N-acetylcysteine and glycyrrhizin combination:Benefit outcome in a murine model of acetaminophen-induced liver failure

BACKGROUND Acetaminophen overdose is the most frequent cause of drug-induced liver failure in developed countries.Substantial progress has been made in understanding the mechanism of hepatocellular injury,but N-acetylcysteine remains the only effective treatment despite its short therapeutic window.Thus,other hepatoprotective drugs are needed for the delayed treatment of acetaminopheninduced hepatotoxicity.Our interest focused on glycyrrhizin for its role as an inhibitor of high mobility group box 1(HMGB1)protein,a member of the family of damage-associated molecular pattern,known to play an important pathological role in various diseases.AIM To investigate the efficacy of the N-acetylcysteine/glycyrrhizin combination compared to N-acetylcysteine alone in the prevention of liver toxicity.METHODS Eight-week-old C57BL/6J wild-type female mice were used for all our experiments.Mice fasted for 15 h were treated with acetaminophen(500 mg/kg)or vehicle(phosphate-buffered saline)by intraperitoneal injection and separated into the following groups:Glycyrrhizin(200 mg/kg);N-acetylcysteine(150 mg/kg);and N-acetylcysteine/glycyrrhizin.In all groups,mice were sacrificed 12 h following acetaminophen administration.The assessment of hepatotoxicity was performed by measuring plasma levels of alanine aminotransferase,aspartate aminotransferase and lactate dehydrogenase.Hepatotoxicity was also evaluated by histological examination of hematoxylin and eosin-stained tissues sections.Survival rates were compared between various groups using Kaplan-Meier curves.RESULTS Consistent with data published in the literature,we confirmed that intraperitoneal administration of acetaminophen(500 mg/kg)in mice induced severe liver injury as evidenced by increases in alanine aminotransferase,aspartate aminotransferase and lactate dehydrogenase but also by liver necrosis score.Glycyrrhizin administration was shown to reduce the release of HMGB1 and significantly decreased the severity of liver injury.Thus,the co-administration of glycyrrhizin and N-acetylcysteine was investigated.Administered concomitantly with acetaminophen,the combination significantly reduced the severity of liver injury.Delayed administration of the combination of drugs,2 h or 6 h after acetaminophen,also induced a significant decrease in hepatocyte necrosis compared to mice treated with N-acetylcysteine alone.In addition,administration of N-acetylcysteine/glycyrrhizin combination was associated with an improved survival rate compared to mice treated with only N-acetylcysteine.CONCLUSION We demonstrate that,compared to N-acetylcysteine alone,co-administration of glycyrrhizin decreases the liver necrosis score and improves survival in a murine model of acetaminophen-induced liver injury.Our study opens a potential new therapeutic pathway in the prevention of acetaminophen hepatotoxicity.

Topical application of glycyrrhizin preparation ameliorates experimentally induced colitis in rats

AIM:To examine the efficacy of glycyrrhizin preparation(GL-p) in the treatment of a rat model of ulcerative colitis(UC).METHODS:Experimental colitis was induced by oral administration of dextran sodium sulfate.Rats with colitis were intrarectally administered GL-p or saline.The extent of colitis was evaluated based on body weight gain,colon wet weight,and macroscopic damage score.The expression levels of pro-inflammatory cytokines and chemokines in the inflamed mucosa were measured by cytokine antibody array analysis.The effect of GL-p on myeloperoxidase(MPO) activity in the inflamed mucosa and purified enzyme was assayed.RESULTS:GL-p treatment significantly ameliorated the extent of colitis compared to sham treatment with saline.Cytokine antibody array analysis showed that GL-p treatment significantly decreased the expression levels of pro-inflammatory cytokines and chemokines,including interleukin(IL)-1β,IL-6,tumor necrosis factor-α,cytokine-induced neutrophil chemoattractant-2,and monocyte chemoattractant protein-1 in the inflamed mucosa.Furthermore,GL-p inhibited the oxidative activity of mucosal and purified MPO.CONCLUSION:GL-p enema has a therapeutic effect on experimental colitis in rats and may be useful in the treatment of UC.

