Apmis

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Online ISSN: 1600-0463
Print ISSN: 0903-4641
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(±) trans-Dihydronarciclasine, isolated from Chinese medicinal plant Zephyranthes candida, has been shown to possess quite potent anti-tumoral effect against selected human cancer cell lines. However, little is known about the anti-tumoral effect of (±) trans-dihydronarciclasine in acute myeloid leukemia (AML). This study was performed to investigate the effect of a novel synthetic (±) trans-dihydronarciclasine (code name; HYU-01) in AML. The HYU-01 inhibited the proliferation of various AML cell lines including HL-60 as well as primary leukemic blasts in a dose-dependent manner. To investigate the mechanism of the anti-proliferative effect of HYU-01, cell-cycle analysis was attempted in HL-60 cells, resulting in G1 arrest. The expression levels of CDK2, CDK4, CDK6, cyclin E, and cyclin A were decreased in a time-dependent manner. In addition, HYU-01 up-regulated the expression of the p27, and markedly enhanced the binding of p27 with CDK2, 4, and 6, ultimately resulting in the decrease of their kinase activities. Furthermore, HYU-01 induced the apoptosis through the induction of proapoptotic molecules and reduction of antiapoptotic molecules in association with the activation of caspase-3, -8, and -9. These results suggest that HYU-01 may inhibit the proliferation of HL-60 cells, via apoptosis, as well as G1 block in association with the induction of p27.
 
Illustration of four different modes of dendritic cell (DC) loading and immunization. (A) All seven peptides were loaded onto the DCs simultaneously and injected intracutaneously (i.c.) in one flank of the mouse. (B) The same number of DCs was aliquoted in seven vials and pulsed separately with one peptide for each vial and mixed in one syringe followed by i.c. injection in one flank of the mouse. (C) Peptides were loaded separately onto DCs as described in group B, but prior to injection, epitopes ranked 1–3 (Table 1) were pooled and administered in the left flank of the mouse and epitopes ranked 4–7 (Table 1) were pooled and administered in the right flank of the mouse. (D) Epitopes ranked 1–3 and 4–7 (Table 1) were loaded onto DCs in two separate vials followed by i.c. injection in the left and right flanks of the mouse, respectively.
Two of seven epitopes dominate the T-cell responses. Chromium release cytotoxic T lymphocyte (CTL) assay and IFN-γ ELISpot assay were used to measure the breadth and magnitude of T-cell responses. (A) (B) (C) and (D) are illustrated in A–D, respectively, and explained in the legend to . Groups of six mice were used for immunization and pooled splenocytes were tested against all seven CTL epitopes and one irrelevant negative control peptide (mock). SFU, spot-forming unit. One representative experiment of three is shown.
Recent human immunodeficiency virus type 1 (HIV-1) vaccination strategies aim at targeting a broad range of cytotoxic T lymphocyte (CTL) epitopes from different HIV-1 proteins by immunization with multiple CTL epitopes simultaneously. However, this may establish an immune hierarchical response, where the immune system responds to only a small number of the epitopes administered. To evaluate the feasibility of such vaccine strategies, we used the human leukocyte antigen (HLA)-A*0201 transgenic (tg) HHD murine in vivo model and immunized with dendritic cells pulsed with seven HIV-1-derived HLA-A*0201 binding CTL epitopes. The seven peptides were simultaneously presented on the same dendritic cell (DC) or on separate DCs before immunization to one or different lymphoid compartments. Data from this study showed that the T-cell response, as measured by cytolytic activity and gamma-interferon (IFN-gamma)-producing CD8(+) T cells, mainly focused on two of seven administered epitopes. The magnitude of individual T-cell responses induced by immunization with multiple peptides correlated with their individual immunogenicity that depended on major histocompatibility class I binding and was not influenced by mode of loading or mode of immunization. These findings may have implications for the design of vaccines based on DCs when using multiple epitopes simultaneously.
 
The active metabolite of vitamin D, 1,25-dihydroxyvitamin D3 (1,25D), has been shown to induce monocyte-to-macrophage maturation in vitro as well as monocytic differentiation of bone marrow precursors and monocytic leukaemic cell lines. In this study we assessed whether 1,25D could improve the maturation defect we have previously demonstrated in monocytes from AIDS patients. In vitro growth and maturation of monocytes from 10 controls, 15 asymptomatic HIV positives (CDC group II or III) and 13 symptomatic HIV positives (CDC group IV) was examined by assessing cellular morphology, differentiation, adherence and protein content. Cells were cultured for 10 days with or without addition of 1,25D at a concentration of 100 pg/ml. In addition, patients were monitored clinically and by immunological parameters and HIV p24 antigen in serum. The present study showed that addition of 1,25D significantly improved the growth and maturation in both patient and control groups. There was a significant negative correlation between response to 1,25D and CD4+ lymphocyte count in blood in HIV-infected patients. A greater response to 1,25D was seen in monocytes from patients with advanced immunodeficiency and symptomatic disease than in monocytes from asymptomatic patients. However, in the most advanced cases of HIV infection with serious ongoing opportunistic infections the response to 1,25D was very poor, possibly reflecting profound and incorrigible dysfunction of monocytes.
 
1,25-Dihydroxyvitamin D3 (calcitriol), transforming growth factor-beta (TGF-beta), and insulin-like growth factors (IGFs) are all important bone regulatory factors known to affect proliferation and differentiation of human bone-forming cells (osteoblasts). We have previously shown that TGF-beta1 increased IGF-I and IGF-binding protein (IGFBP)-3 production in human bone marrow stromal (hMS) osteoblast progenitors and calcitriol stimulated IGFBP-3 and IGFBP-4 production. As interaction between signaling pathways of these factors has been reported, the present study aimed at examining the concerted actions on components of the IGF-system. We report that co-treatment with TGF-beta1 and calcitriol resulted in a synergistic increase in IGFBP-3 production, thereby suggesting that the effects of these factors on hMS osteoblast differentiation may involve the observed increase in IGFBP-3.
 
T helper (Th)1 and Th17 cells play a critical role in inflammatory bowel disease (IBD). 1,25-dihydroxyvitamin D3 (1,25(OH)2 D3 ) has emerged as a direct regulator of immune system function. Mice were grouped as follows: Control group (received PBS, n = 10), DSS group (received 2% DSS and PBS, n = 10), and DSS+VD group (received 2% DSS and 1,25(OH)2 D3 , n = 10). The disease activity index (DAI), colon length, and damage store of the mice were observed; the spleen length, weight, spleen index, and mononuclear cells of spleen were measured; mononuclear cells from mesenteric lymph nodes (MLN) were measured, and the levels of Th 1 and Th17 cytokines in the colon mucosa and spleen were measured. Mice in the DSS group developed severe diarrhea, rectal bleeding, and marked BW loss. Histological examination revealed extensive ulceration and inflammatory cell infiltration in the colon, and the structure of the spleen was disordered, infiltrated with inflammatory cytokines in red pulp. In the DSS group, mononuclear cell numbers from MLN and spleen were increased, and enhanced proteins and mRNA levels of Th 1 and Th17 cytokines were detected. In the DSS+VD group, 1,25(OH)2 D3 ameliorated the inflammation of the colon and spleen. In addition, 1,25(OH)2 D3 down-regulated the levels of Th 1 and Th17 cytokines. 1,25(OH)2 D3 represents a novel therapeutic drug for UC, which may correlate to inhibit the activation of Th1 and Th17 cells. © 2015 APMIS. Published by John Wiley & Sons Ltd.
 
