lung cancer (NSCLC) in a phase I open-label, uncontrolled, and dose-escalation trial. Moreover, we evaluate the potential efficacy of this modified DC vaccine as secondary aim.MethodsThe patients were treated with the vaccine at 1 × 106, 1 × 107and the maximum dose 8 × 107 at day 7, 14, and 21 after characterization of the vaccine phenotype by flow cytometry. The safety of the vaccine was assessed by adverse events, and the efficacy by the levels of several specific tumor markers and the patient quality of life.ResultsThe vaccine was well tolerated without dose-limiting toxicity even at higher doses. The most common adverse event reported was just grade 1 flu-like symptoms without unanticipated or serious adverse event. A significant decrease in CD3 + CD4 + CD25 + Foxp3+ T regulatory (Treg) cell number and increase in TNF-α and IL-6 were observed in two patients. Two patients showed 15% and 64% decrease in carcino-embryonic antigen and CYFRA21, respectively. The vaccination with the maximum dose significantly improved the patients’quality of life when administered at the highest dose. More importantly, in the long-term follow-up until February 17, 2017, 1 patient had no recurrence, 1 patients had a progressive disease (PD), and 1 patient was died in the low dose group. In the middle dose group, all 3 patients had no recurrence. In the high dose group, 1 patient was died, 1 patient had a PD, and the other 7 patients had no recurrence.ConclusionsWe provide preliminary data on the safety and efficacy profile of a novel vaccine against non-small cell lung cancer, which was reasonably well tolerated, induced modest antitumor activity without dose-limiting toxicity, and improved patients’ quality of life. Further more, the vaccine maybe a very efficacious treatment for patients with resected NSCLC to prevent recurrence. Our findings on the safety and efficacy of the vaccine in this phase I trial warrant future phase II/III clinical trial.

Effects of Pogostemon cablin on gastrointestinal function of rats with syndrome of damp retention in middle-jiao

To investigate the effects of Pogostemon cablin(patchouli) on gastrointestinal function of rats with the syndrome of damp retention in middle-jiao, and explore its therapeutic mechanism. In this study, gastrointestinal function of rats with the syndrome of damp retention in middle-jiao was evaluated by multiple assays including gastric remnant rate, small intestine propelling rate, gastric juice quantity, pepsin activity and gastrointestinal tissue morphology. ELISA was used to detect gastrointestinal hormones including MTL, GAS, VIP and cytokines including TNF-α and interleukin 10 in rat serum. Real-time fluorescent quantitative PCR technique was used to detect relative mRNA expression of AQP3, AQP4 and AQP8 in gastric and colonic tissues to explore the mechanism of P. cablin in treatment of gastrointestinal functions. The results showed that middle and high dose of P. cablin (3.24, 6.48 g•kg⁻¹) could obviously decrease the gastric remnant rate, promote gastric emptying, increase the small intestine propelling rate(P<0.05), speed up the propulsive movement of gastrointestinal tract, increase the secretion and acidity of gastric juice, increase the activity of pepsin, and improve the injury of gastrointestinal tissue. All the doses of P. cablin could increase the concentration of MTL and GAS in serum, reduce the concentration of VIP, TNF-α and IL-10 in serum, decrease the mRNA expression of AQP3 in gastric and colonic tissues, and increase the expression levels of AQP4 and AQP8 in colonic tissues. The regulatory effects were better in middle and high dose groups. In conclusion, regulation of the levels of gastrointestinal hormones, inflammatory cytokines and aquaporins may be the paths for P. cablin to maintain normal gastrointestinal function of rats with the syndrome of damp retention in middle-jiao. The results of the study laid a foundation for clarifying the treatment mechanism of aromatic damp-resolving drugs for indications including damp retention in middle-jiao and transformation failure of spleen.

Mechanism of triptolide in treating ankylosing spondylitis through the anti‑ossification effect of the BMP/Smad signaling pathway.