Glycyrrhizin attenuates HMGB1-induced hepatocyte apoptosis by inhibiting the p38-dependent mitochondrial pathway

AIM： To examine how High-mobility group box I （HMGB1） regulates hepatocyte apoptosis and, furthermore, to determine whether glycyrrhizin （GL）, a known HMGB1 inhibitor, prevents HMGBl-induced hepatocyte apoptosis.

Effect of alprostadil combined with Diammonium glycyrrhizinate on renal interstitial fibrosis in SD rats

Objective:To observe effect of alprostadil combined with Diammonium glycyrrhizinate on renal interstitial fibrosis in SD rate.Methods:A total of 75 SD rate were randomly divided into A,B,C,D,E groups with 15 in each group.Rats in group A served as the control group received just only but tissue separation without modeling operation,while model of unilateral ureteral obstruction(UUO) was established in B,C,D,E groups.Rats in A,B group were given saline lavage placebo treatment,while rats in C,D,E groups were given dianunonium glycyrrhizinate and alprostadil injection.Five rats were sacrificed 1,2,3 weeks after modeling,serum creatinine level of femoral venous blood was determined.Transforming growth factor- β1(TCF- β1) and concentration of connective tissue growth factor(CTGF) were also detected by using ELISA.Line renal interstitial tissue was taken after HE staining,renal interstitial TGF- β1 and CTGF expression were detected by using immunohistochemical method.Results:Serum creatinine levels of B,C,D,E group at different time points in were significantly higher than that of group A(P<0.05);serum creatinine levels in group B were significantly higher than that of C,D,E group at each time point(P<0.05).Serum creatinine level of Croup E was significantly lower than C,D group after 2,3 weeks(P<0.05).Rate in A group at each time point showed no significant changes in TGF- β1 and CREA concentration in serum and kidney tissues(P>0.05);while serum and kidney tissue TGF- β1,concentration of CREA.expression of rats in B,C,D,E groups showed a gradual increasing trend over time.TCF- β1 and CREF of Group B in serum and kidney tissues at each time point were significantly higher than that of the other groups(P<0.05).TCF- β1 and CREF of Group E in serum and kidney tissues at each time point were significantly lower than that of B,C,D group at all time points in serum and kidney tissues(P<0.05).Conclusions:Alprostadil combined with diammonium glycyrrhizinate can significantly lower the expression of TGF- β1 and CTGF in serum and tissues of SD rat with renal interstitial fibrosis,thus inhibit rat renal interstitial fibrosis process.It has synergy protective effect.

Anti-inflammatory effect of Diammonium Glycyrrhizinate in a rat model of ulcerative colitis

AIM： To explore the anti-inflammatory mechanism of Diammonium Glycyrrhizinate in a rat model of ulcerative colitis induced by acetic acid. METHODS： Spragur-Dawley female rats were divided into four groups： Diammonium Glycyrrhizinate group, dexamethasone group, acetic acid control and normal control group. Colonic inflammation was evaluated by disease activity index, gross morphologic damage, histological injury and colonic myeloperoxidase activity. Immunohistochemistry was used to detect the expression of NF-κB, TNF-α and ICAM-1 in colonic mucosa. RESULTS： Compared to the acetic acid control, both Diammonium Glycyrrhizinate and dexamethasone showed a significant anti-inflammatory effect （P 〈 0.01）. The expression of NF-κB, TNF-α and ICAM-1 in colonic mucosa was significantly lower in the Diammonium Glycyrrhizinate group and dexamethasone group than in the acetic acid group. CONCLUSION： Diammonium Glycyrrhizinate could reduce inflammatory injury in a rat model of ulcerative colitis. This may occur via suppression of NF-κB, TNF-αand ICAM-1 in colonic mucosa.

