Comparison of expression of apoptosis regulatory proteins in the adipose tissue of tumor-bearing and diet-restricted rabbits

We investigated the expression of apoptosis regulatory proteins in the adipose tissue of tumor-bearing and diet-restricted rabbits, and analyzed the differences between the two groups. The apoptotic index in the tumor-bearing group was 25.1+/-12.4 on day 10 and increased to 27.2+/-14.4 by day 20. Thereafter, however, it gradually decreased, falling to 11.2+/-7.8 on day 30 and 0.67+/-0.88 on day 40. By contrast, no apoptotic cells were detected in the diet-restricted group at any of the times examined. Bcl-2 immunoreactivity was either not detected at all, or was only weakly observed in both groups. Bax expression, on the other hand, gradually increased after implantation in the tumor-bearing group. In 2 of the 5 tumor-bearing rabbits, Bax expression in adipocytes was moderate 10 days after tumor implantation, and strong by day 20, but steadily decreased thereafter. By contrast, hardly any Bax-immunopositive cells were detected in the diet-restricted group. These results suggested that loss of body weight in the tumor-bearing group was different from that in the diet-restricted group, and that it was related to adipocyte apoptosis.     

Bax,Bcl-2, fas and Fas-L antigen expression in human seminoma: correlation with the apoptotic index

Apoptosis plays a crucial role in the regulation of spermatogenesis in male germ cells and is, at least in part, modulated by Bcl-2, Bax, and the Fas pathway. Seminomas have a favourable outcome and respond to radio-/chemotherapy with an increased rate of apoptosis. The expression of Bax, Bcl-2, Fas and Fas-ligand (Fas-L) in human seminoma was evaluated and correlated with the apoptotic index. Twenty-nine classical seminomas were examined by immunohistochemistry and Western blotting using antibodies against Bax, Bcl-2, Fas and Fas-L. Apoptosis was detected by in-situ end-labeling of fragmented DNA and the apoptotic index (AI) was determined. Expression of Fas was found in 26 (89.7%) of Fas-L in 24 seminomas (82.2%); none of the tumours expressed Bcl-2. No correlation between the AI and Fas, Fas-L or Bcl-2 expression was found. Bax was demonstrated in 20/29 tumours (69%). Bax-positive tumours showed an increased AI of 4.75 +/- 2.38% in contrast to 2.60 +/- 1.23% of the Bax-negative tumours (P = 0.002). The number of Bax-positive tumour cells and apoptotic cells revealed a significant correlation using chi2-test (P = 0.04) and linear regression (r = 0.54, P = 0.001). Therefore, Bax seems to play a determinant role in the modulation of apoptosis in human seminoma that may be linked to a favourable outcome.     

Apoptosis and the expression of Bax and Bcl-2 in hyperplasia and adenocarcinoma of the uterine endometrium

BACKGROUND: Apoptosis plays a crucial role in carcinogenesis in various tumours. This study was designed to investigate the occurrence of apoptosis and the expression of Bcl-2 and Bax proteins in endometrial tumours of corpus uteri. METHODS: Endometrial tissues were obtained from 20 patients with endometrioid adenocarcinoma, 16 patients with endometrial hyperplasia, and 4 patients with myoma uteri (which were used as controls). The occurrence of apoptosis was examined by using molecular biochemical techniques. The expression of Bcl-2 and Bax proteins was also investigated using immunohistochemical staining with appropriate antibodies. RESULTS: The labelling of DNA in situ indicated that apoptotic cells were sporadically seen in postmenopausal endometrium (5.2 +/- 2.1, n = 4) and endometrial hyperplasia without atypia (2.6 +/- 0.5, n = 9). In contrast, labelled cells were detected in atypical endometrial hyperplasia (15.9 +/- 2.2, n = 7), and their numbers increased intensely in adenocarcinoma (29.3 +/- 3.7, n = 20). Autoradiographic analysis revealed DNA laddering in many cases of carcinoma. Bcl-2 was highly immunopositive in hyperplasia without atypia (36.2 +/- 6.5%, n = 9), but was decreased in the atypical endometrial hyperplasia (16.3 +/- 4.8%, n = 7). Large fractions of the carcinoma (6.3 +/- 1.8%, n = 20) and normal endometrium (2.8 +/- 1.4%, n = 4) were immunonegative or slightly immunopositive to Bcl-2. In contrast, Bax immunoreactivity was more frequent and stronger in adenocarcinoma (43.6 +/- 4.1%, n = 20) than that in normal endometrium (17.6 +/- 6.7%, n = 4) and hyperplasia (7.2 +/- 2.2%, n = 16). CONCLUSIONS: These results suggest that cells in hyperplasia expressing Bcl-2 might have prolonged survival ability. Neoplastic cells in adenocarcinoma might show apoptosis in association with a decreased expression of Bcl-2 and an increased expression of Bax. Therefore, the frequency of apoptosis and the expression of Bcl-2 and Bax might be correlated with carcinogenesis in the uterine endometrium of humans.     