Critical to the prevention of xenograft loss is the prevention of delayed xenograft rejection (DXR), due to its resistance to conventional immunosuppression. The role of the carbohydrate galactose-α1,3-galactose (α1,3Gal) has been a matter of great debate and it has been proposed that the reaction between α1,3Gal epitopes on donor endothelial cells and recipient anti-α1,3Gal antibodies (Abs) may damage the graft during DXR. Recipient anti-α1,3Gal Abs are produced by CD4-dependent B cells. To test the above-mentioned hypothesis, hearts from α1,3Gal-free mice (GT-Ko mice), generated by α1,3-galacto-syltransferase gene disruption, were transplanted to anti-α1,3Gal antibody-free Lew/Mol rats. This model consists of an α1,3Gal/α1,3Gal-antibody-free environment, eliminating a possible influence of this specific system on DXR. A subgroup of recipients were furthermore CD4 depleted in order to inhibit CD4-dependent B-cell antibody production. Rejected hearts were evaluated by light- and immunofluorescence microscopy. Treatment effects on recipient T-cell subsets and cytokine expression were analyzed by flow cytometry, while antibody production was measured by ELISA. All recipients developed DXR with no differences among the groups. DXR was related to thrombosis with IgG and IgM desposition in vessel walls, as well as macrophage and granulocyte accumulation in the myocardium. No complement C3, CD4 cells or NK cells were found. Flow cytometric analysis confirmed peripheral blood CD4 depletion and IFN-γ suppression in CD4 Ab-treated recipients. Finally, ELISA showed that specific anti-α1,3Gal Ab production was absent. However, Ab(s) against an unidentified Galα 1 were found among recipients. In our model, DXR is resistant to α1,3-galactosyltransferase gene inactivation and CD4 depletion. However, other Galα 1 epitopes and antibodies may play a role during DXR. Further studies are needed to elucidate the precise pathways leading to DXR.
 
Bellanger A-P, Grenouillet F, Henon T, Skana F, Legrand F, Deconinck E, Millon L. Retrospective assessment of β-d-(1,3)-glucan for presumptive diagnosis of fungal infections. APMIS 2011; 119: 280–6. β-d-(1,3)-glucan (BG) is a component of the cell walls of many fungal organisms. Our aims were to investigate the feasibility of the BG assay and its contribution to early diagnosis of different types of invasive fungal infections (IFI) commonly diagnosed in a tertiary care centre. The BG serum levels of 28 patients diagnosed with six IFI [13 probable invasive aspergillosis (IA), 2 proven IA, 2 zygomycosis, 3 fusariosis, 3 cryptococcosis, 3 candidaemia and 2 pneumocystosis] were retrospectively evaluated. The kinetic variations in BG serum levels from the 15 patients diagnosed with IA were compared with those of the galactomannan antigen (GM). In 5/15 cases of IA, BG was positive earlier than GM (time lapse from 4 to 30 days), in 8/15 cases, BG was positive at the same time as GM and, in 2/15 cases, BG was positive after GM. For the five other fungal diseases, BG was highly positive at the period of diagnosis except for the two cases of zygomycosis and one of the three cases of fusariosis. This study, which reflects the common activity of a tertiary care centre, confirms that BG detection could be of interest for IFI screening in patients with haematological malignancies.
 
Deficiencies in adhesion molecules or their counter-receptors in humans may have severe consequences as exemplified by leukocyte adhesion deficiency (LAD) I or II syndromes. Because such diseases occur with great rarity, animal models are valuable for studying the role of particular adhesion molecules and their natural ligands in immunity. We studied selected immune parameters and general health in mice with a defect in the sialyl-Lewis X antigen (selectin ligand) caused by disruption of the gene encoding alpha(1,3)fucosyltransferase VII (Fuc-TVII). Leukocytes from Fuc-TVII -/- and control mice were tested for adherence to cellophane membranes or polymer particles in vivo and phagocytic activity in vitro. While no difference in adherence was found, the number of neutrophil granulocytes in exudate induced by intraperitoneal injection of polymer beads was reduced in knock-out mice. Moreover, the phagocytic activity in Fuc-TVII -/- mice was significantly reduced. These animals have splenomegaly due to increased hematopoiesis and reduced weight but do not exhibit clinical signs of immunodeficiency. In conclusion, the lack of Fuc-TVII activity leads to several morphological and functional abnormalities without an impact on survival rate.
 
Survival rates during a follow-up period of more than seven years were analyzed in 1,349 women with breast cancer in relation to the histo-pathological classification of female breast cancer proposed by Ackerman and to other commonly used histo-pathological criteria, including the axillary node status. The information was collected prospectively during a case-control study. Major emphasis was placed on multivariate evaluation. In analyses based on the histo-pathological findings in the mastectomy specimen alone, the Ackerman grouping was found to be of prognostic value, but apart from nuclear polymorphism the other recorded characteristics (histo-pathological type, histological grading, lymphocyte infiltration, and sinus histiocytosis) did not give any prognostic information. When the axillary node status was included in the multivariate models, the presence of axillary metastases correlated well with the prognosis and the Ackerman classification provided no significant additional information. The results indicate that in cases where the histopathological axillary status is known, little additional prognostic information can be gained from traditional histo-pathological evaluation over and above this status. However, the Ackerman classification and the degree of nuclear polymorphism separate distinct prognostic groups with the same degree of difference in observed survival rates as one discriminated by the axillary node status.
 
Hypoxic lesions in the subendocardium and long-standing dilatation of the interstitium have been described after exposure of the heart to glycine 1.5%, which is a widely used irrigating medium in endoscopic surgery. To study if the hypo-osmotic properties of the fluid cause these morphological changes, whether their long duration can be explained by rupture of the histoskeleton and whether they promote sudden death, 75 mice received an intravenous infusion of 200 ml/kg or 300 ml/kg over 60 min of either glycine 1.5%, glycine 1.5% in normal saline, normal saline, or no fluid (controls). The animals were decapitated when they were dying from the infusion, or else 7 days later, and 69 hearts were examined by light microscopy and 4 by electron microscopy. Rupture of the histoskeleton and hypoxic lesions in the subendocardium were observed in 47% and 35%, repectively, of the mice given glycine 1.5%, while the incidence averaged 20% in the two other groups. Rupture occurred in 38% of the mice that died, in 23% of those that survived an infusion, and in 0% of the controls. Hypoxic changes correlated with bradycardia, which is a sign of glycine absorption in the clinic. In conclusion, rupture of the histoskeleton and hypoxic changes occur in the subendocardium of mice after volume loading. These morphological changes were aggravated by the hypo-osmotic properties of glycine 1.5%.
 
We tested the capacity of the Sysmex UF-1000i system to detect yeasts in urine by screening a total of 22 132 urine samples received for culture in our microbiology laboratory during 1 year. We also analyzed different dilutions of previously filtered urine inoculated with a strain of Candida albicans. With clinical samples, a single cut-off point of 50 yeast-like cells (YLCs)/μL detected candiduria ≥10 000 colony forming units (CFU)/mL and >100 000 CFU/mL with a sensitivity of 87.3%/95.4%, a specificity of 97%, a negative predictive value of 95.9%, and a positive predictive value of 9.3%/5.7%. With the simulated samples, a linear relationship was observed between the dilution factor and the number of cells detected by UF-1000i. This instrument appears to be able to reliably rule out candiduria of a magnitude of at least 10 000 CFU/mL and facilitate urine sample screening, thereby providing fast results. The Sysmex UF1000i system can be adapted for candiduria screening by the use of an appropriate YLCs/μL cut-off point that takes account of the prevalence of candiduria in the population.
 
