Immunohistochemical analysis of MIB-1 proliferative activity in human endometrial cancer. Correlation with clinicopathological parameters, patient outcome, retinoblastoma immunoreactivity and K-ras codon 12 point mutations

To test the prognostic utility of MIB-1 in human endometrial neoplasias, the proliferative activities of fifty-two endometrial carcinomas obtained from Polish women were assessed. We also investigated the relationship between the MIB-1 Proliferative Index and the well-known clinicopathological features of cancer (clinical stage, histological type, histological grade, depth of myometrial invasion), patient's age, overall survival, retinoblastoma immunostaining and K-ras codon 12 point mutations. The mean MIB-1 Proliferation Index was 43.8%, with a median of 36.0%. Due to the great intratumour heterogeneity of the immunoreaction, the Index ranged from 0% to 98%. A significant relationship was noted between MIB-1 expression and histological grading (p=0.0004) and myometrial invasion of cancer (p=0.01). Multivariate Cox regression demonstrated that the clinical stage was the only independent prognostic factor during follow-up (p=0.025). There was a tendency towards a poorer outcome for women with a Proliferative Index of >31% than for patients whose Index was 31%; the difference, however, did not reach significance (p=0.25; log-rank test). Interestingly, uterine cancers lacking retinoblastoma protein expression had a mean MIB-1 Proliferation Index that was nearly twice as high as in those neoplasias that stained positively for retinoblastoma (70.33% and 42.14%, respectively; p=0.09; Mann-Whitney-U test). There were no significant differences between K-ras codon 12 point mutation-positive and -negative endometrial carcinomas regarding the proliferative activity of the cancer (mean Indexes 47.6% and 43.8%, respectively; p=0.66, Mann-Whitney-U test). Our data support the view that MIB-1 proliferative activity was significantly increased with a decrease of the histological grading and with the myometrial invasion of human endometrial cancer.     

Nucleolar spindle-associated protein 1 promotes tumorigenesis and predicts poor prognosis in human esophageal squamous cell carcinoma

The microtubule binding protein, nucleolar spindle-associated protein 1 (NUSAP1), has a crucial function in mitosis and its expression is closely associated with carcinogenesis. Herein, we aimed to determine the function of NUSAP1 in the development of human esophageal squamous cell carcinoma (ESCC), and the association of NUSAP1 expression with ESCC. Immunohistochemical staining of ESCC tissue sections indicated that NUSAP1 was expressed to a higher degree in tumor tissues than in adjacent nontumor tissues. NUSAP1 levels were relevant closely to histological differentiation (P = 0.049). Overall survival was longer in patients with lower NUSAP1 levels ( P < 0.001). NUSAP1 expression ( P = 0.002), histological differentiation ( P < 0.001), tumor depth ( P = 0.045), lymph node metastases ( P < 0.001), and tumor-node-metastasis staging ( P = 0.008) were greatly associated with overall survival using univariate analysis. Multivariate analysis suggested that histological differentiation ( P = 0.014) and NUSAP1 expression ( P = 0.026) could be independent prognostic markers for ESCC. Additionally, the biological behavior of ESCC cells was investigated in vitro and in vivo. Suppression of NUSAP1 inhibited cellular proliferation and invasion, and induced cell cycle arrest and apoptosis in vitro. More importantly, knockdown of NUSAP1 led to inhibition of tumor formation in nude mice. These findings indicated that NUSAP1 is a potential prognostic biomarker in ESCC, and is an ESCC oncogene. Thus, NUSAP1 could represent a therapeutic target for ESCC.     