Antropyloric G-cell hyperplasia in hypercalcemic rabbits bearing the VX2 carcinoma.

The number of distribution and the numbers of G cells in the antropyloric region of the rabbit stomach were mapped employing immunoperoxidase localization and morphometric quantitation and compared to similar analyses in hypercalcemic rabbits bearing the VX2 carcinoma. In normal animals, G cells were confined to the lower third of the antropyloric mucosa, where they were randomyly distributed within the mucosal glands. In contrast, tumor-bearing animals showed an extension of these cells into the middle third of the antropyloric mucosa. The absolute counts of G cells in control rabbits were 5.3 +/- 0.78 (mean +/- SE) per unit area, while those in hypercalcemic tumor-bearing rabbits were 11.9 +/- 0.46, a statistically significant increase. It is concluded that rabbits bearing VX2 carcinoma have G-cell hyperplasia.     

Changed Bcl:Bax ratio in endometrium of patients with unexplained infertility

Apoptosis has been shown to be an important regulator of endometrial function during the menstrual cycle and implantation. Recently, some possible implantation defects were identified in patients with unexplained infertility. In this study, we investigated the role of spontaneous apoptosis, which is regulated by death regulatory genes, such as Bcl-2, Bax, p53, and isoenzymes of nitric oxide synthases; eNOS and iNOS during the implantation window in women with unexplained infertility. Endometrial samples were evaluated from fertile (n=15) and unexplained-infertile women (n=15) during post-ovulatory 7th or 8th day of their menstrual cycles. Apoptotic cells were detected using the dUTP nick-end labelling assay and Bcl-2, Bax, p53, iNOS and eNOS were assessed immunohistochemically. Reduced apoptotic cells, weak immunoreactivity of p53 and strong immunoreactivity of Bcl-2 were observed in the unexplained-infertile group compared with the fertile group (p<0.001). Bax intensity was similar in both groups. While weak iNOS immunoreactivity was detected in both groups, moderately increased eNOS immunoreactivity was observed in infertile cases. Spontaneous apoptosis is reduced in the endometrium of unexplained-infertile women, and is associated with the changed Bcl-2:Bax ratio. This finding may be a contributing factor to defective implantation causing infertility in this group of patients.     

Involvement of angiogenesis in weight-loss in tumor-bearing and diet-restricted animals

The body's weight loss mechanism while in a tumor-bearing state is still unclear. In this study, we investigated expressions of angiogenic factors in the adipose tissue of tumor-bearing and diet-restricted rabbits evaluating the differences between the two groups. We postulated that low nutrition induced vasculogenesis to transport nutrition in the adipose tissues of diet-restricted rabbits, unlike in tumor-bearing rabbits, and that vascular endothelial growth factor (VEGF) and platelet-derived endothelial cell growth factor (PD-ECGF) were related to angiogenesis of the adipose tissues. Although we investigated the expressions of VEGF and PD-ECGF immunohistochemically in tumor-bearing and diet-restricted rabbits, there was no significant difference between the two groups. Whether angiogenesis of the adipose tissue in the diet-restricted animals may be observed during the nutritional recovery period should be investigated.     

