白三烯受体拮抗剂抑制小鼠急性哮喘呼吸道炎症的研究

目的应用白三烯受体1拮抗剂孟鲁斯特(montelukast,MK)治疗小鼠急性哮喘呼吸道炎症,探讨MK对白细胞介素-5(IL-5)和嗜酸性粒细胞趋化因子(eotaxin)表达及血清总IgE水平的影响。方法卵清蛋白(OVA)致敏并激发Balb/c小鼠建立急性哮喘模型。静脉予MK(25 mg/kg)或Saline 3 d。瑞氏染色观察小鼠支气管肺泡灌洗液(BALF)炎症细胞,酶联免疫吸附法(ELISA)检测肺组织、BALF和血清IL-5和eotaxin及血清总IgE水平。半定量逆转录聚合酶链式反应(RT-PCR)检测IL-5和eotaxin的mRNA表达。液相色谱方法检测血MK水平。结果OVA激发后24 h,MK组BALF白细胞总数、嗜中性和嗜酸性粒细胞(EOS)分别为(26.0±18.9)×107/L(、5.92±8.09)×107/L和(0.74±0.88)×107/L,较Saline组相应数明显减少(P均<0.05)。EOS减少达80%以上。MK组BALF和肺组织IL-5分别为(48.52±14.45)ng/L和(27.40±9.62)ng/g蛋白,较Saline组相应值明显降低(P均<0.05);MK组血清IL-5和总IgE水平明显减低。MK组BALF中eotaxin水平也明显降低(P<0.05);MK组肺组织IL-5 mRNA显著降低,肺eotaxin及其mRNA水平无明显下降。结论MK可能通过抑制IL-5和IgE产生和分泌起抗炎作用。     

哮喘儿童不同时期血清白细胞介素-8、白细胞介素-17及痰液中性粒细胞的变化

目的 探讨白细胞介素(interleukin,IL)-8、IL-17及气道中性粒细胞在儿童哮喘发病中的作用.方法 2007年1月至2009年1月,常州市儿童医院收治的符合儿童哮喘诊断,经过诱导痰液检测的12例非嗜酸粒细胞哮喘患儿为哮喘组;以12例健康儿童为对照组.对哮喘组患儿急性期、恢复期及对照组进行诱导痰的细胞分类检查和ELISA方法检测血清IL-8和IL-17浓度.结果 哮喘急性期患儿血清中IL-8[(357.84 ±215.36) pg/ml]、IL-17[(62.76 ±44.13) pg/ml]及诱导痰的中性粒细胞百分比[(43.14±5.79)％]明显高于缓解期[(164.95 ±60.22) pg/ml、(34.57±11.82) pg/ml、(23.25±3.75)％]及对照组[(88.68±38.76) pg/ml、(20.35±10.02) pg/ml、(13.34±3.21)％],差异有统计学意义(P＜0.01);缓解期IL-8、IL-17及诱导痰的中性粒细胞百分比仍高于对照组,差异有统计学意义(P＜0.05,P＜0.01).结论 IL-8、IL-17趋化中性粒细胞聚集于气道,参与加重支气管哮喘发作.     

