A suite of microsatellite markers for genetic management of captive cracids (Aves, Galliformes).

Cracids are medium to large frugivorous birds that are endemic to the Neotropics. Because of deforestation and overhunting, many species are threatened. The conservation of several species has relied on captive breeding and reintroduction in the wild, but captive populations may be inbred. Microsatellite tools can permit the construction of genetic pedigrees to reduce inbreeding, but only a few loci are available for this group of birds. Here, we present 10 novel polymorphic microsatellite loci and the cross-amplification of these and of 10 additional loci available in the literature in a panel of 5 cracid species, including 3 species with high conservation concern. We provide the first polymorphic loci for the jacutinga, Aburria jacutinga (N = 8), and red-billed curassow, Crax blumenbachii (N = 9), and additional loci for bare-faced curassow, C. fasciolata (N = 8), Alagoas curassow, Pauxi mitu (N = 5), and razor-billed curassow, P. tuberosa (N = 5). The average number of alleles was 2.9 for A. jacutinga, 2.7 for C. blumenbachii, 3.5 for C. fasciolata, 2.6 for P. mitu, and 5.7 for P. tuberosa. The mean expected heterozygosities were 0.42, 0.40, 0.48, 0.37, and 0.59, respectively. The average probabilities that the set of loci would not exclude a pair of parents of an arbitrary offspring were 2.9% in A. jacutinga, 1% in C. blumenbachii, 0.5% in C. fasciolata, 0.4% in P. mitu, and 0.002% in P. tuberosa suggesting that these loci may be adequate for parentage analysis and to implement ex situ genetic management plans.

Observer variability in the Goseki grouping of gastric adenocarcinoma in resection and biopsy specimens.

AIMS: The Goseki grouping of gastric adenocarcinoma has been suggested as a possible prognostic factor. In those centres where it is used, it may be valuable to assess the Goseki grouping of a tumour on the initial diagnostic biopsy as well as on the resection specimen since it may in theory influence management. We examined the robustness of Goseki grouping of gastric adenocarcinoma in representative sections from resection and biopsy specimens in order to assess the consistency of agreement among a group of pathologists. METHODS: A single representative block from 100 gastric resection specimens was studied using a haematoxylin and eosin and mucin (alcian blue/periodic acid-Schiff) stain. These were circulated in batches to members of a group of 12 pathologists who each completed a simple proforma confirming the presence of carcinoma and assigning a Goseki group. In a second circulation the diagnostic biopsy specimen taken prior to resection was examined in the same way. This allowed comparison of the Goseki group of the biopsy and resection specimens. RESULTS: In both studies kappa statistics showed good agreement on tubular differentiation of the carcinoma, but only moderate agreement for the intracellular mucin production, resulting in moderate agreement for the final Goseki group. Correlation between the Goseki group assigned on the biopsy and resected specimens was seen in 62% of the cases. However, the reproducibility was low (kappa 0.375). CONCLUSIONS: The Goseki grouping of resected gastric adenocarcinoma is reproducible and can be used in prognostication. Goseki grouping of biopsy specimens is of limited value in predicting the Goseki group assigned to the resected carcinoma.

The Scottish doctor--learning outcomes for the medical undergraduate in Scotland: a foundation for competent and reflective practitioners.

This paper describes a set of learning outcomes that clearly define the abilities of medical graduates from any of the five Scottish medical schools. The outcomes are divided into 12 domains that fit into one of three essential elements for the competent and reflective medical practitioner.

Consistency of histopathological reporting of laryngeal dysplasia. The Scottish Pathology Consistency Group.

AIMS: Clinical management of premalignant and malignant lesions of the larynx is dependent on histopathological evaluation. The Scottish Pathology Consistency Group assessed interobserver variation in the evaluation of laryngeal dysplasia. METHODS AND RESULTS: One hundred laryngeal biopsies ranging from normal to invasive carcinoma were assessed. The overall Kappa result of 0.32 was disappointing. However, agreement on those categories which dictate significantly different management was more favourable. The Kappa figure for mild dysplasia versus severe dysplasia/CIS was 0.7, the Kappa figure for mild dysplasia versus severe dysplasia/CIS and invasive carcinoma was 0.77. The Kappa figure for mild and moderate dysplasia versus severe dysplasia/ CIS and invasive carcinoma was 0.57. An attempt to use a two grade system gave a Kappa figure of 0.52. CONCLUSIONS: Our group had a satisfactory agreement on the distinction of mild from severe dysplasia and on microinvasive carcinoma without any discussion as to histopathological criteria to be used. Clinical management--review endoscopy, repeat cord stripping, radiotherapy and laryngectomy--is in general dependent on histological assessment. Thus the agreement on categories which underpin clinical management is reassuring. However, assessment of moderate dysplasia remains problematic. An attempt to utilize a two grade system--low grade from high grade dysplasia/CIS--may have merit. The implications of the terminology used must be agreed among pathologists and clinicians working closely within clinicopathological cancer groups.