In this mini-review, progress in our understanding of the link between miRNAs and cytokines in inflammatory skin diseases is highlighted with focus on miR-155 in mycosis fungoides (MF). MF is the most common variant of cutaneous T-cell lymphoma (CTCL) and is characterized by malignant T-cell proliferation in a chronic inflammatory environment in affected skin. Recent data show that miR-155 is expressed in situ by both malignant and non-malignant T cells, induced via the STAT5 signal pathway and IL-2Rbeta/gamma cytokines, and thus suggest the importance of miR-155 in malignant proliferation, clinical diagnosis, and prognosis in CTCL.
 
Soft tissue sarcomas (STS) are characterized by deregulated proliferation. Ki-67 is a cell cycle antigen which may be elevated in proliferative states. We analysed Ki-67 expression in fixed and embedded tissues from STS in order to examine associations between proliferation, primary tumour characteristics, and metastasis. One hundred and eighty-two adult patients with trunk wall or extremity STS were treated at our institution between 1980 and 1992 (35 developed local recurrence and 56 developed metastases). Median follow-up time for survivors was 6 years (1-13). We used a semiquantitative score to the assess percentage of Ki-67-positive cells: < or = 10% (n = 86), > 10-25% (n = 57), > 25-50% (n = 30), > 50-75% (n = 7), > 75-100% (n = 2). Increasing Ki-67 expression correlated positively with tumour size, malignancy grade, necrosis, vascular invasion, S-phase fraction, and metastasis. A Ki-67 index Ki-D < or = 10% (n = 86) and > 10% (n = 96) defined two groups who had 84% and 56% 3-year metastasis-free survival (p = 0.0001), respectively. Tumours with Ki-D > 10 were typically large, high grade, necrotic, DNA aneuploid, and had intravascular invasion and a higher S-phase fraction. Ki-67 expression may be helpful in predicting survival of patients with soft tissue sarcomas.
 
Current understanding of the immunological mechanisms involved in the pathogenesis of venous leg ulcers is insufficient. In this study the cellular composition of skin biopsies taken from the center, the edge, and 2 cm distant from the edge of venous leg ulcers was characterized quantitatively by immunohistochemical staining. In the epidermis the mean numbers of Langerhans cells (CD1a+) were four times lower at the edge of the ulcer compared to clinically intact epidermis 2 cm distant from the edge. In the dermis a statistically significant increase in the mean numbers of macrophages (CD68+) and neutrophils (NP57+) from the distant area towards the center of the ulcer was observed. No significant differences were observed in the distribution of T cells nor in the ratio of CD4+/CD8+ T-cell subsets between the different regions of the ulcer. About 30% of T lymphocytes were CD8+ in all microenvironments. The center and the edge of the ulcer were dominated by macrophages comprising 63% and 53% of the cells respectively, while T lymphocytes dominated the distant area. The area 2 cm distant from the edge was also heavily infiltrated by macrophages and neutrophils. B cells (CD22+) and NK cells (CD56+) were relatively rare in all areas, comprising less than 3% of the dermal infiltrate. In conclusion, local microenvironments each with a different cellular composition can be defined within venous leg ulcers.
 
Angiogenesis is a key process in tumour growth and metastasis, and Factor-VIII microvascular density has been found to influence prognosis among endometrial carcinoma patients. The CD105/endoglin antibody has been reported to preferentially bind to activated endothelial cells in tissues participating in angiogenesis, and we therefore wanted to compare the prognostic significance of CD105/endoglin to that of Factor-VIII. In a population-based endometrial carcinoma study with long (median 11.5 years) and complete patient follow-up, mean intratumour microvascular density (MVD) assessed using CD105/endoglin was investigated and compared with previous data for MVD assessed using Factor-VIII. MVD by CD105/endoglin was significantly correlated with MVD by Factor-VIII (p=0.001). However, tumours within the two groups defined by the upper and lower quartiles for CD105/endoglin-MVD were both significantly more often metastatic (FIGO-stage III/IV; p=0.03), with high tumour cell proliferation by Ki67 (p=0.007) and with reduced survival (p=0.036) as compared with the intermediate groups. In Cox regression analysis, CD105/endoglin-MVD showed independent prognostic influence when analysed together with patient age, FIGO stage, histologic subtype, histologic grade and Factor-VIII-MVD, while the latter lost its prognostic impact when CD105/endoglin was included. In the subgroup with high MVD, there was a tendency towards improved response to radiation therapy. In conclusion, CD105/endoglin-MVD is significantly associated with FIGO stage, tumour proliferation and prognosis in endometrial carcinoma, indicating that this is a better angiogenic marker in these tumours.
 
Pheochromocytomas are rare sympathoadrenal tumors that are highly vascular. Their malignancy is extremely difficult to estimate on the basis of histopathological features. Vascular endothelial growth factor (VEGF) is one of the most important angiogenic factors involved in both tumor growth and metastasis. In our search for new prognostic markers, we investigated the expression of VEGF in normal adrenal gland, in 105 primary pheochromocytomas, and in 6 metastases by using immunohistochemistry and Northern blot analysis. We also calculated the microvessel density of these tumors by staining the endothelial cells with monoclonal CD34 antibody. VEGF messenger ribonucleic acid was found in all pheochromocytomas studied. Immunohistochemically, VEGF was not found in normal adrenal medullary cells. Interestingly, all malignant pheochromocytomas (n=8), regardless of their primary location, had strong or moderate VEGF immunoreactivity, while most benign adrenal pheochromocytomas (26 of 37, 70.3%) were either negative or only weakly positive. The staining was heterogenous in extraadrenal pheochromocytomas as well as in a group of tumors that had histologically suspicious features but had not metastasized, here called borderline tumors (n=29). The microvessel density varied greatly in all of the tumor groups, and no statistical difference was found between these groups. Here we report moderate to strong VEGF expression in malignant pheochromocytomas, and negative or weak expression in benign adrenal pheochromocytomas. Normal medullary cells are immunohistochemically negative. Thus, low VEGF expression in pheochromocytomas favors a benign diagnosis.
 
Interferons alpha/beta (IFNs-alpha/beta) are the first cytokines to be produced by recombinant DNA technology. They regulate growth and differentiation, affecting cellular communication, signal transduction pathways and immunological control. This review focuses on the relationships between the structure and biological activities of IFNs-alpha/beta induced as a result of specific interactions with different types of polypeptide receptors as well as on the role of glycolipids in the modulation of these activities. The discovery of the primary structure homology of HuIFNs-alpha and thymus hormone-thymosin alpha 1 (TM alpha 1), the experimental finding of the competition between IFN-alpha and TM alpha 1 for common receptors and the reproduction by reHuIFN-alpha 2 of TM alpha 1 immunomodulating activities create the basis of reHuIFN-alpha therapeutics instead of TM alpha 1, and potentiation of vaccines by reHuIFN-alpha. The first successful attempt at grafting of the HuIFN-alpha 2s TM alpha 1-like immunomodulating site to the designed de novo protein albeferon is described. This article also aims at reviewing recent data concerning the structure of other cytokines and their receptors. Their reciprocal structure-function taxonomy is proposed. The place of IFNs-alpha/beta and their receptors in the hierarchy of cytokines is determined.
 