Glycyrrhizinate reduces portal hypertension in isolated perfused rat livers with chronic hepatitis

AIM:To investigate the effects of diammonium glycyrrhizinate(Gly)on portal hypertension(PHT)in isolated portal perfused rat liver(IPPRL)with carbon tetrachloride(CCl4)-induced chronic hepatitis.METHODS:PHT model was replicated with CCl4 in rats for 84 d.Model was identified by measuring the ascetic amounts,hepatic function,portal pressure in vivo,splenic index,and pathological alterations.Inducible nitric oxide synthase(iNOS)in liver was assessed by immunohistochemistry.IPPRLs were performed at d0,d28,d56,and d84.After phenylephrine-induced constriction,Gly was geometrically used to reduce PHT.Gly action was expressed as median effective concentration(EC50)and area under the curve(AUC).Underlying mechanism was exploited by linear correlation between AUC values of Gly and existed iNOS in portal triads.RESULTS:PHT model was confirmed with ascites,splenomegaly,serum biomarkers of hepatic injury,and elevated portal pressure.Pathological findings had shown normal hepatic structure at d0,degenerations at d28,fibrosis at d56,cirrhosis at d84in PHT rats.Pseudo lobule ratios decreased and collagen ratios increased progressively along with PHT development.Gly does dose-dependently reduce PHT in IPPRLs with CCl4-induced chronic hepatitis.Gly potencies were increased gradually along with PHT development,characterized with its EC50at 2.80×10-10,3.03×10-11,3.77×10-11and 4.65×10-11mol/L at d0,d28,d56and d84,respectively.Existed iNOS was located at hepatocyte at d0,stellate cells at d28,stellate cells and macrophages at d56,and macrophages in portal triads at d84.Macrophages infiltrated more into portal triads and expressed more iNOS along with PHT development.AUC values of Gly were positively correlated with existed iNOS levels in portal triads.CONCLUSION:Gly reduces indirectly PHT in IPPRL with CCl4-induced chronic hepatitis.The underlying mechanisms may relate to rescue NO bioavailability from macrophage-derived peroxynitrite in portal triads.

Effects of Dietary β-Glucan and Glycyrrhizin on Non-Specific Immunity and Disease Resistance of the Sea Cucumber (Apostichopus japonicus Selenka) Challenged with Vibrio splendidus

Sea cucumbers, Apostichopus japonicus Selenka, were fed diets containing non-immunostimulant (basal diet), 0.2% β-glucan and 0.02% glycyrrhizin in a recirculatory water system for 45 days, and subsequently challenged with Vibrio splendidus by injection at 1.0×108 cfu/sea cucumber for 15 days.Phagocytic capacity (PC), intracellular superoxide anion production (ISAP), ly-sozyme (LSZ) activity and superoxide dismutase (SOD) activity in the coelomic fluid were analyzed on the 0th, 5th, 10th and 15th days after injection.Results showed that after the 45-day feeding period, PC, ISAP, LSZ activity and SOD activity in sea cucumbers fed with dietary β-glucan or glycyrrhizin were significantly higher than in those fed with the basal diet.On the 5th day after infection, all the immune parameters examined in the sea cucumbers injected with V.splendidus decreased in value significantly.On the 15th day, PC, ISAP and LSZ activity returned to levels similar to those on the 0th day.For the sea cucumbers injected with saline, there were no significant differences in all the immune parameters examined and in the cumulative morbidity during the 15-day challenging trial.After injecting with V.splendidus, the cumulative morbidity of sea cucumbers fed with the basal diet was significantly higher than those fed with dietary β-glucan or glycyrrhizin when challenged with V.splendidus challenged sea cucumber fed with the basal diet was significantly higher than those fed with dietary β-glucan or glycyrrhizin.There was no significant difference in cumulative morbidity between the dietary β-glucan and glycyrrhizin treatments over time.

Uptake of albumin nanoparticle surface modified with glycyrrhizin by primary cultured rat hepatocytes

