Microenvironmental effects on growth response of Pinus massoniana to climate at its northern boundary in the Tongbai Mountains,Central China

The Tongbai Mountains is an ecologically sensi-tive region and the northern boundary of Pinus massoniana Lamb.To analyze the effect of different microenvironments on tree growth response to climate factors,we developed standard chronologies for earlywood width(EWW),late-wood width(LWW),and total ring width(TRW)of P.massoniana at two sampling sites on slopes with different orientations,then analyzed characteristics of the chronolo-gies and their correlations with climate variables from five stations in the region and with a regional normalized differ-ence vegetation index(NDVI).Statistical results showed that the TRW/EWW/LWW chronology consistency and charac-teristics(mean sensitivity,signal to noise ratio,expressed population signal)for trees growing on the southeastern slope were much higher than for trees on the northeastern slope.Correlations indicated that temperature in current March and August has a significant positive effect on TRW/EWW/LWW formation,and the effect on the northeastern slope was weaker than on the southeastern slope.Compared to temperature,precipitation has more complicated effects on tree growth,but the effect on the northeastern slope was also generally weaker than on the southeastern slope.Step-wise linear regression analyses showed that temperature in August was the main limiting factor at the two sampling sites.Similarly,the response of tree growth on the southeast-ern slope as determined by the NDVI is better than on the northeastern slope,and the TRW/EWW/LWW chronologies for the southeastern slope explained over 50%of the total NDVI variances in June.Overall,the results indicate that the difference in the climate response of P.massoniana at two sampling sites is clearly caused by differences in the microenvironment,and such differences should be properly considered in future studies of forest dynamics and climate reconstructions.

Old Pinus massoniana forests benefit more from recent rapid warming in humid subtropical areas of central-southern China

Trees progress through various growth stages,each marked by specific responses and adaptation strate-gies to environmental conditions.Despite the importance of age-related growth responses on overall forest health and management policies,limited knowledge exists regarding age-related effects on dendroclimatic relationships in key subtropical tree species.In this study,we employed a den-drochronological method to examine the impact of rapid warming on growth dynamics and climatic sensitivity of young(40–60 years)and old(100–180 years)Pinus mas-soniana forests across six sites in central-southern China.The normalized log basal area increment of trees in both age groups increased significantly following rapid warming in 1984.Trees in young forests further showed a distinct growth decline during a prolonged severe drought(2004–2013),whereas those in old forests maintained growth increases.Tree growth was more strongly influenced by temperature than by moisture,particularly in old forests.Spring tem-peratures strongly and positively impacted the growth of old trees but had a weaker effect on young ones.Old forests had a significantly lower resistance to extreme drought but faster recovery compared to young forests.The“divergence problem”was more pronounced in younger forests due to their heightened sensitivity to warming-induced drought and heat stress.With ongoing warming,young forests also may initially experience a growth decline due to their heightened sensitivity to winter drought.Our findings underscore the importance of considering age-dependent changes in forest/tree growth response to warming in subtropical forest man-agement,particularly in the context of achieving“Carbon Peak&Carbon Neutrality”goals in China.

Study on Antibacterial Effects of Extracts from Pinus massoniana Lamb.Needles

[Objectives]To study the antibacterial effects of extracts from Pinus massoniana Lamb.needles.[Methods]In this experiment,the components from Pinus massoniana Lamb.needles were extracted by systematic solvent extraction method,and the antibacterial activity against four common bacteria,Escherichia coli,Staphylococcus aureus,Bacillus subtilis,Aspergillus flavus and the antibacterial active component were examined for by punch method.[Results]Different solvent extraction rate was different,the rates of petroleum ether,chloroform,ethyl acetate,n-butanol,water extracts were 4.2%,16.7%,17.4%,21.1%,40.6%.All extracts showed inhibitory effect against test bacteria.It was observed that the inhibition of G+was stronger than G-,and the extracts had the best antibacterial activity to Staphylococcus aureus while the weakest to Aspergillus flavus.The antibacterial activity of the components decreased in the order:ethyl acetate extract>n-butanol extract>chloroform extract>petroleum ether extract>aqueous phase.The extracts were stable under ultraviolet radiation(UV)light and long term storage.The antibacterial activity of the extracts was weaker with the increase of pH value when the pH value≤8.[Conclusions]It is inferred that the antibacterial components in the extract of Pinus massoniana needles are widely distributed,and the components with medium polarity or above are the main antibacterial components.

