唐成芳,曾 輝,張 莎,王丹楊,楊黎艷,謝 娜

(1西安醫(yī)學(xué)院口腔醫(yī)學(xué)系,西安 710021;2西安醫(yī)學(xué)院藥學(xué)院)

PMBE穩(wěn)定牙本質(zhì)基質(zhì)及封閉牙本質(zhì)小管的雙重功效研究

唐成芳1,曾 輝1,張 莎1,王丹楊1,楊黎艷2,謝 娜1

(1西安醫(yī)學(xué)院口腔醫(yī)學(xué)系,西安 710021;2西安醫(yī)學(xué)院藥學(xué)院)

目的 觀察馬尾松樹皮提取物(pinus massoniana bark extract,PMBE)穩(wěn)定牙本質(zhì)基質(zhì)和封閉牙本質(zhì)小管效果,為其防治牙本質(zhì)過敏提供實(shí)驗(yàn)依據(jù)。 方法 制備牙本質(zhì)塊60個(gè),隨機(jī)分為6組(n=10):乙醇組,氟涂膜組,1%,2%,4%和8%PMBE組。酸蝕牙本質(zhì)使牙本質(zhì)小管開放,各組分別用無水乙醇,氟涂膜,1%,2%,4%和8%PMBE乙醇溶液涂刷牙本質(zhì)表面。采用場發(fā)射掃描電鏡觀察牙本質(zhì)表面及剖面的顯微形貌,并測量牙本質(zhì)小管口開放視野面積(ADTO)、牙本質(zhì)小管口開放相對(duì)面積(PADTO)和小管封閉深度(SD)。 結(jié)果 8%PMBE組ADTO及PADTO顯著低于乙醇組(P<0.05),但與氟保護(hù)漆組比較差異無統(tǒng)計(jì)學(xué)意義(P>0.05)。4%PMBE組ADTO及PADTO顯著低于1%，2%PMBE組及乙醇組,但顯著高于8%PMBE組。掃描電鏡見乙醇組牙本質(zhì)表面牙本質(zhì)小管完全開放,管壁疏松;氟保護(hù)漆組牙本質(zhì)小管完全封閉;PMBE組管壁膠原基質(zhì)清晰可見、致密,小管口封堵程度隨PMBE濃度增高而增加,8%PMBE組小管口基本完全封堵,剖面可見封堵物。 結(jié)論 PMBE乙醇溶液局部涂布使用具有穩(wěn)定膠原基質(zhì)和封閉牙本質(zhì)小管的雙重功效,8%PMBE溶液效果最佳,PMBE具備一定治療牙本質(zhì)過敏的作用。

馬尾松樹皮提取物; 牙本質(zhì)過敏; 牙本質(zhì)小管; 氟保護(hù)漆

牙本質(zhì)過敏癥是指牙齒在受到冷熱等溫度、酸等化學(xué)物質(zhì)以及機(jī)械作用摩擦或咬硬物等所引起的一種異常酸痛癥狀,具有發(fā)作迅速、疼痛尖銳、時(shí)間短的特點(diǎn),好發(fā)于老年患者,嚴(yán)重影響患者身心[1,2]。目前普遍接受的牙本質(zhì)發(fā)生的機(jī)制是外界刺激作用于開放的牙本質(zhì)小管,并將刺激傳遞至近髓處作用于牙髓神經(jīng)導(dǎo)致的疼痛[2]。因而,封閉牙本質(zhì)小管是治療牙本質(zhì)敏感的根本措施之一[2,3]。

馬尾松樹皮提取物(pinus massoniana bark extract,PMBE)是從我國廣泛分布的松科植物馬尾松中提取的天然物質(zhì),其含有原花青素(procyanidins, PA)、兒茶素及表兒茶素等多酚類物質(zhì)及鉬、鈣等微量元素,其中PA可達(dá)90%,具有抗炎、抗腫瘤等多種功能[4-7]。PMBE已被課題組前期通過顯微CT及理化性能分析證實(shí)可增強(qiáng)牙本質(zhì)抗酸能力,抑制根面牙本質(zhì)脫礦,從而具備預(yù)防牙本質(zhì)過敏和根面齲的作用[8]。本研究擬采用PMBE乙醇溶液涂布處理因脫礦所致牙本質(zhì)小管開放的牙本質(zhì),通過掃描電鏡及形態(tài)計(jì)量學(xué)觀察易于臨床局部應(yīng)用的PMBE乙醇溶液對(duì)開放牙本質(zhì)小管的封閉效果及對(duì)脫礦牙本質(zhì)基質(zhì)的穩(wěn)定作用,進(jìn)一步為其治療牙本質(zhì)過敏提供一定的實(shí)驗(yàn)依據(jù)。

