Primary vaginal leiomyosarcoma: A case report with complete morphological,immunohistochemical and ultrastructural study

ObjectivePrimary vaginal leiomyosarcomas (LMS) are rare, easily recurrent tumours with an unknown etiology; the prognosis is poor and there is no consensus guideline on their management.Case reportA nodular, 25 × 23 x 28 mm-mass, infiltrating the urethra, was found in a 58-year-old woman. A biopsy showed a LMS of the vagina that was positive for vimentin, alpha-smooth muscle actin, caldesmon, desmin, p16 and p53. An anterior pelvic exenteration was performed. The sample was fixed and prepared for light microscopy, transmission and scanning electron microscopy, confirming the diagnosis of LMS.ConclusionsBest outcomes occur when the tumour is small, localized, and can be removed surgically with wide, clear margins, as in this case. As there are different kinds of malignant mesenchymal tumours, biopsy followed by immunohistochemistry and electron microscopy still represents a good diagnostic choice and surgical resection is generally the gold standard in these cases.     

Human fetal endometrium -light and electron microscopic study

Summary Human fetal endometrium was examined by light and electron microscopy. Our study shows the following new morphological aspects: (1) Glands are already present. (2) Endometrium undergoes a maturation process during gestation and at late gestational age resembles late proliferative endometrium. (3) The nuclear bodies are present in cell nuclei throughout gestation. (4) Nucleolar channel systems (NCS) sometimes appear at a late gestational age. (5) Cells with the same morphology as that of endocrine cells are found in the basal layers of endometrium at a late gestational age. The significance of these morphological aspects is discussed.Supported by the Stiftung Volkswagenwerk, Hannover, FRGDedicated to Prof. Dr. J. Schneider (Director of the Frauenklinik der Medizinischen Hochschule Hannover) on the occasion of his 60th birthday     

Three-dimensional microanatomical dynamics of the ovary in postreproductive aged women

OBJECTIVE: To evaluate the microanatomical dynamics of the ovary during postreproductive life. DESIGN: Retrospective, observational research study.SETTING: Institutional (university). PATIENT(S): Eleven (43- to 72-year-old) women in perimenopause, menopause, or postmenopause. INTERVENTION(S): Biopsies of ovaries obtained from patients undergoing gynecologic surgery or diagnostic procedures. MAIN OUTCOME MEASURE(S): Analysis of the 3-dimensional microanatomy of the ovary by transmission and high-resolution scanning electron microscopy. RESULT(S): The surface epithelium gets gradually flatter and is always present, even in advanced age. The surface appears smoother because of a smaller number of papillae and crypts as well as a decreased number and shortening of microvilli on surface epithelial cells. Signs of atrophy and fibrosis are evident. Primordial follicles are usually absent in postmenopause, whereas corpora atretica, hemorrhagica, and albicantia, scar tissue, and simple follicular cysts are common after menopause. Apoptotic and necrotic cells appear frequently within the surface epithelium. Major common features are a marked reduction in number and caliber of blood vessels with thickening of the vascular walls and changes in endothelial cells. CONCLUSION(S): Scanning electron microscopy studies are a useful complement to ordinary gynecologic diagnostic methods. Variations among patients of the same age range or functional status should be considered.     

盆腔静脉瘀血症患者瘀血静脉超微结构观察

为探讨盆腔静脉瘀血症患者瘀血静脉形态学改变,本文对11例瘀血症患者和6例结扎术后无任何症状和体征的妇女盆腔静脉进行了扫描及透射电镜观察.发现瘀血静脉壁间渗出增多;内皮细胞数量增多、体积变小且外形肿胀;细胞表面粗糙,突起增粗,小孔及凹陷增多、变大;细胞质致密并含大量饮液泡,线粒体肿胀而数量增多;染色质呈块状分布于核的周边;内皮细胞间嵌合连接和紧密连接正常存在.这些变化说明缺氧是引起本症盆腔静脉超微结构改变的主要原因,提示本症是一种功能性疾病.     

Second harmonic generation microscopy of fetal membranes under deformation: Normal and altered morphology

Introduction

Insight into the microstructure of fetal membrane and its response to deformation is important for understanding causes of preterm premature rupture of the membrane. However, the microstructure of fetal membranes under deformation has not been visualized yet. Second harmonic generation microscopy, combined with an in-situ stretching device, can provide this valuable information.

