HE-stained Sections Research Articles

Pathologic study of diffuse pulmonary interstitial fibrosis caused by chronic hypersensitivity pneumonitis

To study the pathologic characteristics of chronic hypersensitivity pneumonitis, especially the pattern of pulmonary interstitial fibrosis; and to compare the histologic features with those of idiopathic interstitial pneumonitis. The HE-stained paraffin sections of 10 cases of chronic hypersensitivity pneumonitis encountered during the period from 2000 to 2008 were retrospectively analyzed. There were altogether 6 males and 4 females, with age of patients ranging from 23 to 59 years (mean=47.2 years). Clinically, the patients presented with chronic cough and shortness of breath for 4 months to 6 years. Histologically, 7 cases showed usual interstitial pneumonitis (UIP)-like fibrosis. Patchy fibrosis was observed under the pleura, adjacent to interlobular septa and around bronchioles. In all of the 7 cases, foci of fibroblastic proliferation, as well as bronchiolar metaplasia of peribronchiolar alveoli and mild bronchiolitis, were noted. Three cases presented with mild honeycomb changes of lung and 3 cases showed non-specific interstitial pneumonitis (NSIP)-like fibrosis, in which the alveolar septa were expanded by fibrous tissue and collagen, with relative preservation of alveolar architecture. Bronchiolitis and lymphocytic infiltrates in alveolar septa were seen. Schaumann bodies were identified in 1 case. In general, patients with chronic hypersensitivity pneumonitis were younger than patients with idiopathic UIP. Computed tomography often showed upper and middle lobar involvement and mosaic attenuation. Compared with idiopathic UIP, the UIP-like fibrosis of chronic hypersensitivity pneumonitis often occurred not only under the pleura and adjacent to interlobular septa, but also around bronchioles and was accompanied by bronchiolar metaplasia. Chronic hypersensitivity pneumonitis can mimic other types of lung conditions with interstitial fibrosis, especially UIP and NSIP. As a result, some cases of chronic hypersensitivity pneumonitis may be misdiagnosed as such.

Some biological effects of scorpion envenomation in late pregnant rats

Scorpion envenoming is less studied during gestation; however, it may induce various biological disturbances in maternal organism and hypothetical ones on their fetuses. The scope of this report was to elucidate some biological effects of such poisoning in late pregnant rats. Hence, TBARS levels in maternal lung, placental and fetal pulmonary and hepatic tissues and dam's biochemical blood parameters (glucose, creatinine, 17-β estradiol, progesterone, blood nitrogen urea, sodium and potassium maternal plasma concentrations) had been evaluated after saline (G1), and scorpion venom (G2: 30 min and G3: 60 min) injections in 22nd day pregnant rats. Histological microscopic examination of these tissues was also carried out in HE-stained paraffin sections. In addition, the mean arterial blood pressure following the envenomation variations was measured in three rats from the same pool. Our results showed that Buthus occitanus tunetanus crude venom induced significant increase in maternal, placental and fetal tissues lipid peroxidation, concomitant with blood pressure elevation. Maternal plasma creatinine, estradiol and progesterone concentrations levelled up significantly after 30 min or later (60 min) after the venom injection. Except for a probable pronounced oedema and few congestions in maternal lungs and degenerative aspects of trophoblast cells, all examined tissues showed a conserved structure. These results suggest that scorpion envenomation may induce gestation process disturbances and threatens both mother's and fetus’ well-being.

Moderne morphologische Diagnostik und aktuelle Klassifikation von Weichteilsarkomen

Malignant soft tissue tumors are somewhat rare, and thus sufficient experience in diagnostics and therapy of these sarcomas is available as a rule only at specialist centers. The gold standard of morphological diagnosis is still represented by evaluation of HE-stained histological sections. However modern methods of examination are also helpful in diagnosis. Because immunohistochemistry is now used routinely, emphasis in this article is laid on molecular methods with special reference to fluorescence in-situ hybridization (FISH) and reverse transcriptase polymerase chain reaction. Principles of the WHO soft tissue tumor classification are explained, and the changed or expanded interpretation of some tumor entities is illustrated using the example of atypical lipomatous tumors, so-called malignant fibrous histiocytomas and hemangiopericytomas, fibrosarcomas, and inflammatory myofibroblastic tumors.

