FRET multiphoton spectral imaging microscopy of 7-ketocholesterol and Nile Red in U937 monocytic cells loaded with 7-ketocholesterol

OBJECTIVE: To show the effect of 7-ketocholesterol (7KC) on cellular lipid content by means of flow cytometry and the interaction of 7KC with Nile Red (NR) via ultraviolet fluorescence resonance energy transfer (FRET) excitation of NR on U937 monocytic cells by means of 2-photon excitation confocal laser scanning microscopy (CLSM). STUDY DESIGN: Untreated and 7KC-treated U937 cells were stained with NR and analyzed by flow cytometry and CLSM. 3D sequences of images were obtained by spectral analysis in a 2-photon excitation CLSM and analyzed by the factor analysis of medical image sequences (FAMIS) algorithm, which provides factor curves and images. Factor images are the result of the FAMIS image processing method, which handles emission spectra. In FRET analysis, preparations are screened at selected UV wavelengths to avoid emission of NR in the absence of 7KC. RESULTS: During 7KC-induced cell death,flow cytometry and CLSM revealed a modification of the cellular lipid content. Factor images show FRET occurrence and subsequent colocalization of 7KC and NR. CONCLUSION: This investigation established the utility of 2-photon excitation CLSM to assess colocalization of 7KC with NR by FRET and to identify and distinguish polar and neutral lipids stained by NR that accumulate from the effect of 7KC.     

DNA image cytometry in the differential diagnosis of endometrial hyperplasia and adenocarcinoma

OBJECTIVE: To test the value of DNA image cytometry in the differential diagnosis of hyperplastic endometrial lesions and endometrial carcinoma on a series of 153 cases of simple hyperplasia (n = 71), complex hyperplasia (n = 28), complex atypical hyperplasia (n = 11) and endometrial adenocarcinoma (n = 43). STUDY DESIGN: Monolayer smears were prepared from three 50-micron-thick sections by a cell separation technique and were stained according to Feulgen. The DNA content of 250 epithelial cells, chosen randomly, was determined using a TV image analysis system (CM-1, Hund, Wetzlar, Germany). The DNA content of 30 lymphocytes served as an internal standard for the normal diploid value in every case. Different DNA cytometric parameters and the mean nuclear area were calculated. RESULTS: Cases of adenocarcinoma and complex atypical hyperplasia (n = 54) were defined as clinically "positive" as these patients are normally treated by hysterectomy. The remaining cases of simple and complex hyperplasia (n = 99) were interpreted as clinically "negative" as conservative therapy is usually preferred. Requesting a specificity of > 90%, high sensitivity rates were calculated for ploidy imbalance (94%), mean ploidy (91%), diploid deviation quotient (91%), DNA stemline ploidy (87%) and 2c deviation index (85%), based on suitable thresholds. Entropy (76%), 5c exceeding events (63%), mean nuclear area (48%) and 9c exceeding events (6%) revealed lower sensitivity values. 5c Exceeding events (P = .0117) and mean nuclear area (P = .0392) were helpful in differentiating between atypical hyperplasia and endometrial carcinoma as the data distribution was significantly different with the U test. CONCLUSION: Our results indicate that DNA single cell cytometry is a highly relevant tool in the differential diagnosis of endometrial lesions and could be used as a complementary diagnostic method, especially in histomorphologically difficult cases.     

Digital image analysis of silver-stained nucleolar organizer region--associated proteins in endometrial cytologic samples

