陳尚勤， 鄭亞兵， 李昌崇， 陳 超， 林振浪△

(溫州醫(yī)學(xué)院附屬育英兒童醫(yī)院 1新生兒科，2呼吸科，浙江 溫州 325027；3復(fù)旦大學(xué)附屬兒科醫(yī)院新生兒科，上海 200032)

產(chǎn)前不同劑量地塞米松暴露對(duì)新生鼠肺泡成熟的影響

陳尚勤1， 鄭亞兵1， 李昌崇2， 陳 超3， 林振浪1△

(溫州醫(yī)學(xué)院附屬育英兒童醫(yī)院1新生兒科，2呼吸科，浙江 溫州 325027；3復(fù)旦大學(xué)附屬兒科醫(yī)院新生兒科，上海 200032)

目的探討孕期肌注不同劑量地塞米松對(duì)新生鼠肺泡成熟的影響。方法24只SD孕大鼠分為對(duì)照組、產(chǎn)前1次注射組(孕19 d肌注地塞米松0.8 mg·kg-1·d-1)和產(chǎn)前3次注射組(孕18、19、20 d均肌注地塞米松 0.8 mg·kg-1·d-1)，每組8只，取新生1、4、10、14 d鼠肺，病理切片，HE染色，觀察肺泡形態(tài)學(xué)的改變，計(jì)算機(jī)圖像分析。電鏡檢查觀察產(chǎn)前3次注射組和對(duì)照組肺上皮細(xì)胞基底膜的連續(xù)性。結(jié)果對(duì)照組從1-14日齡鼠肺泡間隔漸變薄，肺泡面積變小，肺泡表面積/肺泡體積(S/V)增大，肺(囊)泡數(shù)漸增多，4日齡鼠肺次級(jí)隔開(kāi)始長(zhǎng)出，至14日齡隔數(shù)目漸增多。產(chǎn)前3次注射組的1日齡囊泡隔變薄，肺囊泡面積增大，差異非常顯著(P<0.01)，肺囊泡數(shù)明顯減少(P<0.05)；4、10、14日齡肺泡間隔明顯變薄(P<0.05)，肺泡明顯增大，S/V較對(duì)照組小，肺囊泡數(shù)明顯減少，次級(jí)隔的長(zhǎng)出減少，相對(duì)較短，數(shù)目較少，差異非常顯著(P<0.01)。產(chǎn)前1次注射組1日齡鼠的肺泡間隔厚度較對(duì)照組變薄，差異顯著(P<0.05)。電鏡檢查發(fā)現(xiàn)，正常1、4日齡鼠外周肺上皮細(xì)胞基底膜變薄，多處不連續(xù)，上皮細(xì)胞與成纖維細(xì)胞靠近、接觸。產(chǎn)前3次注射組肺上皮細(xì)胞基底膜的不連續(xù)現(xiàn)象較少，上皮間質(zhì)的直接接觸較少。結(jié)論產(chǎn)前3次注射地塞米松影響新生鼠肺泡形態(tài)發(fā)育，產(chǎn)前1次注射地塞米松對(duì)新生鼠肺泡形態(tài)發(fā)育影響不大。

產(chǎn)前暴露; 地塞米松； 肺泡； 新生大鼠

自從1972年報(bào)道產(chǎn)前糖皮質(zhì)激素治療(antenatal corticosteriod therapy,ACT)能促進(jìn)肺表面活性物質(zhì)合成和分泌，ACT廣泛應(yīng)用于臨床預(yù)防早產(chǎn)兒呼吸窘迫綜合征(neonatal respiratory distress syndrome,NRDS)。近40年來(lái)，NRDS發(fā)病率大為降低[1]。但是并不減低早產(chǎn)兒支氣管肺發(fā)育不良的發(fā)生(bronchopulmonary dysplasia,BPD)[2]。而B(niǎo)PD 的發(fā)生與肺泡成熟障礙關(guān)系密切[3]。哺乳動(dòng)物肺泡發(fā)育從孕26周開(kāi)始，其關(guān)鍵階段在圍產(chǎn)期，決定了生后甚至成年后的肺形態(tài)結(jié)構(gòu)及功能[4]。圍產(chǎn)期的肺發(fā)育易受物理、化學(xué)、生物、病理等不良因素的影響，近年來(lái)隨著醫(yī)療技術(shù)的進(jìn)步，早產(chǎn)兒、超早早產(chǎn)兒的出生率和存活率大大提高，慢性肺疾病(chronic lung disease,CLD)成為影響早產(chǎn)兒生存質(zhì)量的重要并發(fā)癥，BPD是CLD的主要表現(xiàn)。