卡介苗干预对哮喘小鼠支气管肺泡灌洗液干细胞因子、巨噬细胞集落刺激因子表达的影响

目的 观察哮喘小鼠支气管肺泡灌洗液(BALF)干细胞因子(SCF)、巨噬细胞集落刺激因子(M-CSF)的表达,及卡介苗(BCG)干预对其表达的影响.方法 昆明小鼠30只随机分为哮喘组、BCG干预组、对照组,每组10只.以卵清蛋白(OVA)致敏激发建立哮喘模型.BCG干预组小鼠在OVA致敏前7、3、1 d以BCG皮内注射干预.末次激发24h后收集小鼠BALF,进行BALF中细胞总数及嗜酸性粒细胞(EOS)计数,酶联免疫吸附(ELISA)法检测各组BALF中SCF、M-CSF水平.结果 哮喘组小鼠BALF中细胞总数[(27.06±4.25)×107L-1]、EOS计数[(7.58±1.30)×107L-1]较对照组[(4.93±1.43)×107L-1、0]明显增高(Pa<0.01);BCG干预组小鼠BALF中细胞总数[(18.87±2.96)×107L-1]、EOS计数[(3.78±1.44)×107L-1]较哮喘组显著降低(Pa<0.01);哮喘组SCF水平[(280.25±14.20)ng/L]与对照组[(223.59±15.61)ng/L]比较显著增高(P<0.01),而BCG干预组SCF水平[(266.77±31.15)ng/L]与哮喘组比较无显著差异(P>0.05);哮喘组M-CSF水平[(204.30±10.39)ng/L]与对照组[(181.33±8.63)ng/L]比较显著增高(P<0.01),而BCG干预组M-CSF水平[(189.97±8.50)ng/L]与哮喘组比较显著下降(P<0.01).结论 致敏前多次给予BCG干预能显著抑制哮喘小鼠呼吸道炎性反应.哮喘小鼠BALF中SCF及M-CSF的表达增高,BCG干预能降低哮喘小鼠M-CSF的表达.抑制SCF或M-CSF的表达可能成为治疗哮喘的潜在靶点.     

生命早期补充维生素D对哮喘大鼠白细胞介素10和细胞间黏附分子1表达的影响

目的 研究生命早期补充维生素D对大鼠哮喘模型的影响与机制.方法选用性成熟的雌性Wistar大鼠32只.随机分为4组:对照组、低、中、高剂量组各8只.于受孕第7天起隔天以灌胃的方式给予各组大鼠不同剂量的1,25(OH)_2D_3,对照组以DMSO-PBS代替.子代大鼠离乳后,从每组中随机取出8只制备哮喘模型.HE染色肺组织切片;ELISA法检测血清及肺泡灌洗液(BALF)中IL-10浓度;免疫组化法检测细胞间黏附分子I(ICAM-1)在肺组织中的表达.结果 (1)肺组织病理变化:低剂量组和中剂量组气道炎症和嗜酸性细胞浸润均低于对照组,高剂量组则明显高于对照组.(2)血清及BALF中IL-10浓度:血清中,低剂量组[(30.2±2.8)pg/ml]和中剂量组[(51.5±6.6)pg/ml]IL-10浓度均高于对照组[(18.7±4.7)pg/ml](P<0.05),且中剂量组高于低剂量组(P<0.05),而高剂量组[(15.4±3.5)pg/ml]无明显变化(P>0.05);BALF中,与对照组[(59.1±14.4)pg/ml]相比,低剂量组[(58.1±3.4)pg/ml]无显著性变化(P>0.05),中剂量组[(90.0±14.3)pg/ml]IL-10浓度高于对照组(P<0.05),高剂量组[(45.3±6.5)pg/ml]低于对照组(P<0.05).(3)ICAM-1在支气管上皮细胞中的表达:低剂量组与对照组比较差异无统计学意义(P>0.05);中剂量组低于对照组(P<0.05);高剂量组高于对照组(P<0.05).结论 生命早期适量的1,25(OH)_2D_3干预可改善大鼠肺部炎症,减少嗜酸性细胞浸润,而过量则加重气道炎症.其机制可能与1,25(OH)_2D_3对IL-10及支气管上皮ICAM-1表达的影响有关.     

