Conventional Hematoxylin Research Articles

Bergmann glia are reduced in spinocerebellar ataxia type 1

Non-cell-autonomous pathology involving glial cells has been implicated in Purkinje cell degeneration. We reported previously that mutant ataxin-1, a causative gene product of spinocerebellar ataxia type 1 (SCA1), prevents Bergmann glia proliferation in mutant ataxin-1 knockin mice and that suppressed Bergmann glia function leads to Purkinje cell degeneration. However, because reactive astrocytes are produced in response to brain injuries and diseases, Bergmann glia are also suspected to proliferate and increase in response to Purkinje cell degeneration, including during SCA1 pathogenesis. However, little is known about reactive Bergmann glia (Bergmann gliosis) and its beneficial or detrimental role. Given the lack of quantitative studies of Bergmann glia using specific molecular markers, we quantified Bergmann glia in human SCA1 brains with Bergmann glia-specific Sox2 staining and conventional hematoxylin and eosin staining. Our results showed reduced numbers of Bergmann glia in SCA1 patient brains and support the hypothesis that Bergmann glia loss contributes toward Purkinje cell degeneration in human SCA1.

Differences in Tumor Thickness Between Hematoxylin and Eosin and Melan-A Immunohistochemically Stained Primary Cutaneous Melanomas

Tumor thickness (Breslow thickness) represents the main prognostic factor in primary melanoma. Potential differences in melanoma tumor thickness measurements between conventional hematoxylin and eosin (H&E) and Melan-A immunohistochemical staining were evaluated. Ninety-nine excisional biopsies were included in the study. From each sample, 2 consecutive histological sections were stained with H&E and Melan-A, respectively. Tumor thickness was measured from both sections by 2 independent observers. In 59 biopsy specimens (59.6%), higher tumor thickness measurements were recorded in Melan-A-stained than in H&E-stained sections. In 42.4% of such cases (25 biopsies), the observed differences were ≥0.2 mm. After Melan-A evaluation, 33% of in situ melanoma cases were reclassified as invasive melanoma, with thickness measurements ranging from 0.15 to 0.35 mm. In 23 biopsies, identical values were recorded with both techniques, whereas in 17 cases, measurements obtained with H&E staining were slightly higher (from 0.01 to 0.18 mm) than those obtained with Melan-A staining. A high rate of interobserver agreement was noted, and significant intertechnique measurement differences were detected. Significant discrepancies (≥0.2 mm) in thickness measurements between the 2 techniques were mainly attributed to the presence of individual or small clusters of melanocytic cells in the papillary dermis. These melanocytic cells could be easily overlooked in H&E-stained sections, especially in sections showing dense lymphohistiocytic inflammatory infiltrates, numerous melanin-containing histiocytic cells in the upper dermis, or extensive fibrotic changes or regression phenomena. This study confirms the practical interest of immunohistochemical staining with Melan-A in evaluating primary melanoma and, specifically, in situ melanoma cases.

Endocytoscopic visualization of squamous cell islands within Barrett’s epithelium

To study the endocytoscopic visualization of squamous cell islands within Barrett's epithelium. Endocytoscopy (ECS) has been studied in the surveillance of Barrett's esophagus, with controversial results. In initial studies, however, a soft catheter type endocytoscope was used, while only methylene blue dye was used for the staining of Barrett's mucosa. Integrated type endocytoscopes (GIF-Q260 EC, Olympus Corp, Tokyo, Japan) have been recently developed, with the incorporation of a high-power magnifying endocytoscope into a standard endoscope together with narrow-band imaging (NBI). Moreover, double staining with a mixture of 0.05% crystal violet and 0.1% of methylene blue (CM) during ECS enables higher quality images comparable to conventional hematoxylin eosin histopathological images. In vivo endocytoscopic visualization of papillary squamous cell islands within glandular Barrett's epithelium in a patient with long-segment Barrett's esophagus is reported. Conventional white light endoscopy showed typical long-segment Barrett's esophagus, with small squamous cell islands within normal Barrett's mucosa, which were better visualized by NBI endoscopy. ECS after double CM staining showed regular Barrett's esophagus, while higher magnification (× 480) revealed the orifices of glandular structures better. Furthermore, typical squamous cell papillary protrusion, classified as endocytoscopic atypia classification (ECA) 2 according to ECA, was identified within regular glandular Barrett's mucosa. Histological examination of biopsies taken from the same area showed squamous epithelium within glandular Barrett's mucosa, corresponding well to endocytoscopic findings. To our knowledge, this is the first report of in vivo visualization of esophageal papillary squamous cell islands surrounded by glandular Barrett's epithelium.