缺血再灌注对在体兔窦房结细胞凋亡及凋亡相关基因表达的影响

目的:探讨缺血再灌注对在体兔窦房结细胞凋亡及凋亡相关基因表达的影响。方法:取家兔90只随机分为对照组, 缺血10 min、30 min、60 min、120 min组及缺血10 min、30 min、60 min、120 min再灌注4h组, 每组10只。通过结扎及放松右冠状动脉起始部制作窦房结缺血再灌注损伤模型, 当达各预定时点后, 迅速切取窦房结组织固定, 用TUNEL法检测窦房结细胞凋亡, 用免疫组化法检测窦房结细胞Fas-L、Bax及Bcl-2表达。结果:①对照组、缺血10 min、30 min组均未观察到明显的窦房结细胞凋亡现象;缺血60 min、120 min组及缺血再灌注4组中共有68.3%(41/60)的兔窦房结细胞出现不同程度的凋亡现象, 其细胞凋亡率分别为8.6%、16.1%、23.5%、34.5%、44.7%与31.2%。②Fas-L、Bax表达量随缺血时间延长逐渐增加, 以缺血120 min组表达最强;而Bcl-2表达则以缺血60 min组最强。③缺血再灌注各组中, Fas-L、Bax表达均明显强于对照组, 并以缺血60 min再灌注4h组最强;Bcl-2表达以缺血30 min再灌注4h组最强。④缺血再灌注组窦房结细胞Fas-L、Bax表达明显高于相同时间缺血组(P＜0.01)。结论:缺血及缺血再灌注均可诱导在体兔窦房结细胞凋亡, 凋亡相关基因Fas-L、Bax、Bcl-2可能参与了细胞凋亡的调控过程, 缺血再灌注损伤对窦房结细胞凋亡的发生具有促进作用。     

不同制动时相兔骨骼肌萎缩与细胞凋亡的实验研究-原位缺口末端标记法(TUNEL)的观察检测

探讨制动后骨骼肌萎缩的发生机制与骨骼肌细胞凋亡的关系。按照 Sievanen法制备不同时相的制动实验组 ,2 4只实验兔随机分为 4组 ,每组 6只 ,实验侧用管形石膏固定 ,自身未固定侧做对照组。在制动后 3、7、14和 2 8d取材 ,应用末端转移酶介导的 U TP缺口末端标记法 (Td T— mediated d UTP nick end labeling,TUNEL )检测萎缩骨骼肌中有无凋亡的骨骼肌细胞 ,并与形态学的观察结果相对照。对各组的数据进行统计学处理。结果表明不同制动时相所致的萎缩骨骼肌中均可发现有程度不一的骨骼肌细胞发生凋亡 ,以 14 d组最为显著。不同时相凋亡的肌细胞在空间的分布上呈不一致性。制动所致萎缩的骨骼肌中有时可见呈假阳性 TU NEL染色 ,但与凋亡的肌细胞的表现是不同的。我们认为 (1)制动后骨骼肌萎缩有骨骼肌细胞凋亡的参与。 (2 )骨骼肌细胞凋亡的多少与制动时相密切相关 ,以制动后 14 d最为明显。 (3)凋亡的骨骼肌细胞在空间分布上随制动时相不同而不同 ,呈不一致性。 (4 )制动后骨骼肌萎缩的程度与肌细胞凋亡的程度密切相关。     

Age-related activation of mitochondrial caspase-independent apoptotic signaling in rat gastrocnemius muscle

Mitochondria-mediated apoptosis represents a central process driving age-related muscle loss. However, the temporal relation between mitochondrial apoptotic signaling and sarcopenia as well as the regulation of release of pro-apoptotic factors from the mitochondria has not been elucidated. In this study, we investigated mitochondrial apoptotic signaling in skeletal muscle of rats across a wide age range. We also investigated whether mitochondrial-driven apoptosis was accompanied by changes in the expression of Bcl-2 proteins and components of the mitochondrial permeability transition pore (mPTP). Analyses were performed on gastrocnemius muscle of 8-, 18-, 29- and 37-month-old male Fischer344 x Brown Norway rats (9 per group). Muscle weight declined progressively with advancing age, concomitant with increased apoptotic DNA fragmentation. Cytosolic and nuclear levels of apoptosis inducing factor (AIF) and endonuclease G (EndoG) increased in old and senescent animals. In contrast, cytosolic levels of cytochrome c were unchanged with age. Mitochondrial Bcl-2, Bax and Bid increased dramatically in 37-month-old rats, with no changes in the Bax/Bcl-2 ratio in any of the age groups. Finally, expression of cyclophilin D (CyPD) was enhanced at very old age. Our findings indicate that the mitochondrial caspase-independent apoptotic pathway may play a more prominent role in skeletal muscle loss than caspase-mediated apoptosis.     