The present study aimed to examine the mechanism of triptolide in the treatment of ankylosing spondylitis (AS) through the anti‑ossification effect of bone morphogenetic protein (BMP)/small mothers against decapentaplegic (Smad) pathway. Male rats were randomly divided into five groups: Normal rat group; model group; triptolide low dose group (10mg/kg); triptolide middle dose group (20mg/kg); triptolide high dose group (40mg/kg). The spinal joint capsules of each group of rats were collected to perform primary cell culture to determine the levels of cell proliferation. Western blot and reverse transcription‑polymerase chain reaction analyses, and ELISA were used to detect the mRNA and protein expression levels of core‑binding factor α1 (Cbfal), BMP receptor type II (BMPRII), Smad1, Smad4, Smad5 and Smad6, the protein expression of phosphorylation indicators, including phosphorylated (p)Smad1 and pSmad5, the mRNA expression of tumor necrosis factor‑α (TNF‑α), interleukin (IL)‑1β and IL‑6 in rat plasma, and the mRNA expression of BMP/Smads in fibroblasts induced by recombinant human (rh)BMP‑2. The effects on AS in the rats were also examined. The results revealed that, following intervention with triptolide, inflammation was suppressed, and the mRNA expression levels of TNF‑α, IL‑1β and IL‑6 were reduced. The expression levels of Cbfal, BMPRII, Smad1, Smad4 and Smad5 were also reduced, whereas the expression of Smad6 was increased. Following induction by rhBMP‑2, the effects of triptolide weakened, with the most marked effects observed at the highest concentration, suggesting that triptolide functions through the BMP/Smad signaling pathway. Taken together, the results suggested that triptolide may be used to treat AS, the mechanism of which may be through the BMP/Smad pathway.

Metabonomics analysis of serum from rats given long-term and low-level cadmium by ultra-performance liquid chromatography–mass spectrometry

1. This study evaluated the toxicity of chronic exposure to low-level cadmium (Cd) in rats using ultra-performance liquid chromatography–mass spectrometry (UPLC–MS). Forty male Sprague–Dawley rats were randomly assigned to four groups, namely, the control group, low-dose group (0.13 mg/kg·bw), middle-dose group (0.8 mg/kg·bw) and high-dose group (4.89 mg/kg·bw). The rats continuously received CdCl2 via drinking water for 24 weeks. Serum samples were collected for metabonomics analysis. The data generated from the UPLC–MS was analysed using principal components analysis (PCA) and partial least-squares discriminant analysis (PLS-DA). PLS-DA model with satisfactory explanatory and predictive ability is capable of discriminating the treatment groups from the control group.2. Finally, the 10 metabolites were identified and showed significant changes in some treatment groups compared with that in the control group (p < 0.0167 or p < 0.003). Exposure to Cd resulted in increased intensities of lysophosphatidic acid (P-16:0e/0:0), glycocholic acid, bicyclo-prostaglandin E2, lithocholyltaurine, sulfolithocholylglycine, lysophosphatidylethanolamine (20:5/0:0) and lysophosphatidylcholine (20:0), as well as decreased intensities of 3-indolepropionic acid, phosphatidylcholine (18:4/18:0) and 15S-hydroxyeicosatrienoic acid in rat serum.3. Results suggest that exposure to Cd can cause disturbances in the lipid metabolism, amino acid metabolism, nervous system, antioxidant defence system, liver and kidney function.

Preharvest Ultraviolet C Irradiation Increased the Level of Polyphenol Accumulation and Flavonoid Pathway Gene Expression in Strawberry Fruit.

Preharvest ultraviolet C (UV-C) irradiation is an innovative approach for increasing the bioactive phytochemical content of strawberries to increase the disease resistance and nutritional value. This study investigated the changes in individual flavonoids in strawberry developed with three different cumulative doses of preharvest UV-C treatment (low, 9.6 kJ m-2; middle, 15 kJ m-2; and high , 29.4 kJ m-2). Significant accumulation (p < 0.05) of phenolics (25-75% increase), namely, cyanidin 3-glucoside, pelargonidin 3-glucoside/rutinoside, glucoside and glucuronide of quercetin and kaempferol, and ellagic acid, was found in the fruit subjected to low and middle supplemental doses of UV-C radiation. The expression of the flavonoid pathway structural genes, i.e., FaCHS1, FaCHI, FaFHT, FaDFR, FaFLS, and FaFGT, was upregulated in the low- and middle-dose groups, while the early stage genes were not affected by the high dose. FaMYB1 was also relatively enhanced in the low- and middle-dose groups, while FaASR was upregulated in only the low-dose group. Hormetic preharvest UV-C dose ranges for enhancing the polyphenol content of strawberries were established for the first time.