AIM: To investigate the uptake difference between bovine serum albumin nanoparticle (BSA-NP) and bovine serum albumin nanoparticles with their surface modified byglycyrrhizin (BSA-NP-GL) and to develop a novel hepatocyte targeting BSA-NP-GL based on active targeting technology mediated by specific binding site of GL on rat cellular membrane. METHODS: Calcein loaded bovine serum albumin nanoparticles (Cal-BSA-NP) were prepared by desolvation process. Glycyrrhizin was conjugated to the surface reactive amino groups (SRAG) of Cal-BSA-NP by sodium periodate oxidization, which resulted in calcein-loaded bovine serum albumin nanoparticles with their surface modified by glycyrrhizin (Cal-BSA-NP-GL). The morphology of the two types of prepared nanoparticles (NP) was observed by transmission electron microscopy. The diameter of NP was measured with a laser particle size analyzer. The interaction between Cal-BSA-NP-GL and primary cultured hepatocytes was studied through cellular uptake experiments. The uptake amount of Cal-BSA-NPGL and Cal-BSA-NP by rat hepatocytes was determinedby fluorospectrophotometry. Uptake characteristics were investigated through experiments of competitive inhibition of specific binding site of GL. RESULTS: Both Cal-BSA-NP-GL and Cal-BSA-NP had regular spherical surfaces. The average diameter of CalBSA-NP-GL and Cal-BSA-NP was 77 and 79 nm respectively. The uptake amount of the two NP by hepatocytes reached its maximum at 2 h after incubation. The uptake amount of Cal-BSA-NP-GL by rat hepatocytes was 4.43-fold higher than that of Cal-BSA-NP. There was a significant difference in the uptake of Cal-BSA-NP-GL and Cal-BSA-NP by hepatocytes (P＜0.01). The uptake of Cal-BSA-NP-GL was inhibited when GL was added previously to isolated rat hepatocytes, and the uptake of Cal-BSA-NP was not affected by GL.CONCLUSION: A binding site of GL is present on the surface of rat hepatocytes, BSA-NP-GL may be internalized via this site by hepatocytes and can be used as a drug carrier for active targeting of delivery drugs to hepatocytes.

Effect of Discontinuous Administration of β -glucan and Glycyrrhizin on the Growth and Immunity of Pacific White Shrimp Litopenaeus Vannamei

A six-week growth trial was conducted to compare the effects of different feeding strate- gies of dietary immunostimulants on the growth and immunity of white shrimp Litopenaeus vannamei （4.70 ±0.20g）. Shrimps were fed with diet containing glycyrrhizin continuously, containing β -glucan continuously, discontinuously （seven days with diet containing β -gluseven days with diet without -glucan; two days with diet containing β-glucan following five days with diet without -glucan）,

Pseudo-hyperaldosteronism caused by compound glycyrrhizin tablets: a case report

Compound glycyrrhizin tablets(CGT)is a glycyrrhizin-containing preparation for the treatment of chronic hepatic diseases,it contained Glycyrrhizin,Monoammonium Glycyrrhizinate,Aminoacetic Acid and Methionine.CGT has been shown to have anti-inflammatory,anti-oxidative,and anti-viral effects.Although pseudo-hyperaldosteronism was reported as its important side effects,the frequency was unknown.A 59-year-old male patient with tuberculous pleurisy received CGT to prevent impairment in liver function which was caused by antituberculosis drugs.After more than one month,it appeared edema in the lower extremity,blood pressure increased,serum potassium lowed,serum sodium raised at the normal high limit.Finally,it was improved by suspending the CGT and diuretic treatment.

Influence of compound glycyrrhizin on liver functions, liver fibrosis indexes and inflammatory factors of patients with chronic hepatitis B