Development and integration of CAL: a case study in medicine.

CONTEXT: The Faculty of Medicine and Medical Sciences at the University of Aberdeen viewed the use of Computer Assisted Leaning (CAL) and other IT based learning resources as a possible way of coping with an increase in student numbers whilst maintaining or increasing the quality of medical teaching. OBJECTIVES: Our primary objective was to develop and integrate Computer Assisted Learning (CAL) applications into the undergraduate medical curriculum. SUBJECTS/MATERIALS: A wide spectrum of CAL applications were developed dealing with many topics in the curriculum. METHODS: We formulated a structured approach to CAL development by establishing a team of professionals (forming a CAL Unit), using existing expertise and by implementing a process to ensure that the CAL had a maximum impact upon the curriculum. The CAL included multimedia tutorials, learning guides, computer aided assessment (CAA) and Model Patients. RESULTS: There are now over 150 IT based learning resources in our curriculum and course evaluation has showed that these have been well received by students. CONCLUSIONS: We conclude that with the wise use of the many skills and facilities usually available within an institution and by promoting collaborative projects with others, the production of high quality CAL is possible within most institutions.

The expression of beta 1 integrins in human coronary artery.

The beta 1 integrin adhesion receptors mediate the binding of cells to extracellular matrices, facilitating their growth, migration, and capacity to deposit matrix proteins: important factors in arterial restenosis and atherosclerosis. The expression of integrins in human coronary artery is, however, unexplored. The aim of the current study was, therefore, to define the expression of beta 1 integrins by cultured human coronary artery vascular smooth muscle cells (hCAVSMC) and in normal human coronary artery; confirming whether or not this differs from the repertoire found in other species and human vessels. The expression of beta 1 integrins by hCAVSMC was assessed by immuno-precipitation and the alkaline phosphatase anti-alkaline phosphatase (APAAP) immunochemical technique. In addition, mRNA expression was defined by reverse transcription polymerase chain reaction (RT-PCR). Normal adult human coronary arteries (n = 4) were also stained by the APAAP method. In vitro hCAVSMC express alpha 2 beta 1 (a collagen and occasional laminin receptor) and alpha 5 beta 1 (a fibronectin receptor) with lesser expression of alpha 3 beta 1 (a multifunctional receptor). They do, however, possess mRNA for several other integrins. Cells within the media of human coronary artery wall express alpha 3 beta 1 and alpha 5 beta 1 but not alpha 2 beta 1: instead the alternative collagen/laminin receptor, alpha 1 beta 1, is expressed in vivo. This pattern of expression differs subtly from that described in rats through it closely parallels that found in other human arteries.

Upregulation and co-localization of connexin43 and cellular adhesion molecules in inflammatory renal disease.

Connexin43 (Cx43) is a major component of gap junctions. These are widely distributed in the human kidney and are thought to be involved in the inflammatory response and in the regulation of cell growth. Cellular adhesion molecules (CAMs) are also thought to be important in these processes, where they possibly facilitate gap junction formation. The aims of the current study were to define for the first time the expression of Cx43 in inflammatory glomerulonephritis and to compare the localization of this connexin with that of the intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin. Human renal biopsies and control sections of normal human kidney were stained using the alkaline phosphatase/anti-alkaline phosphatase immunohistochemical technique, demonstrating that Cx43 was strongly expressed on inflammatory cells, on damaged tubular cells, and on interstitial cells. This pattern of expression was paralleled closely by that of ICAM-1 and, to a lesser extent, by that of VCAM-1. Cx43 is therefore primarily implicated in tubulointerstitial inflammation.

Scleroderma, D-penicillamine treatment, and progressive renal failure associated with positive antimyeloperoxidase antineutrophil cytoplasmic antibodies.

Progressive renal failure in patients with scleroderma is a sinister development that is usually attributed to impaired renal blood flow. In some exceptional cases, the underlying pathology is a crescentic glomerulonephritis, which has been associated with positive antineutrophil cytoplasmic antibodies, and in particular antimyeloperoxidase antibodies. The prognosis in such cases has been very poor. We report such a patient whose renal function has improved and stabilized on immunosuppressive therapy.