1 材料與方法

1.1 主要材料和儀器

PMBE(其中PA>95%,奧賽生物科技有限公司,西安),無水乙醇(分析純,西安化學(xué)試劑廠),37%磷酸凝膠(DenFilTM,韓國),柯伯脂氟保護(hù)漆(PULDENT?,美國Puldent公司,美國),以無水乙醇為溶劑配制1%,2%,4%,8%的PMBE溶液,取上清液備用。

慢速切割機(jī)(UNIPOL-830,科晶有限公司,沈陽),場發(fā)射掃描電鏡(FESEM S-4800,Hitachi,日本),分析天平(塞多利斯,塞多利斯斯泰帝公司,德國)。

1.2 試樣制備

西安交通大學(xué)口腔醫(yī)院新近拔除的無病變隱裂第三磨牙(均來源于20-30歲患者)30顆,去凈軟組織,垂直于牙長軸截取2 mm厚的冠方牙本質(zhì)片,進(jìn)一步將其制備為寬3 mm×長4 mm的牙本質(zhì)塊,2塊/牙,共60塊。每個(gè)試件近咬合面?zhèn)扔锰蓟枭凹埩魉麓蚰テ秸?近髓腔面制備一垂直于牙片深約1.7 mm的引導(dǎo)溝,以便于劈開牙本質(zhì)片觀察牙本質(zhì)剖面形態(tài)。

1.3 分組及處理

將60個(gè)試件隨機(jī)分到6個(gè)組:乙醇組,氟保護(hù)漆組,1%,2%,4%,8% PMBE組,每組10個(gè),其中乙醇組和氟保護(hù)漆分別作為陰性和陽性對(duì)照組。

試件經(jīng)超聲清洗干凈后用37%磷酸凝膠酸蝕30 s,蒸餾水沖洗3次,10 s/次,氣槍輕吹去除多余水分,分別用小毛刷蘸取無水乙醇、氟保護(hù)漆,1%,2%,4%,8% PMBE乙醇液均勻涂刷2層,自然干燥。

1.4 表面及剖面形貌觀察

樣本自然干燥后,沿制備的引導(dǎo)溝劈開分為兩半,其中一半用于觀察牙本質(zhì)表面特征,另一半用于觀察樣本的縱剖面。樣本噴金,FE-SEM掃描電鏡觀察表面牙本質(zhì)小管封堵情況以及剖面牙本質(zhì)小管管壁形貌;每個(gè)試樣表面和剖面分別采集3張圖像。

1.5 牙本質(zhì)小管開口面積、開口相對(duì)面積和小管封閉深度測定

采用Image-Pro Plus 6.0圖像分析軟件,對(duì)牙本質(zhì)表面和縱剖面圖像分別進(jìn)行測量分析。導(dǎo)入采集圖像,牙本質(zhì)表面測量牙本質(zhì)小管開口面積(area of dentin tubule openings,ADTO),剖面測量牙本質(zhì)小管封堵的深度(sealed depth, SD),每個(gè)樣本結(jié)果為3張圖像測量的平均值。按照如下公式計(jì)算牙本質(zhì)小管開口放相對(duì)面積(percentage area of dentin tubule openings,PADTO),PADTO=(ADTO/牙本質(zhì)面積)×100%。

1.6 統(tǒng)計(jì)學(xué)分析

2 結(jié)果

2.1 表面及剖面形貌

乙醇組牙本質(zhì)表面所有小管口均呈圓形或類圓形開放(見圖1A),剖面見牙本質(zhì)小管清晰可見,管壁較疏松且夾雜空隙,膠原纖維斷裂(見圖2A);氟保護(hù)漆組牙本質(zhì)表面基本被膜狀物覆蓋,牙本質(zhì)小管口完全封閉(見圖1B);剖面見管壁內(nèi)及其管間牙本質(zhì)均被覆厚薄不一的膜狀物,管腔內(nèi)無明顯的樹脂突形成(見圖2B)。