Galectin-3 in preneoplastic lesions of glioma

Gliomas are the most common primary brain tumors in adults and have a poor prognosis. Galectin-3 is a β-galactosidase-binding lectin which is important in pre-mRNA splicing, regulation of cell proliferation, cell adhesion and apoptosis. Although galectin-3 has been shown as a glioma related marker and expression of galectin-3 has been reported to correlate with WHO grade in human gliomas, expression of galectin-3 in early neoplastic lesions such as early neoplastic proliferation (ENP) and microtumor is still far from fully understood. In the present study, expression of galectin-3 in ethylnitrosourea-induced rat gliomas including preneoplastic and neoplastic lesions was examined by immunohistochemistry for galectin-3, Iba-1 (a specific microglial cell marker), GFAP (a specific astrocyte cell marker), and conventional hematoxylin and eosin staining (for morphological observation). The results showed that exact location of ENP was detected clearly by galectin-3 immunohistochemistry whereas normal brain tissues were negative. In ENP and microtumor, galectin-3 was expressed in neoplastic astrocytic cells but rarely in microglia. In malignant glioma, however, galectin-3 was expressed in both neoplastic astrocytic cells and microglia. This suggests that galectin-3 is activated in microglia and macrophages according to the progression of glioma. Galectin-3 was not expressed in oligodendrocytic cells. Our results indicate that galectin-3 is a good specific marker indicating the early stage of glioma tumorigenesis.

Periodontal Findings in Patients With Oral Submucous Fibrosis and Comet Assay of Affected Gingival Epithelial Cells

Oral submucous fibrosis (OSF) involves large sections of population and is related to certain chewing practices that involve direct exposure of the gingiva to noxious chemicals and additives. The impact of such practices, although studied extensively in relation to the cheek mucosa, is not elucidated as far as gingival tissues are concerned. The possible DNA damage on affected gingiva is also one of the objectives of the present study. One hundred cases of individuals with OSF and 89 without OSF were examined for periodontal parameters and compared. Biopsies of gingival tissue from both categories were examined with conventional hematoxylin and eosin staining for epithelial, connective tissue, and inflammatory changes. Comet assay of epithelial cells obtained from scraping the gingival surface of both patients with OSF and control individuals was performed. Patients with OSF had poorer oral hygiene and greater loss of attachment, probing depth, and recession compared with controls. Significant histopathologic changes in the form of loss of rete ridges with gingival epithelium reduced to a flattened surface was observed. All the patients with OSF showed comet cells in larger numbers. The controls also showed comet cells but in fewer numbers, and the number of individuals showing comet cells was significantly less. OSF showed significant changes in affected gingiva, and the presence of comet cells in all the patients with OSF is a significant indicator of possible pathognomonic developments.

Direct Molecular Tissue Analysis by MALDI Imaging Mass Spectrometry in the Field of Gastrointestinal Disease

*Institute of Pathology, Research Unit Analytical Pathology, Helmholtz Zentrum Munchen, German Research Center for Environmental Health, Neuherberg, Germany; ‡ § Department of Medicine II, Klinikum rechts der Isar, Technische Universitat Munchen, Munich, Germany; Department of Medicine II, Universitatsklinikum Mannheim, University of Heidelberg, Mannheim, Germany; and Institute of Pathology, Technische Universitat Munchen, Munich, Germany

DisseminatedNocardia farcinica: Literature Review and Fatal Outcome in an Immunocompetent Patient