Diagnostic problems in the subtyping of renal tumors encountered by five pathologists

The diagnostic problems in the subtyping of renal tumors were evaluated by a panel of five pathologists studying a set of selected tumors. Five pathologists independently assessed a single hematoxylin-and-eosin (HE)-stained slide from 28 selected renal tumors. After this independent assessment, the pathologists reevaluated and discussed all discordant cases. Additional HE-stained sections and immunohistochemically (IHC) stained slides were available. The generalized kappa for interobserver agreement was calculated. After independent assessment of the HE-stained slides, the five pathologists unanimously reached an agreement in the decision between malignant and benign in 82% of the cases. Fifty percent of the cases were correctly subclassified. The overall generalized kappa value for the five pathologists was 0.320 (CI 95% 0.090–0.551), which is considered a moderate agreement. A 100% agreement was reached for all 28 cases after examination of more slides from different tumor areas and IHC-stained sections. An accurate histologic distinction between benign and malignant renal tumors is possible on one HE-stained section. Correct assignment of the subtype is difficult on one slide alone and relies on IHC-markers and additional slides. Tumors composed of an eosinophilic cell type and tumors with a papillary growth pattern were the major causes of an incorrect diagnosis on an HE-stained section alone.

Siatic nerve regeneration in rats stimulated by fibrin glue containing nerve growth factor: An experimental study

To study the effects of fibrin sealant containing nerve growth factor on the peripheral nerve regeneration. A four-group experimental design with repeated measures on one factor was used. Fibrin glue (FG) containing NGF was injected into the site of end-and-end sutured peripheral nerve (sciatic nerve) (group I: NGF+FG), meanwhile three control groups were set-up: group II (NGF), group III (FG), and group IV (normal saline). Observation to the function and morphology of the sciatic nerve was carried out at the end of 4, 8, 12 weeks postoperation. Data were analyzed using ANOVA, with the appropriate post hoc between-groups comparison test. Electrophysiological testing. The NAP and NCV of group I (NGF+FG) were greater than those of group II (NGF), group III (FG), or group IV (normal saline) (p<0.05). Sciatic functional index (SFI). It began to ameliorate at 4 weeks postoperation and SFI increased as time went on. And the SFI in group I (NGF+FG) was better than those in group II (NGF), group III (FG), or group IV (normal saline) (p<0.05). Morphology. In the MGF-stained sections, dissociated myelin debris was less and regenerated nerve fibres were in larger quantities in group I (NGF+FG) than in other groups. In the HE-stained sections, regenerated nerve fibres distal to anastomosis significantly increased, and axon and myelin had a clearer outline in group I (NGF+FG) than in other groups. Electron microscopy indicated that the regenerated nerve fibres were more mature and the development of the axons was greater in group I than in other control groups. FG can be used as carrier of exogenous NGF, and they can provide synergistic effects for the peripheral nerve regeneration after the integration of the two.

Gastrointestinal stromal tumor in Brazil: Clinicopathology, immunohistochemistry, and molecular genetics of 513 cases

The aim of the present study was to evaluate the clinicopathological, immunohistochemical, and molecular genetic features of gastrointestinal stromal tumors in Brazil and compare them with cases from other countries. Five hundred and thirteen cases were retrospectively analyzed. HE-stained sections and clinical information were reviewed and the immunohistochemical expression of CD117, CD34, smooth-muscle actin, S-100 protein, desmin, CD44v3 adhesion molecule, p53 protein, epidermal growth factor receptor, and Ki-67 antigen was studied using tissue microarrays. Mutation analysis of KIT and platelet-derived growth factor receptor-alpha genes was also performed. There was a slight female predominance (50.3%) and the median age at diagnosis was 59 years. The tumors were mainly located in the stomach (38.4%). Immunohistochemistry showed that CD117 was expressed in 95.7% of cases. Epidermal growth factor receptor expression was observed in 84.4% of tumors. p53 protein expression was found only in 2.6% of cases but all belonged to the high-risk group for aggressive behavior according to the National Institutes of Health consensus approach. No CD44v3 adhesion molecule expression was detected. KIT exon 11 mutations were the most frequent (62.2%). The present data confirm that gastrointestinal stromal tumors in Brazilian patients do not differ from tumors occurring in other countries.