Objective:To investigate influence of Compound Glycyrrhizin on liver functions, liver fibrosis indexes and inflammatory factors of patients with chronic hepatitis B.Methods:A total of 96 cases of patients with chronic hepatitis B treated in our hospital from Jan2015 to Jun2016 were selected as subjects, and randomly divided to be 48 cases of observation group and 48 cases of control group. Patients in both of the two groups were received routine liver protecting drug treatment. For observation group, Compound Glycyrrhizin injection was given on the basis of routine treatment. Variations of liver function indexes, liver fibrosis indexes and inflammatory factors between the two groups before and after treatment were compared and observed.Results:No obvious difference showed on AST, ALT, ALB TBIL levels between two groups of patients before treatment;After treatment, AST, ALT, TBIL in two groups of patients were significantly decreased, ALB were significantly increased. Significant difference showed comparing with prior treatment;After treatment, AST, ALT and TBIL levels in observation group were (29.53±9.44) U/L, (32.36±10.93) U/L and (10.12±3.22) μmol/L, which were significantly lower than in control group. ALB levels in observation group were (43.57±12.42) g/L, which were significantly higher than ALB levels in control group. Before treatment, no statistical difference showed on HA, LN, IV-C and PCIII levels between two groups of patients. After treatment, HA, LN, IV-C and PCIII in two groups of patients were significantly decreased, which showed significant difference comparing with prior treatment;After treatment, HA, LN, IV-C and PCIII levels in observation group were (97.33±31.75) μg/L, (77.52±23.72) μg/L, (82.92±24.55) μg/L, (15.33±5.11) μg/L, which were significantly lower than in control group. Before treatment, no significant difference showed on IL-2, IL-6 and TNF- levels between two groups of patients;After treatment, IL-2 levels in two groups of patients were significantly increased, IL-6 and TNF-α were significantly decreased, the differences showed significance;After treatment, IL-2 levels in observation group were (131.48±30.63) U/mL, which were higher than IL-2 levels in control group. IL-6 and TNF-αlevels in observation group were (45.23±16.45) μg/L, (41.75±17.53) ng/L, which were lower than IL-6 and TNF-α levels in control group, differences showed significance.Conclusion:Compound Glycyrrhizin could effectively release liver fibrosis and inflammatory reactions for patients with chronic hepatitis B, and could further improve liver functions.

甘草苷抑制吗啡引起的神经损伤作用及机制

目的:探讨甘草苷对吗啡引起神经损伤的调节作用及机制。方法:采用侧脑室注射吗啡建立神经损伤模型;腹腔注射甘草苷5 d和10 d,采用病理学观察、蛋白免疫印迹、细胞活力、凋亡及原代神经元分化检测。结果:吗啡处理之后,皮层组织中神经元减少,原代神经元细胞相对活力下降,凋亡数目增加,轴突断裂和胞体缩小;甘草苷给药后,细胞活力显著改善;轴突、树突及胞体结构逐渐完整,细胞凋亡情况减轻;蛋白Akt在473位点和PKA在197位点的磷酸化水平下降,自噬相关蛋白Becline和LC3B1/2不变。结论:甘草苷显著抑制吗啡引起的神经元分化抑制和神经元凋亡,这些作用可能是通过甘草苷协同吗啡对Akt通路。

高效液相色谱法测定化妆品中3种甘草类功效成分

建立高效液相色谱法测定化妆品中3种甘草类功效成分。样品经75%(体积分数)甲醇溶液超声提取,采用Luna C_(18)(2)100A色谱柱(250 mm×4.6 mm,5μm),以0.05%(体积分数)磷酸溶液-乙腈为流动相,梯度洗脱,流量为1.0 mL/min,进样体积为10μL,柱温为30℃,采用二极管阵列检测器,在275、251、372 nm波长下分别测定甘草素、甘草酸、甘草查尔酮A,以保留时间和紫外吸收光谱定性,外标法定量。甘草素、甘草酸、甘草查尔酮A的质量浓度分别在1~20、2~40、0.5~10μg/mL范围内与对应的色谱峰面积线性关系良好,相关系数均为0.9999。平均回收率为95.8%~111.8%,测定结果的相对标准偏差均不大于3.6%(n=6)。检出限为0.05~0.6μg/g。该方法操作简便,灵敏度高,适用于化妆品中甘草素、甘草酸、甘草查尔酮A的检测。