A suggestion of a new approach to the theory of some biological periodicities

Previous studies of L. Danziger and G. Elmergreen (Bull. Math. Biophysics,16, 15–21, 1954;18, 1–13, 1956) of possible biochemical periodicities in organisms assumed non-linear biochemical interaction between different metabolites, because linear systems do not lead to undamped ocsillations. They treated homogeneous systems. Later N. Rashevsky generalized their results to a more realistic case where the non-homogeneity due to the histological structure is considered. (Some Medical Aspects of Mathematical Biology, Springfield, Illinois: Charles C. Thomas, Publisher, 1964;Bull. Math. Biophysics,29, 389–393, 1967.) As long as the histological structure remains constant, the existence of sustained periodicities requires the assumption of non-linearity of biochemical interactions. If, however, the secretions of an endocrine gland affect the histological structure of the target organ, notably as in the menstrual cycle, and if there is a feed-back, the equations become non-linear and may admit sustained periodic solutions even if the purely biochemical interactions are linear.     

A simple method for the production of bacterial controls for immunohistochemistry and fluorescent in situ hybridization

Immunohistochemical and fluorescent in situ hybridization (FISH) assays are useful diagnostic methods for the identification of bacteria on formalin fixed paraffin embedded histological sections. To validate an anti-bacterial antibody or an oligonucleotide probe and to ensure fidelity during subsequent analyses, suitable positive and negative controls are necessary. Suspensions of fixed bacteria are often used, but ideally, these controls should be fixed, embedded and processed in the same way of tissue samples under analysis. Herein, we describe a simple method for the production of bacterial histological control samples: the sandwich. The sandwich is composed of two external layers of equine lung parenchyma and a central layer of the target bacterium. We prepared sandwiches containing Escherichia coli, Campylobacter jejuni, and Arcanobacterium pyogenes and tested them with appropriate antibodies and Eub338 FISH probe. The sandwich is an effective and simple method to prepare bacterial histological controls fixed and processed in the same way as the diagnostic tissues.     

Effect of aberrant promoter methylation of FHIT and RASSF1A genes on susceptibility to cervical cancer in a North Indian population

As current evidence suggests the involvement of epigenetic modification of tumour suppressor genes in human cancer, we investigated the aberrant promoter methylation of FHIT and RASSF1A genes in human papillomavirus (HPV)-mediated cervical cancer in Indian women. We analysed 60 cervical cancer tissue biopsies of different clinical stage and histological grading and 23 healthy control samples with normal cervical cytology. Methylation-specific polymerase chain reaction (MSP) was performed to analyse the methylation status of FHIT and RASSF1A genes and confirmed by sequencing. Both patients and controls were screened for HPV infection and 98% of the HPV-infected cases showed positivity for HPV type 16. Aberrant promoter methylation of the FHIT gene was found in 28.3% (17/60) of cases and of the RASSF1A gene in 35.0% (21/60) of cases; promoter methylation of both the genes was found in 13.3% (8/60) of cervical cancer cases. Methylation was significantly (p<0.01) associated with the cervical cancer cases compared with controls. None of the 23 controls was found to be methylated in either of these genes. This is the first study indicating a correlation between the promoter methylation of FHIT and RASSF1A genes and the clinical stage and histological grading of cervical carcinoma in Indian women. Future studies are underway to examine the practical implications of these findings for use as a biomarker.     

Factors influencing extract of Hibiscus sabdariffa staining of rat testes

Abstract

Some plant extracts can be used in biology and medicine to reveal or identify cellular components and tissues. We investigated the effects of time and concentration on staining of histological sections of rat testes by an acidified extract of Hibiscus sabdariffa. An ethanolic extract of H. sabdariffa was diluted using 1% acetic acid in 70% ethanol to stain histological sections of testes at concentrations of 0.2, 0.1 and 0.05 g/ml for 5, 10, 15, 30, 45 and 60 min. The sections of testes were stained deep red. The staining efficiency of H. sabdariffa was greater at a high concentration and required less time to achieve optimal staining. H. sabdariffa is a strongly basic dye that can be used for various diagnostic purposes. Staining time and concentration must be considered to achieve optimal results.     