Applications of Laser Scanning Cytometry in Immunohistochemistry and Routine Histopathology

Laser scanning cytometry (LSC) is a powerful tool for qualitative and quantitative analysis of tissue sections in preclinical drug development. LSC combines the strengths of flow cytometry with tissue architecture retention. This technology has been used predominantly with immunofluorescent techniques on cell culture and tissue sections, but recently LSC has shown promise in evaluating chromogenic immunohistochemistry (IHC) and histochemical products in paraffin-embedded and/or frozen tissue sections. Inverted light scatter measurements or a combination of inverted scatter and fluorescence allows automated determination of cell/nuclear counts (e.g., proliferation labeling indices), cell area (e.g., cellular hypertrophy), stromal elements, and labeling intensity (e.g., cytoplasmic/organellar proteins) in chromogen-labeled IHC or histochemical stained sections that correlates well with standard manual quantification methods. Segmentation with autofluorescence or dual immunolabeling facilitates capture of labeling data from specific cell populations. LSC evaluation of HE-stained sections is accomplished using autofluorescence/eosin fluorescence and inverse scatter. A standardized fluorescent approach with archivability, a lack of fluorescence quenching (photobleaching), and amenability to evaluation of multiple markers in a section has been demonstrated using Qdot nanocrystals. Examples of LSC use in chromogenic IHC, routine histopathology, and Qdot labeling will be reviewed, and advantages and disadvantages of this technology will be discussed.

Core Biopsies of Renal Tumors: A Study on Diagnostic Accuracy, Interobserver, and Intraobserver Variability

ObjectiveThe diagnostic accuracy of in-bench core biopsies (CBs) from renal masses, and the interobserver and intraobserver variability in pathological subtyping of renal tumors were assessed. MethodsWe performed two CBs in 62 consecutive renal masses suspected for renal cell carcinoma (RCC), obtained after radical or partial nephrectomy and, in one case, after autopsy. Routine hematoxylin-eosin (HE)–stained sections from each CB were evaluated by five pathologists on two occasions. The surgical specimen was the reference standard. Diagnostic accuracy and the generalized kappa for intraobserver and interobserver agreement were calculated. ResultsFive tumors were benign and 57 malignant. Eight percent to 16% of the CBs were considered inadequate for diagnosis. In 0–8% of the cases, the pathologist could not discriminate between a benign or malignant tumor. Overall accuracy ranged from 77% to 90%. Sensitivity (79–100%) and positive predictive value (100%) were high with narrow 95% confidence interval (95%CI). Specificity (100%) was high but negative predictive value (29%–100%) varied, with wide 95% CI. Interobserver agreement was fair to almost perfect (κ=−0.010 to 0.830) for the different subtypes. In 64–81% of the CBs, the subtype was correctly classified. Intraobserver agreement was substantial (mean κ=0.628) for all pathologists. ConclusionDiagnostic accuracy of CBs was high, with a diagnostic yield varying between 84% and 92%. Pathological subtyping of CBs was highly reproducible with the exception of the chromophobe renal cell carcinoma, which was problematic on HE-stained sections only.

Usefulness of Fungiflora Y to detect fungus in a frozen section of allergic mucin

Allergic fungal sinusitis is a non-invasive disease characterized by pansinusitis with nasal polyps, with most causative agents being members of the dematiaceous fungi. Reported herein is the ninth Japanese case of allergic fungal sinusitis; an Alternaria species was culturally identified as the etiological agent. The present patient, a 32-year-old man, presented with multiple sinusitis and nasal polyps. Allergic mucin, which is a histological hallmark of this disease, was examined intraoperatively on frozen section. Although characteristic histology, including laminated masses of mucin and aggregates of eosinophils, was seen in HE-stained sections, fungal hyphae were scarce and recognized merely as clear spaces. Because fluorescent stilbene derivatives seem useful for the rapid detection of fungi, Fungiflora Y, a commercially available staining kit that includes a fluorescent stilbene derivative, was applied to the sections, and hyphae were successfully visualized using a fluorescence microscope. This stain takes only several minutes to perform. It is concluded that Fungiflora Y should be used when fungal elements are hard to find on HE stain, especially in cases in which rapid detection of fungus is requested.