甘草甜素对喉癌Hep-2细胞侵袭和迁移的影响及机制研究

目的:探究甘草甜素(Gly)对喉癌Hep-2细胞侵袭和迁移的影响及作用机制。方法:体外培养正常人喉黏膜上皮细胞和喉癌Hep-2细胞,实时荧光定量PCR(RT-qPCR)法和蛋白印迹实验检测微小RNA-205-5p(miR-205-5p)、沉默信息调节因子2相关酶3(SIRT3)mRNA和蛋白表达水平。将喉癌Hep-2细胞分为对照组、Gly低浓度组、Gly中浓度组、Gly高浓度组、Gly高浓度+空载质粒组、Gly高浓度+miR-205-5p抑制剂组。各组给予相应浓度的Gly干预或转染对应质粒培养48 h。Transwell实验和划痕实验分别检测各组Hep-2细胞侵袭和迁移能力。RT-qPCR法和蛋白印迹实验检测各组Hep-2细胞miR-205-5p、SIRT3 mRNA和蛋白表达水平。双荧光素酶报告基因实验分析miR-205-5p与SIRT3的靶向关系。结果:与人喉黏膜上皮细胞比较,喉癌Hep-2细胞miR-205-5p水平降低,SIRT3 mRNA和蛋白水平升高(均P<0.05)。与对照组比较,Gly低、中、高浓度组穿膜细胞数和划痕愈合率、SIRT3 mRNA和蛋白水平依次降低,miR-205-5p水平依次升高(均P<0.05)。与Gly高浓度组和Gly高浓度+空载质粒组比较,Gly高浓度+miR-205-5p抑制剂组穿膜细胞数和划痕愈合率、SIRT3 mRNA和蛋白水平升高,miR-205-5p水平降低(均P<0.05)。miR-205-5p可靶向调控SIRT3表达。结论:Gly能够抑制Hep-2细胞侵袭和迁移,其机制可能与上调miR-205-5p表达,进而靶向下调SIRT3表达有关。

复方甘草酸苷结合硫酸羟氯喹片治疗玫瑰痤疮的临床效果分析

目的观察复方甘草酸苷结合硫酸羟氯喹片治疗玫瑰痤疮的临床效果。方法选取本院皮肤科收治的98例玫瑰痤疮患者(2022年1月~2024年1月)作为研究对象。患者均采用复方甘草酸苷结合硫酸羟氯喹片治疗,统计治疗前、治疗后总积分,评估临床疗效,并观察患者是否出现胃肠功能紊乱、皮疹等不良反应。结果98例患者治疗后主要症状(2.60±1.09分)、次要症状评分(1.91±1.21分)和总积分(4.51±1.68分)均显著低于治疗前(8.01±1.17分、5.37±1.14分)(P<0.05)。98例患者治疗后总有效率为94.90%(93/98)。患者不良反应发生率为7.14%(7/98)。结论两者联合治疗效果可靠,可有效改善患者潮红、红斑、毛细血管扩张等症状,总有效率较为理想,且不良反应风险较低,该用药方案安全可靠。

清化复肝汤联合复方甘草酸苷注射液治疗急性早幼粒细胞白血病化疗后肝损伤患者的效果

目的:观察清化复肝汤联合复方甘草酸苷注射液治疗急性早幼粒细胞白血病(APL)化疗后肝损伤患者的效果。方法:选取2021年1月至2022年1月该院收治的152例化疗后出现肝损伤的APL患者进行前瞻性研究,按照随机数字表法将其分为对照组和研究组各76例,两组均采用维A酸联合亚砷酸化疗,对照组采用复方甘草酸苷注射液进行保肝治疗,研究组在对照组基础上联合清化复肝汤治疗。比较两组临床疗效,治疗前后血清炎性因子[超敏C反应蛋白(hs-CRP)、白细胞介素-6(IL-6)]水平、肝功能指标[丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)]水平、细胞免疫指标(CD4^(+)、CD8^(+)、CD4^(+)/CD8^(+))水平,以及不良反应发生率。结果:研究组治疗总有效率为98.68%(75/76),高于对照组的86.84%(66/76),差异有统计学意义(P<0.05);治疗后,研究组血清hs-CRP、IL-6、AST、ALT、CD8^(+)水平均低于对照组,CD4^(+)、CD4^(+)/CD8^(+)水平均高于对照组,差异有统计学意义(P<0.05);两组不良反应发生率比较,差异无统计学意义(P>0.05)。结论:清化复肝汤联合复方甘草酸苷注射液治疗APL化疗后肝损伤患者效果显著,可降低ATL、AST、hs-CRP、IL-6水平,改善细胞免疫指标水平,效果优于单用复方甘草酸苷注射液。