Characterisation of the muscular organs of paramphistomes from African freshwater fishes

The histological characteristics of the pharynx, terminal genitalium and acetabulum of four paramphistomes from African freshwater fishes are described. New pharyngeal types are described for Basidiodiscus ectorchis, Sandonia sudanensis, Panamphistomum benoiti and Brevicaecum niloticum, new terminal genitalium types for P. benoiti and B. niloticum, and a new acetabular type for P. benoiti. Amendments are made to the basidiodiscus, sandonia and brevicaecum acetabular types of Sey (1987). P. benoiti is reported from Synodontis nigrita in Nigeria, a new host and geographical record.     

Interactions Among Gastric Somatostatin, Interleukin-8 and Mucosal Inflammation in Helicobacter pylori-Positive Peptic Ulcer Patients

Background. To investigate whether Helicobacter pylori infection, but not drugs, affects gastric somatostatin, interleukin‐8 (IL‐8), histological inflammation through eradication therapy, and interactions among these parameters. Methods. Twenty‐eight H. pylori‐positive patients (21 males; mean age 47.0 years) with either gastric ulcer (GU: n = 11) or duodenal ulcer (n = 17) diagnosed endoscopically were treated with dual therapy. Eradication was defined as negative microbiologic tests and ¹³C‐urea breath test. Levels of antral and gastric juice somatostatin and mucosal IL‐8 were measured by radioimmunoassay and enzyme‐linked immunosorbent assay, respectively. Histology was assessed by the Sydney system. Results. H. pylori was eradicated in 15 patients (10 males, 6 GU) out of 28 (54%). The patients’ backgrounds did not affect the eradication of H. pylori. Successes in eradication significantly increased antral and juice somatostatin contents, and dramatically decreased IL‐8 levels and histological gastritis. In contrast, persistent H. pylori infection did not affect somatostatin and histological gastritis. An inverse correlation was present between changes in somatostatin levels and histological activity. No relationship was observed in changed values between antral somatostatin and IL‐8. Conclusions. These results indicate that eradication of H. pylori, but not the drugs used, induced an increase in somatostatin levels in the antrum and gastric juice, suggesting a close relationship between H. pylori and gastric somatostatin regulation. A close correlation between an increase in gastric somatostatin levels and the normalization of histological activity was present, suggesting that certain peptide‐immune interactions in the gastric mucosa exist in H. pylori infection.     

The scotopic electroretinogram of the sugar glider related to histological features of its retina

The flash electroretinogram (ERG) was used to characterize the scotopic retinal function in a marsupial. Key parameter values of the a- and b-waves of adult male sugar gliders, Petaurus breviceps breviceps, elicited with ganzfeld flashes were determined under dark- and light-adapted conditions. Using standard histological methods, the thicknesses of the major layers of the retina were assessed to provide insight into the nature of the ERG responses. The ERG and histological results were compared to corresponding data for placental C57Bl/6 mice to establish whether the functional retinal specialization that underlies scotopic visual function in a marsupial parallels that of a placental mouse. The sensitivity of the a-wave assessed with the Lamb and Pugh (Invest Ophthalmol Vis Sci 47:5138–5152, 2006) “model” and that of the b-wave assessed with standard methods were lower in the sugar glider compared to the mouse. The thickness of the sugar glider retina was two-third of that of the mouse. The high-intensity flash ERG of the sugar glider substantially differed in shape from that of the mouse reflecting perhaps structural and functional differences between the two species at the level of the inner retina.     

<Emphasis Type="Italic">Batrachochytrium</Emphasis><Emphasis Type="Italic">dendrobatidis</Emphasis> and Chytridiomycosis in Anuran Amphibians of Colombia

In order to investigate the possible presence of the chytrid fungus Batrachochytrium dendrobatidis (Chytridiomycota: Chytridiales) in frogs (Amphibia: Anura) of Colombia, we made a retrospective examination of formalin-fixed specimens preserved in natural history collections. Using the staining technique of hematoxylin and eosin to identify B. dendrobatidis in histological slices, we found evidence of the fungus in 3 of the 53 frog species examined from a total of 672 specimens collected in 17 departments within Colombia between 1968 and 2006. The infected specimens were found dead or dying in recent years in high elevation sites, suggesting that chytridiomycosis (the disease caused by the fungus) may represent a significant threat to Colombian amphibians. We conclude that a more extensive search for B. dendrobatidis in museum specimens and wild-caught frogs should be undertaken as soon as possible, using both histological and molecular genetic techniques, in order to further characterize the geographic and taxonomic extent of infections of B. dendrobatidis.     