This paper reviews current knowledge about the effect of testicular germ cell cancer (TGCC) on gonadal function and of cancer treatment on spermatogenesis and Leydig cell function. It is well documented that testicular cancer is associated with impaired spermatogenic function and some patients already have impairment of Leydig cell function before orchidectomy. The degree of spermatogenic dysfunction is higher than what can be explained by local tumour effect and by a general cancer effect, since patients with other malignant diseases have normal, or only slightly decreased, semen quality. Furthermore, sperm counts after orchidectomy are further reduced to less than half of the values in healthy men, even in patients cured from the cancer disease after orchidectomy alone. These observations are supported by histological investigations which have shown a high prevalence of abnormalities of spermatogenesis in the contralateral testis in patients with unilateral TGCC. The association between testicular cancer and poor gonadal function is very interesting both from a biological and from a therapeutic point of view. Firstly, the increase in incidence of testicular cancer has been suggested to be associated with a general decline in male reproductive health and it seems likely that the development of TGCC shares common aetiologic factors with development of other types of testicular dysfunction. This suggestion is supported by the observation that men with various types of gonadal dysfunction such as testicular dysgenesis, androgen insensitivity syndrome, and cryptorchidism have increased risk of testicular cancer. Secondly, the general cure rate in patients with testicular cancer exceeds 90% and the quality of life, including fertility aspects, is therefore important in the management of these patients. Spermatogenesis is already so severely impaired before treatment that fertility is lower than in healthy men. Moreover, radiotherapy and chemotherapy both induce dose-dependent impairment of spermatogenesis and recovery of spermatogenesis after treatment may be long lasting even more than five years in some patients. Sufficient androgen production is seen in the majority of the patients, but some patients suffer from testosterone deficiency. The effect of chemotherapy on Leydig cell function also seems to be dose-dependent. In conclusion there is no doubt that testicular cancer is associated with poor gonadal function even before treatment. Furthermore, the treatment of testicular cancer may have a serious impact on the gonadal function in these patients, most of whom are in the reproductive age. Moreover, the epidemiological and clinical data indicate a common aetiology between testicular germ cell cancer and other abnormalities in male reproductive health (such as infertility and cryptorchidism). These observations are in agreement with the suggestions of hormonal involvement in the aetiology of testicular cancer. Generally, men with TGCC need counselling about their reproductive function with respect to semen cryopreservation, chance of recovery of spermatogenesis, fertility, and the possible need for androgen replacement.
 
Based on immunohistochemistry (IHC) and DNA ploidy, different paths of carcinogenesis have been suggested for spermatocytic seminoma (SS) and classical seminoma (CS). The present study extends current knowledge on the above parameters. Seventeen SSs and twenty-two CSs were assessed by IHC for placental-like alkaline phosphatase (PLAP), c-kit, cytokeratin and adhesion carbohydrate molecyles. All SSs and 11 CSs were also analysed for DNA ploidy. All CSs, but none of the SSs, were positive for PLAP. C-kit positivity was found in 7 of 17 SSs and in all CSs. The other IHC parameters were similarly distributed among the evaluated SSs and CSs. Fourteen SSs were diploid or polyploid, and three were aneuploid. All CSs were aneuploid. The new observation of c-kit positivity in about 40% of SSs suggests that at least some of the SSs originate from primordial cells. The predominantly diploid or polyploid DNA pattern indicates that SSs follow a pathogenetic pathway which is most probably different from that of CSs.
 
Matrix metalloproteinases (MMP) degrade components of extracellular matrix (ECM), and thereby regulate formation, remodeling and maintenance of tissue. Abnormal function of cell surface proteases associated with malignant tumors may contribute directly to the invasive and malignant nature of the cells. Among the MMP's associated with the tumor cell surface, gelatinase A is believed to be particularly important, since it degrades type IV collagen, and is activated in a tumor specific manner, correlating with tumor spread and poor prognosis. Activation of pro-gelatinase A is uniquely regulated by a cell-mediated mechanism. This study describes an in vitro model that mimics the cell-surface activation mechanism. The expression of MT-MMP could not be detected in normal epithelial cells, but can be seen in transformed epithelial carcinoma cells.
 
Model for tumor cell adhesion to microvascular vessel walls within the circulation. Laminar fluid flow causes shear forces that act against adhesive bindings. Flowing tumor cells can interact with microvascular endothelial cells mediated mainly by selectins that have a high elasticity, but the strength of these bonds is relatively low and not sufficient for stable cell adhesions resulting in rolling of the tumor cells with a reduced speed along the endothelium. This results in enhanced probability for integrin-mediated adhesion. However flowing tumor cells can also achieve direct integrin-binding. Initial integrin-mediated adhesion is characterized by a low affinity state. If high affinity of integrin-binding can be established rapidly stable tumor cell adhesion occurs with subsequent signaling for cell spreading, migration and extravasation. However, failure of the conformational affinity switch in integrins results in breaking of these bonds and recirculation of tumor cells or cell damage by shear forces. 
Current evidence indicates that tumor cell adhesion to the microvasculature in host organs during formation of distant metastases is a complex process involving various types of cell adhesion molecules. Recent results have shown that stabilization of tumor cell adhesion to the microvascular vessel wall is a very important step for successful tumor cell migration and colonization of host organs. We are beginning to understand the influences of fluid flow and local shear forces on these adhesive interactions and cellular responses within the circulation. Mechanosensory molecules or molecular complexes can transform shear forces acting on circulating tumor cells into intracellular signals and modulate cell signaling pathways, gene expression and other cellular functions. Flowing tumor cells can interact with microvascular endothelial cells mediated mainly by selectins, but the strength of these bonds is relatively low and not sufficient for stable cell adhesions. Integrin-mediated tumor cell adhesion and changes in the binding affinity of these adhesion molecules appear to be required for stabilized tumor cell adhesion and subsequent cell migration into the host organ. Failure of the conformational affinity switch in integrins results in breaking of these bonds and recirculation or mechanical damage of the tumor cells. Various cell signaling molecules, such as focal adhesion kinase, pp60src or paxillin, and cytoskeletal components, such as actin or microtubules, appear to be required for tumor cell adhesion and its stabilization under hydrodynamic conditions of fluid flow.
 
Comparative studies of DNA isolated from adult C3H mouse liver, immortalized mouse embryo cells (C3H/10T1/2 Cl 8) and the tumorigenic methylcholanthrene transformed C3H/10T1/2 Cl 16 cell line have been carried out in order to analyze possible structural changes in the ribosomal genes associated with the immortalization and tumorigenic transformation of mouse cells. Southern blot hybridization experiments revealed a mutation hotspot within repetitive sequences 13 kb upstream from the 18S rRNA genes in the non-transcribed spacer (NTS). Other DNA changes were localized near the initiation and termination regions of rRNA transcription. The differences found in the restriction maps of the 5'-region resided 5 to 6 kd upstream from the 18S 5'-end and the changes located in the 3'-end mapped approximately 5 kb downstream from the 28S 3'-end. Thus, oncogenic transformation of the C3H/10T1/2 Cl 8 cells by methylcholanthrene treatment was associated with stable genetic changes in the 18S rRNA gene. There was no evidence for rRNA gene amplification.
 
All malignant primary tumours of the thyroid gland submitted for histological diagnosis in Iceland during the 30 years 1955-1984, and available for review, were typed histologically according to the World Health Organization classification but also taking into account the more recent well recognized follicular variant of papillary carcinoma. A total of 480 thyroid tumours were classified with a female--to male ratio of 2.8 (367 females, 129 males). The age distribution is much what would be expected, the anaplastic type of carcinomas occurring in the elderly while papillary and follicular tumours occur over a much wider age range. The incidence of thyroid carcinomas in Iceland is about 2-3 times higher than in the other Nordic countries. This is largely due to an unusually high incidence of the papillary type of carcinoma. Overall, the papillary carcinoma accounted for 80% of thyroid malignancies. The tumours diagnosed incidentally at autopsy were about 20% of the entire material, and these tumours were only of the differentiated types of thyroid carcinoma. Even if the incidentally diagnosed tumours are excluded, the percentage of papillary tumours is 77% which is unusually high. The papillary type of carcinomas occasionally occurred in familial clusters in Iceland but not sufficiently to account for the unusually high incidence. Some of the possible etiological factors are discussed.
 