甘草酸二铵对大鼠心肌缺血再灌注损伤后细胞凋亡的影响

目的 :观察甘草酸二铵 (DG)对大鼠心肌缺血再灌注 (IR)损伤后心肌细胞凋亡及凋亡相关基因表达的影响。方法 :雄性wistar大鼠2 4只 ,随机分为假手术组 ;IR组 ;DG组 ( 2 0mg/kg)。每组 8只。采用末端脱氧核苷酸转移酶介导的带荧光的dUTP缺口末端标记(TUNEL)法、westernblot法和免疫组化法检测大鼠心肌缺血 3 0min再灌注 6h心肌凋亡细胞及凋亡相关基因Bcl 2、Bax蛋白表达的变化。结果 :与假手术组比较 ,IR组心肌细胞凋亡指数 (AI)、Bax蛋白的阳性表达明显增加 (P均 <0 .0 1) ;与IR组比较 ,DG治疗后AI和Bax蛋白的阳性表达明显下降 (P <0 .0 1) ,而Bcl 2蛋白的阳性表达明显上调 (P <0 .0 1)。结论 :DG具有保护大鼠心肌缺血再灌注损伤的作用 ,其作用机制可能与其下调Bax蛋白表达 ,上调Bcl 2蛋白表达抑制心肌细胞凋亡有关     

人非小细胞肺癌组织中自发性癌细胞凋亡的研究

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长期置入骶神经根刺激电极的兔组织学变化及其安全性

背景：有研究表明基于阳极阻滞技术的骶神经根刺激器能有效重建脊髓损伤兔的膀胱排尿功能,但符合此技术的刺激电极至今未见报道。 目的：设计并研制既与兔骶神经根匹配又符合阳极阻滞技术的刺激电极,观察长期植入刺激电极的兔骶神经根超微结构及病理形态学变化,评估刺激电极安全性。 方法：纳入新西兰兔30只,随机抽取10只兔切取双侧S2及S3神经前根,光镜下测量其直径后,制成与其直径相匹配的套筒型刺激电极。将剩余20只兔随机分为对照组及植入组,每组10只。植入组麻醉后将刺激电极植入S2及S3神经根前处,饲养半年后处死取材,观察植入处骶神经根超微结构变化。 结果与结论：长期植入该刺激电极后,光学显微镜下见植入组植入处骶神经根神经细胞结构保存良好,轴突无明显变性,无炎症细胞浸润及胶质瘢痕形成;透射电镜下观察,植入组髓鞘排列紧密,无脱髓鞘现象,神经元无核萎缩、核凹陷和异染色质增多等现象。免疫组织化学染色显示,与对照组相比,植入组植入处神经根中胶质纤维酸性蛋白、Bax,Bcl-2和Caspase-3蛋白表达差异无显著性意义。结果说明实验成功研制了兔骶神经根刺激电极,长期植入骶神经根未出现组织病理学改变及无细胞凋亡现象,安全性好。     

退行性变椎间盘组织中TGF-β1和Bax的表达及意义

目的探讨退行性变椎间盘组织中TGF-β1和Bax的表达及其意义。方法收集正常与退变椎间盘组织,并根据病理改变将退变椎间盘组织分成4级,采用HE、免疫组化、TUNEL染色和RT-PCR法进行研究。结果免疫组化和RT-PCR均显示在正常组织中有TGF-β1表达,Bax只有微量表达;在病变组织中随病理分级加大TGF-β1随之增加,与正常组相比,差异有显著性;Bax也逐步增加,与正常组相比,差异有显著性,Bax表达升高与凋亡指数(AI)呈正相关性。结论退变椎间盘的病理分级与TGF-β1的表达增高相关,由此调控了Bax的表达,导致了细胞凋亡,促进了椎间盘的退变。     