各PMBE組牙本質(zhì)表面牙本質(zhì)小管不同程度封閉,小管口封閉程度隨PMBE濃度增加而增大,但僅有8%剖面見小管內(nèi)有封堵物,長約2-3 μm(見圖1)。1%PMBE和2%PMBE組牙本質(zhì)表面致密,小管邊緣可見膜狀物形成,小管口較乙醇組減小(見圖1);剖面見管壁致密,似有薄層物質(zhì)覆蓋,管壁和管間膠原纖維較清晰(見圖2)。4%PMBE組表面牙本質(zhì)表面呈致密顆粒狀,部分小管口被封堵,其余小管開口明顯減小,剖面形貌與2%相似(見圖1E);8%PMBE組表面牙本質(zhì)小管基本完全封閉且部分小管口輪廓可見,僅個(gè)別牙本質(zhì)小管未完全封堵(見圖1F),剖面見牙本質(zhì)小管內(nèi)徑較其余組明顯縮小,且自管口向下約2-3 μm長度的管腔完全封堵,管壁及管間牙本質(zhì)均致密,未見明顯的膠原纖維暴露(見圖2F)。

A-F分別為乙醇組，氟涂膜組，1%PMBE組，2%PMBE組，4%PMBE組，8%PMBE組;圖中白箭頭示牙本質(zhì)小管開口圖1 牙本質(zhì)表面微觀形貌 (FE-SEM,×5 000)Figure 1 Micro morphology of dentin surface in different groups (FE-SEM,×5 000)

2.2 ADTO,PADTO結(jié)果

ADTO乙醇組最大,乙醇組ADTO及PADTO顯著高于氟保護(hù)漆組,差異有統(tǒng)計(jì)學(xué)意義(P<0.05);1%PMBE和2%PMBE組ADTO及PADTO均低于乙醇組,但組間差異均無統(tǒng)計(jì)學(xué)意義(P>0.05);8%PMBE組ADTO及PADTO略高于氟保護(hù)漆組,差異無統(tǒng)計(jì)學(xué)意義(P>0.05);4%PMBE組ADTO及PADTO顯著低于2%PMBE組,且顯著高于8%PMBE組,差異均有統(tǒng)計(jì)學(xué)意義(P<0.05,見表1)。

組別ADTO(μm)PADTO(%)乙醇組 321157±62478 2950±574 氟保護(hù)漆組 001±003? 000±000?1%PMBE組277857±46419#△2552±426#△2%PMBE組241853±55354#△2221±509#△4%PMBE組106430±27709?# 978±255?#8%PMBE組21179±4410?195±040?

與乙醇組比較,*P<0.05;與氟保護(hù)漆組比較,#P<0.05;與4%PMBE比較,△P<0.05

3 討論

牙本質(zhì)過敏癥根本上是牙本質(zhì)暴露,牙本質(zhì)小管開放所致的牙齒敏感癥狀[1,2]。目前臨床上可用激光、樹脂或氟化鈉、鉀鹽等藥物的方法,通過直接的神經(jīng)抑制或改變神經(jīng)沖動(dòng)來降低敏感性或機(jī)械地封閉牙本質(zhì)小管來降低牙本質(zhì)通透性并阻止小管液的流動(dòng)從而防治牙本質(zhì)過敏[1-3,9-13]。氟保護(hù)漆因含有效成分氟已被用于牙本質(zhì)過敏和齲齒的預(yù)防治療[13]。氟保護(hù)漆是以樹脂為基質(zhì)的含氟緩釋劑,能在口腔內(nèi)快速成膜,并持續(xù)釋放氟離子的有機(jī)溶液,對(duì)牙體組織具有良好保護(hù)作用[13]。本實(shí)驗(yàn)以NaF作為主要有效成分的柯伯脂氟保護(hù)漆作為對(duì)照觀察富含天然交聯(lián)劑PA的PMBE封閉因酸脫礦而開放的牙本質(zhì)小管的功效,為防治牙本質(zhì)過敏提供一定的實(shí)驗(yàn)參考。本實(shí)驗(yàn)也觀察到柯伯脂含氟涂膜能在牙本質(zhì)表面形成一層致密膜狀物從而阻止外界的刺激傳入,并具有較強(qiáng)滲透性可進(jìn)入脫礦的管間牙本質(zhì)膠原纖維間,但無明顯樹脂突形成。