护肝布祖热颗粒治疗药物性肝损伤的疗效观察

目的探讨护肝布祖热颗粒治疗药物性肝损伤的疗效及安全性。方法选取我院61例抗精神病药致药物性肝损伤患者,随机分为观察组(30例)和对照组(31例)。对照组在常规抗精神病用药治疗的基础上服用甘草酸二铵胶囊,观察组在常规抗精神病用药治疗基础上服用护肝布祖热颗粒,比较两组患者治疗6周后的临床疗效、肝功能及不良反应发生情况。结果观察组治疗6周后,肝功能谷丙转氨酶(ALT)、谷草转氨酶(AST)降低,指标优于对照组(P<0.05)。治疗后两组中医症候积分均有明显改善(P<0.05),且两组患者均未见明显不良反应。结论观察组在降低抗精神病药所致药物性肝损伤患者肝功能ALT、AST指标上优于对照组,起到较好的保肝护肝的作用,可有效帮助患者改善恶心呕吐、口干口苦等临床症状,且用药安全,值得临床推广应用。

甘草甜素调节TGF-β/Smad信号通路对肺癌荷瘤小鼠放射性肺损伤的影响

目的 研究甘草甜素(GL)是否可以通过调节TGF-β/Smad信号通路对肺癌荷瘤小鼠放射性肺损伤(RILI)产生影响。方法 取48只小鼠,先随机选出12只小鼠作为NC组,其余小鼠构建肺癌荷瘤小鼠模型并采用X射线照射全胸建立RILI模型,再将小鼠随机分为照射+Model组、照射+GL组(10 mg/kg)、照射+GL+SRI-011381组(10 mg/kg+30 mg/kg TGF-β激动剂)。使用酶联免疫吸附(ELISA)法检测小鼠血清中白细胞介素6(IL-6)、肿瘤坏死因子α(TNF-α)的水平;HE染色法检测小鼠肺组织病理损伤;试剂盒检测小鼠肺组织中羟脯氨酸(HYP)、髓过氧化物酶(MPO)含量;免疫组化法(IHC)检测肺组织中TGF-β、α-SMA表达;Western Blot检测小鼠肺组织中TGF-β1、p-Smad2/3、Smad7蛋白表达。结果 与NC组比较,照射+Model组小鼠血清中IL-6、TNF-α水平、HYP、MPO、TGF-β和α-SMA表达、TGF-β1、p-Smad2/3蛋白表达升高,肺组织病理损伤加重,Smad7蛋白表达降低(P<0.05);与照射+Model组比较,照射+GL组小鼠血清中IL-6、TNF-α水平、HYP、MPO、TGF-β和α-SMA表达、TGF-β1、p-Smad2/3蛋白表达减少,肺组织病理损伤减轻,Smad7蛋白表达增加(P<0.05);照射+GL+SRI-011381组中实验检测结果则与照射+GL组结果趋势相反,表明SRI-011381干预后减弱了GL对肺癌荷瘤小鼠RILI的改善作用。结论 甘草甜素可以通过抑制TGF-β/Smad信号通路对肺癌荷瘤小鼠放射性肺损伤进行修护改善。