Nocardia farcinica is a gram-positive, partially acid-fast, methenamine silver-positive aerobic actinomycete. Nocardia spp. are opportunistic pathogens, and N. farcinica is the least common species of clinical importance. Review of the recent literature and description of a immunocompetent patient with no known risk factors who contracted fatal N. farcinica sepsis. Positive pre-mortem and post-mortem cultures from the lung and synovium correlated with acute bronchopneumonia and synovitis at autopsy. Colonies of filamentous bacteria, which were not apparent in conventional hematoxylin and eosin-stained sections, were observed with gram and methenamine silver stains, but acid-fast stains were negative. A literature review revealed that disseminated N. farcinica often is associated with an underlying malignant tumor or autoimmune disease (88% of patients). Chemotherapy or corticosteroid treatments are additional risk factors. Trimethoprim-sulfamethoxazole typically is the first-line therapy for N. farcinica; treatment with amikacin and imipenem-cilastatin is used less often (7% of patients). Despite aggressive therapy, we observed that the death rate (39%) associated with N. farcinica in recent publications was eight percentage points higher than reported in a review from 2000.

Abstract 4374: Combination of Bevacizumab and oncolytic virus hRr3 in an experimentalmodel of pancreatic carcinoma

Abstract Aim: Proangiogenic growth factors stimulate endothelial cells in nearby vessels to proliferate and migrate into the tumor. Vascular endothelial growth factor (VEGF) is an endothelial cell specific growth factor that is known to facilitate angiogenesis. Excess levels of VEGF are linked with poor prognosis. The effect oncolytic herpes viruses (OHSV) on VEGF A are not accurately determined. Nevertheless; limited number of studies on wild type herpes simplex type 1 (HSV1) have been shown to enhance VEGF A. The aim of the present study was to evaluate the effect of combination of OHSV hrR3 with a novel monoclonal VEGF A antibody Bevacizumab in a xenograft experimental model. Materials and Methods: Several cell were tested for expression of VEGFA. BxPC3 cell line had the highest and MIA PaCa-2 had the lowest expression of VEGFA among the other cell lines. 20 BALB-C nu/nu mice were divided into 4 experimental groups. All animals were implanted 5 mm diameter of BxPC-3 tumor xenograft on each side of their flank. After 2 week of observation; Control group (n=5) received no therapy; oncolytic viral therapy group (OVT) (n=5) received 5x106 pfu hrR3 injected intratumorally per week for two weeks; Bevacizumab treated group (BVT) (n=5) received 5mg/kg Bevacizumab intraperitoneally two times a week for 3 weeks; Combination therapy group (CT) (n=5) received both Bevacizumab and hRr3 virus injected intratumorally the same periods as other groups. The same was performed for MIA PaCa-2 cell lines as In order to confirm our results in a low expression cell line. Tumor diameters were evaluated every 3 days by caliper for 8 weeks. Tumor specimen was collected for conventional hematoxylin eosin stain, immunohistochemistochistry for VEGFA, Thrombospondin1, CD 31, HSV type 1 antigen and TUNEL. In MIA PaCa-2 cell line; VEGFA antigen distribution was especially evaluated on day 1, 3, 5, 7, 14 following hRr3 treatment. Results: The BxPC3 tumor volumes of the animals in the combination therapy were significantly lower than the other groups (p<0.05). Nevertheless; we were unable to show a statistically significant difference among OVT and BVT groups in terms of tumor volume. Histopathologically the CT and the OVT group had degenerated cells and low cellular density and expressed HSV antigen. Interestingly we found highest VEGFA expression in OVT group when compared to other groups (p<0.001). Tumor response was highest in combination group in the MIA PaCa-2 tumors as well (p<0.05). In Mia PACA2 tumors VEGFA expression was increasingly expressed starting from day 5 (p<0.05). Conclusion: Oncolytic virus seems to induce the VEGFA and this may be the proangiogenic stimulus in OHSV to induce angiogenesis. If a combination therapy towards blocking the VEGF A can be administered with OHSV; it may enhance the effect of the virus. Combination of OHSV with an antiangiogenic agent such as Bevacizumab seems to be a safe and effective treatment modality. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4374. doi:1538-7445.AM2012-4374