Effect of Niuhuang-Xingnao pill on apoptosis in hippocampus of Alzheimer's disease model mice

Objective To study the the treatment effects of Niuhuang-Xingnao pill on Alzheimer’s diseases mice and its influence on the levels of Bcl-2, caspase-3 and C-caspase-3. Methods 50 APP/V717 mice were divided into the positive control group, the model group, Niuhuang-Xingnao pill high, medium and low dosage groups. 10 C57BL/6 mice were selected as blank control. The high, medium and low dosage groups were given 142, 71 and 35.5 mg/kg Niuhuang-Xingnao pill, the positive control group was given 10 mg/kg donepezil, the blank group and model group were given same volume normal salin. 1 time/day, and continuous administration for 60 d. Morris water maze experiment was performed to test the learning and memory ability in mice. The levels of CAT, SOD and GSH-Px were detected by the method of ELISA, while Bcl-2, caspase-3 and C-caspase-3 in the groups were detected by western-blotting. The apoptosis of hippocampus of mice were observed by the method of TUNEL. Results Compared with the model group, the third square time (36.58 ± 4.57 s, 32.46 ± 4.25 s, 29.71 ± 4.26 s vs. 25.48 ± 3.91 s) of high, middle and low dose groups were significantly longer, cross time of the table (5.82 ± 0.87, 4.59 ± 0.76, 3.96 ± 0.75 vs. 3.27 ± 0.53) were significantly higher (P<0.05). In the high, middle and low dose groups, the levels of GSH-Px (161.14 ± 14.01 U/mg, 150.76 ± 13.16 U/mg, 143.17 ± 12.54 U/mg vs. 120.78 ± 10.92 U/mg), SOD (14.25 ± 1.82 U/mg, 11.17 ± 1.65 U/mg, 7.24 ± 1.02 U/mg vs. 3.12 ± 0.31 U/mg), CAT (17.95 ± 2.16 U/mg, 16.72 ± 1.83 U/mg, 15.54 ± 1.47 U/mg vs. 13.25 ± 2.60 U/mg) were significantly higher (P<0.05); caspase-3 (1.13 ± 0.13, 1.25 ± 0.15, 1.41 ± 0.17 vs. 1.49 ± 0.20), C-caspase-3 (1.17 ± 0.14, 1.27 ± 0.16, 1.42 ± 0.18 vs. 1.52 ± 0.23) significantly lower(P<0.05), Bcl-2 (0.88 ± 0.08, 0.79 ± 0.06, 0.67 ± 0.04 vs. 0.59 ± 0.04) significantly higher(P<0.05). Conclusions Niuhuang-Xingnao pill treatment of Alzheimer's disease in mice can effectively promote the expression of Bcl-2, caspase-3 and C-caspase-3 in the hippocampus of mice, inhibit the apoptosis of the cells in hippocampus. Key words: Alzheimer disease; Apoptosis; Niuhuang-Xingnao pill; Mice

Preliminary study of intervention in effect of Bushen Huoxue recipe on calcification of lumbar vertebra cartilage endplate of the aging gerbils