國(guó)際認(rèn)可的ACT用法指產(chǎn)前單一療程使用，但目前全球產(chǎn)科醫(yī)師對(duì)高危早產(chǎn)孕婦每周1次的多療程反復(fù)用藥[5]的現(xiàn)象相當(dāng)普遍，這一現(xiàn)象在國(guó)內(nèi)也相當(dāng)突出。同時(shí)ACT的不良反應(yīng)近期日漸受到重視，隨著 ACT重復(fù)應(yīng)用增多，糖皮質(zhì)激素對(duì)胎兒、新生兒的不良反應(yīng)也相應(yīng)增加，但尚無(wú)定論[6]。對(duì)腦發(fā)育的不良影響研究報(bào)道較多，ACT可導(dǎo)致發(fā)生腦癱的危險(xiǎn)性增加，影響智力性格的發(fā)育等；對(duì)肺研究少，而對(duì)肺發(fā)育的不良影響，尤其與BPD發(fā)生的關(guān)系正引起國(guó)內(nèi)外學(xué)者的關(guān)注。國(guó)外研究發(fā)現(xiàn)ACT影響肺泡形態(tài)發(fā)育，甚至持續(xù)到成年，影響成年期肺功能[7]。為進(jìn)一步探討不同劑量ACT對(duì)肺泡發(fā)育的影響，我們通過(guò)對(duì)不同孕齡鼠注射不同次數(shù)的地塞米松(dexamethasone，DXM)，觀察新生1、4、10、14日齡鼠的肺泡結(jié)構(gòu)形態(tài)發(fā)育及肺泡上皮細(xì)胞基底膜的變化，并探討其可能機(jī)制。

材 料 和 方 法

1材料

SD孕大鼠(復(fù)旦大學(xué)醫(yī)學(xué)院動(dòng)物實(shí)驗(yàn)中心提供)共24只，分3種情況用藥。

1.1產(chǎn)前3次注射組 隨機(jī)取8只孕18 d(足月21-22 d)大鼠，于孕18、19、20 d均肌注DXM 0.8 mg·kg-1·d-1(稀釋至0.5 mL)[8]。足月(21-22 d)自然分娩，每窩隨機(jī)取2只，共16只；4日齡，每窩隨機(jī)取2只，共16只；10日齡每窩隨機(jī)取1-2只(因有4窩孕鼠產(chǎn)仔較少)，共13只處死取肺；14 d亦每窩隨機(jī)取1-2只，共12只處死取肺。

1.2產(chǎn)前1次注射組 隨機(jī)取8只孕18 d大鼠，于孕19 d肌注DXM 0.8 mg·kg-1·d-1。孕18、20 d相應(yīng)肌注0.5 mL生理鹽水，后續(xù)分組似產(chǎn)前3次注射組。

1.3對(duì)照組 對(duì)照組孕鼠8只，于孕18、19、20 d各肌注0.5 mL生理鹽水，后續(xù)分組如產(chǎn)前3次注射組。

2方法

2.1動(dòng)物模型的建立 1、 4、10、14 d新生鼠，稱重，10%水合氯醛6 mL·kg-1腹腔注射麻醉。氣管插管給20 cmH2O的氣道壓力擴(kuò)張肺泡，1 min后逐漸調(diào)低至10 cmH2O維持,同時(shí)剖開(kāi)腹腔，刺破膈肌，打開(kāi)胸腔，右心室穿刺留置灌注生理鹽水，剪開(kāi)左心房放血，待左心房流出液體變清后改用10%中性緩沖福爾馬林右心灌注,持續(xù)20 min。絲線結(jié)扎氣管，小心分離出氣管和肺，浸入10%中性緩沖福爾馬林溶液固定48 h,取出放在含50%乙醇的福爾馬林中。24 h后,置于75%乙醛中,并放于4 ℃冰箱。取左肺近肺門(mén)，邊緣部肺組織石蠟包埋，5 μm切片，HE染色。