Importance of the tubulointerstitium in human glomerulonephritis. II. Distribution of integrin chains beta 1, alpha 1 to 6 and alpha V.

Accumulation of extracellular matrix is important in the progression of glomerulonephritis. Since adherent cell types utilize integrins to bind and organize extracellular matrix proteins, we have assessed expression of the beta 1 integrins in sequential sections from 85 human renal biopsies and 4 normal kidneys by immunohistochemical staining. Our results demonstrate strong correlations between expression of the alpha 5 chain within the interstitium, the alpha V chain on proximal and distal tubular epithelium and the presence of chronic histological damage. Moreover, staining for interstitial alpha 5 and proximal and distal tubular alpha V were also strongly associated with expression of certain adhesion molecules (ICAM-1, VCAM-1, E-selectin and L-selectin) and the presence of macrophages within the interstitium, which have been linked, in an earlier study, with the degree of chronic histological damage and disease progression. However, in contrast to our earlier study of adhesion molecules, there were also associations between expression of integrin chains within the glomerulus and tubulointerstitium. For example, there were strong positive associations between staining for alpha 5 on glomerular endothelium and its expression on extraglomerular vascular endothelium and between both mesangial alpha 1 and podocyte alpha 3 and tubular staining for the common beta 1 subunit. While the functional significance of these associations is obscure, they suggest some kind of communication between cells in different sites in the kidney. There were also positive associations between staining for different integrins within the glomerulus, notably mesangial cell staining for alpha 2, glomerular endothelial cells staining for alpha 5 and glomerular epithelial cell alpha 3. These results suggest that there is a coordinated upregulation of integrin expression both within the tubulointerstitium and the glomerulus and that at least some of these integrins (interstitial alpha 5 and distal tubular alpha V) are associated with the expression of other adhesion molecules, macrophage infiltration and the presence of markers of disease progression (interstitial fibrosis and tubular atrophy).

Apoptosis in atypical fibroxanthoma and pleomorphic malignant fibrous histiocytoma.

Explanations for the disparate behavior of atypical fibroxanthoma (AFX) as compared with pleomorphic malignant fibrous histiocytoma (MFH) have included the proposition that the former is a pseudosarcoma. Nonetheless, these tumors are now widely regarded as the same process, but with AFX behaving benignly by virtue of its superficial location. However, a neoplasm's metastastatic potential has been proposed to be related to apoptosis. Therefore, the aim of the present study was to examine apoptotic counts, in conjunction with two important regulators of apoptosis: p53 and bcl-2, to determine if a distinction exists that may account for the different outcomes of these lesions. There was no significant statistical difference between eight AFX and nine pleomorphic MFH in terms of apoptotic behavior, proliferative indexes, p53 protein expression, or presence of bcl-2 product. Therefore, our results further support the contention that AFX should be regarded as a form of pleomorphic MFH, which demonstrates low malignant potential by virtue of its location in readily accessible sites.

Observer variability in the histopathological reporting of needle biopsy specimens of the prostate.

The Scottish Pathology Consistency Group has in previous studies examined the consistency of histopathological reporting of biopsies from the cervix, bladder, bronchus, and rectum. In the current study, consisting of 100 needle biopsy specimens of the prostate, a single hematoxylin-eosin (H&E) slide from each case was circulated in batches of 10 to the 12 pathologists, who filled in a simple proforma. This had two sections: a diagnostic category (benign; suspicious or malignant) along with a standard Gleason score for those regarded as malignant. The majority diagnosis of the 100 cases was benign, 53; suspicious, 1; and malignant, 46. The Kappa value for benign cases versus others was 0.86 and for malignant cases versus others was 0.91. Analysis of the data on Gleason scores showed a value of 0.54 when cases were divided into two categories (2 to 6 v 7 to 10) and 0.41 when three categories were used (2 to 4; 5 to 6; 7 to 10). Although not initially part of the design of the study, the majority diagnosis was compared with the original reported diagnosis. In a small subset, examination of further levels, basal cell antibody staining, along with further clinical information, was obtained. With this added information, it appears that there were probably 52 benign and 48 malignant cases. Of the 48 malignant cases, the group majority diagnosis was malignant, 46; suspicious, 1; and benign, 1. The original reported diagnosis was 56 benign, 1 suspicious, and 43 malignant. The group therefore appeared to perform better than the original reporting pathologists. When compared with the results of our previous studies, this study has shown that the diagnosis of carcinoma of the prostate on a needle biopsy is robust.