To study the effect of Bushen Huoxue decoction on calcification of cartilage endplate in lumbar vertebrae. Six healthy male gerbils with 2-month-old were selected as normal control group, and 24 7-month-old healthy male gerbils were fed to 12-month-old to establish the aged gerbil model. Thirty gerbils were randomly divided into five groups as follow: the normal control group (n=6), model group (n=6, normal saline 4 ml/kg, intragastric 30 d), Bushen Huoxue low dose group(n=6, 1.9× 10⁻ ³ ml/g given Bushen Huoxue recipe orally, 30 d), Bushen Huoxue middle dose group(n=6, 3.8× 10⁻ ³ ml/g given Bushen Huoxue recipe orally, 30 d), Bushen Huoxue high dose group(n=6, 7.6× 10⁻ ³ ml/g given Bushen Huoxue recipe orally, 30 d), the intervention group administered for 1.36 g from 7-month-old age, 30 d. The animals were sacrificed at the age of 2 months in the normal control group and 12 months of age in the other groups. The morphology of the lumbar vertebral cartilage endplate, the area of vascular bud, the ratio of non-calcified/calcified layer were analysis by HE chromosome visual method. The expression of type X collagen and BMPs in cartilage endplates were detected by rabbit monoclonal immunohistochemical staining. The relative area of the vascular buds cartilage endplate measurements showed that compared with the model group, middle dose group and normal control group increased (P<0.05), high and low dose groups all had different degrees of increase, but no statistical significance(P>0.05). The ratio of cartilage endplate thickness of non-calcified/calcified showed that compared with the model group, Bushen Huoxue middle dose, normal control group increased, with statistical significance(P<0.05), and high and low dose groups all had different degrees of increase, but there were no statistical significance(P>0.05). Compared with the model group, the expression of type X collagen in the cartilage endplate of the normal group, the Bushen Huoxue low, middle and high dose groups decreased, and had statistical significance(P<0.01); compared with the model group, the expression of BMPs in the normal group, Bushen Huoxue middle dose group increased, with statistically significant(P<0.01), while the high and low dose groups increased in different degrees, but there was no statistical significance(P>0.05). Bushen Huoxue prescription can delay the calcification of cartilage endplate in the process of aging, suggesting that it can be used as a preventive medicine for early disc degeneration.

DPPH Radical Scavenging and Postprandial Hyperglycemia Inhibition Activities and Flavonoid Composition Analysis of Hawk Tea by UPLC-DAD and UPLC-Q/TOF MSE.

Hawk tea (Litsea coreana Lévl. var. Lanuginosa (Migo) Yen C. Yang & P.H. Huang), a very popular herbal tea material, has attracted more and more attention due to its high antioxidant properties and possible therapeutic effect on type II diabetes mellitus. The raw materials of Hawk tea are usually divided into three kinds: bud tea (BT), primary leaf tea (PLT) and mature leaf tea (MLT). In this study, the DPPH radical scavenging activity and the antimicrobial properties of these three kinds of Hawk tea from different regions were comparatively investigated, and a ultra-high performance liquid chromatographic coupled with a photodiode array detector (UPLC-DAD) method was employed for comparison of the three major flavonoid constituents, including hyperoside, isoquercitrin and astragalin, in different samples of Hawk tea. At the same time, the effect of methanol extract (ME) of PLT on the mouse postprandial blood glucose and the effect of ME and its different fractions (petroleum ether fraction (PE), ethyl acetate fraction (EA), n-butanol fraction (n-BuOH), and water fraction (WF)) on the activity of α-glucosidase were studied. The results showed that Hawk BT and Hawk PLT possessed the higher radicals scavenging activity than Hawk MLT, while the antibacterial activity against P. vulgaris of PLT and MLT was higher than Hawk BT. The contents of the three major flavonoid constituents in samples of Hawk PLT are higher than Hawk BT and Hawk MLT. The mouse postprandial blood glucose levels of the middle dose (0.5 g/kg) group and the high dose (1 g/kg) group with oral administration of the ME of PLT were significantly lower than the control group. What’s more, the inhibitory effect of ME of PLT and its EA and n-BuOH fractions on α-glucosidase was significantly higher than that of acarbose. Rapid ultra-high performance liquid chromatography/quadrupole time-of-flight-mass spectrometry (UPLC-ESI-QTOF-MS) was used to identify the flavonoids in Hawk PLT, and a total of 20 flavonoids were identified or tentatively identified by comparing their retention times and accurate mass measurements with reference compounds or literature data. The bioactive flavonoid composition and DPPH radical scavenging activities present in different Hawk tea raw materials are quite different due to the different ontogenesis of these raw materials. Further studies on PLT showed that the substances in PLT ME could reduce the level of mouse postprandial blood glucose through inhibiting the activity of α-glucosidase.