Reduction in Purkinje Fiber Number in Rats Undergone Fatal Electrocution

Purkinje fibers in cardiac conduction tissue during fatal electrocution. A total of 16 Sprague Dawley rats were divided into 2 groups as follows: the electrocution group and the control group.Animals were deeply anesthetized with sodium pentobarbital and, in the electrocution group, all 8 rats underwent a fatal electrical shock (220 v,50 Hz) followed by cervical dislocation. In the control group, all 8 rats underwent execution by cervical dislocation. Following death, hearts were rapidly excised and perfused with 1% paraformaldehyde before tissues of the left ventricular anterior wall (LVAW) were isolated. The microscopic structure of the Purkinje fibers were subsequently analyzed using conventional hematoxylin and eosin staining. A majority of the Purkinje fibers were located in groups among the cardiac muscle of the LVAW. A significant reduction in Purkinje fiber expression was displayed in the electrocution group compared with the control group (P G 0.05).The mean total number of Purkinje fibers for the electrocution and control groups were 59 T 11 and 3287 T 19 cells, respectively (P G 0.05).The estimated number of Purkinje fibers in the LVAW of the control group was significantly greater than observed in the electrocution group(41.09 T 0.24 vs. 0.7375 T 0.14, P G 0.05). The findings of the current study suggest that such a reduction would be reflected in abnormal cardiac conduction and a possible cause of sudden death.

Immunohistochemical dual staining as an adjunct in assessment of mitotic activity in melanoma

The mitotic rate was introduced as a major prognostic criterion for the staging of thin (≤1.0 mm) melanoma by the 2009 American Joint Committee on Cancer Staging and Classification (seventh edition). The detection of a single mitotic figure changes the tumor stage in thin melanoma. We sought to address the value of a dual staining to facilitate the determination of the mitotic rate and to assign the mitotic activity to melanocytes. The mitotic rate of melanoma cells was determined by dual phosphohistone-H3 (PHH3)/Melan-A immunohistochemistry. Results were compared with PHH3 staining alone and conventional hematoxylin and eosin (H&E)-stained slides of 15 melanomas with a tumor thickness <1.0 mm. PHH3 staining clearly labeled cells in the mitotic cell cycle. The mitotic rate in the PHH3/Melan-A dual stain was equal to that derived by H&E staining. Time required for counting mitotic figures was significantly reduced. The evaluation of mitotic rate with an immunohistochemical dual stain is faster (mean 63.0%) and more reliable than evaluation by routine H&E staining alone. Dual staining immunohistochemistry may be a useful additional tool to standardize the determination of mitotic rate and may be helpful in evaluation of challenging cases.

Tri-modal confocal mosaics detect residual invasive squamous cell carcinoma in Mohs surgical excisions

For rapid, intra-operative pathological margin assessment to guide staged cancer excisions, multimodal confocal mosaic scan image wide surgical margins (approximately 1 cm) with sub-cellular resolution and mimic the appearance of conventional hematoxylin and eosin histopathology (H&E). The goal of this work is to combine three confocal imaging modes: acridine orange fluorescence (AO) for labeling nuclei, eosin fluorescence (Eo) for labeling cytoplasm, and endogenous reflectance (R) for marking collagen and keratin. Absorption contrast is achieved by alternating the excitation wavelength: 488 nm (AO fluorescence) and 532 nm (Eo fluorescence). Superposition and false-coloring of these modes mimics H&E, enabling detection of cutaneous squamous cell carcinomas (SCC). The sum of mosaic Eo+R is false-colored pink to mimic the appearance of eosin, while the AO mosaic is false-colored purple to mimic the appearance of hematoxylin in H&E. In this study, mosaics of 10 Mohs surgical excisions containing invasive SCC, and five containing only normal tissue were subdivided for digital presentation equivalent to 4 × histology. Of the total 50 SCC and 25 normal sub-mosaics presented, two reviewers made two and three type-2 errors (false positives), respectively. Limitations to precisely mimic H&E included occasional elastin staining by AO. These results suggest that confocal mosaics may effectively guide staged SCC excisions in skin and other tissues.