Application of fluorescence microscopy on hematoxylin and eosin‐stained sections of healthy and diseased teeth and supporting structures

Destruction of dental tissue and supporting structures is usually microscopically assessed by routine hematoxylin and eosin (HE)-stained sections. This short communication is concerned with the potential role of fluorescence microscopy of HE-stained sections to study morphological aspects of intact and pathological teeth in dental research. This methodology improves the visualization of the anatomical structures of the intact teeth, especially anatomical features and periodontal ligament spatial distribution. This technique also improves the visualization of the root and bone resorption and the delineation of the periapical lesion extension. The fluorescence microscopy technique of HE-stained sections is an easy, reliable and inexpensive method that seems to be a useful tool for evaluating morphological aspects of intact and pathological teeth in dental research.

Effect of Particle Size of Intratracheally Instilled Crystalline Silica on Pulmonary Inflammation

Crystalline silica, known as a causal substance of silicosis, has been carefully evaluated for its carcinogenicity and fibrogenicity. In this study, we instilled crystalline silica of two different size (S(1.8) :1.80 microm (S.D. 2.0), S(0.7) :0.74 microm (S.D. 1.5)) into the trachea of rats to evaluate the size effects of the particles on pulmonary inflammation. S(1.8) and S(0.7) samples were administered to rats by a single intratracheal instillation (2 mg/ 0.4 ml saline). At three days, 1 wk and 1, 3 and 6 months after the instillation, the blood, bronchoalveolar lavage fluid (BALF), and pulmonary tissues were analyzed. Six images per HE-stained section were digitally captured and examined by the point counting method (PCM). Polymorphonuclear leukocyte (PMN)-in-blood specimens and cytospin specimens from BALF were stained immunohistochemically with BrdU. At six months after the instillation, the effects on inflammatory cells in the pulmonary tissues and BALF tended to be more marked in the rats instilled with S(1.8) than those instilled with S(0.7). Particularly, clear differences were observed in the number of inflammatory cells in BALF. Even if the particles are of the same chemical composition, the results suggest that, their biological effects vary depending on their particle size. Therefore, when such particles are used in workplaces, strict control systems should be established according to the risks present by different sizes of particles.

Histometría de la glándula sublingual de ratones (Mus musculus) machos y hembras infectados con la cepa RAL del parásito de Chagas, Trypanosoma cruzi

The aim of this work was to analyze histologically and histometrically the sublingual gland of mice infected with the RAL strain of T. cruzi, according to the sex. Swiss mice (Mus musculus) were inoculated with 2 x 10(4) blood trypomastigotes of the RAL strain of T. cruzi. In the peak of the parasitemia (12th day) the mice were sacrificed, and the sublingual glands were fixed in ALFAC. HE-stained histological sections were evaluated histometrically. The parasitemia was higher in females. Histopatologically, acini of the infected animals were smaller, with scanty production of secretion, and smaller striated ducts. The nuclei of the demilunes were smaller and showed amastigote nests in the cytoplasm. Karyometrically, nuclei of the acini, demilunes and striated ducts were smaller in the infected mice. Stereologically, it was observed that relative volumes of acini and ducts were smaller and, inversely, relative volumen were greater for the conjunctive tissue in the infected males. The surface densities of acini and ducts were bigger and the diameter and thickness of the wall were smaller in this group. On the other hand, relative volume of acini was smaller and those of the ducts and conjunctive tissue were bigger in the infected females. The diameter and thickness of the wall of acini were smaller, and those of the striated ducts were bigger in this group. The RAL strain of T. cruzi caused general atrophy in the sublingual gland, with numerous nests of parasites in the glandular parenchyma.