PMBE是松科植物馬尾松的樹皮中提取出的天然物質(zhì),富含由單聚體、寡聚體及多聚體組成的PA[4,5]。PA具有抗氧化、抗菌、抗過敏、抗炎鎮(zhèn)痛及調(diào)節(jié)免疫等多種生物學(xué)活性[6,14]。葡萄籽等來源的PA也被用于脫礦牙本質(zhì)和抗致齲菌研究,Gregoire等[15]和畢玲[16]發(fā)現(xiàn)PA直接抑制變形鏈球菌、遠(yuǎn)緣鏈球菌等致齲菌活性,還可穩(wěn)定膠原基質(zhì)而影響礦化平衡,從而相對(duì)減少齲損深度和礦物損失量。可可、葡萄籽等不同來源的PA可通過氫鍵、離子鍵、共價(jià)鍵等與膠原相互作用,不同程度地增加膠原交聯(lián)度,提高膠原強(qiáng)度和耐酶解能力,提高牙本質(zhì)粘接穩(wěn)定性[17-21]。課題組前期研究表明,PMBE以及與之同源的馬尾松針提取物經(jīng)pH循環(huán)短暫預(yù)處理,可抑制根面牙本質(zhì)鈣離子逸出,延緩礦物密度減低,預(yù)防牙本質(zhì)過敏和根面齲[8,22]。

本實(shí)驗(yàn)以無毒、滲透性好、易揮發(fā)的無水乙醇作為溶劑配制PMBE溶液,以期將PMBE乙醇溶液作為涂布劑或含潄劑局部應(yīng)用于口腔。實(shí)驗(yàn)結(jié)果顯示4%PMBE組和8%PMBE組ADTO及PADTO顯著低于乙醇組,提示PMBE可有效封閉暴露牙本質(zhì)的小管開口。FE-SEM結(jié)果也證實(shí)了該結(jié)論。8%PMBE組ADTO及PADTO高于氟保護(hù)漆組,但兩者間的差異無統(tǒng)計(jì)學(xué)意義,結(jié)合FE-SEM結(jié)果則提示8%濃度的PMBE乙醇溶液的小管即刻封閉效果與氟保護(hù)漆相當(dāng),因而治療牙本質(zhì)過敏效果最佳。此外,FE-SEM剖面形貌見1%,2%PMBE組管壁較乙醇組致密,管間和管壁膠原纖維較清晰,提示PMBE與葡萄籽提取物一樣具有穩(wěn)定膠原的作用[18,19],其結(jié)果可能與PMBE富含PA有關(guān)。

PMBE封閉牙本質(zhì)小管和穩(wěn)定膠原纖維的作用可能與以下幾方面因素有關(guān):①PMBE富含PA[4,5,8],其可通過共價(jià)鍵、離子鍵、氫鍵等與脫礦的膠原纖維作用,增加交聯(lián)度,從而保持了膠原纖維完整;②本實(shí)驗(yàn)所用溶劑采用的是無水乙醇,其具有滲透性強(qiáng)、揮發(fā)快等特點(diǎn),該特點(diǎn)可使PMBE乙醇溶液能快速滲透至脫礦的牙本質(zhì)基質(zhì)中,PA使裸露的膠原快速交聯(lián),乙醇快速揮發(fā)后的有形物質(zhì)覆蓋于牙本質(zhì)表面,從而使該組牙本質(zhì)小管壁致密甚至封堵牙本質(zhì)小管口;③原花青素B環(huán)上酚羥基還可螯合因酸蝕脫礦膠原纖維間殘余的鈣離子[8,21,23],從而沉積于牙本質(zhì)表面;④PMBE組還含有微量的鈣元素和鉬元素[8],其也可沉積脫礦牙本質(zhì)表面;⑤PMBE濃度越大,單位體積內(nèi)的有效成分PA含量越多,上述作用就越大,從而使低濃度1%,2%PMBE乙醇溶液較無水乙醇明顯保存了膠原纖維,并使管壁致密,而高濃度4%及8%組則進(jìn)一步使牙本質(zhì)小管封堵。本實(shí)驗(yàn)中8%PMBE組還觀察到封堵物進(jìn)入管腔且完全封堵約2-3 μm,表明8%PMBE組可能還有一定的耐磨損性?？谇恢械牟牧匣蛐迯?fù)體還要經(jīng)受刷牙及自然磨耗等過程,然而本實(shí)驗(yàn)僅僅就不同濃度PMBE的即刻封堵效果做了觀察,其耐磨損性還有待進(jìn)一步研究。