Chitosan Oligosaccharide Induces Plant Resistance Gene Expression in Pinus massoniana

Chitosan oligosaccharides(COSs)are the main degradation products from chitosan or chitin and have been reported to induce resistance to diseases in herbaceous plants like cucumber and Arabidopsis.Concomitantly,pine wilt disease(PWD)is a devastating disease of conifer tree species.Here,we hypothesized that COSs induce plant resistance gene(PRG)expression in the woody plant Masson pine,Pinus massoniana.COSs were inoculated into P.massoniana seedlings and the BGISEQ-500 platform was used to generate transcriptomes from COSs-treated P.massoniana and control seedlings.A total of 501 differentially expressed genes(DEGs)were identified by comparing the treatment and control groups.A total of 251(50.1%)DEGs were up-regulated in the treatment relative to the control seedlings and 250(49.9%)were down-regulated.Inoculation of COSs induced the expression of 31 PRGs in P.massoniana seedlings and the relative expression levels of six of the PRGs were verified by RT-qPCR.This is the first study to demonstrate that COS induces the expression of PRGs in a tree species.These results provide important insights into the function of COSs and further the prospects of developing a COS-based immune inducer for controlling PWD.

模拟氮沉降和接种外生菌根真菌对马尾松(Pinus massoniana)幼苗营养元素的影响

量化植物地上部和地下部元素含量对于理解和预测植物养分平衡如何响应大气氮沉降的变化至关重要。通过盆栽试验研究了氮沉降增加背景下外生菌根真菌对马尾松幼苗营养元素的影响。对马尾松幼苗进行了接种两种外生菌根真菌:(彩色豆马勃(Pisolithus tinctorius,Pt)与厚环乳牛肝菌(Suillus grevillei,Sg))以及4种氮素浓度添加:0 kg N hm^(-2)a^(-1)(N0)、正常氮沉降30 kg N hm^(-2)a^(-1)(N30)、中度氮沉降60 kg N hm^(-2)a^(-1)(N60)、重度氮沉降90 kg N hm^(-2)a^(-1)(N90),共12个处理,测定了马尾松地上部和地下部大量元素和微量元素的含量。结果表明:施氮改变了营养元素在马尾松幼苗地上部和地下部的含量,马尾松幼苗磷(P)、钙(Ca)、铁(Fe)、锰(Mn)等元素均在N60时达到临界值,而当输入的量超过了马尾松对氮的需求时,氮沉降会使马尾松营养元素含量较最适浓度时降低,地上部碳(C)随施氮浓度的升高先升高后降低,N随施氮浓度的升高而升高,根系和叶片钾(K)、Ca、镁(Mg)均随施氮浓度的升高而降低,施氮也降低了根系C及微量元素的含量。但在同一施氮浓度下,接种外生菌根真菌(EMF)后能够提高大多数元素的含量,N90时接种厚环乳牛肝菌(Sg)和彩色豆马勃(Pt)的叶片N含量与对照相比分别提高112.6%和138.6%,根系N含量分别提高73.1%、71.6%;N60时接种Sg和Pt的植株叶片P含量比不施氮未接种对照分别提高了166.3%、132.9%,根系P含量分别提高了40.8%、38.5%。EMF能够维持植物养分平衡,从而降低高施氮量对植物的影响效果。这为未来气候变化情景中氮沉降增加下接种EMF可以调节植物元素含量,从而达到更适应环境的元素平衡来促进生长提供理论依据。

Differential aluminum tolerance and absorption characteristics in Pinus massoniana seedlings colonized with ectomycorrhizal fungi of Lactarius deliciosus and Pisolithus tinctorius

Plant tolerance to aluminum(Al)toxicity can be enhanced by an ectomycorrhizal(ECM)fungus through biological filtering or physical blockage.To understand the roles of ECM colonization in Al absorption with regard to Al tolerance,Pinus massoniana seedlings were inoculated with either Lactarius deliciosus(L.:Fr.)Gray isolate 2 or Pisolithus tinctorius(Pers.)Coker et Couch isolate 715 and cultivated in an acid yellow soil with or without 1.0 mM Al^(3+)irrigation for 10 weeks.Biomass production,Al bioaccumulation and transport in seedlings colonized by the two ECM fungi were compared,and the three absorption kinetics(pseudo-first order,pseudo-second order and intraparticle diffusion)models used to evaluate variances in root Al^(3+)absorption capacity.Results show that both fungi increased aboveground biomass and Al tolerance of P.massoniana seedlings,but L.deliciosus 2 was more effective than P.tinctorius 715.Lower Al absorption capacity,fewer available active sites and decreased affinity and boundary layer thickness for Al^(3+),and higher Al accumulation and translocation contributed to the increased Al tolerance in the ECM-inoculated seedlings.These results advance our understanding of the mechanisms and strategies in plant Alto lerance conferred by ECM fungi and show that inoculation with L.deliciosus will better enhance Al tolerance in P.massoniana seedlings used for forest plantation and ecosystem restoration in acidic soils,particularly in Southwest China and similar soils worldwide.