Methods

Eight fetal membranes were marked over the cervix with methylene blue during elective caesarean section. One sample per membrane of reflected tissue, between the placenta and the cervical region, was cyclically stretched with a custom built inflation device. Samples were mounted on an in-situ stretching device and imaged with a multiphoton microscope at different deformation levels. Microstructural parameters such as thickness and collagen orientation were determined. Image entropy was evaluated for the spongy layer.

Results

The spongy layer consistently shows an altered collagen structure in the cervical and cycled tissue compared with the reflected membrane, corresponding to a significantly higher image entropy. An increased thickness of collagenous layers was found in cervical and stretched samples in comparison to the reflected tissue. Significant collagen fibre alignment was found to occur already at moderate deformation in all samples.

Conclusions

For the first time, second harmonic generation microscopy has been used to visualize the microstructure of fetal membranes. Repeated mechanical loading was shown to affect the integrity of the amnion–chorion interface which might indicate an increased risk of premature rupture of fetal membrane. Moreover, mechanical loading might contribute to morphological alterations of the fetal membrane over the cervical region.     

Automated detection of rare fetal cells in maternal blood: eliminating the false-positive XY signals in XX pregnancies

OBJECTIVE: The purpose of this study was to develop a new method to help differentiate XX from XY signals in maternal blood from women carrying XY fetuses. STUDY DESIGN: We have developed a system to scan automatically for cells that bear X and Y fluorescence in situ hybridization signals. These XY target cells are identified by scans at low (x20) magnification, and all identified targets are revisited and verified at high (x100) magnification. The viewer software component of the system displays x20 images of all cells and intracellular fluorescence in situ hybridization signals that are present in each of the 4000 optical fields per slide, along with x100 images of automatically detected target cells. RESULTS: We initially examined 36,000 fields from 18 slides in 12 pregnancies (6 male and 6 female) using our system that is based on fluorescence in situ hybridization with a single probe for the X-chromosome and a single probe for the Y-chromosome and found XY nuclei in all samples, regardless of fetal gender. In the second phase of the study, a refinement of the approach that incorporated 2 independent probes for the Y-chromosome resulted in a false-positive rate for detection of XY nuclei in XX cases <0.00005%. CONCLUSION: Our data suggest that this system may allow for excellent "signal to noise" separation, which is required absolutely for fetal cell methods to differentiate aneuploid from normal pregnancies. Quantitation of fetal cells in the maternal circulation and standardization of processes that have been developed for their enrichment are crucial to moving fetal cell assessment from esoteric basic science to applied new technology.     

子宫内膜异位症与正常子宫内膜黏附过程的形态学分析

目的:分析子宫内膜异位症(endometriosis,EM)和正常对照子宫内膜与腹膜黏附过程的特征。方法:将EM和非EM患者的子宫内膜与腹膜自身及交叉接种共培养后,用组织学方法和透射电镜观察EM患者和非EM患者的子宫内膜组织和腹膜组织的特征。结果:患者和正常对照来源的子宫内膜与腹膜之间的黏附过程在黏附发生的时间和组织学检查中无明显区别。透射电镜检查表明,EM患者来源的子宫内膜表现出代谢活跃的特征。结论:子宫内膜和腹膜自身的特征决定了异位内膜能够很快与腹膜黏附。EM和非EM患者的黏附过程无明显区别。     

No ancillary finding is valid to distinguish a primary ovarian hepatoid carcinoma from metastatic hepatocellular carcinoma

Primary ovarian hepatoid carcinomas (POHC) are extremely rare. Especially rare are those with phenotypic properties of hepatocellular carcinoma (HCC) and an absence of clinical evidence of hepatic tumor. We report a case of a POHC with a common microscopic, immunophenotypic, and ultrastructural property of HCC in the absence of a liver mass. It is extremely difficult to differentiate POHC from metastatic HCC using any kinds of ancillary studies, with the exception of clinical identification of a hepatic tumor.     