Investigation on Radiofrequency and Laser (980 nm) Effects after Endoluminal Treatment of Saphenous Vein Insufficiency in an Ex-vivo Model

An ex-vivo model for the experimental evaluation of endoluminal thermal procedures for occlusion of saphenous veins was developed. Radiofrequency obliteration (RFO) and endovenous laser therapy (ELT) were compared using this model. Experimental ex-vivo treatment study. The model consists of the subcutaneous foot veins from freshly slaughtered cows which were reperfused in situ with heparinised bovine blood. The veins were treated with either radiofrequency (RFO n=5) or with endoluminal 980 nm laser light (ELT n=5) using a continuous pull-back for RFO and a stepwise illumination and pull-back protocol for ELT. Immediately after treatment perivenous tissue and veins were examined macroscopically. In a second study the same treatment parameters were used in four further vein segments with RFO (n=2) and ELT (n=2). These vein segments were examined microscopically in HE-stained histological sections. Induration of the vessel wall and contraction of the vessel lumen were observed after RFO. Laser treatment produced carbonised lesions of the vein wall. After 12-24 laser exposures these lesions often became transmural, causing complete perforation of the vessel wall. Histological evaluation after radiofrequency treatment demonstrated homogenous circular thermal tissue alteration with disintegration of intima and media structures. Histological evaluation after endovenous laser treatment showed large variations of thermal tissue effects. Tissue effects ranged from major tissue ablation and vessel wall disruption to minor effects located between laser exposures and on the opposite vessel wall. Our model is suitable for systematic scientific evaluation of endovenous thermal occlusion procedures. Our first results and theoretical considerations indicate that endovenous laser treatment should be modified in order to ensure controlled homogenous circular thermal damage, avoiding vessel wall perforation and damage to perivascular structures.

Molecular verification of stereotactic radiotherapy in rats using ATMpS1981 immunofluorescence

Background and purposeStereotactic radiation therapy and radiosurgery (STS) might be useful for focused treatment in orthotopic tumor models or to evaluate normal tissues. Spatial accuracy of dose-delivery is crucial and needs to be proven. This study intended to show on a molecular level precision of STS using the radiation activated molecule phospho-serine 1981-ATM (ATMpS1981). Patients and methodsWistar rats underwent MRI fused with CT for treatment with a NOVALIS™ (BrainLab) STS device. Rats (body weight=350g, respectively) were irradiated with 2Gy to selected areas of the brain, the lung, the liver, and the pancreas. Animals were sacrificed 1h after STS and organs were explanted for pathohistological workup. Coronal HE-stained sections of treated organs were matched with coronal MRI and CT reconstructions and isodose distributions of STS. Immunofluorescence signal expression in the nuclei after incubation with anti-ATMpS1981 antibody was compared to the target volume of STS. ResultsThree consecutive animals were treated with STS using 2Gy single dose which resulted in no apparent changes on HE staining but obvious immunofluorescence signals in irradiated regions. In concordance with isodose distributions these signals had a maximum intensity within the 90% isodose and gradually decreased to a background level with very low nuclear immunofluorescence signals for ATMpS1981. These results were consistent throughout all of the four investigated organs (brain, lung, liver, pancreas) and could be repeated in all of the investigated animals. ConclusionsATMpS1981 immunofluorescence confirms accuracy of STS allowing for partial organ irradiation in small animals. It can be used for quality assurance in STS treatment of normal tissue or orthotopic tumor models.

Gastric Helicobacter-like Organisms in Stray Cats

Ten adult domestic shorthaired stray cats (Felis catus) were investigated for the presence and localization of different species of gastric Helicobacter-like organisms (GHLOs) using Warthin-Starry silver staining, immunohistochemistry and transmission electron microscopy (TEM); the severity and distribution of lesions in different regions of the stomach were assessed in HE-stained sections. GHLOs were present in all areas of the stomach in all of 10 cats. Three morphologically different types of spiral-shaped bacteria were demonstrated; in silver-stained sections, H. pylori like organisms (HPLO) were easily differentiated from other GHLOs. Eight of the cats had H. heilmannii-like organisms (HHLOs) and one cat had HPLO. Mixed H. heilmannii and H. felis infection was seen in only one cat. GHLO infection was associated with a mild to severe gastritis in 8 of 10 cats. GHLOs colonized the cardia, corpus and antrum in similar density. The most striking histopathological changes consisted of accumulation of lymphocytes and neutrophilic granulocytes, fibrosis of the lamina propria mucosae, lymphoid follicles and lymphocytic infiltrates. There was no obvious relation between the degree of colonization by GHLOs and the extent of histopathological changes. GHLOs were present on the mucosal surface, in the lumen of gastric glands, and in the cytoplasm of parietal cells. These findings indicate that immunohistochemistry and silver staining are useful for detecting GHLO infections, particularly with different Helicobacter species present. Stray cats are frequently colonized by HHLOs without any significant correlation between the degree of infection and gastritis score; in contrast HPLOs and HFLOs infections are not very common.