OBJECTIVE: Digital image analysis was applied to determine the number, area and size of silver-stained nucleolar organizer regions (AgNORs) in cytologic samples from curettage in normal, hyperplastic and malignant endometrium. STUDY DESIGN: Thirty-two archival cytologic smears from curettage (previously stained by the Papanicolaou method) with the histologic diagnosis (4 inactive endometrium, 5 secretion, 5 proliferation, 5 simple hyperplasia, 5 complex hyperplasia, 3 atypical hyperplasia, 5 adenocarcinoma, grade 1) were analyzed with the AgNOR technique. Count, area and size of AgNORs were analyzed in 50 cells per sample using a magnification of 1,000x. Quantitative analysis was performed on an SFORM digital imaging system. Data were analyzed with the SPSS/PC+ program. Mann-Whitney and chi 2 tests were performed. RESULTS: The average value of AgNOR count increased from normal to hyperplastic endometrium and well-differentiated adenocarcinoma. Differences were significant except between atypical hyperplasia and adenocarcinoma. Four, five and more AgNORs in 40% or more of the nuclei were found in complex and atypical hyperplasia and adenocarcinoma. Proliferation, and simple and atypical hyperplasia had similar mean values of AgNOR area. The mean total AgNOR area value increased from normal to hyperplastic had similar mean values of AgNOR area. The mean total AgNOR area value increased from normal to hyperplastic and well-differentiated adenocarcinoma. Differences were statistically significant. AgNOR size in well-differentiated adenocarcinoma was significantly different from that in normal endometrium and different grades of hyperplasia. CONCLUSION: Digital image analysis of AgNOR count, area and size enabled a distinction to be made between normal, hyperplastic and malignant endometrium.     

Confocal DNA cytometry: a contour-based segmentation algorithm for automated three-dimensional image segmentation

BACKGROUND: Confocal laser scanning microscopy (CLSM) presents the opportunity to perform three-dimensional (3D) DNA content measurements on intact cells in thick histological sections. So far, these measurements have been performed manually, which is quite time-consuming. METHODS: In this study, an intuitive contour-based segmentation algorithm for automatic 3D CLSM image cytometry of nuclei in thick histological sections is presented. To evaluate the segmentation algorithm, we measured the DNA content and volume of human liver and breast cancer nuclei in 3D CLSM images. RESULTS: A high percentage of nuclei could be segmented fully automatically (e.g., human liver, 92%). Comparison with (time-consuming) interactive measurements on the same CLSM images showed that the results were well correlated (liver, r = 1.00; breast, r = 0.92). CONCLUSIONS: Automatic 3D CLSM image cytometry enables measurement of volume and DNA content of large numbers of nuclei in thick histological sections within an acceptable time. This makes large-scale studies feasible, whereby the advantages of CLSM can be exploited fully. The intuitive modular segmentation algorithm presented in this study detects and separates overlapping objects, also in two-dimensional (2D) space. Therefore, this algorithm may also be suitable for other applications.     

P63在子宫内膜样腺癌的表达及意义

目的检测正常子宫内膜、子宫内膜增生症和子宫内膜样腺癌（endometriod adenocarcinoma,EC）组织中P63的表达,探讨P63与子宫内膜样腺癌发生、发展及预后的关系。方法采用免疫组化SP法检测正常子宫内膜（20例）,子宫内膜增生症（20例）,子宫内膜样腺癌（50例）组织中P63蛋白的表达。结果（1）正常子宫内膜中仅1例增生期内膜中有P63表达,阳性细胞零星分布于极个别腺体的基底部。子宫内膜增生症和子宫内膜样腺癌组织中,P63阳性细胞常相对集中分布于某一区域的腺体或实性巢,染色强。（2）EC组和子宫内膜增生症组P63的阳性率分别为46％和50％,与正常子宫内膜组（5．0％）比较差异有显著性护〈0．05）。子宫内膜增生症组P63的阳性率与EC组比较差异无显著性（P〉0．05）。（3）P63的表达与EC的分化程度无关（P〉0．05）。结论（1）EC组织中P63阳性细胞可能来源于胚胎时期的未分化细胞,具有多向分化的潜能。（2）P63与EC的发生发展有关,可能起癌基因的作用。（3）P63在子宫内膜增生症组织中高表达,表明P63与子宫内膜的异常增生相关。     

Three-dimensional reconstruction by confocal laser scanning microscopy in routine pathologic specimens of benign and malignant lesions of the human breast