苦参碱对人髓母细胞瘤D341细胞凋亡及相关蛋白表达的影响*

 目的：  探讨苦参碱在体外对人髓母细胞瘤D341细胞凋亡及Bax、Bcl-2、丝氨酸/苏氨酸激酶Akt和磷酸化Akt（p-Akt）表达的影响。方法: 体外培养的D341细胞分为实验组（加不同浓度的苦参碱）和对照组（不加苦参碱）,用CCK-8法检测苦参碱对D341细胞增殖的影响,Annexin V-FITC/PI双染法检测细胞凋亡,Western blotting检测苦参碱对D341细胞Bax、Bcl-2、Akt和p-Akt蛋白表达的影响。结果: 苦参碱在体外能明显地抑制D341细胞的生长,作用呈量效与时效关系;随着作用药物浓度的增高和作用时间的延长,其凋亡率逐渐升高,在透射电镜下观察,细胞可见染色质趋边固缩,胞浆中空泡增多、变大,并且出现凋亡小体;Western blotting结果表明,随着药物浓度的增加,药物能使瘤细胞Bax蛋白的表达水平增强,却使Bcl-2和p-Akt蛋白的表达水平下降。结论: 苦参碱可诱导D341细胞的凋亡而发挥体外抗肿瘤作用,其诱导瘤细胞的凋亡与上调Bax蛋白、下调Bcl-2及PI3K /Akt 信号通路中p-Akt蛋白的表达水平有关。     

肾母细胞瘤经动脉化疗栓塞术疗效机制探讨

目的： 探讨肾母细胞瘤术前经动脉化疗栓塞的疗效机制。方法: 17例肾母细胞瘤患儿介入治疗后再行肿瘤切除，另 22例行单纯手术切除。对照分析2组肾母细胞瘤样本中细胞和间质的变化，采用原位细胞凋亡(TUNEL 法) 检测瘤细胞凋亡，并通过免疫组化法检测瘤细胞P53、Bcl-2 和Bax蛋白的表达。术后随访2年以上。结果: 介入组与单纯手术组的肿瘤坏死区域平均面积分别为60%、15 %（Uc = 2.84）；肿瘤组织呈X₃、X₄退变者分别为58.8%（10/17）和4.55%（1/22）（2c= 11.4）、间质纤维组织增生程度中、重度变化者分别为64.7%（11/17）和18.2%（4/22）（Uc = 2.72）、淋巴细胞侵润者分别为70.6%（12/17）和18.2%（4/22）（2c=10.9），2组有显著差异（P<0.01）；介入组肿瘤细胞分裂指数的中位数为0.2/10高倍视野，明显低于单纯手术组的1.4/10高倍视野（Uc = 54.50 ，P<0.01）；94.9(37/39)的病例中均有不同数量的阳性凋亡细胞出现。介入组肿瘤细胞凋亡指数的中位数为25.9/10高倍视野，明显高于单纯手术组的12.8/10高倍视野（Uc = 117.00，P<0.05）。P53和Bcl-2蛋白表达与瘤细胞凋亡及分化程度均无相关性（P>0.05），但Bax 蛋白在介入组瘤细胞表达率（80.0%）明显高于单纯手术组（40.0%），差异显著（P<0.05）。介入组2年无瘤生存率为73.3%（11/15），单纯手术组为42.9%（6/14）。结论: 经动脉化疗栓塞治疗能够有效杀伤肿瘤细胞、抑制肿瘤生长，诱导由Bax蛋白介导的肿瘤细胞凋亡。     