Experimental study on the inhibitory effect of epigallocatechin gallate on renal calcium oxalate crystal formation

Objective To compare the effects of different dosages of epigallocatechin gallate (EGCG) in calcium oxalate stone rate, to expolre content differences in malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT) and calcium oxalate crystal deposition. Methods Fifty rats were randomly divided into 5 groups: normal control group, calcium oxalate kidney stone model group and different doses of EGCG intervention group. The control group was permitted to drink purified water freely; model group was given 2 ml stone liquid that was prepared with 1% ethylene glycol and 2% ammonium chloride by the lavage. In the EGCG intervention groups, daily doses of 0.1% EGCG (10, 30, and 50 mg/kg) were given introgastrically on the basis treatment in the model group. Twenty-eight days later, the rats were sacrificed and the right kidney was prepared into homogenate. The thiobarbituric acid (TBA) method was used for the determination of MDA. The xanthine oxidase method was applied to detect SOD activity. CAT activity was tested by ultraviolet spectrophotometry. The DTNB method was used to determine GSH-Px activity. The left kidney tissue was stained by HE method, and crystal deposition was observed by polarizing microscopy. Results In the blank control group, MDA concentration was (2.74±0.47) μmol/L, SOD activity (541.71±10.61) Nu/g, CAT activity (44.49±3.45) U/g and GSH-Px activity (41.61±2.50)×103 U/g·protein. As compared with the blank control group, MDA concentration of calcium oxalate kidney stones in the model group was increased significantly, while the activity of SOD (332.44±12.01) Nu/g, CAT (32.15±1.99) U/g and GSH-Px (21.36±1.58)×103 U/g·protein was significantly decreased (P=0.009). In the low-dose EGCG group, the activity of SOD, CAT and GSH-Px was (517.41±28.37) Nu/g, (34.42±2.55) U/g and (23.94±2.95)×103 U/g·protein respectively, that in the middle-dose EGCG group (545.04±23.84), (35.42±2.61) and (25.10±2.64) respectively, and that in the high-dose EGCG group (550.58±16.55) Nu/g, (35.40±2.39) U/g and (26.67±1.93)×103 U/g·protein respectively. As compared with the model group, SOD, GSH-Px and CAT actively was increased in the EGCG groups (P=0.005). The calcium oxalate crystal deposition in the renal tissue was negatively correlated with the dose of EGCG interference (r=-0.483, P= 0.008). Conclusion There were significant differences in MDA, SOD and GSH-Px contents in calcium oxalate stone rats treated with different doses of EGCG, which indicated that EGCG could reduce the formation of calcium oxalate kidney stones by reducing the oxidative damage. Key words: Calcium oxalate; Epigallocatechin gallate; Oxidative damage

Intervention effects of Dan-fang capsule in rats with hepatic fibrosis

Objective: To investigate the interventional effect of Dan-fang capsule on liver fibrosis in rats. Methods: Sixty one-week aged male healthy SD rats [weight (180±20) g] were randomly divided into normal control group (group A), hepatic fibrosis model group (group B), Fu-Fang-Bie-Jia-Ruan-Gan tablet group (group C), Dan-fang capsule groups at high, middle and low dose group (group D, E, F, respectively). Except for the normal control group, hepatic fibrosis was induced in other groups by intraperitoneal injection of porcine serum.Simultaneously, rats in Dan-fang capsule groups were administered by gavage with Dan-fang capsule at doses of 4.32, 2.16, 0.54 g/kg, respectively.Rats in Fu-Fang-Bie-Jia-Ruan-Gan tablet group were orally administered by gavage with Fu-Fang-Bie-Jia-Ruan-Gan tablet (0.54 g/kg) every day and the normal control group received saline alone.All rats were killed at the end of the 12th week. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) and PⅢnp clia (PⅢNP) were measured in the groups.Pathology changes of hepatic tissue were evaluated by hematoxylin-eosin (HE) and Masson staining.The proteinic expressions of alpha-smooth muscle actin (α-SMA), collagen-Ⅰ (COL-Ⅰ) and collagen-Ⅲ (COL-Ⅲ) were observed with the method of immunohistochemistry.Analysis of variance was applied when data were compared among groups. Results: Compared with those in the group A, the levels of ALT, AST and PⅢNP in serum and the expressions of α-SMA, COL-Ⅰ and COL-Ⅲ in liver tissues were significantly higher in group B [(68.3±3.4) vs (51.5±6.3) U/L, (205±52) vs (135±24) U/L, (3.1±1.4) vs (1.6±0.6) μg/L and 0.35±0.02 vs 0.13±0.02, 0.37±0.02 vs 0.13±0.02, 0.43±0.13 vs 0.13±0.01, t=17.020, 71.053, 1.552, 0.214, 0.241, 0.292, all P<0.01], and the degree of liver fibrosis significantly increased in group B than that in group A. Compared with those in group B, the levels of ALT, AST, PⅢNP and the expressions of α-SMA, COL-Ⅰ, COL-Ⅲ were all significantly lower in group D, E and F (t=-58.232--0.104, all P<0.01). The degree of liver fibrosis significantly reduced in group D, E and F than that in group B (Z=3.82, 3.76, 3.90, all P<0.05). Conclusion: Dan-fang capsule has certain preventive effect on liver fibrosis that caused by porcine serum in rats.