2.2肺組織病理學(xué)觀察 光鏡下觀察肺泡形態(tài)改變，IMS細(xì)胞圖像分析系統(tǒng)的醫(yī)學(xué)圖像分析軟件(上海申騰信息技術(shù)有限公司)作圖像分析，使顯微鏡處于同一放大倍數(shù)(100倍)及電壓，每張切片隨機(jī)選擇6個(gè)視野，分別測(cè)量以下指標(biāo)，測(cè)量時(shí)避開(kāi)大、中支氣管及血管 。(1)肺泡間隔厚度：每個(gè)視野隨機(jī)選6個(gè)肺泡，各測(cè)量4處間隔厚度，取平均值。(2)肺泡面積，肺泡壁周長(zhǎng)，肺泡等效內(nèi)徑：反映肺泡的大小。(3)肺泡數(shù)：計(jì)數(shù)每個(gè)視野(放大100倍視野)的肺泡數(shù)，反映肺泡密度。(4)次級(jí)隔高度和次級(jí)隔數(shù)目：測(cè)量次級(jí)隔頂至基底的高度及每個(gè)視野次級(jí)隔數(shù)目，反映肺呼吸膜面積的擴(kuò)大程度。(5)肺泡表面積/肺泡體積(surface area/volume,S/V)：以每個(gè)視野的肺泡面積計(jì)算出等效半徑，得出平均等效半徑，再換算出S/V, S/V=4πr2/(4 πr3/3) =3/r，此指標(biāo)反映有限的肺泡體積下呼吸膜面積。

2.3電鏡檢查 取0.1 cm×0.1 cm×0.1 cm的外周新鮮肺組織，2.5%戊二醛磷酸緩沖液固定2 h，0.1 mol/L磷酸漂洗液漂洗15 min×3次，1%鋨酸作后固定2 h，再0.1 mol/L磷酸漂洗液漂洗15 min×3次，50%、70%、90%、90%乙醇與90%丙酮1∶1混合液、90%丙酮在4 ℃冰箱內(nèi)依次脫水15 min，再100%丙酮室溫脫水15 min×3次，室溫包埋過(guò)夜，再37 ℃包埋2 h，固化后LKB-Ⅰ型超薄切片機(jī)切片50-60 nm，3%醋酸鈾-枸櫞酸鉛染色，日本電子JEM—1200EX透射電鏡(復(fù)旦大學(xué)附屬醫(yī)學(xué)院電鏡室)觀察3次注射組和對(duì)照組上皮細(xì)胞基底膜的連續(xù)性，并攝片。

3統(tǒng)計(jì)學(xué)處理

結(jié) 果

1ACT大鼠的大體表現(xiàn)、體重改變

與對(duì)照組相比，3次注射組的孕鼠有死產(chǎn)，其新生大鼠個(gè)頭較小，體重偏輕，皮膚較干燥，毛色光澤度差，膚色欠紅潤(rùn)，活動(dòng)偏少，易激惹，吸吮力較弱，喂養(yǎng)較對(duì)照組困難，有夭折；1次注射組與對(duì)照組差異不大。3次注射組1、4日齡鼠體重較對(duì)照組非常顯著下降(P<0.01，1日齡鼠下降10.3%，4日齡鼠下降23.9%)；1次注射組僅1日齡的體重較對(duì)照組顯著下降(P<0.05，下降幅度7.3%),見(jiàn)表1。