海洛因、麻黄素对小鼠仔鼠下丘脑、海马组织结构及胆碱乙酰基转移酶活性的影响

目的 探讨海洛因和麻黄素对仔鼠下丘脑、海马组织结构及胆碱乙酰基转移酶(ChAT)活性的影响,观察Bax蛋白和角质细胞生长因子(KGF)在下丘脑、海马的表达。 方法 108只昆明小鼠仔鼠,采用递增剂量连续腹腔注射海洛因和麻黄素,利用吉姆萨染色及免疫组织化学方法观察下丘脑和海马凋亡细胞的变化及Bax蛋白和KGF的表达,利用比色法检测下丘脑和海马ChAT活性的变化。 结果 仔鼠注射海洛因、麻黄素5d、10d、15d、20d,下丘脑和海马凋亡细胞及Bax蛋白和KGF阳性表达细胞的数量均高于对照组,ChAT的活性低于对照组,差异显著或极显著(P<0.05或P<0.01);海洛因组仔鼠下丘脑和海马的上述4项指标与麻黄素组相比,差异显著或极显著(P<0.05或P<0.01)。随着海洛因和麻黄素剂量的递增,仔鼠下丘脑和海马凋亡细胞及Bax蛋白和KGF阳性表达细胞的数量呈增多趋势。 结论 海洛因和麻黄素影响仔鼠下丘脑和海马的组织结构及ChAT活性,其机制可能与下丘脑和海马组织的细胞凋亡有一定相关性。     

Activated status of tumour-infiltrating lymphocytes and apoptosis in testicular seminoma.

Testicular seminoma is characterized by a prominent lymphoid infiltrate and an excellent prognosis. Cytotoxic T-lymphocytes (CTLs) infiltrating seminoma tumour nests constitute a major subset of the lymphoid infiltrate. The objective of this study was to determine whether CTLs express markers of cytotoxic potential and activity and whether the number of activated CTLs correlates with the extent of apoptosis in testicular seminomas, as opposed to non-seminomatous testicular germ cell tumours (NSTGCTs). Twenty cases of pure seminoma as well as 20 cases of NSTGCTs including 16 mixed germ cell tumours (MGCTs) were studied. Immunohistochemistry for the cytotoxic markers TIA-1 (cytotoxic potential) and granzyme B (cytotoxic activity) and the T-cell markers CD3 and CD8 was performed on formalin-fixed, paraffin-embedded sections. The apoptotic index (AI) was determined by the TUNEL method. The number of CD3(+), CD8(+), TIA-1(+), and granzyme B(+) cells in tumour cell nests was markedly increased in testicular seminomas, compared with NSTGCTs (p<0.01). Activated granzyme B(+) cells numbered 25.6+/-5.2 per high power field in seminomas and 8.9+/-3.2, 8.1+/-3.9, and 0.4+/-0.2 for embryonal carcinomas, yolk sac tumours, and immature teratomas, respectively. Double immunohistochemical staining for granzyme B and CD8 revealed that 82.6+/-8.5% of granzyme B-expressing cells were CD8(+). The tumour cell AI was significantly increased in embryonal carcinoma, compared with the seminoma, yolk sac tumour, and immature teratoma subgroups (6.7+/-1.3, 2.3+/-0.3, 3.0+/-1.1, and 2.3+/-1.1, respectively, p<0.001). TUNEL/CD3 double immunostaining revealed that a significant proportion of the apoptotic seminomatous tumour cells were in direct contact with one or more CD3(+) lymphocytes (47.2+/-6.2%). The number of activated granzyme B(+) CTLs showed a strong linear correlation with the AI in the seminoma group (r=0.71, p<0.0001) but not in other subgroups. TUNEL/granzyme B double immunolabelling revealed that a proportion of activated granzyme B(+) lymphocytes (20%) were often seen in close contact with apoptotic tumour cells. The presence of increased numbers of activated cytotoxic lymphocytes in testicular seminomas suggests that apoptotic tumour cell death in this neoplasm may be triggered by cytotoxic granule effectors. This phenomenon may be one of the key host immune mechanisms leading to the excellent prognosis in this tumour.     