穿心莲内酯对哮喘小鼠嗜酸性粒细胞表达的影响及其作用机制

目的：穿心莲内酯是中药穿心莲的主要有效成分,在体内外均具有较强的抗炎作用。该研究观察穿心莲内酯对哮喘小鼠支气管肺泡灌洗液（BALF）中嗜酸性粒细胞、eotaxin及IL-5表达的影响。方法：32只BALB/c小鼠随机分为对照组、哮喘组、布地奈德组、穿心莲内酯组,每组8只。以卵清蛋白（OVA）致敏激发建立哮喘小鼠模型。ELISA法检测BALF及外周血中eotaxin、IL-5的水平,实时荧光定量PCR法检测肺组织eotaxin、IL-5 mRNA的表达。结果：穿心莲内酯明显抑制哮喘小鼠BALF中嗜酸性粒细胞（P0.05）。结论：穿心莲内酯通过降低哮喘小鼠eotaxin、IL-5的表达,抑制哮喘气道嗜酸性粒细胞浸润。     

巨噬细胞炎性蛋白1α及其mRNA在哮喘小鼠气道炎症中的作用

目的探讨巨噬细胞炎性蛋白1α(MIP-1α)及其mRNA在哮喘小鼠气道炎症中的作用.方法 70只二级雄性BALB/C小鼠随机分为2大组7小组(1)正常小鼠组,分成灌洗组(A24组)和未灌洗组(A0组);(2)哮喘小鼠组,分成灌洗组(根据末次激发时相不同分成B3组、B8组、B24组和B36组)和未灌洗组(B0组).对支气管肺泡灌洗液(BALF)进行嗜酸性粒细胞(EOS)计数和分类计数;应用双抗体夹心酶联免疫吸附试验法测定血清和BALF中MIP-1α的浓度;采用免疫组化法和原位杂交法检测MIP-1α蛋白和MIP-1α mRNA的表达.结果 (1)BALF和血清中MIP-1α的浓度B3组[(30.2±4.2)pg/ml,(30.8±4.6)pg/ml]、B8组[(35.3±4.9)pg/ml,(34.9±5.1)pg/ml]、B24组[(42.9±5.8)pg/ml,(41.7±6.3)pg/ml]和B36组[(37.8±4.7)pg/ml,(35.7±4.9)pg/ml]均高于A24组[(20.9±3.8)pg/ml,(22.4±4.3)pg/ml](P均<0.01); MIP-1α浓度3 h开始升高,24 h达峰值,36 h已下降;(2)免疫组化和原位杂交显示,B0组支气管MIP-1α蛋白和MIP-1α mRNA的阳性表达率[(26.4±6.2)%, (23.9±4.2)%]均高于A0组[(10.3±2.5)%, (8.7±1.8)%](P均<0.01),其表达的主要细胞是上皮细胞;(3)哮喘小鼠BALF中MIP-1α的浓度和EOS绝对值之间呈显著正相关(r=0.890, P＜0.01),MIP-1α的浓度和EOS占细胞总数百分比(EOS %)之间呈显著正相关(r=0.897, P＜0.01).结论哮喘小鼠MIP-1α蛋白和mRNA较强表达,上皮细胞是主要表达细胞,MIP-1α动力学特点为3 h开始升高,24 h达峰值,36 h已下降,并与EOS聚集密切相关.     

哮喘大鼠肺泡Ⅱ型上皮细胞损伤及机制研究

目的观察哮喘大鼠肺泡Ⅱ型上皮细胞(AT_Ⅱ)损伤情况并探讨其可能机制。方法用卵白蛋白(OVA)制备Wistar大鼠哮喘模型,分别测定支气管肺泡灌洗液(BALF)中一氧化氮(NO)、肿瘤坏死因子_α(TNF_α)浓度并检测AT_Ⅱ细胞的凋亡率和坏死率,将AT_Ⅱ细胞凋亡和坏死率之和(总损伤率)与NO和TNF_α浓度进行相关分析。结果哮喘组AT_Ⅱ细胞的凋亡率和坏死率[(11.55±2.82)%和(10.69±3.99)%]分别显著高于正常对照组[(2.81±2.22)%和(1.84±0.95)%],P均<0.01;BALF中的TNF_α和NO[(93.23±19.84)pg/ml和(7.36±0.63)μmol/L]也显著高于正常对照组[(26.97±5.85)pg/ml和(3.99±0.42)μmol/L],P均<0.01;BALF中TNF_α和NO浓度分别与AT_Ⅱ细胞总损伤率呈显著正相关(P均<0.01)。结论哮喘大鼠AT_Ⅱ细胞损伤明显,而TNF_α及NO的生成和释放增加可能是导致AT_Ⅱ细胞损伤增加的重要原因。     