Sentinel Node Biopsy in Early Gastric Cancer: Constant Exertion for Clinical Application

Lymph node status is the most important prognostic factor in gastric cancer. Standard surgery, therefore, is radical subtotal or total gastrectomy with wide dissection of regional lymph nodes. Recently, the proportion of early gastric cancer has increased to more than 60% in Korea and Japan because of screening gastroscopy. The rate of lymph node metastasis in early gastric cancer ranges from 5 to 20%, which means that more than 80% of patients with early gastric cancer undergo unnecessarily wide excision of the stomach with aggressive lymphadenectomy. If there is a tool to predict the exact status of lymph nodes, radical gastrectomy with lymphadenectomy can be substituted with limited resection of stomach without lymph node dissection. However, preoperative endoscopic ultrasound and computed tomography have limited accuracy for nodal staging in gastric cancer patients. The sentinel node (SN) concept has been introduced into various solid tumors, including stomach cancer, to predict lymph node status. Ideally, if the SN has no metastasis, extensive lymphadenectomy can be avoided, the extent of resection of the stomach reduced, function preserved, complications lessened, recovery accelerated, and the quality of life for the patient improved. SN navigation surgery has been widely performed for the treatment of breast cancer and malignant melanoma. Regarding gastric cancer, it seemed that SN concept is difficult to apply because the lymphatic flow is multidirectional. However, many studies reported that SN mapping is feasible in gastric cancer especially in early gastric cancer even though many issues still remain to be solved. Sensitivity of SN biopsy is highly important in gastric cancer compared with breast cancer and malignant melanoma. When false-negative cases develop in patients with malignant melanoma or breast cancer, other treatment options, such as chemotherapy and radiotherapy exist. Because gastric cancer has no other treatment option except surgery, a gastric cancer patient with false-negative SN loses an opportunity for cure. The detection rate is not as important as sensitivity in gastric cancer SN biopsy. Surgeons should perform just conventional lymphadenectomy for detection failure cases. Sensitivities of SN mapping in gastric cancer reported until now are not adequate for clinical application. Until now, the sensitivities were generally low and even the studies with high sensitivity had small number of cases with lymph node metastasis. There are several methods to increase the sensitivity of SN biopsy in gastric cancer. As pathologic methods, multiple sections of the SN may raise the rate of detecting lymph node metastases. Immunohistochemistry (IHC) staining with anti-cytokeratin antibody helps to detect micrometastases in SNs in addition to conventional hematoxylin and eosin (HE) staining. Reverse transcription-polymerase chain reaction (RT-PCR) using tumor markers, such as carcinoembryonic antigen, cytokeratin, or CD44, also is good tool for molecular detection of SN micrometastases. Rapid IHC or real-time RT-PCR is needed so that IHC or RT-PCR is applicable to SN biopsy for gastric cancer. Takeuchi et al. 1 introduced a novel realtime multiplex RT-PCR assay that is suitable for intraoperative detection of micrometastasis in SNs for the patients with clinical N0 gastric carcinoma. In the present study, 31% of patients with no metastasis over histological diagnosis were RT-PCR positive and the diagnostic accuracy is significantly higher in their multiplex RT-PCR assay than in intraoperative histopathology. Because the time taken for the assay is just 80 minutes, real-time multiplex RT-PCR is thought to be applicable for use in the intraoperative diagnosis of SN. Society of Surgical Oncology 2011

Prognostic Impact of Morphologic and Phenotypic Features of Childhood ALK-Positive Anaplastic Large-Cell Lymphoma: Results of the ALCL99 Study