Expression of beta 1 integrins in IgA nephropathy.

AIM: To compare the expression of beta 1 integrins in renal biopsies from patients with IgA nephropathy with that found in normal human kidney. METHODS: Thirty renal biopsies from patients with IgA disease plus six control specimens were stained with monoclonal antibodies directed against the alpha 1, alpha 2, alpha 3, alpha 4, alpha 5, alpha 6, alpha v, and beta 1 integrin chains using the alkaline phosphatase anti-alkaline phosphatase (APAAP) technique. The intensity of integrin expression was graded semiquantitatively by a pathologist unaware of the antibody used. RESULTS: Glomerular crescents stained strongly for alpha 3, alpha v, and beta 1, but integrin expression was greatly reduced or absent in fibrotic glomeruli. There were no alterations in the intensity of mesangial cell staining for any of the integrins tested. There was accentuated staining for the alpha 2, alpha 5, alpha v, and beta 1 chains in areas of interstitial scarring plus alpha 2, alpha 3, alpha v, and beta 1 on damaged tubules. Inflammatory cells expressed alpha 4, alpha 5, and beta 1. CONCLUSIONS: In IgA nephropathy the interstitium is the main site of altered beta 1 integrin expression. Glomerular crescents also express several beta 1 integrins, but we found no differences in the intensity of integrin expression on mesangial cells. Altered beta 1 integrin expression may play a role in tubulointerstitial scarring in IgA disease. Thus modulation of integrin expression might attenuate this process.

The expression of connexin 43 in human kidney and cultured renal cells.

Gap junctions enable intercellular communication and play an important role in a variety of vital cellular functions including differentiation and the control of growth. These junctions are formed by a hexameric of proteins known as connexins. We investigated the distribution of the connexin 43 (Cx43) gap junction protein in renal cells and human kidney using the alkaline phosphatase anti-alkaline phosphatase immunohistochemical technique with a monoclonal antibody directed against the cytoplasmic domain of this antigen. Strong staining was demonstrated on the vascular endothelium, the smooth muscle of larger vessels and on glomerular epithelial cells. In addition, Cx43 was expressed on proximal tubular cells, glomerular endothelial cells and occasional cells infiltrating the interstitium. In areas of tubular atrophy there was increased staining for Cx43. Using reverse transcription-polymerase chain reaction we have also demonstrated that cultured human and rat mesangial cells and human proximal tubular cells express Cx43 messenger RNA. In summary, we have described for the first time the distribution of Cx43 in human kidney and cultured renal cells.

Endoglin, a transforming growth factor-beta-binding protein, is upregulated in chronic progressive renal disease.

Endoglin is a non-signalling receptor for TGF-beta. In view of the importance of transforming growth factor-beta (TGF-beta) in the pathogenesis of renal disease, we have determined the distribution of TGF-beta in human glomerulonephritis. Endoglin was present within the glomerular mesangium and interstitium in normal kidneys. In diseased biopsies, there was a weak but significant correlation between staining for endoglin in the interstitium and the extent of chronic histological damage (r = 0.3343, p = 0.003). This was supported by division of biopsies into those showing mild damage and those with moderate to severe damage, where the latter group had significantly increased interstitial staining for endoglin (p = 0.0035). However, there was no correlation between mesangial staining for endoglin and specific types of glomerular pathology, such as IgA nephropathy, suggesting that the interstitial expression of endoglin is associated with increased renal damage independent of the specific type of glomerular lesion which initiates the process. There was also a positive correlation between mesangial cell staining for endoglin and interstitial endoglin expression (r = 0.3104, p = 0.003), although the former was not independently associated with chronic histological damage. These data suggest that the response of interstitial fibroblasts and mesangial cells may be linked in glomerulonephritis. Both could contribute to renal scarring by increased binding of TGF-beta which would be independent of the type of initial glomerular damage.

CD44 in glomerulonephritis: expression in human renal biopsies, the Thy 1.1 model, and by cultured mesangial cells.