Pai R, Samuel P, Nehru AG, Manipadam MT, Thomas SN. Comparison of 11 endogenous control genes for normalization of mRNA obtained from paraffin-embedded tissues. APMIS 2009; 117: 886–92. Real-time reverse transcriptase PCR (RT-PCR) based assays are being increasingly used in characterization of gene expression. Good quality mRNA is an essential prerequisite for such assays. While fresh tissues provide quality mRNA, the same may not be true of tissues which are formalin-fixed and paraffin-embedded (FFPE). This emphasizes the need to identify a good endogenous control gene to normalize for differences in quality and RNA recovery. We attempted to characterize gene expression patterns of 11 commonly used endogenous control genes among 20 FFPE tissues (both neoplastic and normal). Pearson’s coefficient of correlation was determined by comparing the expression of each gene against the mean expression of all other genes. β2 microglobulin (β2M) and β-actin (βA) (r = 0.95 and 0.94, respectively) were found to be stably expressed across all tissues. However, βA had greater accuracy (2 × SD) than β2M and therefore may be a better choice of an endogenous control for experiments that require normalization while using FFPE tissues.
 
A perianal bowenoid papulosis was examined for the presence of HPV types 2, 6, 11, 13, 16, 18, 32 and 33 by DNA in situ hybridization. Positive signals were seen for HPV types 6/11, 13, 16 and 33. HPV type 13 is strongly related to oral focal epithelial hyperplasia and has not been reported outside the oral cavity before.
 
Bonagura VR, Hatam LJ, Rosenthal DW, DeVoti JA, Lam F, Steinberg BM, Abramson AL. Recurrent respiratory papillomatosis: a complex defect in immune responsiveness to human papillomavirus-6 and -11. APMIS 2010; 118: 455–470. Recurrent respiratory papillomatosis (RRP) is a rare disease of the larynx caused by infection with human papillomaviruses (HPV) -6 or -11, associated with significant morbidity and on occasion mortality. Here we summarize our current understanding of the permissive adaptive and innate responses made by patients with RRP that support chronic HPV infection and prevent immune clearance of these viruses. Furthermore, we provide new evidence of TH2-like polarization in papillomas and blood of patients with RRP, restricted CD4 and CD8 Vβ repertoires, the effect of HPV-11 early protein E6 on T-cell alloreactivity, enriched Langerhans cell presence in papillomas, and evidence that natural killer cells are dysfunctional in RRP. We review the immunogenetic mechanisms that regulate the dysfunctional responses made by patients with RRP in response to HPV infection of the upper airway. In addition, we are identifying T-cell epitopes on HPV-11 early proteins, in the context of human leukocyte antigen (HLA) class II alleles enriched in RRP that should help generate a therapeutic vaccine. Taken together, RRP is a complex, multigene disease manifesting as a tissue and HPV-specific, immune deficiency that prevents effective clearance and/or control of HPV-6 and -11 infection.
 
Unemo M, Shipitsyna E, Domeika M and the Eastern European Sexual and Reproductive Health (EE SRH) Network Antimicrobial Resistance Group. Gonorrhoea surveillance, laboratory diagnosis and antimicrobial susceptibility testing of Neisseria gonorrhoeae in 11 countries of the eastern part of the WHO European region. APMIS 2011. Quality-assured worldwide surveillance of antimicrobial resistance (AMR) in Neisseria gonorrhoeae is crucial for public health purposes. In the countries of the eastern part of the WHO European region the knowledge regarding gonococcal AMR is limited, and antimicrobials of many different types, sources and quality are used for gonorrhoea treatment. This study surveyed gonorrhoea incidence, laboratory diagnosis and gonococcal AMR testing in 11 independent countries of the former Soviet Union. The national gonorrhoea incidences remain mainly high. In general, gonococcal culture and AMR testing were rarely performed, poorly standardized and rarely quality assured. To establish a gonococcal AMR surveillance programme in Eastern Europe, i.e. the geographical area of the former Soviet Union, several actions have recently been undertaken by the Eastern European Sexual and Reproductive Health (EE SRH) Network and the WHO. The information provided herein will be useful in this respect.
 
The predictive value of cadherin-11, tenascin, fascin, and mucin-1 as markers for the likelihood of recurrence in pleomorphic adenoma of the parotid gland was examined. In this retrospective study we analysed 20 tumours from16 patients by immunohistochemistry. Staining intensities were measured using a semiquantitative scoring approach; localisation (tumour centre vs border) as well as clinical data were analysed and correlated with follow-up. Cadherin-11 was increased in recurrent tumours. However, no changes of fascin, tenascin or mucin-1 were observed. Cadherin-11 and fascin were increased in primary tumours of patients with later recurrence, with fascin upregulation restricted to the tumour border. In conclusion, cadherin-11 and fascin should be further analysed for their value as markers for later recurrence in pleomorphic adenoma. Our observations might reflect dysregulation of cellular pathways contributing to cellular dissemination, which might potentially result in later recurrence.
 
All cases of gastrointestinal (GI) non-Hodgkin's lymphoma diagnosed in Finland between 1972 and 1977 were histologically reexamined and immunostained in order to study the value of histological classification. One hundred and eleven cases were found. The crude annual incidence was 0.51/10(5) and the age-adjusted (world standard population) incidence 0.23/10(5). The male-to-female ratio of age-adjusted incidence rates was 2.7. The most common histological type was large B-cell lymphoma comprising 61% of all classifiable cases. Low-grade mucosa-associated lymphoid tissue (MALT) lymphoma comprised 12%, centrocytic lymphoma 9%, peripheral T-cell lymphoma 9%, Burkitt's lymphoma 7% and large-cell anaplastic lymphoma 3% of the total. In the jejunum, almost one half of the cases were T-cell lymphomas and there were no lymphomas with definite MALT features. Gastric lymphomas had higher survival rates than intestinal lymphomas, B-cell lymphomas slightly higher survival rates than T-cell lymphomas, and low-grade MALT lymphomas higher survival rates than other B-cell lymphomas. The other types of lymphomas differed only slightly from each other in prognosis. The histological grade according to the Working Formulation correlated with survival rates, but a great majority of cases were classified as intermediate grade. Classification of GI lymphomas into the types mentioned above appears to correlate with several clinical and pathological parameters.
 
The field of dendritic cell (DC) biology is robust, with several new approaches to analyze their role in vivo and many newly recognized functions in the control of immunity and tolerance. There also is no shortage of mysteries and challenges. To introduce this volume, I would like to summarize four interfaces of DC research with other lines of investigation and highlight some current issues. One interface is with hematopoiesis. DCs constitute a distinct lineage of white blood cell development with some unique features, such as their origin from both lymphoid and myeloid progenitors, the existence of several distinct subsets, and an important final stage of differentiation termed "maturation," which occurs in response to inflammation and infection, and is pivotal for determining the subsequent immune response. A second interface is with lymphocyte biology. DCs are now known to influence many different classes of lymphocytes (B, NK, NKT) and many types of T cell responses (Th1/Th2, regulatory T cells, peripheral T cell deletion), not just the initial priming or induction of T cell-mediated immunity, which was the first function to be uncovered. DCs are sentinels, controlling many of the afferent or inductive limbs of immune function, alerting the immune system and controlling its early decisions. A third interface is with cell biology. This is a critical discipline to understand at the subcellular and molecular levels the distinct capacities of DCs to handle antigens, to move about the body in a directed way, to bind and activate lymphocytes, and to exert many quality controls on the type of responses, for both tolerance and immunity. A fourth interface is with medicine. Here DCs are providing new approaches to disease pathogenesis and therapy. This interface is perhaps the most demanding, because it requires research with humans. Human research currently is being slowed by the need to deal with many challenges in the design of such studies, and the need to excite, attract and support the young scientists who are essential to move human investigation forward. Nonetheless, DCs are providing new opportunities to study patients and the many clinical conditions that involve the immune system.
 