马尾松(Pinus massoniana)、湿地松(Pinus elliottii)挥发性化学物质的昼夜节律释放

用TCT-GC/MS分析了马尾松、湿地松挥发性有机化合物的昼夜节律变化。结果表明:马尾松昼夜节律中检测到的挥发物主要是萜烯类化合物,其中单萜种类最多,且α-蒎烯和β-蒎烯含量约占整个挥发物的80%,其次是含氧化合物等。这些挥发物的释放高峰多数在10:30,少数在1:30;整个变化中有两个低峰期,即13:30和22:30。湿地松中检测到的挥发物组分与马尾松相似,多数释放高峰在12:00~15:00之间;另一些在6:00;α-蒎烯的释放高峰在3:00,而此时其它挥发性化合物的释放量最低。挥发物的释放也受到外界环境的影响,一定范围内随着温度的升高、湿度的减小,其释放量增加。

虫害马尾松(Pinus massoniana Lamb)邻枝针叶挥发物及其内源茉莉酸甲酯的快速变化

对盆栽马尾松针叶进行接虫咬食危害处理后,通过TCT-GC/MS分析了同一株受害枝相邻的健康枝针叶挥发物的成分及相对含量的时序变化。结果表明:萜烯类化合物是邻枝针叶挥发物的主要成分,其次是含氧化合物、含氮化合物等。与对照相比,多数挥发物的相对含量1h略高于对照,2h维持较高水平。同时,用GC/MS分析了邻枝针叶不同时间序列中茉莉酸甲酯的含量,结果表明:虫害马尾松邻枝针叶1h茉莉酸甲酯含量就有所升高,2h显著高于对照,是对照的近1倍。证明马尾松受到虫害后,启动了体内的防御系统,并诱导邻枝产生抗性。

被害马尾松(Pinus massoniana)针叶挥发性物质的提取、鉴定及蚕饰腹寄蝇(Blepharipa zebina)的电生理活性

试验使用动态顶空吸附的方法,提取了被害马尾松(Pinus massoniana)针叶挥发物,并通过气相色谱-质谱联用仪(GC-MS)及标准样品进行鉴定。结果表明,被害马尾松针叶挥发性物质主要包括绿叶挥发物、单萜、含氧单萜及倍半萜等四大类物质;挥发物中以单萜的含量最高,其它3类物质含量较低。通过触角电位图(EAG)技术,测定了23种挥发性物质标准样品的石蜡油溶液对蚕饰腹寄蝇(Blepharipa zebina)雌蝇的电生理活性。结果表明,蚕饰腹寄蝇雌蝇对α-蒎烯、β-蒎烯、长叶烯、莰烯、乙酸冰片酯等的相对触角电位反应值与湿润空气对照相比并无显著差异;雌蝇对其它刺激物的相对触角电位反应值与湿润空气对照相比均存在显著差异。同时测定了雌蝇对8种标样,在5个浓度系列下的剂量-反应,并分析了反应阈值。结果表明,蚕饰腹寄蝇雌蝇对顺-3-己烯-1-醇和里哪醇的反应阈值为0.1μg/μl,对壬醛的反应阈值为1μg/μl,对柠檬烯、α-水芹烯、2,6-二甲基-2,4,6-辛三烯、β-石竹烯4种物质的反应阈值为100μg/μl,而对α-蒎烯,在测试的剂量范围内不存在反应阈值。在确定对蚕饰腹寄蝇有生理活性的物质成分和分析反应阈值的基础上,对寄生蝇类定位寄主的机制进行了讨论。