A comprehensive characterisation of the fibre composition and properties of a limb (<Emphasis Type="Italic">Flexor digitorum superficialis,membri thoraci</Emphasis>) and a trunk (<Emphasis Type="Italic">Psoas major</Emphasis>) muscle in cattle

Background  

The fibre type attributes and the relationships among their properties play an important role in the differences in muscle capabilities and features. Comprehensive characterisation of the skeletal muscles should study the degree of association between them and their involvement in muscle functionality. The purposes of the present study were to characterise the fibre type composition of a trunk (Psoas major, PM) and a limb (Flexor digitorum, membri thoraci, FD) muscle in the bovine species and to study the degree of coordination among contractile, metabolic and histological properties of fibre types. Immunohistochemical, histochemical and histological techniques were used.     

Phylogenetic signal in bone histology of amniotes revisited

Legendre, L, Le Roy, N, Martinez‐Maza, C, Montes, L, Laurin, M & Cubo, J. (2012). Phylogenetic signal in bone histology of amniotes revisited. —Zoologica Scripta, 42, 44–53. There is currently a debate about the presence of a phylogenetic signal in bone histological data, but very few rigorous tests have fuelled the discussions on this topic. Here, we performed new analyses using a larger set of seven histological traits and including 25 taxa (nine extinct and 16 extant taxa), using three methods: the phylogenetic eigenvector regression, the tree length distribution and the regressions on distance matrices. Our results clearly show that the phylogenetic signal in our sample of bone histological characters is strong, even after correcting for multiple testing. Most characters exhibit a significant phylogenetic signal according to at least one of our three tests, with the phylogeny often explaining 20–60% of the variation in the histological characters. Thus, we conclude that the phylogenetic comparative method should be systematically used in interspecific analyses of bone histodiversity to avoid problems of non‐independence among observations.     

Journal of Cellular Physiology: Volume 228, Number 7, July 2013

Cover: Photography Wwox allelated and control mice on a background of a histological section of femoral bone for a Wwox deficient animals. Please see article by Abdeen et al., pages 1377–1382. Cover designed by Lucía Sagredo Sánchez.     

K-ras Gene Mutation Related to Histological Atypias in Human Colorectal Adenomas

To investigate the relationship of oncogene analysis to morphology, we analyzed K-ras gene mutations by dot-blot hybridization with and without consideration of histological atypias in individual colorectal adenomas. Each of 54 colon polyps were divided into two parts after fixation. One part was used as a mass to assess point mutations; the remaining portion of each polyp was paraffin-embedded, stained with hematoxylin and eosin, and examined for point mutations related to histological atypias. In the first part of our study, K-ras gene mutations at codon 12 were detected in 13 cases (24%). In the second part of our study, 12 cases had distinctly different histological atypias. From each of these 12 cases, two areas, one with higher or one with lower grade atypia in the same polyp were excised to analyze for K-ras gene mutation. Two of these 12 cases (17%) had the mutation in different areas of the same tumor. These two cases contained the mutation only in the areas with higher grade atypia, and only one case added information regarding ras mutation upon microdissection when compared to the entire biopsy. These results suggest that oligonucleotide hybridization can identify the majority of cases containing ras mutations despite regional morphologic variation. Individual cases, however, may contain clonal subpopulations within adenomas with different ras sequences from other regions within the same adenoma.     

Histological Localization of Phosphorothioate Oligodeoxynucleotides in Normal Rodent Tissue

Abstract

The distribution of phosphorothioate oligodeoxynucleotides (P=S ODN) in rodent tissues was studied in vivo using three histological methods: direct fluorescence microscopy; immunohistochemistry; and autoradiography. All three methods gave essentially the same pattern of oligonucleotide localization in the tissues studied, and the histological results correlate well with those from radiochemical and biochemical studies of P=S ODN distribution.     

