细胞HE染色涂片与细胞蜡块检测晚期非小细胞肺癌胸腔积液标本中表皮生长因子受体基因突变

目的:伴发胸腔积液的晚期非小细胞肺癌(non-small cell lung cancer,NSCLC)患者已失去手术机会,表皮生长因子受体(epithelial growth factor receptor,EGFR)酪氨酸激酶抑制剂(tyrosine kinase inhibitors,TKIs)是EGFR敏感突变的晚期NSCLC患者的一线用药。但EGFR-TKIs治疗时或治疗后出现的疾病进展以及药物的更新迭代给临床诊治带来了挑战,需要对存档的胸腔积液细胞样本进行EGFR复检或比对研究。本研究采用细胞HE染色涂片与细胞蜡块两种制作方法对NSCLC胸腔积液样本的EGFR突变基因进行检测,分析两种保存方式、保存时间对胸腔积液细胞DNA质量的影响,以期探索出一条保存细胞学资料及充分利用细胞学档案资料的可靠途径。方法:选取2014年10月—2021年4月航天中心医院病理科接收的胸腔积液样本57例,所有样本同时制作细胞HE染色涂片与细胞蜡块。将57例配对的细胞HE染色涂片与细胞蜡块采用扩增阻碍突变系统-聚合酶链反应(amplification refractory mutation system-polymerase chain reaction,ARMS-PCR)技术进行EGFR基因突变检测。DNA检测浓度为2 ng/μL,细胞HE涂片与配对细胞蜡块DNA样本并排进行扩增。结果判读按试剂说明书要求,待测样本的8号孔外控循环阈值(cycle threshold,Ct值)在13~21之间为样本合格,EGFR基因突变Ct值<26时,判断为阳性;26≤Ct值<29为临界阳性,Ct值≥29为阴性。ΔCt值为突变Ct值与外控信号Ct值的差值。比较细胞HE染色涂片与细胞蜡块检测为阳性的ΔCt值。同时将57例患者按时间段分为4组:2014—2015年(n=10),2016—2017年(n=20),2018—2019年(n=17),2020—2021年(n=10)。比较不同时间段的检测结果。结果:57例晚期NSCLC患者中以胸腔积液为首发症状入院者42例,占73.7%;57例中37例发生EGFR突变,突变率64.9%;19del突变和L858R突变为主要的突变类型,分别占37.8%(14/37)和48.6%(18/37);37例突变患者中女性占56.7%(21/37);细胞涂片与配对的细胞蜡块突变一致率为100%;细胞HE染色涂片的ΔCt值小于配对细胞蜡块的ΔCt值(t=4.526,P<0.001)。37例配对细胞蜡块与细胞HE染色涂片的突变Ct值均<26。4个时间段中,2014—2015年、2016—2017年、2018—2019年的细胞蜡块突变基因Ct均值均高于2020—2021年(P<0.05);4个时间段的细胞HE染色涂片突变Ct均值差异无统计学意义(P>0.05)。57例配对细胞蜡块与细胞HE染色涂片的外控Ct值均在13~21之间。2014—2015年、2016—2017年的细胞蜡块与细胞HE染色涂片的外控Ct值均高于2018—2019年、2020—2021年(均P<0.05)。结论:细胞HE染色涂片与细胞蜡块检测NSCLC胸腔积液样本EGFR突变具有极好的一致性,且细胞HE染色涂片的DNA质量优于细胞蜡块,但细胞HE染色涂片的局限在于不可复制性,建议在胸腔积液标本制作中进行多张涂片,以满足多次检测的需求。

细胞HE染色褪色处理后免疫组化染色体会

免疫组化在病理诊断中广泛而深入的推广和应用,使之在辅助诊断、鉴别诊断方面进入了一个传统病理学极少涉及的领域,使病理诊断提高到了新的水平。最近几年发展的PV法又称为两步法使得免疫组化染色技术更加简单快速,背景更加清晰。本院病理科细胞涂片均采用中性福尔马林液固定,本文对细胞学检查疑似阳性的涂片,将HE切片褪色处理后行免疫组化染色的体会介绍如下。

Histological Structure Difference of Dog's Olfactory Bulb Between Different Age and Sex

The purpose of this article is to detect sex and age difference in the structure of the olfactory bulb in dogs by histological methods. The thickness of the olfactory bulbs layers and its main cells were analyzed comparatively with the methods of HE-staining and statistics, through which we studied the development course of dogs＇ olfactory bulb and the structural differences which affect the olfaction in both males and females. The results showed that between both male and female juveniles and adult males and females, the difference in thickness of each layer is not significant. But the difference in quantity of mitral cells between adult males and females was significant. Meanwhile, the structure of every layer in juvenile dogs was apparent while the volume and the weight of adult dogs＇ olfactory bulb and each layer＇s width increased significantly. On the other hand, the density of each layer＇s cells decreased apparently. Our results demonstrated that the olfactory bulb developed with age, and the apparent differences in morphology and quantity of mitral cells between males and females may be one of the reasons leading to the sexual variations of olfactory sensitivity.

Effect of osteoprotegerin in combination with interleukin-6 on inhibition of osteoclast differentiation

Objective： To observe the effect of recombinant interleukin-6 （IL-6） and osteoprotegerin （OPG） on inhibiting bone absorption induced by receptor activa- tor for nuclear factor- rB ligand （RANKL） in murine osteo- clast precursor cells （OCPs） model. Methods： RAW 264.7 cells were solely treated with 50 ng/ml RANKL for 1 day, and then they were divided into three groups： RANKL （control group）, RANKL＋IL-6 （IL-6 group） and RANKL＋IL-6＋OPG （combination group）. These cells were harvested and investigated by means of HE stain- ing under light microscope after consecutive 9 days. Furthermore, staining tartrate-resistant acid phosphatase （TRAP）-positive multinucleated cells were detected by in- verted phase contrast microscope. The absorption pits of bone slices were observed under scanning electron microscope. Results： The number of mature osteoclast cells in control group was more than that in IL-6 alone or IL-6 com- bined with OPG group （P〈0.05）. Interestingly, this experi- ment has also demonstrated that there was a large number of TRAP-positive multinucleated osteoclasts （more than 3 nuclei） and several bone absorption formation in the con- trol group, whereas the outcome was completely different in both IL-6 group and IL-6＋OPG group （P〈0.05）. Conclusion： IL-6 can suppress the differentiation of mature osteoclasts as directly adding it into the RAW 264.7 cells induced by 50 ng/ml RANKL, and further the effect of osteolysis is remarkably reduced. When treatment with IL- 6 combined with OPG, a more effective strategy for the treat- ment of osteoporosis is reached.