We investigated the expression of thioredoxin and thioredoxin reductase in a large set of breast invasive and in situ carcinomas by immunohistochemistry. Additionally, NF-kappa B, p53 and proliferating cell nuclear antigen (PCNA) expression was studied. Thioredoxin and thioredoxin reductase expression was located in both cytoplasmic and nuclear compartments of the cell. Cytoplasmic thioredoxin positivity was found in 67 % and nuclear in 59 % of the cases, while thioredoxin reductase was found in 55 % and 6 % of cases, respectively. Ductal carcinomas showed stronger cytoplasmic thioredoxin immunoreactivity than lobular ones. Nuclear thioredoxin positivity was more often found in in situ lesions, and lobular carcinomas were more often negative than ductal ones. Both cytoplasmic and nuclear thioredoxin-positive cases had a high proliferation measured by PCNA staining. Positive nuclear immunostaining was associated with negative estrogen and progesterone receptor status. Cases with high p53 expression showed significantly higher nuclear thioredoxin positivity, but lower thioredoxin reductase positivity. Whilst thioredoxin or thioredoxin reductase was not associated with patient survival, cases showing both cytoplasmic and nuclear thioredoxin reductase-positive tumours had a shorter disease-free interval than those with negative immunostaining.
 
One hundred and twelve clinical strains of Acinetobacter were collected during a 7-month period in 1990-1991 from Danish clinical microbiology departments. According to the old notation, 37 strains were biovar anitratus and 75 b.lwoffii. Using the newest terminology, 35 strains were unambiguously identified as the species A. haemolyticus, A. johnsonii, A. radioresistens, and as the unnamed phenons 6, 10, and 14, by a numerical approach using a simplified phenotypical identification scheme. Most of the remaining strains were identified to the genotypically heterogeneous A. calcoaceticus-A. baumannii complex and the A. lwoffii phenotype. Sixteen strains (14%) were left unidentified. Eight A. lwoffii strains were glucose-oxidizing and were thus classified as b. anitratus using the old terminology. The strains were tested for susceptibility to ampicillin, piperacillin, ticarcillin, cefotaxime, ceftazidime, imipenem, gentamicin, tetracycline, sulphonamide, and nalidixic acid. All strains were susceptible to imipenem. The susceptibility to the remaining beta-lactams was variable, the A. lwoffii isolates being the most and the A. calcoaceticus-A. baumannii complex strains the least susceptible. All non-A. calcoaceticus-A. baumannii complex strains were susceptible to all other antibiotics tested, except for one A. lwoffii strain that was sulphonamide resistant. Twenty-two percent of the strains in the A. calcoaceticus-A. baumannii complex showed reduced susceptibility or resistance to gentamicin, but only sporadic resistance to sulphonamides, tetracyclines, and nalidixic acid. Eight infections were recorded: three cases of septicaemia, three cases of peritonitis related to continuous ambulatory peritoneal dialysis, and two cases of recurrent urinary tract infection. No epidemics were detected.
 
Alterations of caspases, the main executioners of apoptosis, have been described in human cancers. Caspase-9 plays a crucial role in the initiation phase of the intrinsic apoptosis pathway. Caspase-9 is phosphorylated at Thr125 through the mitogen-activated protein kinase (MAPK) pathway, and this phosphorylation is associated with inhibition of caspase-9 activation. The aim of this study was to explore whether phosphorylated caspase-9 (p-caspase-9) expression could be a characteristic of gastric carcinomas. We analyzed expression of p-caspase-9 protein in 60 gastric adenocarcinomas by immunohistochemistry using a tissue microarray approach. p-caspase-9 was detected in 33 of the 60 carcinomas (55%). Both early and advanced gastric carcinomas expressed p-caspase-9. There was no significant association of p-caspase-9 expression with clinocopathological characteristics, including invasion, metastasis and stage. In contrast to gastric cancer cells, epithelial cells in normal gastric mucosa showed no or only weak expression of p-caspase-9. Taken together, these results indicate that caspase-9 is frequently phosphorylated in gastric carcinomas, and that the phosphorylation of caspase-9 might be an inhibitory mechanism of caspase-9-mediated apoptosis in gastric carcinomas. Increased expression of p-caspase-9 in malignant gastric epithelial cells compared to normal mucosal epithelial cells suggests that p-caspase-9 expression might play a role in gastric carcinoma development.
 
Lajer CB, von Buchwald C. The role of human papillomavirus in head and neck cancer. APMIS 2010; 118: 510–519. Over the last 20 years, there has been increasing awareness of a subset of squamous cell carcinomas of the head and neck (HNSCC), i.e. HPV-positive HNSCC. These cancers seem to differ somewhat from HPV-negative HNSCC. Patients with HPV-positive HNSCC tend to be younger and have a lower intake of tobacco and alcohol. Distinct molecular profiles separate them from HPV-negative cancers and show similarities with HPV-positive cervical SCC. There is evidence that HPV-positive HNSCC is a sexually transmitted disease. Patients with HPV-positive HNSCC are often diagnosed at a late stage with large cystic lymph nodes in the neck. HPV-positive HNSCC show an affinity for the oropharynx, especially the tonsils and the base of the tongue, and tend to show low differentiation histopathologically. There is a better prognosis regardless of the treatment regimen for HPV-positive HNSCC compared with HPV-negative HNSCC, and this seems to be related to the immune system. Whether the new vaccines for HPV will protect not only against cervical cancer but also against HPV-positive HNSCC remains unknown.
 
In a previous study we have defined a subgroup of human malignant extragonadal germ cell tumours that is characterized by complex translocations involving chromosomes 6 and 11 (Echten et al. 1995). Here we report (i) the use of fluorescent in situ hybridization, pulsed field gel electrophoresis and direct visual hybridization techniques to localize the tumour-associated breakpoint within band 11q13, and (ii) the construction of a phage library enriched for this region to facilitate genomic walks towards the breakpoint. Extensive breakpoint-flanking contigs were generated and within these contigs six candidate genes could be identified.
 
We have studied MAbs* for their ability to detect SCLC and differentiate this tumor type from the other lung tumor histotypes in cryostat sections of biopsy specimens taken at bronchoscopy from patients with suspected primary lung tumor disease. MAb F12, specific for the ganglioside fucosyl-GM1, reacted with 58% of the cases with SCLC (n = 19) and with less than 3% of those with non-SCLC (n = 38). MAb 123C3, specifically reactive with NCAM, reacted with 78% of the SCLC cases (n = 23). With this MAb no positive staining was seen in the non-SCLC cases (n = 41). None of the two MAbs reacted with tissue sections without tumor. In combined analysis with MAbs F12 and 123C3, all SCLC cases (n = 15) were positive with either and 47% with both of the MAbs. Our results show that both MAbs F12 and 123C3 are highly specific for SCLC in bronchoscopic biopsy tissue specimens, whereas the sensitivity for this histotype tends to be higher with MAb 123C3 than with F12 (P = 0.14). When used in combination, all SCLC cases could be identified. These MAbs may therefore be valuable as complements to current histopathologic characterization and differentiation of lung cancer.
 