The prognostic value of pathologic characteristics of childhood ALK-positive anaplastic large-cell lymphomas (ALCL), such as histologic subtypes, immunophenotype, and presence of the t(2;5) translocation or its variants, was assessed. All 375 patients with systemic ALK-positive ALCL included in an international trial launched by the European Intergroup for Childhood Non-Hodgkin's Lymphoma were reviewed by an international panel of pathologists based on conventional hematoxylin and eosin-stained and immunostained sections and classified according to the 2001 WHO classification. A small-cell (SC) or lymphohistiocytic (LH) component was observed in 114 (32%) of 361 patients, whereas ALCL of common type was diagnosed in 235 (65%) of 361 patients. Regarding the histologic subtyping of patients within the two categories of ALCL (with v without SC/LH component), the concordance between the national and international reviews was quite good, with a κ index equal to 0.67 (95% CI, 0.57 to 0.75). The presence of an SC/LH component was significantly associated with a high risk of failure (hazard ratio [HR], 2.0; 95% CI, 1.3 to 3.0; P = .002) in the multivariate analysis controlling for clinical characteristics, as well as the perivascular pattern (HR, 1.7; 95% CI, 1.1 to 2.7; P = .01), whereas CD3 positivity was significantly associated with a high risk of failure only in univariate analysis. Our study, which to our knowledge includes the largest series of childhood systemic ALK-positive ALCL so far, demonstrates the adverse prognostic value of SC and/or LH morphologic features. Combining these histologic characteristics with other biologic or clinical factors might have a high potential for future risk stratification and treatment.

Recent advancement of observing living cells in the esophagus using CM double staining: Endocytoscopic atypia classification

Magnification endoscopy enables in vivo evaluation of gastrointestinal mucosa. Furthermore, endocytoscopy (ECS) with ultra-high magnification enables in vivo observation of cellular atypia during routine endoscopic examination. The purpose of this study is to clarify the efficacy of ECS and endocytoscopic atypia (ECA) classification in various types of benign and malignant pathology in the esophagus. Consecutive 110 patients, who underwent ECS in our institution from March 2003 to December 2009, were included in this study. One hundred and forty-six esophageal lesions were classified according to ECA classification, and these endocytoscopic images were compared with histological images. We categorized endocytoscopic images into five categories according to size and uniformity of nuclei, number of cells and regularity of cellular arrangement. Eighty-one out of 89 ECA-1 to ECA-3 lesions (91.0%) corresponded to Vienna categories 1 to 3. Seventy-one out of 84 ECA-4 or ECA-5 lesions (91.2%) corresponded to Vienna category 4 or 5. Overall accuracy of ECS was 91.3%, providing images similar to conventional hematoxylin and eosin staining. In addition, with ECS, we can take an 'optical biopsy' even in patients with cardiovascular disease without interrupting anticoagulant therapy. A newly designed single charge-coupled device endocytoscope allows observation of target tissue noninvasibly from regular magnification to ultra-high magnification. The development of ECS has opened the door to in vivo cellular imaging, enabling endoscopic diagnosis of tissue cytological atypia during routine endoscopic examination.

Distinctive eosinophilic cytoplasmic inclusion bodies in melanocytic nevi: an immunohistochemical and ultrastructural study

We sought to further determine the histochemical, immunohistochemical and ultrastructural properties of eosinophilic cytoplasmic inclusion bodies in melanocytic nevi. Skin specimens from four patients with a known diagnosis of conventional melanocytic nevus (3) or Spitz nevus (1) and containing intracytoplasmic eosinophilic inclusion bodies were selected. In addition, melanomas (25), Spitz nevi (10) and blue nevi (4) were examined to determine the frequency of the inclusions. Inclusions tended to be located in multinucleated melanocytes with abundant vacuolated cytoplasm. In conventional (hematoxylin and eosin-stained) sections, the degree of density and eosinophilia of intracytoplasmic inclusions varied with size. Periodic acid-Schiff, Fontana and Congo red stains showed no reactivity. All bodies were immunoreactive for ubiquitin but negative for tyrosinase, keratin and vimentin. Ultrastructurally, inclusion bodies were non-membrane bound, ranged from 4 to 7 µm, and were comprised of radiating filamentous structures with or without an electron-dense core. Electron probe x-ray microanalysis revealed no significant peaks. None of additional melanomas, Spitz nevi and blue nevi that were evaluated showed similar inclusions. The inclusion bodies described herein bear no resemblance to other cytoplasmic inclusion bodies previously described in melanocytic lesions. There is no discernible relationship to melanosomes by ultrastructural analysis. We postulate a relationship with dysfunction of ubiquitin-mediated protein degradation occurring in melanocytes.