本實(shí)驗(yàn)證實(shí)PMBE乙醇溶液涂布處理具備穩(wěn)定膠原基質(zhì)和封閉牙本質(zhì)小管的雙重功效。低濃度PMBE溶液僅具備明顯的穩(wěn)定膠原作用,4%及以上濃度還具備封閉牙本質(zhì)小管的效能,8%PMBE溶液效果最佳,PMBE具備一定治療牙本質(zhì)過敏的作用。

[1] Davari A, Ataei E, Assarzadeh H. Dentin hypersensitivity: etiology, diagnosis and treatment; a literature review[J]. J Dent (Shiraz), 2013, 14(3):136-145.

[2] Chinese Stomatological Association Expert Committee On Dentin Hypersensitivity. Guideline for the diagnosis and management of dentin hypersensitivity[J]. Chin J Dent Res,2015,18(1):13-16.

[3] Rosenthal MW. Historic review of the management of tooth hypersensitivity[J].Dent Clin North Am, 1990, (34):403-427.

[4] 伍春蓮.馬尾松樹皮提取物生物活性物質(zhì)的檢測及抗炎癥機(jī)理的初步研究[D].廣州:中山大學(xué),2006.

[5] Hang JH, Feng DR, Ma HL,etal. Antitumor effects of Pinus massoniana bark extract in murine sarcoma S180 both in vitro and in vivo[J]. Am J Chin Med, 2012, 40(4):861-875.

[6] Maimoona A, Naeem I, Saddiqe Z,etal. A review on biological, nutraceutical and clinical aspects of French maritime pine bark extract[J]. J Ethnopharmacol, 2011, 133(2):261-277.

[7] Günel C, Demirci B, Erylmaz A,etal. Inhibitory effect of pycnogenol?on airway inflammation in ovalbumin-induced allergic rhinitis[J]. Balkan Med J, 2016, 33(6): 620-626.

[8] 唐成芳,朱勇,阮建平,等.馬尾松樹皮提取物預(yù)處理對(duì)根面牙本質(zhì)耐酸蝕和脫礦能力的影響[J].山西醫(yī)科大學(xué)學(xué)報(bào),2016,47(8):732-736.

[9] Palazon MT, Scaramucci T, Aranha AC,etal. Immediate and short-term effects of in-office desensitizing treatments for dentinal tubule occlusion[J]. Dent Mater J, 2014, 33 (1): 125-132.

[10] Agrafioti A, Tzimpoulas N, Chatzitheodoridis E,etal. Comparative evaluation of sealing ability and microstructure of MTA and Biodentine after exposure to different environments[J]. Clin Oral Invest, 2016, 20(7):1535-1540.

[11] Petersson LG. The role of fluoride in the preventive management of dentin hypersensitivity and root caries[J]. Clin Oral Investig, 2013, 17(Suppl 1): 63-71.

[12] 陳麗薇,顧爽,賈興亞.生物活性玻璃與氟保護(hù)漆聯(lián)合應(yīng)用治療牙本質(zhì)過敏癥臨床療效研究[J].中國實(shí)用口腔科雜志,2015,8(3):162-166.

[13] 陳珊,張露露,柳志文.氟保護(hù)漆防齲的研究進(jìn)展[J].現(xiàn)代醫(yī)藥衛(wèi)生,2016,32(14):2172-2174.

[14] Bagchi D, Bagchi M, Stohs SJ,etal. Free radicals and grape seed proanthocyanidin extract: importance in human health and disease prevention[J]. Toxicology, 2000, 148(2-3):187-197.

[15] Gregoire S, Singh AP, Vorsa N,etal. Influence of cranberry phenolics on glucan synthesis by glucosyltransferases and Streptococcus mutans acidogenicity[J]. J Appl Microbiol, 2007, 103(5): 1960-1968.

[16] 畢玲.葡萄籽原花青素提取物預(yù)防齲病的實(shí)驗(yàn)研究[D].杭州:浙江大學(xué),2009.

[17] Liu R, Fang M, Xiao Y,etal. The effect of transient proanthocyanidins preconditioning on the cross-linking and mechanical properties of demineralized dentin[J]. J Mater Sci Mater Med, 2011, 22(11): 2403-2411.