 Confocal laser scanning microscopy (CLSM) has become an exciting new instrument because of its increased resolution over conventional wide-field microscopy and its high performance three-dimensional (3D) optical sectioning. Although CLSM has been used extensively in cell biology, few applications have been reported in routine clinical pathology. In this study, 3D reconstruction was performed on routine formalin-fixed, paraffin-embedded tissues of normal mammary duct, simple ductal hyperplasia, intraductal papillary hyperplasia, ductal carcinoma in situ, invasive carcinoma, and lymph node metastatic carcinomas of the human breast by using computer-assisted CLSM in conjunction with a 3D reconstruction software package (microVoxel). The selected specimens were sectioned at 30 μm, mounted on glass slides, and stained with the DNA fluorescent probe, YOYO-1 iodide. The nuclear DNA and chromatin texture were clearly demonstrated after pretreatment with RNAase and hydrolysis with 2 N HCl. High quality 3D images were obtained by processing the optical section stacks with volume render and surface display parameters in microVoxel. 3D morphologic characteristics of different breast lesions were examined in various orientations by angular image rotation. The clearly benign lesions (simple ductal hyperplasia and intraductal papillary hyperplasia) revealed similar 3D morphologic features, including: (1) smooth nuclear surface and homogeneous chromatin fluorescence intensity; (2) hyperplastic cell nuclei showing similar shape and volume; and (3) clear-cut margin of basement membrane defined by spindle-shaped myocytes of the ductal outer layer. In contrast, carcinomas displayed remarkably different features in 3D morphology, including: (1) irregular nuclear surface; (2) marked nuclear pleomorphism (irregular, angulated and indented shape of nuclear volume); (3) irregular and coarse chromatin texture; (4) chaotic arrangement of tumor cell nuclei; and (5) absence of myocytes, indicating no clear margin at the site of infiltration of cancer cells. In conclusion, nuclear structure, specifically demonstrated by CLSM of YOYO-1 iodide fluorescently stained cells, used in tandem with 3D volume morphologic reconstruction, may provide a useful research diagnostic tool in pathology. Accepted: 7 November 1996     

Cytological criteria of endometrial lesions with emphasis on stromal and epithelial cell clusters: result of 8 years of experience with intrauterine sampling

Objective:  There are a number of unresolved issues in endometrial cytology. They include the significance of nuclear atypia for the diagnosis of grade1 adenocarcinoma (G1AC) and atypical endometrial hyperplasia (AEH), cytological criteria of endometrial hyperplasia without atypia, and recognition of stromal cell cluster (SC) and its distinction from epithelial cell cluster (EC).
Methods:  We examined nuclear atypia, SC and EC in typical cases of five categories: normal endometrium (NEM), simple endometrial hyperplasia without atypia (SEH), complex endometrial hyperplasia without atypia (CEH), G1AC and grade2 adenocarcinoma (G2AC). We classified EC into four types: simple EC (SPEC), large regular EC (LREC), large irregular EC (LIEC) and small irregular EC (SIEC). Based on the results, we developed criteria of endometrial cytology and have evaluated 13 639 cases over 8 years.
Results:  Nuclear atypia was significantly more frequent in G2AC than in any of the other four categories ( P  <   0.001). SC was significantly more frequent in NEM and SEH than in the other three categories ( P  <   0.001). G1AC and G2AC showed significantly higher frequency of LIEC than the other three categories ( P  <   0.001). CEH exhibited significantly higher frequency of LREC than the four categories ( P  <   0.001). The sensitivity and the specificity was 88.8% and 99.0% respectively.
Conclusions:  We could diagnose G1AC, G2AC and CEH with high accuracy using the established criteria mainly based on SC and EC. We think that the criteria may facilitate an effective screening and an objective interpretation of endometrial samples.     