Dietary effect of Pleurotus eryngii on biochemical function and histology in hypercholesterolemic rats

This work was conducted to investigate diet supplement of king oyster mushroom fruiting bodies on biochemical and histological changes in hypercholesterolemic rats. Six-week old female Sprague–Dawley albino rats were divided into three groups of 10 rats each. The feeding of 5% powder of the fruiting bodies of Pleurotus eryngii to hypercholesterolemic rats reduced their plasma total cholesterol, triglyceride, low-density lipoprotein, total lipid, phospholipids, and LDL/HDL ratio by 24.05%, 46.33%, 62.50%, 24.63%, 19.22%, and 57.14%, respectively. Mushroom also significantly reduced body weight in hypercholesterolemic rats. However, it had no adverse effects on plasma albumin, total bilirubin, direct bilirubin, creatinine, blood urea nitrogen, uric acid, glucose, total protein, calcium, sodium, potassium, chloride, inorganic phosphate, magnesium, and enzyme profiles. Feeding mushroom increased total lipid and cholesterol excretion in feces. The plasma lipoprotein fraction, separated by agarose gel electrophoresis, indicated that P. eryngii significantly reduced plasma β and pre-β-lipoprotein, while increased α-lipoprotein. A histological study of hepatic cells by conventional hematoxylin–eosin and oil red O staining showed normal findings for mushroom-fed hypercholesterolemic rat. The present study suggests that 5% P. eryngii diet supplement provided health benefits by acting on the atherogenic lipid profile in hypercholesterolaemic rats. Therefore, king oyster mushroom could be recommended as a natural cholesterol lowering substance within the human diet.

Decreased incidence of isolated tumor cells in lymph nodes after laparoscopic resection for colorectal cancer

BackgroundLaparoscopic surgery has potential for less tumor cell spread because of the no-touch technique. We assessed the effect of the surgical approach (open versus no-touch laparoscopic) on the presence of tumor cells in sentinel lymph nodes (SN) of patients with stage I and II colorectal cancer.MethodsA single-center consecutive prospective series of patients operated on for colorectal cancer was analyzed. After conventional hematoxylin and eosin (H&E) staining, 107 patients without lymphatic metastases were included; 59 patients had open surgery, and 48 patients underwent laparoscopic resection. Patients in the laparoscopic group underwent a no-touch medial to lateral approach, whereas the conventional lateral to medial approach was applied in open surgery. A SN procedure was performed in all patients. The SNs were immunohistochemically analyzed for presence of occult tumor cells (OTC). According to the American Joint Committee on Cancer (AJCC) these tumor cells were divided into micrometastases (0.2–2 mm) or isolated tumor cells (ITC, < 0.2 mm).ResultsIn ten patients micrometastases were found, equally distributed between the two groups. However, ITC were more often found after open surgery (18 versus 5 patients, p = 0.03). Presence of OTC was related to depth of tumor invasion and tumor diameter > 3.5 cm. Logistic regression analysis identified lymphovascular invasion as a predictor for micrometastases [odds ratio (OR) 18.4], whereas open resection was predictive for presence of ITC (OR 3.3).ConclusionsNo-touch medial to lateral laparoscopic surgery results in less isolated tumor cells in lymph nodes compared with open lateral to medial surgery in patients with stage I and II colorectal cancer.

The Feasibility of anEx-vivoSentinel Lymph Mapping Using Preoperative Radioisotope Injection in Cases of Extraperitoneal Rectal Cancer

PurposeThe purpose of this research was to evaluate the feasibility of sentinel lymph node (SLN) mapping involving transanal injection with an ex-vivo mapping in patients with rectal cancer.MethodsBetween April 2007 and December 2009, 20 consecutive patients with T1-3, N0-1 clinical stage rectal cancer preoperatively underwent a SLN procedure using submucosal 99mTc-phytate injection. All the patients underwent a total mesorectal excision. After the standard surgical resection, all specimens were identified on lymphoscintigraphy, and bench work was done to pick up the sentinel node basin. All the lymph nodes (non-SLNs and SLNs) were examined using conventional hematoxylin and eosin staining and immunohistochemistry with anti-cytokeratin antibodies.ResultsSLNs were identified from 19 of 20 patients with rectal cancer. The total number of sentinel nodes retrieved from the surgical specimens was 29, and the mean number per patient was 1.6 (range, 0 to 4). In three patients, the SLN was the only positive lymph node. There was one false-negative case with a sensitivity of 88.8% and two upstaged cases (20.0%). The SLN samples from rectal cancer are mainly localized in the pararectal region, but aberrant nodes receive direct drainage from the rectal cancer. On planar lymphoscintigraphy, 15.7% of all patients had aberrant lymphatic drainage to the sigmoid mesenteric or sigmoid lymph node station.ConclusionIn conclusion, the intraoperative transanal injection for ex-vivo SLN navigation is a safe, feasible surgical modality in patients with rectal cancer. Large studies are warranted to determine the clinical significance of the SLN concept and micrometastasis in rectal cancer.