The in vitro susceptibility of 124 Xanthomonas maltophilia isolates was tested by four methods: Agar dilution (reference method), E-test, a disk diffusion and a tablet diffusion method. Trimethoprim-sulfamethoxazole had the highest activity against X. maltophilia, followed by a combination of aztreonam-clavulanic acid at different ratios, the ratio 1:1 being the most active with a susceptibility rate of 85% as compared to 2% for aztreonam alone. Addition of the beta-lactamase inhibitor tazobactam to piperacillin enhanced the rate of susceptible isolates from 31% to 53%, Relatively few isolates were susceptible to ciprofloxacin (27%) and gentamicin (9%). Generally, the disk diffusion method had a considerably higher frequency of "very major" discrepancies when compared with the agar dilution method than with the other methods. The susceptibility of X. maltophilia to trimethoprim-sulfamethoxazole and ciprofloxacin could reliably be determined by all the diffusion methods tested, but otherwise the agar dilution method is to be preferred. A standardized and reliable diffusion method for susceptibility testing of X. maltophilia remains to be found. Trimethoprim-sulfamethoxazole must be considered the drug of choice in the treatment of severe X. maltophilia infections. The combination aztreonam-clavulanic acid is promising, but must be proved in a clinical setting.
 
CA-125 is a high molecular weight glycoprotein that is best known as a tumour marker for ovarian carcinoma but has been found to be present on various epithelial surfaces including normal tissues. Elevated serum levels of CA-125 have been described in malignancies other than ovarian carcinoma as well as in inflammatory conditions. The expression of CA-125 was studied in paraffin-embedded tissue from 48 mammary carcinomas and 11 samples of normal mammary gland using two monoclonal antibodies, M2 and M11. CA-125 was detected in all normal tissue samples and 64% of the breast carcinomas. Eight of the thirty CA-125-positive carcinomas reacted with only one of the antibodies, indicating molecular change. In normal mammary tissue, CA-125 was seen on apical surfaces and in ductal contents, whilst the majority of the carcinomas (90%) expressed CA-125 in cytoplasmic granules, often showing membranous staining as well. In 16 samples of lymph node metastases CA-125 expression was similar to that seen in the primary tumour. Elevated serum levels of CA-125 were detected in only 3 out of 41 samples available from this patient group. No significant associations were detected with various clinical parameters. We conclude that CA-125 is normally expressed in the mammary gland and that the expression is frequently altered and sometimes absent in mammary carcinoma, possibly reflecting the loss of cellular polarity. Measuring serum levels of CA-125 is not relevant in breast carcinoma patients since one third of breast carcinomas were CA-125 negative and even patients with strongly CA-125-positive tumors had undetectable CA-125 serum levels.
 
Recently, miR-155 has been implicated in cutaneous T-cell lymphoma (CTCL). Thus, elevated levels of miR-155 were observed in skin lesions from CTCL patients as judged from qPCR and micro-array analysis and aberrant, high miR-155 expression was associated with severe disease. Moreover, miR-155 promoted proliferation of malignant T cells in vitro. Little is, however, known about which cell types express miR-155 in vivo in CTCL skin lesions. Here, we study miR-155 expression using in situ hybridization (ISH) with a miR-155 probe, a negative control (scrambled), and a miR-126 probe as a positive control in nine patients with mycosis fungoides, the most frequent subtype of CTCL. We provide evidence that both malignant and non-malignant T cells stain weakly to moderately positive with the miR-155 probe, but generally negative with the miR-126 and negative control probes. Reversely, endothelial cells stain positive for miR-126 and negative for miR-155 and the control probe. Solitary T cells with a malignant morphology display brighter staining with the miR-155 probe. Taken together, our findings suggest that both malignant and non-malignant T cells express miR-155 in situ in CTCL. Moreover, they indicate heterogeneity in miR-155 expression among malignant T cells.
 
PAF antagonists have been used in xenotransplantation to alleviate the pathogenesis of hyperacute rejection. This study evaluated the ability of the PAF antagonist UR-12670 to improve graft function in late xenograft rejection (LXR) in an orthotopic liver xenotransplantation model, and the involvement of PAF (platelet activating factor) in this type of rejection. The recipients of a hamster xenograft received standard immunosuppression (tacrolimus 0.2 mg/kg/30 days, MMF 25 mg/kg/8 days). Study groups: group A, without UR-12670, group B, UR-12670 (20 mg/kg/8 d) and group C, continuous administration of UR-12670 (20 mg/kg/d). Serum levels of xenoantibodies were evaluated by flow cytometry and tissue deposits by immunofluorescence. Immunoblot and indirect immunofluorescence assessed specificity of xenoantibodies. Conventional histology was performed. Continuous administration of UR-12670 improved the histological pattern of liver xenografts, especially necrosis, loss of hepatocytes, hemorrhage, sinusoidal congestion and lymphocyte infiltration. There was not a shift in specificity of xenoantibodies at different times posttransplantation, as demonstrated by immunoblotting and indirect immunofluorescence. UR-12670 administration had a beneficial effect on graft function and considerably improved the histopathological pattern, but it failed to induce tolerance after withdrawal of immunosuppression. UR-12670 had an immunomodulatory effect on cellular response but not on antibody production. There was not a change in the specificity of xenoantibodies produced at LXR compared with pretransplant antibodies.
 
Compared with conventional culture media, the TB BACTEC system has demonstrated improved isolation rates as well as an earlier detection time for mycobacterial species. However, the identification of M. tuberculosis by the rho-nitro-alpha-acetylamino-beta-hydroxypropiophenone (NAP) test in the TB BACTEC 460 system may require 6 days for interpretable results. We evaluated the usefulness of a polymerase chain reaction (PCR) assay for earlier identification of M. tuberculosis in positive BACTEC 12B cultures. A total of 262 TB BACTEC culture specimens with GIs > or = 10 were assayed by PCR, and the results were compared with those of the NAP test. The aliquot from BACTEC 12B vials was boiled for 10 min, and 2 microliters of the boiled suspension was used for the PCR assay. One set of primers based on the IS 6110 sequence of M. tuberculosis was used to amplify a 457 bp fragment of DNA. Of the 173 TB BACTEC culture specimens which were identified as M. tuberculosis by the NAP test. 171 were PCR positive. Of the 21 TB BACTEC cultures identified as MOTT by the NAP test. 19 were PCR negative, but 2 were PCR positive: these two cultures were shown to grow both M. tuberculosis and MOTT in BACTEC 12B vials. Of the remaining 68 cultures which were contaminated with AFB-negative bacteria, the PCR identified M. tuberculosis in 13, in agreement with the NAP results in the reprocessed specimens. Overall, the PCR results in the 262 BACTEC culture specimens with GIs > or = 10 were sensitive in 99.5% (186/187) and specific in 100% (68/68). The mean time for the identification of M. tuberculosis in TB BACTEC cultures with GIs > or = 10 was 7 h by the PCR compared to 5.9 days by the NAP test. These results suggest that the PCR could be used as an alternative to the NAP test for the rapid identification of M. tuberculosis in BACTEC 12B cultures, particularly in those which contained both M. tuberculosis and MOTT or M. tuberculosis and AFB-negative bacteria.
 