Polarized light microscopy-detectable structures of human oocytes and embryos are related to the likelihood of conception in IVF

Purpose

To clarify if birefringent structures of human oocytes and embryos, measurable by polarized light microscopy, have any value in predicting the chance of pregnancy in human in vitro fertilization and may halp to identify the most competent oocytes and embryos.

Methods

The inner layer of the zona pellucida (IL-ZP) and the meiotic spindle (MS) were analyzed by polarized light microscopy in 258 oocytes and in the 209 embryos deriving from them. Data obtained from 102 ICSI cycles with conception were compared with those obtained in 156 cycles without conception. The retardance and area of the IL-ZP, as well as the retardance, length of the major axis, and area of the MS were measured. Furthermore, polarized light microscopy parameters were related to the embryo morphological score by multiple regression analysis.

Result(s)

The mean area of the IL-ZP of both oocytes and embryos was significantly lower in conception than in non-conception cycles (p = 0.0001 for oocytes and p = 0.002 for embryos). The area of the IL-ZP in embryos was significantly, inversely related to the embryo morphological score (p = 0.011). The area, the major axis length and the retarcance of the MS, as well as the retardance of the IL-ZP in oocytes and embryos were comparable in conception and non-conception cycles.

Conclusion

The area of the IL-ZP of the human oocytes may represent a marker of oocyte competence, as oocytes with a low IL-ZP area are more frequently obtained in conception cycles. When measured in embryos, a low IL-ZP area identifies embryos with a high chance of implantation.     

Redefining 3Dimensional placental membrane microarchitecture using multiphoton microscopy and optical clearing

IntroductionRemodeling of human placental membranes (amniochorionic or fetalmembrane) throughout gestation, a necessity to accommodate increasing uterine volume, involves continuous alterations (replacement of cells and remodeling of extracellular matrix). Methodologic limitations have obscured microscopic determination of cellular and layer-level alterations. This study used a combination of advanced imaging by multiphoton autofluorescence microscopy (MPAM) and second harmonic generation (SHG) microscopy along with tissue optical clearing to characterize the 3Dimensional multilayer organization of placental membranes.MethodsPlacental membranes biopsies (6 mm) collected from term, not-in-labor cesarean deliveries (n = 7) were fixed in 10% formalin (native) or treated with 2,2′-thiodiethanol to render them transparent for deeper imaging. Native and cleared tissues were imaged using MPAM (cellular autofluorescence) and SHG (fibrillar collagen). Depth z-stacks captured the amnion epithelium, underlying matrix layers, and in the cleared biopsies, the decidua layer.ResultsMPAM and SHG revealed fetal membrane epithelial topography and collagen organization in multiple matrix layers. Term amnion layers showed epithelial shedding and gaps. Optical clearing provided full-depth imaging with improved visualization of collagen structure, mesenchymal cells in extracellular matrix layers, and decidua morphology. Layer thicknesses measured by imaging corroborated with histology. Mosaic tiling of MPAM/SHG image stacks allowed large area visualization of entire biopsies.ConclusionMPAM-SHG microscopy allowed for study of this multi-layered tissue and revealed shedding, gap formation, and other structural changes. This approach could be used to study structural changes associated with membranes as well as other uterine tissues to better understand events in normal and abnormal parturition.     

光敏剂竹红菌乙素在卵巢癌细胞中定位定量的荧光显微成像研究

目的:研究国产光敏剂竹红菌乙素(HB)在卵巢癌细胞系SKOV3中的荧光显微成像特点,并分析其细胞内荧光强度。方法:40μg/ml HB与细胞孵育不同时间后,应用激光共聚焦显微镜和配备致冷CCD的倒置荧光显微镜采集细胞中的荧光显微数字图像,计算机进行图像处理分析光敏剂在细胞内的定位,捷达801形态学软件分析其荧光强度;不同浓度(10～80μg/ml)HB与细胞孵育后,行荧光强度分析,确定光敏剂最佳给药浓度。结果:HB呈红色荧光弥漫性分布于细胞浆内,细胞核区域很少荧光分布;细胞内荧光强度于用药4h后达到高峰,当光敏剂浓度增高到≥40μg/ml时,加大药物浓度细胞内荧光强度不随之增强。结论:激光共聚焦显微镜和倒置荧光显微镜结合捷达801形态学分析软件可有效应用于低荧光效率的光敏剂HB细胞内定位与定量研究;HB主要分布细胞浆内,细胞对其吸收存在最大阈浓度。研究结果可为临床应用光动力学治疗卵巢癌提供有用参数。     