[18] Liu RR, Fang M, Zhang L,etal. Anti-proteolytic capacity and bonding durability of proanthocyanidin-biomodified demineralized dentin matrix[J]. Int J Oral Sci, 2014, 6(3): 168-174.

[19] Green B, Yao X, Ganguly A,etal. Grape seed proanthocyanidins increase collagen biodegradation resistance in the dentin/adhesive interface when included in an adhesive[J]. J Dent, 2010, 38(11): 908-915.

[20] Pavan S, Xie Q, Hara AT,etal. Biomimetic approach for root caries prevention using a poanthocyanidin-rich agent[J]. Caries Res, 2011, 45(5): 443-447.

[21] Tang CF, Fang M, Liu RR,etal. The role of grape seed extract in the remineralization of demineralized dentine: micromorphological and physical analyses[J]. Arch Oral Biol, 2013, 58(12):1769-1776.

[22] 唐成芳,阮建平,朱勇,等.馬尾松松針提取物對(duì)根面牙本質(zhì)脫礦影響的體外研究[J].華西口腔醫(yī)學(xué)雜志,2016,34(5):521-525.

[23] Jeon JR, Kim EJ, Kim YM,etal. Use of grape seed and its natural polyphenol extracts as a natural organic coagulant for removal of cationic dyes[J]. Chemosphere, 2009, 77(8):1090-1098.

Effectsofpinusmassonianabarkextractondentinmatrixstabilizationanddentintubuleocclusioninvitro

TANG Chengfang1,ZENG Hui1,ZHANG Sha1,WANG Danyang1,YANG Liyan2,XIE Na1

(1DepartmentofStomatology,Xi’anMedicalUniversity,Xi’an710021,China;2DepartmentofPharmacy,Xi’anMedicalUniversity)

ObjectiveTo investigate the effect of pinus massoniana bark extract(PMBE) on dentin matrix stabilization and dentin tubule occlusion, and provide an experimental basis for the application of PMBE in treatment of dentin hypersensitivity.MethodsSixty dentin blocks were prepared and randomly divided into 6 groups(n=10): alcohol group, fluoride vanish group, 1% PMBE group, 2% PMBE group, 4% PMBE group and 8%PMBE group. The dentin blocks were etched to open the tubules with 37% phosphoric acid in each group, and then treated with absolute alcohol, fluoride vanish, 1%, 2%, 4% and 8%PMBE ethanol solution, respectively. The micro morphology of dentin surface and profile were observed using field emission scanning electron microscope(FESEM), and the area of dentin tubule openings(ADTO), percentage area of dentin tubule openings(PADTO) and sealed depth(SD) were also measured.ResultsThe ADTO and PADTO in 8%PMBE group were significantly lower than that in ethanol group(P<0.05), but there was no significant difference between 8% PMBE group and fluoride vanish group(P>0.05). The ADTO and PADTO in 4% PMBE group were lower than those in 1%, 2% PMBE and ethanol group, and higher than that in 8% PMBE group(P<0.05). The dentin tubules were opened completely in ethanol group, and the walls of tubule were looser. The dentin tubules of dentin surface were sealed completely in fluoride vanish group. The collagen matrix along the tube wall was clear and dense in PMBE groups, and the sealing degree of dentin tubule increased with the increase of PMBE concentration, and the dentin tubules of dentin surface were sealed almost in 8% PMBE group and some sealing substances were seen in profile.ConclusionLocal application of PMBE ethanol solution may stablize the dentin matrix and close the dentin tubules, and 8% PMBE could achieve a better effect of sealing dentin tubules. PMBE has certain therapeutic effects on dentin hypersensitivity.

pinus massoniana bark extract; dentin hypersensitivity; dentin tubule; fluoride vanish

國家自然科學(xué)基金資助項(xiàng)目(81641150)；陜西省科技廳社會(huì)發(fā)展項(xiàng)目(2015SF242)；西安醫(yī)學(xué)院博士啟動(dòng)基金資助項(xiàng)目(2015DOC20)

唐成芳,女,1977-12生,博士,副教授,E-mail:tangchengfang@sohu.com

2017-07-10

R781.2

A

1007-6611(2017)11-1160-05

10.13753/j.issn.1007-6611.2017.11.015