Is there any potential link among caspase-8, p-p38 MAPK and bcl-2 in clear cell renal cell carcinomas? A comparative immunohistochemical analysis with clinical connotations

Background

Management of endometrial precancerous lesions has been of much debate due to inconsistencies in their classification, natural history and histologic diagnosis. Endometrial hyperplasia constitutes a wide range of histomorphologic features associated with high intra and interobserver diagnostic variability. Although traditional microscopic diagnosis is by far the most applicable method and the gold standard for histomorphologic diagnosis, digitized image analysis has been used as a powerful adjunct to maximize the histologic data retrieval and to add some detailed objective criteria for correct diagnosis in difficult cases.

Methods

A series of 100 endometrial curettage specimens with diagnosis of endometrial hyperplasia or well differentiated adenocarcinoma were blindly reviewed by 5 pathologists; their intra and interobserver reproducibility determined and further compared to the objective morphometric data i.e. D-score and volume percent of stroma (VPS).

Results

The results were assessed using the weighted kappa statistics. Mean intraobserver kappa value was 0.8690 (99.44% agreement). Mean interobserver kappa values by diagnostic category were: simple hyperplasia without atypia: 0.7441; complex hyperplasia without atypia: 0.3379; atypical hyperplasia: 0.3473, and well-differentiated endometrioid carcinoma: 0.6428; with a kappa value of 0.5372 for all cases combined. Interobserver agreement was in substantial rate for simple hyperplasia (SH) and well differentiated adenocarcinoma (WDA) but was in fair limit for complex hyperplasia (CH) and atypical hyperplasia (AH). Intraobserver agreement was almost perfect. The specimens were divided in two groups according to the computerized morphometric analysis: Endometrial Hyperplasia (EH) ( D Score ≥ 1 or VPS ≥ 55%) and Endometrial Intraepithelial Neoplasia (EIN) (D-Score < 1 or VPS < 55%). Morphometric findings were closely compatible with routine WHO classification made by one expert pathologist; however; diagnosis of (CH) and (AH) made by other pathologists were not concordant with morphometric data.

Conclusion

It may be necessary to make some revisions in WHO classification for endometrial hyperplasia and precancerous lesions.     

A karyometric approach to the characterization of atypical endometrial hyperplasia with and without co-occurring adenocarcinoma

OBJECTIVE: To develop a karyometric image analysis approach to distinguishing atypical endometrial hyperplasia with and without co-occurring adenocarcinoma. STUDY DESIGN: Six cases of atypical hyperplasia without and 6 cases with co-occurring adenocarcinoma, 4 cases of normal endometrium and 3 cases of adenocarcinoma were identified. From each case 100 nuclei were measured in representative diagnostic areas identified by an experienced pathologist. Discriminant analyses were performed. An unsupervised learning algorithm was applied to define and characterize different nuclear phenotypes, and those data were used to identify cases with co-occurring adenocarcinoma. RESULTS: Discriminant analysis showed that nuclei from atypical hyperplasia and atypical hyperplasia with co-occurring adenocarcinoma are statistically different. The unsupervised learning algorithm revealed differences in nuclear subpopulations that can be used to correctly identify an estimated 85% of individual cases. CONCLUSION: Nuclei from atypical hyperplasia without and with co-occurring adenocarcinoma have statistically different karyometric characteristics that may facilitate case classification.     

Liquid-based endometrial cytology: cyto-histological correlation in a population of 917 women