2肺泡病理組織學(xué)觀察

正常鼠肺從1日齡至14日齡，次級(jí)隔長(zhǎng)出數(shù)目漸增多，肺泡數(shù)增多，肺泡面積變小，S/V增大，肺泡間隔漸變薄，14日齡次級(jí)分隔基本結(jié)束，肺泡發(fā)育基本完成；3次注射組次級(jí)隔的長(zhǎng)出減少，相對(duì)較短，肺泡較對(duì)照組明顯增大，數(shù)目較少，呼吸膜面積較對(duì)照組小，肺泡間隔更薄,見(jiàn)圖1A、2A、3A、4A、圖1C、2C、3C、4C及表2-5。1次注射組除1日齡鼠的間隔厚度較對(duì)照組顯著變薄外(P<0.05)，余改變不明顯,見(jiàn)圖1B、2B、3B、4B及表2-5。

表1 各組新生鼠體重

Figure 1. Effect of antenatal DXM exposure on the lung histology of 1-day-old newborn rats(×100). A: lung section from a rat in thrice-DXM group on day 1, in which the air space was larger and the intersaccular walls were thinner compared with control group on day 1(C)；B: lung section from a rat in once-DXM group on day 1, in which the air space was similar to that in control group(C) and the intersaccular walls were thinner than those in the control rats on day 1(C).

Figure 2. Effect of antenatal DXM exposure on the lung histology of 4-day-old newborn rats(×100).A:lung section from a rat in thrice-DXM group on day 4，in which there were larger air space, thinner alveolar walls and less sprouting crests compared with the control rats on day 4(C)； B:lung section from a rat in once-DXM group on day 4 was similar to that in the control rats on day 4(C).Arrow designates sprouting crests.

Figure 3. Effect of antenatal DXM exposure on the lung histology of 10-day-old newborn rats(×100).A:lung section from a rat in thrice-DXM group on day 10，in which there were much larger air space, much thinner alveolar walls and apparently less and shorter secondary septa compared with the control rats on day 10(C)；B:Lung section from a rat in once-DXM group on day 10 was similar to that in the control rats on day 10(C).Arrows designate secondary septa.

Figure 4. Effect of antenatal DXM exposure on the lung histology of 14-day-old newborn rats(×100).A: lung section from a rat in thrice-DXM group on day 14，in which there were much air space larger, thinner alveolar walls and apparently less secondary septa compared with the control rats on day 14(C) ；B: lung section from a rat in once-DXM group on day14 was similar to that in the control rats on day 14(C). Arrows designate secondary septa.

表2 1日齡鼠肺泡形態(tài)分析結(jié)果

表3 4日齡鼠肺泡形態(tài)分析結(jié)果

表4 10日齡鼠肺泡形態(tài)分析結(jié)果

表5 14日齡鼠肺泡形態(tài)分析結(jié)果

3電鏡結(jié)果

電鏡下基底膜包括近細(xì)胞膜的透明層和近間質(zhì)的致密層，正常1、4日齡鼠外周肺上皮細(xì)胞的BM變薄，致密層多處不連續(xù)，此現(xiàn)象以4日齡鼠肺更為明顯，并見(jiàn)上皮細(xì)胞漿伸出偽足穿越這些基底膜的孔隙進(jìn)入間質(zhì)，上皮細(xì)胞與成纖維細(xì)胞靠近、接觸，見(jiàn)圖5B、6B。3次注射組1、4日齡鼠外周肺上皮細(xì)胞的BM的不連續(xù)現(xiàn)象較少，較少見(jiàn)伸出偽足，上皮間質(zhì)的直接接觸較少，見(jiàn)圖5A、6A。

Figure 5. Transmission electron micrographs of 1-day-old rat lungs. A:from a rat in thrice-DXM group on day 1，in which the basement membrane was continuous and close contact of cells were seldom seen；B:from a rat in control group, in which the basement membrane of lung epithelial cells turned thin and discontinuous,helping the close contact between the epithelial cells and the fibroblasts.