A study to evaluate the efficacy of xylene-free hematoxylin and eosin staining procedure as compared to the conventional hematoxylin and eosin staining: An experimental study

Context:Use of diluted dish washing solution (DWS) has been experimented successfully as a substitute for xylene to deparaffinize tissue sections during hematoxylin and eosin (H and E) staining.Aims:(1) Test the hypothesis that xylene- and methanol-free sections (XMF) deparaffinized with diluted DWS are better than or at par with conventional H and E sections. (2) To compare the efficacy of xylene-free sections with the conventional H and E sections.Settings and Design:Single blinded experimental study.Materials and Methods:Sixty paraffin blocks were considered. One section was stained with conventional H and E method (Group A) and the other with XMF H and E (Group B). Slides were scored for parameters; nuclear staining, cytoplasmic staining (adequate = score1, inadequate = score0), uniformity, clarity, crispness (present = score1, absent = score0). Score >/= 2 was inadequate for diagnosis and 3-5 was adequate for diagnosis.Statistical analysis used:Z test.Results:Adequate nuclear staining, 96.66% sections in group A and 98.33% in Group B (Z = 0.59, P>0.05); adequate cytoplasmic staining, 93.33% in group A and 83.33% in Group B (Z = 1.97, P<0.05); uniform staining, 70% in group A, 50% in group B (Z = 1.94, P<0.05), clarity present in 85% of group A, 88.33% of group B sections (Z = 0.27, P>0.05), crisp staining in 76.66% in group A and 83.33% in Group B (Z = 1.98, P<0.05), 88.33% Group A sections stained adequately for diagnosis as compared with 90% in Group B (Z = 0.17, P>0.05).Conclusion:Xylene- and methanol-free H and E staining is a better alternative to the conventional H and E staining procedure.

Paracrine Loops of Keratinocyte Stimulation in Cholesteatoma Tissue

In relation to the keratinocyte growth factor (KGF) expression in cholesteatoma tissue, the inflammatory infiltrate present in this disease could play a relevant role in the paracrine stimulation of keratinocytes. Cholesteatoma is a temporal bone pathologic disease characterized by active proliferation of epithelial cells, with progressive growth and involvement of the neighboring middle/inner ear structures. The pathogenetic mechanism underlying the hyperproliferation of keratinocytes is not yet fully clarified. It has been suggested that keratinocyte proliferation and migration could be mediated by several autocrine and paracrine growth factors and their receptors. A previous study has reported that the expression of KGF receptor is increased in more differentiated areas of the cholesteatoma tissue, whereas the expression of the epidermal growth factor receptor is associated with proliferative and migratory areas of the lesion. Fresh cholesteatoma samples were collected during surgical procedures. Serial cryosections were examined by conventional hematoxylin and eosin or by immunofluorescence with antipancytokeratin antibodies. The ultrastructural features were analyzed by transmission electron microscopy. The expression of KGF, K1, and filaggrin in the samples was evaluated by quantitative reverse transcription-polymerase chain reaction and correlated with the dermal degree of inflammatory infiltrate and the presence of stromal cells. Increased expression of KGF was associated with strong levels of the inflammatory infiltrate. These results would suggest that KGF up-modulation is a consequence of fibroblast stimulation by inflammatory cells and that this paracrine loop could be responsible not only for the hyperproliferation of keratinocytes in cholesteatoma tissue but also for the deregulation of epidermal differentiation.