OBJECTIVE: Liquid-based cytology, because of its capacity to reduce the obscuring factors and to provide thin-layer specimens, represents an opportunity to reevaluate endometrial cytology. In order to assess the utility of the liquid-based method in endometrial diagnosis, we evaluated its accuracy in comparison with histology. METHODS: Nine hundred and seventeen women scheduled for hysteroscopy were enrolled in the study. After providing informed consent, all the women proceeded sequentially to hysteroscopy, endometrial cytology and then biopsy endometrial sampling. RESULTS: Cyto-histological correlations were possible in 519 cases (57%): in 361 (39%) cases the biopsy was inadequate, in 15 (2%) the cytology was inadequate, and in 22 (2%) both were inadequate. At biopsy 25 (3%) women had adenocarcinoma, 5 (1%) had adenomatous atypical hyperplasia and 21 (2%) had simple non atypical hyperplasia. At cytology two adenocarcinomas and one adenomatous atypical hyperplasia were underrated as atypical hyperplasias and as non-atypical hyperplasia; two simple non-atypical hyperplasias were reported as negative; and eight cases were false positive (non-atypical hyperplasia at cytology, negative at biopsy). In our population, the cytology provided sufficient material more often than biopsy (P < 0.04). Sensitivity was estimated at 96%, specificity at 98%, positive predictive value at 86% and negative predictive value at 99%. CONCLUSIONS: We concluded that endometrial cytology may be an efficient diagnostic method. It could be applied to selected patients solely or in association with ultrasonography. The combination of these two noninvasive procedures may improve their diagnostic accuracy and reduce unnecessary hysteroscopies, thereby producing benefits for women and society.     

Cytologic grading and DNA image cytometry of breast carcinoma on fine needle aspiration cytology smears

OBJECTIVE: To correlate the cytologic grade of breast carcinoma with DNA image cytometry (ICM) and nuclear area on fine needle aspiration cytology (FNAC) smears. STUDY DESIGN: In this prospective study, FNAC material from 28 breast carcinomas were studied for cytologic grade and DNA ICM. Breast carcinomas were classified as grade 1-3 (low to high). DNA histograms were classified by the modified Auer method. Degree of hyperploidy (DH), ploidy balance (PB) and nuclear area (NA) were measured on Feulgen-stained smears by a CAS 200 image cytometer. Cytologic grade was correlated with DNA ICM findings and NA. RESULTS: There were 3 cytologic grade 1, 13 grade 2 and 12 grade 3 breast carcinomas. Seven of eight cases of hypertetraploid aneuploidy were grade 3 tumors. All cytologic grade 1 tumors were diploid. There were significant differences in DH, PB and NA in different grades of breast carcinoma (one-way ANOVA). CONCLUSION: DNA image cytometry in combination with cytologic grading might offer additional information for the characterization of breast carcinomas diagnosed by FNAC. These observations are of particular interest with the introduction of preoperative chemotherapy.     

Flow cytometry and factor analysis evaluation of confocal image sequences of morphologic and functional changes occurring at the mitochondrial level during 7-ketocholesterol-induced cell death

OBJECTIVE: To analyze functional and morphologic alterations that occur at the mitochondrial level by flow cytometry and laser scanning confocal microscopy (CLSM) combined with factor analysis of biomedical image sequences (FAMIS). STUDY DESIGN: Under treatment of U937 cells with 7-ketocholesterol, functional alterations that occur at the mitochondrial level (especially loss of transmembrane mitochondrial potential [delta psi m]) were assessed with 3,3'-dihexyloxacarbocyanine iodide (DiOC6(3)) and mitotracker red (CMXRos), whereas morphologic changes were analyzed with nonyl acridine orange (NAO). By flow cytometry, these different dyes were excited at 488 nm, whereas on CLSM, excitation of NAO and CMXRos was performed by lines of an argon laser. By CLSM, spectral sequences were performed to characterize NAO and CMXRos. FAMIS was used to transform the image sequences in factor images. RESULTS: By flow cytometry, rapid loss of delta psi m induced by 7-ketocholesterol was detected with both DiOC6(3) and CMXRos, which gave similar results. Morphologic alterations of mitochondria were revealed with NAO. The factor images obtained from confocal image sequences confirmed these results. CONCLUSION: The simultaneous use of NAO, CMXRos and FAMIS constitutes a new method to detect morphologic and functional alterations occurring at the mitochondrial level during cell death.     