討 論

在產(chǎn)科臨床重復(fù)多療程的ACT現(xiàn)象相當(dāng)普遍，因此帶來(lái)的對(duì)新生個(gè)體近期和遠(yuǎn)期的副作用也使學(xué)術(shù)界擔(dān)憂。實(shí)驗(yàn)報(bào)道重復(fù)ACT使胎兒宮內(nèi)發(fā)育遲緩，出生體重下降[9]。我們實(shí)驗(yàn)觀察到，產(chǎn)前3次注射DXM治療的1、4 d新生鼠體重明顯下降。糖皮質(zhì)激素抑制蛋白合成，以及3次注射組出生時(shí)喂養(yǎng)較困難影響了體重增長(zhǎng)，至10、14 d產(chǎn)前的外源性糖皮質(zhì)激素的作用已基本消失，喂養(yǎng)亦較前好轉(zhuǎn)，體重與對(duì)照組比較無(wú)差異。1次注射組體重改變不明顯，僅1日齡體重較對(duì)照組輕度下降，14日齡體重反而略高于對(duì)照組，表明產(chǎn)前1次注射DXM對(duì)新生鼠影響不大，ACT對(duì)仔鼠體重的影響有劑量或重復(fù)次數(shù)的依從性。

Figure 6. Transmission electron micrographs of 4-day-old rat lungs. A: from a rat in thrice-DXM group on day 4, in which the basement membrane is continuous and close contact of cells were seldom seen;B: from a rat in control group on day 4, showing the basement membrane of lung epithelial cells turned thin and discontinuous.

哺乳動(dòng)物的肺泡發(fā)育是由體積大、數(shù)目少、壁厚的囊泡向體積小、數(shù)目多、壁薄、次級(jí)隔長(zhǎng)出的成熟肺泡轉(zhuǎn)化的過(guò)程，呼吸膜面積成倍擴(kuò)大。人類肺泡化發(fā)育于孕26周左右開(kāi)始，持續(xù)到生后3歲[4]，但圍產(chǎn)期是肺泡大量發(fā)育的關(guān)鍵期，此期諸多不良因素，如早產(chǎn)、化學(xué)物質(zhì)、感染等會(huì)阻礙肺泡正常發(fā)育，造成BPD。本實(shí)驗(yàn)觀察到對(duì)照組新生大鼠隨日齡增長(zhǎng)，肺泡間隔漸變薄，次級(jí)隔萌出，數(shù)目漸增多，14日齡次級(jí)分隔基本結(jié)束，肺泡面積變小，S/V增大，肺泡數(shù)增多，肺泡基本成熟，有限肺容量下肺氣體交換的表面積增加。

產(chǎn)前3次注射注射DXM組的新生大鼠肺泡成熟化受到抑制，肺泡間隔的變薄現(xiàn)象出現(xiàn)早，使氣體通過(guò)呼吸膜的彌散速度加快，但次級(jí)隔發(fā)育較對(duì)照組明顯受阻，數(shù)目減少，較短，肺泡呈現(xiàn)“大而少”的征象，肺泡化水平降低，阻礙了呼吸膜面積的擴(kuò)大，影響了肺通/換氣功能的儲(chǔ)備，使之在機(jī)械正壓通氣下或/和炎癥、化學(xué)等致肺損傷因素的作用下易于肺氣腫或氣胸的產(chǎn)生。有報(bào)道糖皮質(zhì)激素使發(fā)育期的羊在機(jī)械通氣下易產(chǎn)生肺氣腫[10]。Satoru等[8]研究也發(fā)現(xiàn)類似現(xiàn)象：產(chǎn)前注射DXM的大鼠，10日齡仔鼠肺泡化明顯受到抑制，表現(xiàn)肺泡直徑變大，肺泡次級(jí)間隔發(fā)育受阻，總肺泡數(shù)減少；至13-36 d，出現(xiàn)追趕性肺泡化生長(zhǎng)，先回復(fù)到不成熟狀態(tài)，繼而生出次級(jí)隔，但延遲的肺泡化不能代償完全，至成年60 d治療組肺泡直徑仍大于對(duì)照組，肺泡總數(shù)少于對(duì)照組。

我們實(shí)驗(yàn)產(chǎn)前1次注射組鼠除1日齡的間隔厚度較對(duì)照組變薄外，余改變不明顯，提示ACT對(duì)肺泡成熟的影響存在劑量或重復(fù)次數(shù)的依從性，單劑ACT較安全。因國(guó)外相關(guān)研究產(chǎn)前用DXM的劑量相差較大[8]，并無(wú)定論，故本實(shí)驗(yàn)依據(jù)產(chǎn)科DXM用于孕婦最多重復(fù)5次的總劑量，按人鼠體表面積比折算成大鼠的用量分3次注射做為3次注射組[11]，1次注射組相當(dāng)于重復(fù)1.5次，而介于兩者之間的劑量對(duì)肺泡發(fā)育的影響程度待進(jìn)一步研究。