三峡库区马尾松(Pinus massoniana)林林分结构特征对灌木层物种多样性的影响

林分结构特征对灌木层物种多样性有重要影响,对其之间关系进行研究,可为提高生态系统功能稳定性、进行合理的林分经营管理提供理论依据。以马尾松林为研究对象,采用多元逐步回归、因子分析和典型相关分析等方法分上林层、中林层和下林层研究林分结构对灌木层多样性的影响。结果表明:(1)Pearson相关分析结果表明,上林层是影响灌木层物种多样性的主要层次,而林分总体结构特征与灌木层物种多样性无显著性关系;(2)用13个林分结构参数和灌木层物种多样性各参数进行多元逐步回归,拟合方程都通过显著性检验,R2为0.246-0.630,且仅剔除了树高和大小比数2个变量而保留了其它11个林分结构变量,说明林分结构的综合作用对灌木层物种多样性有较大影响;(3)林分结构可分为3类,分别为林分竞争状况因子、水平结构及冠幅因子和垂直结构及生长因子;(4)上林层林分结构总体上与灌木层物种多样性之间存在着较强的相关性,典型相关系数为0.822(P=0.014),其中垂直结构及生长因子为主要影响因子,权重为0.962。在马尾松林中,上林层林分结构中的垂直结构与生长因子和灌木层物种多样性表现为正相关关系,在经营管理中,可通过调控上林层林木的胸径、冠长及冠层垂直结构等以达到提高林分灌木层物种多样性的目的。

Branching and growth of plantings in fifth year of a seedling seed orchard of Masson pine (Pinus massoniana Lamb.)

The features of branching and growth studied included height, diameter at breast height (DBH), total number of branches, annual height growth, annual branch elongation in the year of elongating, annual branch number for four consecutive years, diameter of branches of different ages, and diameter of stem where branch-whorl originates. For features of total growth and overall branching, no significant differences were found between families, except for DBH. For annual features, no significant differences were found in annual stem height growth, annual branch elongation in the year of elongation and diameter of branches. In the last four years, differences in number of branches were not significant in the first two years but were significant in the last two year; differences in stem diameter where branch-whorls grow were significant for the four consecutive years. Trend of annual growth and branching features of families can be divided into three types as increasing type, stable type and fluctuating type. Most of families have an increasing type with respect of annual height growth and annual branch elongation, while most families belong to a fluctuating type with annual branch number. The results indicated that in the fifth year after planted in seedling seed orchard, differences between families were mostly insignificant. This result may have two main explanations: one is the growth rhyme in early ages of Masson pine, the other one is the complex paternal components to form the open-pollinated families. Family selection seemed to be not useful based on the result. It is suggested to select some of families in the nursery instead of to use all the families when establishing seedling seed orchards with open-pollinated families from plus-trees.

柱前衍生反相高效液相色谱法测定马尾松(Pinus massoniana)针叶中γ-氨基丁酸和17种游离氨基酸

建立了自动在线柱前衍生反相高效液相色谱法同时测定γ-氨基丁酸（GABA）和17种游离氨基酸含量的方法。以邻苯二甲醛-9-芴基甲基氯甲酸酯（OPA—FMOC）为衍生试剂进行衍生，AgilentHypersilAA-ODS-C18色谱柱分离，梯度洗脱，二极管阵列检测器检测，在19min内分离测定了马尾松苗木针叶中GABA和17种游离氨基酸的含量。该方法测定氨基酸的回收率高于90．1％，精密度和重现性均较好（相对标准偏差为0．21％～2．81％），经测定，发现马尾松被马尾松毛虫取食后，所测18种氨基酸总量明显降低，从418．3μg·g^-1降低到310．4μg·g^-1鲜重。

浙江省马尾松(Pinus massoniana)林土壤呼吸的研究

本文研究了浙江省马尾松（Pinusmassoniana）林土壤呼吸速率与土壤温度的关系，建立土壤呼吸速率与地表温度的模型，根据气象资料估算得到马尾松林的年均土壤CO2呼吸量为25．33tCO2·hm-2a-1，浙江省马尾松林每年土壤CO2排放量为3．60×107tCO2·a-1．