Analysis of CD36 expression on human monocytic cells and atherosclerotic tissue sections with quantum dots: investigation by flow cytometry and spectral imaging microscopy

OBJECTIVE: To demonstrate CD36 expression with quantum dots (QDs) 525 and/or 605 on human monocytic U937 cells and atherosclerotic tissue sections by means of flow cytometry (FCM) and/or confocal laser scanning microscopy (CLSM). STUDY DESIGN: U937 cells and tissue sections were analyzed by means of FCM and/or CLSM. FCM was performed, using different ultraviolet (UV) and visible (488/532 nm) excitation modes. In the visible mode, fluorescence intensities of QDs, phycoerythrin (PE) and fluorescein isothiocyanate (FITC) were compared. Three-dimensional (3-D) sequences of images were obtained by spectral analysis in a CLSM and analyzed by the factor analysis of medical image sequences (FAMIS) algorithm, providing factor curves and images. Factor images are the result of the FAMIS image processing method, which differentiates emission spectra from 3D sequences of images. In CLSM analysis, preparations are screened in a UV excitation mode to optimize the possibilities of QDs and have the benefit of 4',6-diamino-2-phenylindole or Hoechst 33342 counterstaining of nuclei. RESULTS: FCM and CLSM revealed CD36 expression by means of QDs 525 and/or 605. Fluorescence intensity of PE and of FITC was higher than that of QDs 525 and of 605. As factor curves and images show the red emission of QDs 605 only, subsequent reliable identification and localization of CD36 was obtained. CONCLUSION: QDs 525 and 605 are useful to analyze antigenic expression. Following FCM, which is well adapted to detect fluorescence emission of QDs in the UV or visible excitation mode, CLSM and subsequent spectral analysis assess more specific characterization of QD fluorescent emissions.     

Concurrent Quantification of Cellular and Extracellular Components of Biofilms

Confocal laser scanning microscopy (CLSM) is a powerful tool for investigation of biofilms. Very few investigations have successfully quantified concurrent distribution of more than two components within biofilms because: 1) selection of fluorescent dyes having minimal spectral overlap is complicated, and 2) quantification of multiple fluorochromes poses a multifactorial problem. Objectives: Report a methodology to quantify and compare concurrent 3-dimensional distributions of three cellular/extracellular components of biofilms grown on relevant substrates. Methods: The method consists of distinct, interconnected steps involving biofilm growth, staining, CLSM imaging, biofilm structural analysis and visualization, and statistical analysis of structural parameters. Biofilms of Streptococcus mutans (strain UA159) were grown for 48 hr on sterile specimens of Point 4 and TPH³ resin composites. Specimens were subsequently immersed for 60 sec in either Biotène PBF (BIO) or Listerine Total Care (LTO) mouthwashes, or water (control group; n=5/group). Biofilms were stained with fluorochromes for extracellular polymeric substances, proteins and nucleic acids before imaging with CLSM. Biofilm structural parameters calculated using ISA3D image analysis software were biovolume and mean biofilm thickness. Mixed models statistical analyses compared structural parameters between mouthwash and control groups (SAS software; α=0.05). Volocity software permitted visualization of 3D distributions of overlaid biofilm components (fluorochromes). Results: Mouthwash BIO produced biofilm structures that differed significantly from the control (p<0.05) on both resin composites, whereas LTO did not produce differences (p>0.05) on either product. Conclusions: This methodology efficiently and successfully quantified and compared concurrent 3D distributions of three major components within S. mutans biofilms on relevant substrates, thus overcoming two challenges to simultaneous assessment of biofilm components. This method can also be used to determine the efficacy of antibacterial/antifouling agents against multiple biofilm components, as shown using mouthwashes. Furthermore, this method has broad application because it facilitates comparison of 3D structures/architecture of biofilms in a variety of disciplines.     