依那西普对哮喘大鼠的抗炎作用研究

目的 探讨抗TNF-α受体单抗--依那西普(益赛普)对哮喘大鼠的抗炎作用及其机制.方法 30只SD雄性大鼠随机分成3组,每组10只.哮喘组:于第1、第8及第15天分别予腹腔注射10%鸡卵清蛋白(OVA)1 ml、氢氧化铝100 mg,于第16、17、18天密闭雾化罐中用1%OVA雾化吸入,每天激发20min致其哮喘发作;益赛普组:诱喘方法 同哮喘组,从第1天开始(致敏或激发后1 h)每隔1 d皮下注射益赛普0.5 mg/kg;对照组:以生理盐水代替OVA和益赛普.运用免疫组织化学方法 测定肺组织标本IL-4阳性细胞数,对支气管肺泡灌洗液(BALF)沉淀细胞进行计数以及运用ELISA方法 对BALF上清中TNF-α水平进行测定.结果 益赛普组大鼠肺组织中IL-4阳性细胞数较哮喘组明显降低(P<0.05);BALF中嗜酸性粒细胞和中性粒细胞数目较哮喘组明显减少(P<0.01),TNF-α水平(196±13)较哮喘组(236±8)明显降低(P<0.01);TNF-α、IL-4阳性细胞数、嗜酸性粒细胞和中性粒细胞数目均明显高于对照组.结论 益赛普可以减少哮喘大鼠BALF及肺组织中TNF-α和Th2细胞因子水平,减轻哮喘气道炎症.     

防哮饮早期干预对哮喘小鼠血清白细胞介素-10水平影响的实验研究

目的观察中药防哮饮早期干预对哮喘小鼠血清白细胞介素10(IL-10)水平以及支气管肺泡灌洗液(BALF)中嗜酸性粒细胞百分比(EOS%)的影响。方法将60只健康昆明小鼠随机分为6组:正常对照组、哮喘模型组、防哮饮小剂量组、防哮饮中剂量组、防哮饮大剂量组、布地奈德组各10只,用卵蛋白致敏激发造模。中药防哮饮组从致敏第1天起,将中药防哮饮按小剂量(含生药0.5 g/mL)、中剂量(含生药1.5 g/mL)、大剂量(含生药2.5 g/mL)灌胃给药,每日1次,每次0.4 mL,共27 d。在最后1次激发后24 h(第28 d)处死小鼠,采用双抗夹心酶联免疫吸附试验检测小鼠血清IL-10的含量,并镜检BALF中EOS%。结果哮喘模型组小鼠BALF中EOS[(2.9±1.4)%]较对照组[(1.3±0.9)%]明显升高(P<0.01);血清IL-10的含量[(7.6±3.0)ng/mL]较对照组[(44.9±33.0)ng/mL]降低(P<0.05)。防哮饮小、中、大剂量组均能显著提高哮喘小鼠血清IL-10水平,与哮喘模型组比较差异有统计学意义(P<0.01),与布地奈德组比较差异无统计学意义(P>0.05)。防哮饮大...     