3次注射組肺泡發(fā)育停滯，呈現(xiàn)“大而少”的征象與早產(chǎn)兒BPD的發(fā)生和發(fā)展關(guān)系密切。ACT 減低了24 周至29周出生的早產(chǎn)兒RDS發(fā)生率，但并無(wú)改善其BPD的發(fā)生率和住院時(shí)間[12]。1999年Jobe[13]提出以肺發(fā)育受阻為特征的 “新型BPD”的說(shuō)法，病理學(xué)上表現(xiàn)為少而大的肺泡，推測(cè)胎肺炎癥，產(chǎn)后暴露于機(jī)械通氣、吸氧等病理刺激，協(xié)同糖皮質(zhì)激素、營(yíng)養(yǎng)不良等均會(huì)致肺泡發(fā)育受抑。BPD與多劑ACT以及生后用激素的實(shí)驗(yàn)肺病理形態(tài)學(xué)改變相似高度提示ACT可能參與或與其它高危因素協(xié)同促進(jìn)BPD的發(fā)生和發(fā)展[3],甚至影響成年后的肺功能[2,7,8]。糖皮質(zhì)激素用于BPD的治療可抑制炎癥反應(yīng)，但也發(fā)現(xiàn)類似損害肺泡化，阻礙肺發(fā)育的現(xiàn)象[14]。因在BPD患兒中ACT以及生后用激素的機(jī)率較高，故對(duì)ACT加重BPD的可能性值得深入研究。

肺的發(fā)育和成熟有賴糖皮質(zhì)激素的作用，促皮質(zhì)釋放激素基因敲除的小鼠血漿糖皮質(zhì)激素水平很低，出現(xiàn)遠(yuǎn)端肺實(shí)質(zhì)解剖上成熟障礙[15]。大鼠在生后10 d內(nèi)血漿皮質(zhì)激素濃度出現(xiàn)一低谷期，恰為肺泡次級(jí)隔生成期，而過(guò)量的糖皮質(zhì)激素可阻礙肺泡化。次級(jí)隔的長(zhǎng)出，毛細(xì)血管迅速長(zhǎng)入形成網(wǎng)狀結(jié)構(gòu)，上皮細(xì)胞、內(nèi)皮細(xì)胞、間質(zhì)、間質(zhì)細(xì)胞間的接觸和相互作用等與肺上皮細(xì)胞基底膜(BM)的重建關(guān)系密切[16]，我們進(jìn)一步觀察了基底膜在肺泡發(fā)育中的變化。1、4日齡鼠外周肺電鏡可見(jiàn)上皮細(xì)胞BM變薄，多處不連續(xù)，細(xì)胞漿伸出偽足穿越這些BM的孔隙進(jìn)入下面的間質(zhì)，與間質(zhì)細(xì)胞，主要是成纖維細(xì)胞靠近、緊密接觸，考慮BM的改變可能與上皮細(xì)胞的分化、增殖等生物活動(dòng)有關(guān)，促進(jìn)肺泡的發(fā)育。產(chǎn)前3次注射DXM作用下外周肺上皮細(xì)胞BM的不連續(xù)現(xiàn)象較少見(jiàn)，鮮見(jiàn)伸出偽足，上皮間質(zhì)的直接接觸較少，說(shuō)明ACT使肺泡發(fā)育中的各個(gè)細(xì)胞接觸、交流受阻，導(dǎo)致肺發(fā)育障礙。