Effect of docosahexenoic acid microalgae oil supplementation during pregnancy and lactation on levels of serum polyunsaturated fatty acid and tolerance to ovalbumin in suckling mice

Objective: To study the tolerance to ovalbumin (OVA) in suckling mice whose mothers had different doses of docosahexenoic acid (DHA) microalgae oil (DMO) supplementation during pregnancy and lactation. Method: According to different doses of DMO fed to mother mice during pregnancy and lactation, 66 suckling mice were divided into four groups. Suckling mice whose mothers were fed with 0.7% DMO were designated as low dose group (group L) (n=16), 2.1% DMO as middle dose group (group M) (n=16), 3.5% DMO as high dose group (group H) (n=17) and no DMO as control group (n=17). Before exposing to OVA, 8 suckling mice were killed in each group at 21-day-old. Remaining suckling mice were killed at 59-day-old after repeated OVA exposure. The serum polyunsaturated fatty acid (PUFA) levels of suckling mice were analyzed by high performance liquid chromatography (HPLC) at the age of 21- and 59-day.Histological examinations of jejunum were performed by HE staining and the mast cells in jejunum were observed by toluidine blue staining. OVA-IgE in serum, total IgA and OVA-IgA in the feces and IL-4 and IFN-γ in the supernatants of splenic mononuclear cells (SMC) were measured by ELISA. Real time PCR was performed to identify the gene expression of IL-10, TGF-β1 mRNA in SMC. Differences among groups were compared by one-way AVOVA and that between each group were compared by LSD. Result: In group M and H, the serum levels of n-3DHA (108±29)μg/ml; (102±34)μg/ml vs.(40±19)μg/ml (F=12.052, P=0.000)and n-3 eicosapentaenoic acid (6.7±2.3)μg/ml; (7.7±2.0)μg/ml vs. (3.9±1.1)μg/ml(F=9.573, P=0.000) were significantly higher than that in control group at the age of 21-day. The serum levels of n-3DHA were higher in group H (17.1±2.9)μg/ml than that in control group (5.9±3.3) μg/ml after repeated OVA exposure at the age of 59-day (F=10.339, P<0.000). Compared with control group (53±12) pg/ml, the levels of IL-4 in SMC in group H (42±9)pg/ml were lower (F=2.484, P<0.05). Conclusion: The serum levels of DHA in baby mice, whose mothers was fed with DMO during pregnancy and lactation, were significantly increased till adulthood. However, the effect on tolerance to OVA was limited.

Bortezomib could down-regulate the expression of RANKL, inhibit cell proliferation and induce cell apoptosis in the human myeloma cell line RPMI 8226 by activating casepase-3.