Development of a simple,cost-effective,semi-correlative light and electron microscopy method to allow the immunoelectron localisation of non-uniformly distributed placental proteins

Immunoelectron microscopy is wrought with technical limitations that complicate its use. However, advances in correlative light and electron microscopy have recently lead to improvements in this field. We report the development of a semi-correlative approach to investigate the ultrastructural location of an antiphospholipid antibody within the syncytiotrophoblast. This method offers several advantages over existing methodologies, since it preserves antigenicity, shows good immunolabel penetrability and does not require specialized equipment. The use of a pre-embedding screen has also allowed us to target individual placental villi and overcome sampling limitations of the electron microscope. This simple, cost-effective method is likely to find widespread application in placental research.     

Ultrastructure of placental hyperplasia in mice: comparison of placental phenotypes with three different etiologies

Hyperplastic placentas have been reported in several experimental mouse models, including animals produced by somatic cell nuclear transfer, by inter(sub)species hybridization, and by somatic cytoplasm introduction to oocytes followed by intracytoplasmic sperm injection. Of great interest are the gross and histological features common to these placental phenotypes - despite their quite different etiologies - such as the enlargement of the spongiotrophoblast layers. To find morphological clues to the pathways leading to these similar placental phenotypes, we analyzed the ultrastructure of the three different types of hyperplastic placenta. Most cells affected were of trophoblast origin and their subcellular ultrastructural lesions were common to the three groups, e.g., a heavy accumulation of cytoplasmic vacuoles in the trophoblastic cells composing the labyrinthine wall and an increased volume of spongiotrophoblastic cells with extraordinarily dilatated rough endoplasmic reticulum. Although the numbers of trophoblastic glycogen cells were greatly increased, they maintained their normal ultrastructural morphology, including a heavy glycogen deposition throughout the cytoplasm. The fetal endothelium and small vessels were nearly intact. Our ultrastructural study suggests that these three types of placental hyperplasias, with different etiologies, may have common pathological pathways, which probably exclusively affect the development of certain cell types of the trophoblastic lineage during mouse placentation.     

Sperm ultrastructure and 18, X,Y aneuploidies in a man with a 46 XY, 47 XY + 18 mosaic karyotype: case report

Purpose To describe sperm morphology and meiotic segregation in the case of a man with a 46 XY/ 47 XY + 18 mosaic karyotype. Materials and methods A 25-year-old man came to our Centre for semen analysis. Morphological sperm evaluation was performed by light and electron microscopy; meiotic segregation was examined by fluorescence in situ hybridisation (FISH) technique using probes for chromosomes 18, X and Y. PCR was carried out on DNA from peripheral blood lymphocytes to analyze Y microdeletions. Results Mathematically elaborated transmission electron microscopy data highlighted a low number of sperm devoid of ultrastructural defects, and the presence of characteristics of apoptosis and immaturity. FISH showed the presence of aneuploidies of chromosome 18 and sex chromosomes. Conclusions In this case of mosaicism morphological and meiotic spermatogenetic impairment is shown, as well as structural chromosomal alterations. We describe sperm morphology and meiotic segregation in a carrier of 46 XY/ 47 XY + 18 mosaic karyotype.     

Submicroscopic mathematical evaluation of spermatozoa in assisted reproduction. 2. In vitro fertilization. (notulae seminologicae. 7)