糖皮質(zhì)激素促進(jìn)Ⅱ型肺泡上皮細(xì)胞成熟，但同時(shí)也減弱了Ⅱ型肺泡上皮細(xì)胞的增殖以及抑制其向Ⅰ型細(xì)胞轉(zhuǎn)化，導(dǎo)致Ⅰ型細(xì)胞減少(后者為擴(kuò)大肺泡交換膜面積的細(xì)胞)，影響上皮細(xì)胞向內(nèi)腔的隆起，即次級(jí)隔的長(zhǎng)出；此外新生的次級(jí)隔必須充滿毛細(xì)血管和成纖維細(xì)胞，此需相應(yīng)細(xì)胞大量復(fù)制增殖，以使次級(jí)隔延長(zhǎng)，糖皮質(zhì)激素抑制細(xì)胞增殖。亦有研究認(rèn)為ACT抑制了機(jī)體的抗氧化系統(tǒng)，使肺易受脂質(zhì)氧化的損傷，而造成肺泡發(fā)育受阻[17，18]。

多中心研究表明，1個(gè)療程的激素與每周重復(fù)使用激素比較，新生兒NRDS發(fā)生率并無(wú)顯著差異[9]，從正面作用角度亦無(wú)必要重復(fù)使用糖皮質(zhì)激素促胎肺成熟。鑒于1次注射組DXM對(duì)新生鼠肺泡形態(tài)發(fā)育影響不大，而3次注射組DXM明顯影響新生鼠肺泡形態(tài)發(fā)育，我們建議產(chǎn)前多療程糖皮質(zhì)激素治療措施不常規(guī)用于有持續(xù)早產(chǎn)危險(xiǎn)的孕婦,而常規(guī)1療程的產(chǎn)前糖皮質(zhì)激素治療是安全的。

[1] Bonanno C,Wapner RJ. Antenatal corticosteroid treatment:what’s happened since Drs Liggins and Howie? [J].Am J Obstet Gynecol,2009,200(4):448-457.

[2] Pérez Pérez G, Navarro Merino M.Bronchopulmonary dysplasia and prematurity.Short-and long-term respiratory changes[J]. An Pediatr(Barc),2010, 72(1):79.e1-e16.

[3] Kramer BW,Lievense S,Been JV,et al. From classic to new bronchopulmonary dysplasia[J].Ned Tijdschr Geneeskd,2010,154: A1024.

[4] Smith LJ, McKay KO, van Asperen PP,et al.Normal development of the lung and premature birth[J]. Paediatr Respir Rev,2010,11(3):135-142.

[5] Koenen SV, Dunn EA, Kingdom JC, et al. Overexposure to antenatal corticosteroids:a global concern[J].J Obstet Gynaecol Can,2007, 29(11):879.

[6] Bevilacqua E,Brunelli R,Anceschi MM. Review and meta-analysis: Benefits and risks of multiple courses of antenatal corticosteroids[J]. J Matern Fetal Neonatal Med,2010, 23(4):244-260.

[7] Dalziel SR,Rea HH,Walker NK,et al. Long term effects of antenatal betamethasone on lung function:30 year follow up of a randomised controlled trial[J]. Thorax,2006,61(8):678-683.

[8] Okajima S,Matsuda T, Cho K,et al. Antenatal dexamethasone administration impairs normal postnatal lung growth in rats[J]. Pediatr Res，2001, 49(6):777-781.

[9] Mazumder P, Dutta S, Kaur J,et al. Single versus multiple courses of antenatal betamethasone and neonatal outcome: a randomized controlled trial[J]. Indian Pediatr,2008,45(8):661-667.

[10]Polk DH, Ikegai M, Jobe AH, et al. Preterm lung function after retreatment with atenatal betamethasone in preterm lambs[J]. Am J Obstet Gynecol,1997,176(2):308-315.

[11]章元沛 主編.藥理學(xué)實(shí)驗(yàn)[M].北京:人民衛(wèi)生出版社,1996.241.

[12]Manktelow BN, Lal MK, Field DJ, et al. Antenatal corticosteroids and neonatal outcomes according to gestational age: a cohort study[J]. Arch Dis Child Fetal Neonatal Ed,2010,95(2):F95-F98.

[13]Jobe AH,The new BPD:an arrest of lung development[J].Pediatr Res,1999,46(6):641-643.

[14]Grier DG, Halliday HL.Effects of glucocorticoids on fetal and neonatal lung development[J]. Treat Respir Med,2004,3(5):295-306.