Use of confocal microscopy in examining fungi and bacteria in wood

When used in conjunction with digital image processing techniques, confocal laser scanning microscopy (CLSM) enables non-invasive optical sectioning, allowing micromorphologies of wood decay to be examined at any depth within a relatively thick (0.05-0.1 mm) wood specimen without incision. In this study, the use of specially tailored multi-fluorescent staining techniques with CLSM produced new information concerning spatial relationships between fungi and bacteria and the wood substrate, particularly in regard to their 3D characteristics. Glutaraldehyde fixation and a chitin fluorescent probe were used to locate fungal hyphae in wood. Bacteria colonising wood were examined using a fluorescent phospholipid probe. By counterstaining wood with this probe and a fluorescent dye specific for Gram-positive bacteria, it was possible to clearly distinguish Gram types through simultaneous, multichannel fluorescent CLSM imaging. The combination of glutaraldehyde fixation and phospholipid probing proved to be reliable for detecting wood-degrading bacteria in wood cell walls.     

Potential of the learning vector quantizer in the cell classification of endometrial lesions in postmenopausal women

OBJECTIVE: To investigate the potential of artificial neural networks for cell identification in endometrial lesions from postmenopausal women. STUDY DESIGN: The study was performed on cytologic material obtained by the Gynoscann endometrial cell samplerfrom 12 cases of atrophic endometrium, 48 cases of hyperplasia without cytologic atypia (18 cases of simple hyperplasia and 30 cases of complex hyperplasia), 12 cases of hyperplasia with cytologic atypia (complex atypical hyperplasia) and 48 cases of adenocarcinoma (30 cases of well-differentiated, 12 cases of moderately differentiated and 6 cases of poorly differentiated carcinoma). From each case approximately 100 cells were examined using a custom image analysis system. A learning vector quantizer (LVQ) identified the collected data. RESULTS: Investigation of cells from Endometrial Alterations with LVQ proved that according to the nuclear characteristics, as expressed by morphometric and textural measures, the endometrial cells from postmenopausal women may be identified as belonging to one of thefollowing three groups: atrophy, hyperplasia without cytologic atypia (simple and complex hyperplasia) and malignant neoplastic lesions (atypical complex and adenocarcinoma). CONCLUSION: The role of nuclear morphologic features in the cytologic diagnosis of endometrial alterations was confirmed. The overlap in thefeature space observed indicates that cell characteristics do not form strictly separate clusters. Thatfact explains the difficulty that morphologists have with the reproducible identification of cells from endometrial lesions in postmenopausal women. Application of LVQ offers a good classification at the cell level and promises to be a powerful toolfor classification on the individual patient level andfor the clarification of the natural history of endometrial pathology.     

子宫内膜癌组织中uPA及Cath-D的表达及意义(英文)

目的:检测子宫内膜癌组织中尿激酶型纤溶酶原激活物(uPA)及组织蛋白酶(Cath-D)的表达并探讨相关性及其临床意义。方法:采用免疫组织化学方法(PV-6000二步法)检测31例子宫内膜癌组织(内膜癌组),17例子宫内膜增生组织(增生组)及10例正常子宫内膜组织(对照组)中uPA及Cath-D的表达,并研究其相关性。结果:1.内膜癌组中uPA和Cath-D的表达均高于增生组及对照组中的表达,差异均有统计学意义(P0.05);在增生组中的表达与对照组差异无统计学意义(P0.05)。2.uPA和Cath-D的阳性表达与子宫内膜癌的临床病理分期、组织学分级及肌层浸润深度有关,差异均具有统计学意义(P0.05)。3.内膜癌组中uPA与Cath-D的表达呈正相关(r=0.673,P0.05)。结论:uPA和Cath-D在子宫内膜癌发生发展及侵袭转移过程中起着协同作用,Cath-D可诱导产生活化的uPA,促进癌细胞的浸润转移,因此,两者的联合检测可有助于成为判断子宫内膜癌的发展及预后的重要指标。     

The combined GnRH-agonist and intrauterine levonorgestrel-releasing system treatment of complicated atypical hyperplasia and endometrial cancer: a pilot study