CD44 is a transmembrane proteoglycan that serves as a cell adhesion receptor and is involved in cell-cell and cell-matrix interactions, both key events in the pathogenesis of clinical and experimental glomerulonephritis. In addition, recent evidence suggests that the binding of cytokines to proteoglycans could regulate cytokine function. We have, therefore, studied the expression of CD44 by mesangial cells in culture and in experimental (Thy 1.1 model) and human glomerulonephritis. Mesangial expression of CD44 detected by immunohistochemistry was markedly increased four days after induction of the Thy 1.1 model, coinciding with the peak of mesangial cell proliferation and macrophage infiltration. Analysis of 92 human renal biopsies by immunohistochemistry showed that CD44 expression by infiltrating cells within the glomerulus, in focal interstitial infiltrates and within the interstitium (interstitial fibroblasts, and extracellular matrix), was significantly increased in biopsies with a greater degree of histological damage. There was, however, no increase in mesangial staining in diseased kidneys as compared with control sections. In contrast, cultured human mesangial cells expressed CD44 strongly when assayed by immunohistochemistry, immunoprecipitation and Northern blotting. CD44, therefore, is an example of a protein strongly expressed by mesangial cells in vitro and weakly or not at all in vivo, but which is up-regulated in a disease model. In human disease, however, little expression was detected within the glomerular mesangium, which may be related to the greater proliferation and more profound disruption of mesangial architecture seen in the Thy 1.1 model. CD44 expression by infiltrating cells and by components of the interstitium could, however, play an important role in the pathogenesis of chronic progressive renal disease in humans.

Observer variability in histopathological reporting of non-small cell lung carcinoma on bronchial biopsy specimens.

AIMS: To evaluate the ability of histopathologists to sub-classify non-small cell lung carcinomas on bronchial biopsy material using the current World Health Organisation (WHO) classification. METHODS: Twelve histopathologists each reviewed 100 randomly selected bronchial biopsy specimens which had originally been reported as showing non-small cell lung carcinoma. For each case, two sections were circulated, one stained by haematoxylin and eosin and the other by a standard method for mucin (alcian blue/periodic acid Schiff). The participants were allowed to indicate their degree of confidence in their classification of each case. A standard proforma was completed and the results were analysed using kappa statistics. RESULTS: Where the participants were confident in their classification, they were actually quite good at sub-classifying the non-small cell carcinoma sections (kappa = 0.71, standard error = 0.058). Overall, however, the results were only fair (kappa = 0.39, standard error = 0.034). CONCLUSIONS: The majority of non-small cell lung carcinomas can be correctly categorised on adequate bronchial biopsy material. Where a confident diagnosis was made, both squamous carcinoma (kappa = 0.73) and adenocarcinoma (kappa = 0.83) were well recognised.

Adhesion molecule interactions in human glomerulonephritis: importance of the tubulointerstitium.

Infiltration of leukocytes into glomerular and interstitial regions of the kidney is a key event in the pathogenesis of human glomerulonephritis. This process is mediated by specific adhesion molecules, some of which are expressed in a coordinated fashion following endothelial cell activation. We have assessed the pattern of expression of the selectins (E, P and L), and the counter-receptors (LFA-1 and ICAM-1, and VLA-4 and VCAM-1 in 119 renal biopsies using sequential sections, and have correlated this with the degree of histological damage (tubular atrophy and interstitial fibrosis) and the intensity of the macrophage infiltrate. Sections were stained with the monoclonal antibodies using a standard alkaline phosphatase anti-alkaline phosphatase (APAAP) technique. There were strong correlations between the following: (1) expression of LFA-1, VLA-4, and L-selectin in the periglomerular region, interstitium and in focal interstitial infiltrates and the presence of macrophages in these regions; (2) de novo tubular expression of ICAM-1 and VCAM-1; (3) staining for ICAM-1 and VCAM-1 on focal cellular infiltrates within the interstitium; and (4) staining for E- and P-selectin on extraglomerular endothelium. These are also strongly correlated with the degree of chronic histological damage. There was, however, no correlation between glomerular expression of adhesion molecules or glomerular macrophage infiltration and chronic histological damage. Although expression of VCAM-1 by the glomerular mesangium was strongly correlated with the presence of cells staining for VLA-4 within the glomerulus, glomerular expression of adhesion molecules correlated poorly with their expression in other sites. These results show that coordinated up-regulation of adhesion molecule expression in the tubulointerstitium is associated with interstitial fibrosis and tubular atrophy and may contribute, therefore, to the progression of renal disease.

Ecstasy induced hepatitis mimicking viral hepatitis.

Three cases of jaundice after ingestion of 3,4-methylenedioxymetamphetamine (MDMA), known as 'ecstasy', are reported and the complications associated with the misuse of this drug, which was initially misrepresented as 'safer than alcohol' are described. Ingestion of 'ecstasy' should be considered when investigating unexplained jaundice in younger patients.