男性不育精液中NO的含量与生殖细胞凋亡的关系

目的:探讨人精液中一氧化氮(nitricoxide,NO)与生殖细胞凋亡的关系。方法:采用镀铜镉还原荧光法检测NO代谢产物硝酸盐(NO-3)。用脱氧核苷酸末端转移酶(TdT)介导的缺口末端标记(TUNEL)法和透射电镜,分别检测和观察生殖细胞的凋亡及凋亡细胞的超微结构。结果:生育组精液中NO的含量为(56.83±11.65)μmol/L,生殖细胞的凋亡率(4.60±1.25)%,与不育组(128.86±23.76)μmol/L和(17.36±3.05)%相比较有非常显著性差异(P<0.01)。不育组NO的含量和生殖细胞的凋亡率呈显著的正相关(r=0.96)凋亡的生殖细胞核染色质浓缩在核周形成新月形,电子密度增高,核膜折叠,核裂解形成凋亡小体。结论:不育者精液中NO的含量与生殖细胞的凋亡率有密切关系。高浓度的NO可能是导致睾丸生殖细胞凋亡率增加而致使男性生育力下降的原因。     

肺泡壁细胞凋亡参与木瓜蛋白酶所致大鼠肺气肿的形成

目的：观察木瓜蛋白酶所致的肺气肿大鼠肺泡壁细胞凋亡，探讨细胞凋亡在肺气肿发病中的作用。方法：大鼠随机分为3组：正常对照组、木瓜蛋白酶组、木瓜蛋白酶联合［60Co］处理组。观察肺组织病理学变化，TUNEL法检测大鼠肺泡壁凋亡细胞，免疫组化法检测肺泡壁细胞PCNA、Bax和SP-C蛋白的表达。TUNEL与SP-C免疫荧光法鉴定凋亡细胞中的II型肺泡上皮细胞。结果： 木瓜蛋白酶组、木瓜蛋白酶联合［60Co］处理组大鼠出现明显肺气肿。木瓜蛋白酶联合［60Co］处理组平均内衬间隔，平均肺泡面积和单位面积肺泡数与木瓜蛋白酶组比较有显著差异。木瓜蛋白酶组，木瓜蛋白酶联合［60Co］处理组大鼠肺泡壁细胞的AI、PI以及Bax染色阳性的细胞百分比显著高于正常对照组；而SP-C染色阳性的细胞百分比显著低于正常对照组。木瓜蛋白酶联合［60Co］处理组大鼠的AI、PI、Bax染色阳性的细胞百分比显著高于木瓜蛋白酶组；而SP-C染色阳性的细胞百分比显著低于木瓜蛋白酶组。部分TUNEL阳性细胞表达II型肺泡上皮细胞标志SP-C。结论：大鼠肺泡壁细胞特别是II型肺泡上皮细胞凋亡参与肺气肿的形成，Bax蛋白的表达上调可能导致肺气肿大鼠肺泡壁细胞的凋亡。     

Apoptosis in normal rat embryo tissues during early organogenesis: the possible involvement of Bax and Bcl-2

Apoptosis commonly occurs in a variety of developmental processes in mammals. In this study, we investigated the relationship between apoptosis and the expression of both Bax and Bcl-2 during the early organogenesis period (9.5-11.5 days of gestation) of rat embryos. Apoptotic cells detected by the terminal deoxynucleotidyl transferase-mediated nick-end labeling (TUNEL) method were extremely abundant in the foregut diverticulum at 9.5 days of gestation, while they largely disappeared at 10.5 and 11.5 days of gestation, although they were detected in newly formed mid- and hindgut diverticulum at these times. Real-time RT-PCR analysis of whole embryos revealed that the expression of bax mRNA was constant at days 9.5 to 11.5, while the expression of bcl-2 mRNA gradually increased. Immunohistochemical studies of Bax and Bcl-2 expression revealed that these apoptotic cells were exactly positive to Bax in mirror sections, while their expression of Bcl-2 was generally too low to be detected. A disappearance of apoptotic cells was associated with strong Bcl-2 expression in the foregut diverticulum at 10.5 and 11.5 days of gestation. It was similarly observed that apoptotic cells detected in the cardiogenic area at 9.5 days of gestation disappeared with the formation of the primitive heart tube--accompanied by a strong expression of both Bcl-2 and Bax--in the developmental process of the primitive heart. Apoptotic cells were also observed in the primitive brain vesicle, optic vesicle, otic vesicle, and thyroid primordium at 10.5 and 11.5 days of gestation during the developmental process, with a strong expression of Bax. These results indicate that the Bax and Bcl-2 may be important in regulating the induction of embryonic cell apoptosis during early organogenesis.