Immunohistochemical study of serum albumin in normal and cadmium-treated mouse testis organs by “in vivo cryotechnique”

The in vivo injection of cadmium (Cd) was reported to induce blood-testis barrier disruption, and assumed to be an experimental model to examine junctional structures in seminiferous tubules. The purpose of this study is to investigate time-dependent changes of albumin permeability in the normal or Cd-treated mouse testis by our "in vivo cryotechnique" with immunohistochemistry, reflecting tight junctional (TJ) barriers of Sertoli cells. The albumin in the seminiferous tubules was firstly immobilized by the cryotechnique, in which normal blood circulation was always kept. The cryofixed testicular tissues were then processed for freeze-substitution, and embedded in the paraffin wax. Serial sections were immunostained by anti-mouse albumin antibody with peroxidase immunostaining, and also stained with hematoxylin-eosine (HE) for morphological observation. In normal seminiferous tubules, the immunoreaction products were localized around peritubular myoid cells and between Leydig cells, as well as in blood vessels. They were also localized as arch-like patterns around some spermatogonia in basal compartments of seminiferous tubules. Twenty-four and 48 hrs after Cd-treatment, some enlarged spaces and vesicular formations in the seminiferous epithelium were observed on the HE-stained sections. The albumin immunolocalization was detected not only in the basal compartments, but also in the adluminal compartments between Sertoli cells and germ cells. Thus, the structural disruptions of inter-Sertoli TJ barriers could be clearly demonstrated by the "in vivo cryotechnique".

Distribution of labelled anti-tenascin antibodies and fragments after injection into intact or partly resected C6-gliomas in rats.

For treatment of malignant glioma, radioimmunotherapy has become a valuable alternative for more than 2 decades. Surprisingly, very little is known about the distribution of intralesionally administered labelled antibodies or fragments. We investigated the migration of labelled antibodies and antibody fragments injected into intact and partly resected C6-glioma in rats at different times after injection. Nine days after induction of a C6-glioma, 5 microl of 125I-labelled murine anti-tenascin antibodies (n = 31) or 125I-labelled fragments of anti-tenascin antibodies (n = 32) was injected slowly into the tumour (group I). In group II the tumour was subtotally resected 9 days after induction of the C6-glioma, and 24 h later the labelled antibodies (n = 30) or fragments (n = 12) were injected into the resection cavity. At 6, 24 or 48 h after the injection, animals were sacrificed, and brains removed. Distribution of labelled antibodies and fragments was determined by superimposing autoradiographs onto frozen sections and HE-stained neighbouring sections using a digital image analysing system. After injection into intact C6-glioma, labelled antibodies covered a maximum distance of 3.2 +/- 1.0, 4.1 +/- 1.9 and 4.8 +/- 0.9 mm after 6, 24 and 48 h, respectively; while labelled fragments were found at a distance of 6.7 mm (+/-1.1) after 24 h and 5.8 mm (+/-0.9) after 48 h (significant in univariate analysis). Following partial tumour resection, the respective distances at 24 h were 3 +/- 0.4 mm for anti-tenascin antibodies and 3.4 +/- 0.3 mm for Fab fragments. After injection into C6-glioma, labelled fragments are able to cover a greater distance than labelled antibodies. Injection of antibodies and fragments 1 day after tumour resection results in reduced velocity of both antibodies and fragments.