In this article, we present the results of organ-preserving treatment applied in 24 patients of reproductive age with atypical endometrial hyperplasia or early-stage endometrial cancer. All of them would like to preserve their reproductive potential. Thirteen women with atypical endometrial hyperplasia were treated with the combination of six intramuscular injections of 3.75 mg gonadotropin-releasing hormone agonist (GnRHa)--leuproreline acetate depot every 4 weeks. After the third injection of 3.75 mg of leuproreline acetate, the levonorgestrel intrauterine hormonal system containing 52 mg levonorgestrel (Mirena(?), Bayer, Germany) was inserted for at least 6 months. In 11 women with stage IA well-differentiated endometrial adenocarcinoma, hormonal therapy included nine intramuscular injections of 3.75 mg of GnRHa every 4 weeks. After the third injection of 3.75 mg of GnRHa, we also inserted a GnRH-IUS (Mirena(?)) for at least 12 months. This type of therapy was effective for all these patients and may be offered to be used as an alternative to surgery in women with atypical endometrial hyperplasia or early stage 1A well-differentiated endometrial cancer in women of reproductive age. Three women with endometrial cancer became pregnant and two of them delivered at term and one has an ongoing pregnancy.     

FAS在子宫内膜样腺癌及其癌前病变中的表达及意义

目的研究脂肪酸合成酶(Fatty acid synthase FAS)在子宫内膜样腺癌及其癌前病变中的表达以及与临床病理意义。方法 37例子宫内膜样腺癌、21例子宫内膜非典型增生、23例子宫内膜复杂性增生、17例子宫内膜单纯性增生及11例增生期子宫内膜中FAS的表达情况,结合临床病理参数并进行统计学分析。结果 FAS在子宫内膜样腺癌、子宫内膜非典型增生、子宫内膜复杂性增生、子宫内膜单纯性增生及增生期子宫内膜中均有表达,其阳性表达率依次为81.1%、57.1%、56.5%、52.9%、45.5%。子宫内膜样腺癌中FAS的阳性表达率明显高于增生期子宫内膜、子宫内膜单纯性增生、子宫内膜复杂性增生及子宫内膜非典型增生(P<0.05)。FAS的阳性表达率在浸润深度≥1/2肌层的子宫内膜样腺癌明显高于浸润深度<1/2肌层者(P<0.05)。FAS的表达与年龄及组织学分级无显著差异性(P>0.05)。在子宫内膜样腺癌中FAS的表达与ER有关(P<0.05)结论本研究提示FAS的表达可能与子宫内膜样腺癌的发生、发展有关。FAS可以作为提示子宫内膜样腺癌预后的一个新的参考指标。部分ER阳性的子宫内膜样腺癌的发生、发展可能与FA...     

Adaptive Geometric Tessellation for 3D Reconstruction of Anisotropically Developing Cells in Multilayer Tissues from Sparse Volumetric Microscopy Images

The need for quantification of cell growth patterns in a multilayer, multi-cellular tissue necessitates the development of a 3D reconstruction technique that can estimate 3D shapes and sizes of individual cells from Confocal Microscopy (CLSM) image slices. However, the current methods of 3D reconstruction using CLSM imaging require large number of image slices per cell. But, in case of Live Cell Imaging of an actively developing tissue, large depth resolution is not feasible in order to avoid damage to cells from prolonged exposure to laser radiation. In the present work, we have proposed an anisotropic Voronoi tessellation based 3D reconstruction framework for a tightly packed multilayer tissue with extreme z-sparsity (2–4 slices/cell) and wide range of cell shapes and sizes. The proposed method, named as the ‘Adaptive Quadratic Voronoi Tessellation’ (AQVT), is capable of handling both the sparsity problem and the non-uniformity in cell shapes by estimating the tessellation parameters for each cell from the sparse data-points on its boundaries. We have tested the proposed 3D reconstruction method on time-lapse CLSM image stacks of the Arabidopsis Shoot Apical Meristem (SAM) and have shown that the AQVT based reconstruction method can correctly estimate the 3D shapes of a large number of SAM cells.