不同林龄马尾松(Pinus massoniana)人工林碳氮磷分配格局及化学计量特征

为了解长江上游低山丘陵区马尾松（Pinus massoniana）人工林生态系统的C、N、P分配格局及化学计量特征，本文采用时空互代的方法，在宜宾高县来复林区选取三种不同林龄（5年生幼龄林、14年生中龄林、39年生成熟林），但立地条件相近、样地情况基本一致的马尾松（Pinus massoniana）人工林作为研究对象，对马尾松针叶、凋落物及土壤中的C、N、P含量及 w（C）？w（N）？w（P）化学计量特征进行测定和分析。结果表明，（1）C、N、P 含量均表现为针叶〉凋落物〉土壤，且在三个库之间差异显著；（2）林龄对针叶、凋落物、土壤的 C、N、P 及 w（C）？w（N）、w（C）？w（P）计量比均有显著影响。（3）土壤 C、N、P含量在成熟林中最高；针叶和凋落物的C含量在成熟林中最低，N、P含量则在中龄林中最高。（4）随林龄增加马尾松对N、P的利用效率降低，针叶、凋落物及土壤的w（C）？w（N）与 w（C）？w（P）均表现为下降。（5）马尾松针叶w（N）？w（P）比值在14.37~15.53之间，说明该地区马尾松人工林受N和P的共同限制，但林龄对N、P养分限制的影响不显著。为提高该区马尾松人工林的生产力，建议在人工林的抚育管理中要适当增加N肥和P肥，同时也可在马尾松人工林引入豆科固氮植物以提高地力。该研究将马尾松针叶、凋落物及土壤结合起来探究随林龄增长C、N、P养分元素的分配格局及化学计量特征的变化，有助于全面、系统地揭示马尾松人工林生态系统的养分循环，对指导马尾松人工林生产，调节和改善林木生长环境，提高系统的养分利用效率及林地生产力具有重要意义。

不同郁闭度控制下马尾松(Pinus massoniana)人工林土壤动物群落特征

采用郁闭度控制方法,形成马尾松人工林郁闭度梯度(0.5,0.6,0.7,0.8,0.9)试验样地,研究不同郁闭度下土壤动物群落特征。运用手捡法、Tullgren法和Baermann法进行土壤动物群落调查。结果显示,1)共捕获土壤动物8 860只,隶属于4门11纲25目111科,其中大型土壤动物589只,以蜚蠊、疣和康叭为优势类群;中小型土壤动物8271只,以等节和丽甲螨为优势类群。2)大型土壤动物以杂食性为主,杂食性在0.9郁闭度显著增加,捕食性显著减少;中小型土壤动物以菌食和腐食性为主,腐食性土壤动物随郁闭度增大逐渐增加,菌食性逐渐减少。中小型捕食性土壤动物在郁闭度0.5—0.7显著增加,0.7—0.9无显著变化。3)大型土壤动物仅5—10 cm土层类群数在0.9郁闭度显著增加;中小型土壤动物总个体数及枯落物层个体数、5—10 cm土层类群数显著增加(0.5—0.7)后减少(0.7—0.9);中小型土壤动物总类群数在郁闭度0.5—0.6显著增加,枯落层类群数在郁闭度0.5—0.8显著增加,0.8—0.9显著减少。4)大型、中小型土壤动物多样性在土层中差异显著。中小型土壤动物Simpson优势度指数随郁闭度增大而减小,Shannon-Wiener多样性指数、Pielou均匀性指数则显著增大,Margalef丰富度指数先显著增高(0.5—0.6)后趋于平稳(0.6—0.9);大型土壤动物Jaccard相似性指数低于中小型。5)CCA分析表明,不同类群土壤动物对环境因子响应不同,郁闭度、含水量、有机质及全P对土壤动物类群影响显著。研究表明,0.7郁闭度马尾松人工林下土壤动物优势度、丰富度适中,林下植被恢复情况较好,该郁闭度可能更有利于土壤动物多样性和群落稳定性。

封育次生马尾松(Pinus massoniana)林优势种群竞争密度效应

以退化红壤区封山育林植被恢复过程中优势种群马尾松为研究对象,研究不同恢复阶段马尾松种群竞争密度效应,并提出一个适用性更强的竞争密度效应模型.研究表明,退化红壤区植被恢复过程中优势种群马尾松竞争密度效应明显,优势种群平均胸径(D)、平均胸高断面积(S)、平均单株材积(V)、林分蓄积量 (M)与种群密度 (ρ)的竞争密度关系密切,达到极显著水平(α<0. 01);新竞争密度效应模型较前人模型描述C-D效应效果更理想,可在森林竞争密度效应研究中应用. 表 4参

Modeling compatible single-tree aboveground biomass equations for masson pine(Pinus massoniana) in southern China

Because of global climate change, it is necessary to add forest biomass estimation to national forest resource monitoring. The biomass equations developed for forest biomass estimation should be compatible with volume equations. Based on the tree volume and aboveground biomass data of Masson pine （Pinus massoniana Lamb.） in southern China, we constructed one-, two- and three-variable aboveground biomass equations and biomass conversion functions compatible with tree volume equations by using error-in-variable simultaneous equations. The prediction precision of aboveground biomass estimates from one variable equa- tion exceeded 95%. The regressions of aboveground biomass equations were improved slightly when tree height and crown width were used together with diameter on breast height, although the contributions to regressions were statistically insignificant. For the biomass conversion function on one variable, the conversion factor decreased with increasing diameter, but for the conversion function on two variables, the conversion factor increased with increasing diameter but decreased with in- creasing tree height.