The effects of the novel selective progesterone receptor modulator (SPRM) asoprisnil on the morphology of the reproductive tract of cycling and ovariectomized guinea pigs

Asoprisnil, its active metabolite J912, and J956 are novel SPRMs that exhibit partial (mixed) agonist/antagonist activities in animal and humans. SPRMs inhibit endometrial proliferation and induce amenorrhea in non-human primates and humans. To evaluate the effects of asoprisnil, J912, and J956 on weights and morphology of the uterus and vagina in intact and ovariectomized (OVX) guinea pigs. The effects of SPRMs were compared with those of promegestone (R5020, pure progestin), progesterone (P) and the antiprogestins mifepristone (RU486) and onapristone (ONA). Intact guinea pigs (n=5/group) were treated subcutaneously (s.c.) with either the SPRMs or the reference compounds RU486, ONA, R5020, P or vehicle from cycle day 8–14 (dose/animal/day: 1.0mg for promegestone and 10.0mg for the other compounds). OVX guinea pigs (n=7/group) were primed s.c. with estradiol (E2) (0.03μg/day for 5 days) and then treated with the SPRMs and the reference compounds for 14 days in the presence of E2 (same dose). Autopsy was performed 24 hours later. Histological evaluation was based on serial HE-stained paraffin sections (6–10μm). SPRMs, progesterone and R5020, but not the antiprogestins, significantly (p<0.05) reduced the uterine and vaginal weights in intact animals. Proliferation and cornification of the vaginal epithelium prevailed after ONA and RU486. The employed E2 dose, when administered alone, led to minor increase of uterine and vaginal weights. ONA enhanced E2 effects on the uterus and vagina. Vaginal and endometrial histology revealed a massive epithelial proliferation, consistent with an “unopposed” estrogenic effect. The endometrium of animals exposed to ONA+E2 was highly hyperplastic with cystic changes. In contrast, the SPRMs and R5020 restored secretory appearance of the endometrium and induced mucification of the vaginal epithelium. Asoprisnil and the structurally related SPRMs exhibited P agonist effects on endometrial and vaginal morphology in both intact and OVX guinea pigs. In contrast, the antiprogestins induced a condition of “unopposed” estrogenic effects in the vagina and uterus. Asoprisnil has the potential to control hormone-dependent gynaecological disorders without inducing “unopposed” estrogen effects.

Successful sentinel node identification in colon carcinoma using Patent Blue V

Aim. The aim of this study was to evaluate the use of Patent Blue V for identification of the sentinel node in patients with colon carcinoma.Method. From May 2002, 35 patients operated for colon carcinoma underwent lymphatic mapping using Patent Blue V as marker. Either directly after resection of the colon or during operation 2 ml of Patent Blue V was injected peritumourally, and the first 1 to 4 blue nodes were marked as sentinel nodes. Pathological evaluation was done on a single HE-stained section of all lymph nodes. Only if all sentinel nodes were negative for metastases, serial sectioning and additional immunohistochemical staining against keratine CK 7/8 was performed to reveal micrometastasis in the sentinel nodes.Results. In 33/35 of patients at least one sentinel node was identified. In 10/33 the sentinel node was positive for metastases, and in 5/10 this was the only node containing metastases. One patient had a false negative sentinel node (accuracy 97%, sensitivity 91%).Conclusion. Using Patent Blue V, it is possible to identify the sentinel node in most patients with colon cancer. The results are comparable with other sentinel node studies using Lymphazurin.

The germ line lineage in ukigori, Gymnogobius species (Teleostei: Gobiidae) during embryonic development

In order to determine the origin and migration of ukigori primordial germ cells (PGCs), we observed the aggregation of vasa mRNA by whole mount in situ hybridization. To observe PGC migration in the germ layers, we analyzed HE-stained paraffin sections. The germ line lineages were derived from the edge of the first, second and third cleavage furrows. During subsequent cleavages, vasa mRNA aggregations were respectively taken into four to eight cells in each embryo and vasa expressing cells proliferated from the sphere stage. At the bud to early somitogenesis period, PGCs aligned from head to tail bud regions on both sides of the embryonic body. During the late somitogenesis period, PGCs mainly aggregated just underneath the body axis. After gut formation, PGCs aligned along both sides of the gut at the 4th- to 8th- somite regions. Finally, PGCs reached the genital ridge via the inside of the lateral plate mesoderm and dorsal peritoneum. These results suggest that localized patterns of vasa transcripts and the migration routes of PGCs are different among fish (Teleost) species, perhaps depending on the amount of germinal cytoplasm derived maternally and the timing of endoderm differentiation.