[15]Muglia LJ,Bae DS,Brown TT,et al.Proliferation and differentiation defects during lung development in corticotropin releasing hormone- deficient mice[J]. Am J Respir Cell Mol Biol,1999, 20(2): 181-188.

[16]Burri PH.Structural aspects of postnatal lung development-alveolar formation and growth[J]. Biol Neonate,2006,89(4):313-322.

[17]Verhaeghe J, van Bree R, van Herck E. Oxidative stress after antenatal betamethasone: acute downregulation of glutathione peroxidase-3[J]. Early Hum Dev,2009,85(12):767-771.

[18]陳佰義，姜 莉，侯顯明.實(shí)驗(yàn)性肺纖維化大鼠肺泡巨噬細(xì)胞脂質(zhì)過(guò)氧化先于其細(xì)胞因子的釋放[J]. 中國(guó)病理生理雜志，2000，16(11)：1228-1230.

Antenatalexposuretodifferentdosesofdexamethasoneleadstodelayedalveolarizationinnewbornrats

CHEN Shang-qin1, ZHENG Ya-bing1, LI Chang-chong2, CHEN Chao3, LIN Zhen-lang1

(1NewbornDivision,2RespiratoryDivision,TheAffiliatedYuyingChildren’sHospital,WenzhouMedicalCollege,Wenzhou325027,China;3NewbornDivision,AffiliatedChildren’sHospital,FudanUniversity,Shanghai200032,China.E-mail:linzhenlang@hotmail.com)

AIM: To evaluate the effects of prenatal dexamethasone(DXM) exposure on growth and lung maturation in rats.METHODSTwenty-four pregnant Sprague-Dawley(SD) rats were divided into 3 groups: 8 pregnant rats in antenatal thrice-DXM group were injected intramuscularly with DXM(0.8 mg·kg-1·day-1) for 3 consecutive days(on gestational day 18, 19 and 20); 8 pregnant rats in antenatal once-DXM group were injected intramuscularly with DXM on gestational day 19. The control rats

equivalent volume of isotonic saline at the same time points. The lungs of the newborn rats on day 1, 4, 10 and 14 were examined. The tissue sections were prepared with HE staining for morphological investigation under light microscope. The data analysis was performed by means of a digital image analysis system, including thickness of interalveolar walls, alveolar(saccules) areas, the number of air spaces, the number and length of secondary septa, and the surface densities of the air spaces. Transmission electronic microscope was used to observe the basement membrane of the lung epithelial cells.RESULTSIn control group, and thickness of interalveolar walls, smaller alveolar(saccules) area,increased surface area/volume and number of air spaces were observed from day 1 to day 14. On day 4, an appreciable number of sprouting crests appeared, then the number and length of secondary septa increased until day 14. In antenatal thrice-DXM group, the air spaces were larger, and the thickness of intersaccular walls were thinner as compared to saccules of the control rats on day 1(P<0.01,P<0.05). There were thinner inter-air space septa(P<0.05), larger and fewer air spaces, and a reduction in the number of sprouting secondary septa on day 4, 10 and 14 than those in the control rats with significant differences(P<0.01). In antenatal once-DXA group, intersaccular walls were thinner than those in the control rats with significant differences(P<0.05). The results of electron microscopy revealed that the basement membrane of lung epithelial cells turned thin and discontinuous on day 1 and day 4 in the control rats, resulting in the close contact between epithelial cells and fibroblasts. In antenatal thrice-DXN group, the discontinuous basement membrane and close contact of the cells were seldom seen.CONCLUSIONAntenatal thrice-injection of DXM applied on gestational day 18, 19 and 20 impairs the alveoli formation in newborn rats. Antenatal once-injection of DXM has no remarkable effect on postnatal alveolarization.

Prenatal exposure; Dexamethasone; Pulmonary alveoli; Newborn rats

R363

A

10.3969/j.issn.1000-4718.2011.02-025

1000-4718(2011)02-0343-07

2010-09-15

2010-11-08

△通訊作者 Tel：0577-88816447；E-mail: linzhenlang@hotmail.com