Improving the assessment method of seed vigor in Cunninghamia lanceolata and Pinus massoniana based on oxygen sensing technology

Oxygen sensing technology was employed to study the rapid methods for seed vigor assessment of Chinese fir （Cunninghamia lanceolata） and Masson pine （Pinus massoniana）. Firstly, seeds of five lots were performed using accelerated aging （AA） into three vigor levels. Then, four oxygen sensing indices, including increased metabolism time （IMT）, oxygen metabolism rate （OMR）, critical oxygen pressure （COP）, relative germination time （RGT） and the control indiees such as labora- tory germination indices, dehydrogenase activity （DA）, and electrical conductivity （EC） were analyzed by the tests of 15 samples. The results of correlation analysis between these indices and field emergence per- formances based on two-year and two-spot data showed that RGT and OMR should be indicated as the optimal oxygen sensing indices to rap- idly and automatically evaluate seed vigor of Chinese fir and Masson pine, respectively. On the basis, one-variable linear regression equations were built to forecast their field emergence performances by the two oxygen sensing indices.

The essential oil components of Cinnamomum cassia: an analysis under different thinning models of plantation Pinus massoniana

Thinning is an important activity employed in forest management. To date, studies have mainly focused on the effects of thinning on the growth of trees during the same thinning period. In this study, plantation Pinus massoniana Lamb. near maturity were thinned at varying intensities and an economically important species, Cinnamomum cassia Presl., was planted beneath the thinned canopy. The aim of the study was to explore the effects of the extent of thinning on the essential oil content and its components of C. cassia in different parts of the plant, as well as the economic feasibility of the P. massoniana-C.cassia management model. Thinning significantly reduced the oil yield in the bark and branches of C. cassia, but hardly impacted the oil yield from C. cassia leaves compared with pure C. cassia forest（CK）. Among the different thinning treatments, both light（T.4） and extensive（T.1）thinning reduced the oil yield of C. cassia bark and new branches. The concentrations of the main aldehydes differed in different parts of the plant and were affected by the extent of thinning. The influence on cinnamaldehyde in the bark was minor, but was much greater in the branches and leaves. Both the oil yield and content of cinnamaldehyde showed that moderate（T.3） thinning was more favorable than other thinning models. These results not only provide a potentially promising model for the transformation of low-yield artificial pure forests of P. massoniana in the future, but also offer a reference for the management of artificial mixed stands.

Effects of Pinus massoniana bark extract on cell proliferation and apoptosis of human hepatoma BEL-7402 cells

AIM: To study the effects of Pinus massoniana bark extract (PMBE) on cell proliferation and apoptosis of human hepatoma BEL-7402 cells and to elucidate its molecular mechanism.METHODS: BEL-7402 cells were incubated with various concentrations (20-200 μg/mL) of PMBE for different periods of time. After 48 h, cell proliferation was determined by 3-(4,5-dimethyl-thiazolyl-2)-2,5-diphenyl tetrazolium bromide (MTT) assay. Apoptosis was evaluated by morphological observation, agarose gel electrophoresis,and flow cytometry analysis. Possible molecular mechanisms were primarily explored through immunohistochemical staining.RESULTS: PMBE (20-200 μg/mL) significantly suppressed BEL-7402 cell proliferation in a time- and dose-dependent manner. After treatment of BEL-7402 cells with 160 μg/mL PMBE for 24, 48, or 72 h, a typical apoptotic 'DNA ladder'was observed using agarose gel electrophoresis. Nuclear condensation and boundary aggregation or split, apoptotic bodies were seen by fluorescence and electron microscopy.Sub-G1 curves were displayed by flow cytometry analysis.PMBE decreased the expression levels of Bcl-2 protein in a time-dependent manner after treatment of cells with 160 μg/mL PMBE.CONCLUSION: PMBE suppresses proliferation of BEL-7402 cells in a time- and dose-dependent manner and induces cell apoptosis by possibly downregulating the expression of the bcl-2 gene.