In spite of bortezomib being developed and demonstrated as a safe drug therapy for multiple myeloma (MM), the role of bortezomib-induced receptor activator of nuclear factor (NF)-κB ligand (RANKL) in the MM cell lines remains to be understood. Thus the present study aims to explore the impact of bortezomib on RANKL expression, cell growth and apoptosis in human myeloma cell line RPMI 8226. Four experiment groups were set according to different concentrations of bortezomib, namely blank group (treated with DMEM solution free of other drugs), low-dose group (treated with 10 nmol/L bortezomib), middle-dose group (treated with 20 nmol/L bortezomib) and high-dose group (treated with 40 nmol/L bortezomib). Western blotting was adopted to detect RANKL protein expression. MTT assay was performed to detect cell proliferation. Flow cytometry was used to analyze cell cycle and apoptosis. Spectrophotometry was applied to determine caspases-3 activity. Compared with the blank group, the RANKL protein expression, cell number at the S stage was reduced while cell inhibition rate, cell apoptosis rate and caspase-3 activity enhanced remarkably in the low-dose, middle-dose and high-dose groups with dose-dependent manner. Compared with those treated with bortezomib (20 nmol/L and 40 nmol/L) for 6 h, the RANKL expression was down-regulated, cell inhibition rate was increased, cells at the S stage were reduced, cell apoptosis rate was enhanced, and caspase-3 activity elevated in the RPMI 8226 cells as treated with bortezomib for 24 h, with a dose- and time-dependent manner. Bortezomib could reduce the RANKL expression, inhibit cell proliferation and activate caspase-3 activity to induce cell apoptosis in RPMI 8266 cells.

Isolation of an Anti-fatigue Preparation from Cellulase-treated Maca by Hydroalcoholic Extraction

The purpose of this study was to evaluate the anti-fatigue activity of maca hydroalcoholic extract (ME), which mainly contains macamides and polysaccharides. ME was prepared by circumfluence extraction with enzymatic pre-treatment. Anti-fatigue activity of ME was investigated in weight-loaded forced swimming mice, with pure macamides and commercially available maca tablet as positive control. Compared with normal group, pure macamides treatment group could prolong the swimming time to exhaustion, but there was no statistically significant difference (P &gt; 0.05); while ME (middle-dose and high-dose groups) could effectively prolong the swimming durations (P &lt; 0.05). Supplementation with pure macamides significantly decreased blood lactic acid (BLA), whereas ME significantly increased hepatic glycogen (HG), decreased BLA, and blood urea nitrogen (BUN) compared with those in normal control (P &lt; 0.05). The results suggested that the anti-fatigue effect of ME was better than that of pure macamides, which can be explained by the increase of glycogen storage and the reduction of metabolites accumulation.

Cardiotoxicity and Mechanism of Particulate Matter 2.5 (PM2.5) Exposure in Offspring Rats During Pregnancy

BackgroundThe aim of this study was to investigate the cardiotoxicity and mechanism of particulate matter 2.5 (PM2.5) exposure on offspring rats during pregnancy.Material/MethodsWistar rats were used to establish a PM2.5 exposure animal model during pregnancy, and they were divided into a control group, a low-dose group, a middle-dose group, and a high-dose group according to PM2.5 exposure dose. The pathological changes of heart tissue, the rate of myocardial cell apoptosis, the levels of LDH, AST, and CM-KB in serum, and the difference in mitochondrial fusion genes (OPA1 and Mfn1) and mitochondrial genes (Drp1 and Fis1) were compared among groups.ResultsThe arrangement of myocardial fibers in offspring mice of PM2.5 exposure groups became disordered, the shape of some cardiomyocytes became irregular, and some staining darker nuclei appeared. The apoptotic rates of myocardium in offspring rats exposed to PM2.5 were (12.61±0.93)% in the low-dose group, (25.14±1.53)% in the middle-dose group, and (30.13±1.50)% in the high-dose group, which were all significantly higher than in the control group (9.12±0.80)% (P<0.05). The levels of LDH, AST, and CM-KB and the expression of OPA1, Mfn1, Drp1, and Fis1 in offspring mice of PM2.5 exposure groups increased with the increase of PM2.5 exposure dose, and were significantly higher than that of the control group (P<0.05).ConclusionsThe mitochondria of the offspring mice were damaged due to the abnormal expression of mitochondrial fusion/splicing gene by PM2.5 exposure during pregnancy, and the hearts of offspring mice were damaged due to damaged mitochondria.