甘草酸苷联合艾司氯胺酮对小鼠围手术期神经认知障碍的影响及其机制

目的探讨甘草酸苷联合艾司氯胺酮通过调节海马区高迁移率族蛋白B1(HMGB1)信号通路对小鼠围手术期神经认知障碍(PND)的影响及相关机制。方法将小鼠分为空白组、PND组、甘草酸苷组、艾司氯胺酮组、甘草酸苷+艾司氯胺酮组,每组24只。本实验通过Morris水迷宫评估小鼠空间学习记忆能力,各组小鼠分别于术前1 d,术后1,3,7 d行定位航行实验记录逃避潜伏期,术后第7天行空间探索实验记录目标象限穿梭次数。ELISA法检测术前1 d,术后1,3,7 d各组小鼠海马区白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)、HMGB1、Toll样受体4(TLR-4)、核因子κB(NF-κB)表达含量。免疫荧光实验检测空白组、PND组、甘草酸苷+艾司氯胺酮组术后第1天小鼠海马区HMGB1表达情况。结果各组术前1 d各指标比较差异无统计学意义(P>0.05)。Morris水迷宫结果显示:与空白组相比,其他组术后同时点逃避潜伏期均延长(P<0.05),术后第7天穿越平台次数均减少(P<0.05);与PND组相比,甘草酸苷组、艾司氯胺酮组和甘草酸苷+艾司氯胺酮组术后同时点逃避潜伏期缩短(P<0.05),术后第7天穿越平台次数增多(P<0.05);与甘草酸苷组和艾司氯胺酮组相比,甘草酸苷+艾司氯胺酮组术后同时点逃避潜伏期缩短(P<0.05),术后第7天穿越平台次数增多(P<0.05)。ELISA结果显示:与空白组相比,其余组术后同时点海马区IL-1β、TNF-α、HMGB1、TLR-4、NF-κB表达含量均明显升高(P<0.05);与PND组相比,甘草酸苷组、艾司氯胺酮组和甘草酸苷+艾司氯胺酮组术后同时点海马区IL-1β、TNF-α、HMGB1、TLR-4、NF-κB表达含量明显降低(P<0.05);与甘草酸苷组和艾司氯胺酮组相比,甘草酸苷+艾司氯胺酮组术后同时点海马区IL-1β、TNF-α、HMGB1、TLR-4、NF-κB表达含量明显降低(P<0.05)。免疫荧光结果显示:与空白组相比,PND组小鼠术后第1天海马CA1区及DG区HMGB1表达明显增多,且在海马CA1区及DG区均检测到HMGB1在细胞质中表达;与PND组相比,甘草酸苷+艾司氯胺酮组术后第1天小鼠海马CA1区及DG区HMGB1表达明显减少。结论甘草酸苷联合亚麻醉剂量艾司氯胺酮可改善小鼠围手术期神经认知功能及海马神经炎症,其机制可能与抑制HMGB1/TLR4/NF-κB信号通路相关。

高迁移率族蛋白B1对甲苯二异氰酸酯诱导的支气管上皮细胞中性粒细胞趋化因子CXCL12、IL-8表达的影响

目的通过体外实验探讨高迁移率族蛋白B1(high mobility group box 1,HMGB1)在甲苯二异氰酸酯(toluene diisocyanate,TDI)所致的职业性哮喘中趋化中性粒细胞的作用机制。方法制备TDI-人血清白蛋白(human serum albumin,HSA)偶联物(TDI-HSA),并通过Gutmann法与BCA法分别测定偶联物中TDI及HSA含量。分别以0、40、80、120 mg/L TDI-HSA染毒人支气管上皮细胞(human bronchial epithelial cells,HBECs)12 h,ELISA法检测细胞培养上清中白细胞介素-8(interleukin-8,IL-8)、趋化因子12(C-X-C motif chemokine 12,CXCL12)的水平;Western blot法检测细胞中HMGB1、CXCL12及核因子κB(nuclear factor kappa-B,NF-κB)相关蛋白的表达;活细胞荧光探针法检测活性氧(reactive oxygen species,ROS)的水平;免疫荧光法观察HMGB1的核转位情况。使用不同浓度(100、200 mmol/L)甘草素(glycyrrhizin,GL)预处理HBECs细胞24 h抑制HMGB1后,继续用TDI-HSA染毒12 h,检测细胞培养上清中IL-8水平,HMGB1、CXCL12及NF-κB相关蛋白表达,ROS释放水平及HMGB1的核转位情况。结果不同浓度TDI-HSA染毒HBECs 12 h后,随着染毒浓度的升高,HBECs细胞内HMGB1发生明显核转位,细胞中ROS和细胞上清中IL-8水平明显增加,差异有统计学意义(P<0.05);与对照组相比,120 mg/L染毒组HBECs细胞HMGB1、磷酸化P65蛋白表达显著增加,差异有统计学意义(P<0.05)。以含有不同浓度GL的培养基联合120 mg/L TDI-HSA处理HBECs细胞12 h后,与对照组相比,GL作用于HBECs细胞可显著抑制TDI-HSA导致的HMGB1的核转位,抑制HBECs细胞中的ROS和上清液中的IL-8水平的升高,显著减少细胞中HMGB1、磷酸化P65蛋白的表达,差异有统计学意义(P<0.05)。结论TDI可通过增加HBECs细胞HMGB1蛋白表达和核转位,激活NF-κB,促进IL-8的释放,不影响CXCL12的表达。GL可通过降低HMGB1表达,抑制NF-κB活化,减少IL-8释放水平。