卡介苗干预对哮喘小鼠干细胞因子表达的影响

目的研究卡介苗(BCG)对哮喘小鼠干细胞因子(SCF)表达的影响。方法昆明小鼠随机分为哮喘组、BCG组和对照组,每组10只,以卵清蛋白(OVA)致敏激发建立哮喘模型。末次激发24 h后取肺组织及支气管肺泡灌洗液(BALF),计数嗜酸性粒细胞(EOS);酶联免疫吸附法检测BALF中SCF水平;免疫组织化学图像分析技术测定肺组织中SCF蛋白表达。结果哮喘组、BCG组和对照组肺组织的EOS计数分别为(10.67±1.94)个/HP、(6.40±1.55)个/HP和(0.37±0.33)个/HP,BALF中EOS计数分别为(7.58±1.30)×107/L、(3.78±1.15)×107/L和0,三组间差异有统计学意义(P均0.01)。哮喘组、BCG组和对照组的肺组织SCF蛋白免疫组化指数分别为(24 787.97±7 214.12)、(20 509.50±4 775.27)和(12 261.66±3 277.65),BALF中SCF水平分别为(280.25±14.20)pg/ml、(266.77±31.15)pg/ml和(223.59±15.61)pg/ml,其中哮喘组和BCG组均高于对照组,差异有统计学意义(P均0.05),BCG组与哮喘组差异无统计学意义(P0.05)。哮喘组肺部SCF表达与EOS计数呈正相关(P0.05)。结论 BCG干预能显著减轻哮喘小鼠的气道炎症,但不能抑制SCF的表达。     

Human milk supplementation. Delivery of energy, calcium, phosphorus, magnesium, copper, and zinc

Human milk when fed to preterm infants is frequently supplemented with human milk fortifiers that provide an additional source of protein, energy, and minerals. Human milk that was provided by the mother of a preterm infant, and that was supplemented with commercially available human milk fortifiers, was assessed under simulated syringe-pump and bolus feeding circumstances for the delivery of energy, calcium, phosphorus, copper, magnesium, and zinc to an infant. In general, the nutrients were not completely delivered with syringe-pump feedings, with the greatest losses occurring in the concentrations of calcium and phosphorus. The losses were more pronounced with the use of a powdered fortifier than with the use of a liquid fortifier. Little or no change in the concentrations of the various nutrients were observed with simulated bolus feeding. We suggest that human milk fortified with supplements be fed with care to assure complete delivery of the nutrients and that infants receiving such feedings be monitored to assure adequate nutritional status.     

Serum protein binding of diazepam, desmethyldiazepam, furosemide, indomethacin, warfarin, and phenobarbital in human fetus, mother, and newborn infant

The protein binding of diazepam, desmethyldiazepam, furosemide, indomethacin, warfarin, and phenobarbital in maternal and fetal cord serum at the time of birth, and in serum of neonates between 1 and 11 days of age was studied. The protein binding of diazepam and desmethyldiazepam was higher in the fetus than in the mother, thus explaining the fetal cumulation of these drugs in vivo. After birth, both drugs were partially displaced from neonatal binding sites. The decreased protein binding capacity in the mother and the neonate related to increased free fatty-acid levels. The pattern of protein binding of warfarin in the groups investigated was a mirror image of those of diazepam and its metabolite. The protein binding of indomethacin progressively decreased in the neonate during the first two postnatal weeks, while that of furosemide remained at lowered levels throughout this time interval. The protein binding of phenobarbital was similar in the groups investigated. Our results suggest that drugs such as diazepam, which can be displaced from binding sites by free fatty acids, may cumulate in the fetus and may exhibit much decreased protein binding and possibly unexpectedly strong effects in the neonate after birth.     

Effect of lactate, pyruvate, acetone, acetoacetate, and beta-hydroxybutyrate on albumin binding of bilirubin

Lactate, pyruvate, acetone, acetoacetate, and beta-hydroxybutyrate were tested for their bilirubin-displacing effect on human serum albumin. Only lactate had a significant effect at levels found in asphyxiated infants (up to 20 mM). The reserve albumin equivalent for binding bilirubin was determined, using the deputy ligand monoacetyldiaminodiphenyl sulfone (MADDS), in adult human serum albumin solution, neonatal serum, and neonatal albumin solution. Twenty mM lactate caused a 23% decrease of reserve albumin when adult albumin was used, but did not cause any change of binding when neonatal serum or neonatal albumin solution was used. It is unlikely that endogenous substances, acting as competitive ligands, cause the low binding affinity of albumin for bilirubin in sick, premature infants.