Expression of a chimeric GUS gene construct as a tool to study nodule morphogenesis in chicory leaves

Leaves from micropropagated chicory plantlets were cultivated in vitro to regenerate organogenic nodules. The nodules were transformed with a 2.3 kb fragment of an apple calmodulin promoter region fused to the coding sequence of uidA reporter gene. Histochemical detection of β-glucuronidase expression in transgenic regenerants showed that vessel-associated cells were strongly stained. Previous histological investigations have shown that the differentiation of vascular elements is essential to nodule and bud-derived-nodule development. Therefore, β-glucuronidase activity was tested in a single transgenic chicory line during nodule morphogenesis and bud regeneration. The vascular connections between leaves and nodules, then between nodules and buds were stained blue, indicating an alternative system for determining the histological origin of nodules and adventitious buds from chicory leaf explants. This revised version was published online in June 2006 with corrections to the Cover Date.     

Germ-line dependence of the maroon-like maternal effect in Drosophila

We have used pole cell transplantations to construct germ-line mosaics for maroon-like (mal), a maternal effect mutation in Drosophila. Such mosaics allow one to determine the cell type in which a gene is active. We find that the maroon-like maternal effect is (1) autonomous to the germ line and (2) dose sensitive in germ-line mosaics. Aldehyde oxidase activity is used as a histological probe to investigate the tissue and temporal distribution of mal⁺ activity in the developing ovary. The adult ovary shows mal⁺ activity in the germ line at all discernible stages of oogenesis but no activity is observed in the mesodermally derived follicle cells. Differential mal⁺ activity is observed even in the ovary of the third-instar larvae.     

Histogenesis of the cuticle of the adult flies,Sarcophaga bullata and S. argyrostoma

Sequential patterns of cuticle deposition and “melanization” in the imaginal cuticle of Sarcophaga argyrostoma in parts of the body darkening before or after emergence are examined on a histological basis. The patterns in the cuticles examined range from a simple absence of “melanization” to a complex of histological changes involving “melanization” and deposition. Ultrastructural changes in the post-emergent cuticle of Sarcophaga bullata during the hardening and darkening process and cuticle deposition are described.     

Development and Characterization of Mucoadhesive <Emphasis Type="Italic">In Situ</Emphasis> Nasal Gel of Midazolam Prepared with <Emphasis Type="Italic">Ficus carica</Emphasis> Mucilage

The objective of the present study was to prepare mucoadhesive in situ nasal gels with mucilage isolated from fig fruits (Ficus carica, family: Moraceae) containing midazolam hydrochloride. Nasal gels of midazolam were prepared using three different concentrations (0.5%, 1.0% and 1.5% w/v) of F. carica mucilage (FCM) and synthetic polymers (hydroxypropylmethyl cellulose and Carbopol 934). Evaluation of FCM showed that it was as safe as the synthetic polymers for nasal administration. In situ gels were prepared with mixture Pluronic F127 and mucoadhesive agents. Evaluation of the prepared gels was carried out, including determination of viscosity, texture profile analysis and mucoadhesive strength. In vitro drug permeation study was conducted with the gels prepared with and without permeation enhancer (0.5% w/v sodium taurocholate) using excised goat nasal mucosa. In vitro permeation profiles were evaluated, and histological study of nasal mucosae before and after permeation study was also conducted to determine histological change, if any. In vivo experiments conducted in rabbits further confirmed that in situ nasal gels provided better bioavailability of midazolam than the gels prepared from synthetic mucoadhesive polymers. It was observed that the nasal gel containing 0.5% FCM and 0.5% sodium taurocholate exhibited appropriate rheological, mechanical and mucoadhesive properties and showed better drug release profiles. Moreover, this formulation produced no damage to the nasal mucosa that was used for the permeation study, and absolute bioavailability was also higher compared to gels prepared from synthetic polymers.