This paper belongs to a series of applications of the Baccetti et al. formula (1) to the submicroscopical mathematical examination of human spermatozoa used for assisted reproduction. The present experiment concerns IVF, a technique requiring careful evaluation of sperm quality to predict the success of the program. Our results demonstrate that the sperm submicroscopic characters introduced in the formula are clearly correlated with the result of IVF. In fact the two numbers concerning sperm quality (percentage of spermatozoa free from structural defects and total number in the ejaculate of spermatozoa free from defects) obtained in successful and unsuccessful IVF groups, showed a large difference. Thet distribution in both cases reached a significance of 0.005. The synthetic parameters obtained are therefore a good tool in the prediction of sperm power in in vitro insemination techniques. The most important characteristics seem to be the quality of the acrosomal complex, the status of the chromatin, the shape of mitochondria, the axonemal pattern, and the membrane integrity. All these characteristics are expressed with largely different means in successful and unsuccessful ejaculates (t distribution significant at 0.005). All these data confirm that submicroscopic mathematical diagnosis offers a convincing evaluation of sperm structure and function, involving all organelles, including acrosome function and cell motility. It is also demonstrated that sperm quality is a major factor in the success of IVF and that it is clearly revealed by the integrity of the majority of the sperm organelles. An erratum to this article is available at .     

光学显微镜和尿液分析仪对儿童血尿定位诊断作用的探讨

目的探讨应用普通光学显微镜检查尿红细胞形态和UF 50尿液全自动分析仪2种方法对儿童血尿定位诊断的作用和影响因素。 方法研究对象为1977 01—2006 01首都儿科研究所肾内科病房收治的临床出现血尿患儿125例，均经肾活检证实为肾小球源性疾病。应用普通光学显微镜对经肾脏病理证实为肾小球疾病和非肾小球疾病患者的晨尿进行红细胞形态的检查；同时应用UF 50尿液全自动分析仪对其中部分肾小球和非肾小球疾病患者的同一标本进行尿红细胞参数测定。 结果(1)肾小球疾病组患者均经病理检查所证实。(2)以RBC严重变形≥30%或变形≥70%两者之一作为肾性血尿的诊断标准,其特异度均达到90%以上，但全自动尿液分析仪的灵敏度较低。(3)晨尿和随意尿标本经上述2种方法检查，其肾性血尿阳性率差异无显著性。 结论资料显示以RBC变形≥70%或严重变形≥30%作为诊断肾性血尿的标准，其灵敏度和特异度较高；光镜检测尿红细胞形态与UF 50全自动尿液分析仪用于肾小球性血尿检测，其特异度均达到90%以上，但UF 50全自动尿液分析仪的灵敏度较低；儿童可选用随意尿作为2种方法的检测标本。     

Quantitative nuclear image analysis: differentiation between normal,hyperplastic, and malignant appearing uterine glands in a paraffin section I. Elementary features for differentiation

This study is of a methodological nature. Using an automatic quantitative image analysis instrument, an attempt is made to differentiate between nuclei in normal, atypical hyperplastic and carcinomatous endometrial glands present in the same tissue section. With subjective judgement, these groups were clearly distinguishable although in each group nuclei occurred, which resembled nuclei in the other groups.The results not only confirm the deviations in nuclear features in the three groups as observed subjectively, but also reveal other features, which are even more significantly different. On the other hand, the results reveal, that single features of the three groups show a great deal of overlapping. Thus, the present study is another example of the fact, that quantitative microscopical techniques have the capacity to equal subjective observations. Therefore, further application of these techniques is promising.     

Effects of aging on the zona pellucida surface of mouse oocytes

In a scanning microscopy study of mouse oocytes from immature, young, and aged females and of oocytes agedin vivo orin vitro, we have observed four types of zona pellucida, which we classify as types A, B, C, and D. Oocyte aging gives rise to a significant increase in predegenerative (type C) oocytes; this type of zona pellucida surface could result from a zona hardening effect and decrease the rate of fertilization of the oocytes affected.     

Quantitative nuclear image analysis: differentiation between normal, hyperplastic, and malignant appearing uterine glands in a paraffin section. II. Computer assisted recognition by discriminant analysis.

Quantitative image analysis was applied to data from microscope photometry of nuclei in a paraffin section. The data were essentially the same as described in a previous publication (Baak and Diegenbach, 1977). The results from measurements on the nuclei of morphologically normal, atypical hyperplastic and carcinomatous uterine glands were used in discriminant analysis. With this method it is possible to discriminate between the three groups of nuclei. Depending on the (sub)set of the variables used, 60-70% of all nuclei are correctly classified in one of the three groups. Discrimination of one of the groups against the other two results in up to 81% correct classifications. Therefore, discriminant analysis offers a possibility of assisting diagnosis in an objective way.