Within the human testis, three entities of germ cell tumours are distinguished: the teratomas and yolk sac tumors of newborn and infants, the seminomas and nonseminomas of adolescents and young adults, referred to as testicular germ cell tumours (TGCT), and the spermatocytic seminomas. Characteristic chromosomal anomalies have been reported for each group, supporting their distinct pathogenesis. TGCT are the most common cancer in young adult men. The initiating pathogenetic event of these tumours occurs during embryonal development, affecting a primordial germ cell or gonocyte. Despite this intra-uterine initiation, the tumour will only be clinically manifest after puberty, with carcinoma in situ (IS) as the precursor. All invasive TGCT, both seminomas and nonseminomas, as well as CIS cells are aneuploid. The only consistent (structural) chromosomal abnormalities in invasive TGCT are gains of the short arm of chromosome 12, mostly due to isochromosome (i(12p)) formation. This suggests that an increase in copy number of a gene(s) on 12p is associated with the development of a clinically manifest TGCT. Despite the numerous (positional) candidate gene approaches that have been undertaken thus far, identification of a causative gene(s) has been hampered by the fact that most 12p gains involve rather large genomic intervals, containing unmanageable numbers of candidate genes. Several years ago, we initiated a search for 12p candidate genes using TGCT with a restricted 12p-amplification, cytogenetically identified as 12p11.2-p12.1. This approach is mainly based on identification of candidate genes mapped within the shortest region of overlap of amplification (SROA). In this review, data will be presented, which support the model that gain of 12p-sequences is associated with suppression of apoptosis and Sertoli cell-independence of CIS cells. So far, DAD-R is one of the most likely candidate genes involved in this process, possibly via N-glycosylation. Preliminary results on high through-put DNA- and cDNA array analyses of 12p-sequences will be presented.
 
The antibiotic susceptibility of consecutive isolates of the upper respiratory tract pathogens Streptococcus pyogenes, Streptococcus pneumoniae, Haemophilus influenzae, Branhamella catarrhalis, and Staphylococcus aureus, (100 strains of each species collected each year during March through April 1985, 1988 and 1992) to penicillin V, amoxycillin, cefaclor, cefuroxime, doxycycline, erythromycin, and cotrimoxazole was investigated by MIC determination on PDM and PDM II agar. The MICs of the upper respiratory isolates from 1992 supplemented with 100 isolates each of Escherichia coli, Klebsiella spp., Enterobacter cloacae, Proteus mirabilis and Staphylococcus saprophyticus collected during 1992 were determined against the above antibiotics plus cefadroxil, cefpodoxime, roxithromycin, ciprofloxacin, ofloxacin, and BAY Y 3118. Beta-lactamase production was found in 10% of H. influenzae and 80-90% of S. aureus and B. catarrhalis in 1992. Among H. influenzae isolates, non-beta-lactamase-induced resistance to all beta-lactam antibiotics was first detected in 1988 and amounted to 3% of isolates in 1992. Decreased susceptibility of S. preumoniae to penicillin (> or = 0.12 mg/l), co-trimoxazole > or = 32 mg/l, doxycycline (> or = 2 ml/l) and erythromycin (> or = 1 mg/l) was detected in 11%, 7%, and 8%, respectively, in 1992, which is significantly higher than in previous years at the same laboratory. Decreased susceptibility of S. pyogenes to doxycycline and erythromycin was detected in 11% and 9% in 1992. The two most recently developed antibiotics, cefpodoxime and BAY Y 3118, showed high antibacterial activity. The study emphasizes the need to screen for resistance mechanisms such as beta-lactamase production and lowered penicillin affinity.
 
In stratified squamous epithelia, altered expression of keratins (Ks) is one possible marker of malignant potential. In the epithelium of the uterine cervix, presence of human papillomaviruses (HPVs) is increasingly regarded as a marker of risk for cervical cancer. However, a similar role in oral cancer and precancer remains controversial. To address these questions, formalin-fixed, paraffin-embedded oral carcinomas from Sudanese snuff dippers (n=14) and oral carcinomas from Sudanese (n=14), Swedish (n=19) and Norwegian (n=41) non-snuff dippers were examined by immunohistochemistry for expression of K types 13, 14 and 19 using monoclonal antibodies. HPV infection was searched for in all the carcinomas by in situ hybridization (ISH) using the cocktail HPV OmniProbe and the ViraType probe. Carcinomas from Sudanese (snuff dippers/non-snuff dippers) were also examined for HPV infection by polymerase chain reaction (PCR) using the general HPV primers GP5+/GP6+. For the oral carcinomas from snuff dippers, moderate to intense expression of K13 (71%; 10/14), K14 (86%; 12/14) and K19 (93%; 13/14) was found. For the oral carcinomas from non-snuff dippers, weak to moderate expression of K13 (64%; 47/74), K14 (43%; 32/74) and K19 (45%; 33/74) was found. HPV DNA was not detected in any of the carcinomas from three countries when examined by ISH. The Sudanese (from snuff dippers/non-snuff dippers) oral carcinomas were also negative for HPV DNA with the PCR. The present study shows that (i) there is a high level of expression of K13, K14 and K19 in oral carcinomas from snuff dippers compared to those from non-snuff dippers, (ii) this high level of expression may arise from dysregulation of keratinocyte proliferation and maturation caused by damaging effects of snuff, (iii) the HPV genome is not found in Sudanese (snuff dippers/non-snuff dippers), Swedish or Norwegian oral carcinomas, and (iv) this may suggest that these viruses do not play a prominent role in the aetiology of oral carcinomas from these countries.
 
Data from 76 patients with malignant tumours of the testis registered with the Icelandic Cancer Registry during the period 1955-1984 were analyzed and classified histologically according to the WHO criteria. In 55 patients the tumours were of one histological type (72% of total). In the order of frequency, seminoma was diagnosed in 35 (46%), embryonal carcinoma in 14 (18%) and teratoma in 6 patients (8%). Twenty patients had tumours of more than one histological type (26%), i.e. a combination of embryonal carcinoma and teratoma (teratocarcinoma) in nine (12%) and combined tumours (seminoma component with other tumours) in seven (9%). Two patients had choriocarcinomas and three had yolk sac (endodermal sinus) tumours, both of them in combination with other neoplasms. Embryonal rhabdomyosarcoma was the only malignant non-germ cell tumour diagnosed. The age-adjusted incidence of malignant testis tumours increased by 48% from 1.6 per 100,000 per annum in 1955-1964 to 3.1 per 100,000 in 1975-1984. The increase affected both seminoma and non-seminomatous tumours (NSGCT). There has been an improvement in survival with time. Sixteen deaths were related to testis tumours and all occurred within four years of diagnosis. The incidence of testicular tumours in Iceland is intermediate between "high risk" and "low risk" countries.
 
From 1952 to 2005, 13 cases of cryptococcosis confirmed by postmortem examination were diagnosed in autopsy material from the University Hospital in Hradec Králové, the Czech Republic. Histologically, Cryptococcus was found in multiple organs (brain and spinal cord, lungs, lymph nodes, spleen, bone marrow, liver, kidneys and adrenal glands). The lungs and CNS were the organs most often involved. Only in two cases was the diagnosis of cryptococcal infection established during the patient's lifetime, in both presenting clinically as meningitis, with positive result of CSF cultivation. Data and issues of diagnostics and treatment of cryptococcosis are discussed.
 
The aim of this study was to compare colorectal adenocarcinoma with mucinous component, ordinary adenocarcinoma (OA) and mucinous adenocarcinoma (MA) regarding clinicopathological parameters, survival, EGFR, MMP-13, and E-cadherin. We studied tumor tissue specimens from 28 patients with adenocarcinoma with mucinous component, 47 with OA, and 56 with MA, who underwent radical surgery from January 2007 to January 2012 at the Gastroenterology Centre, Mansoura University, Egypt. High density manual tissue microarrays were constructed and immunohistochemistry for EGFR, MMP-13, and E-cadherin was done. Colorectal adenocarcinoma with mucinous component (AWMC) was significantly associated with more perineural invasion, lower EGFR, and MMP-13 expressions than OA, with no difference in E-cadherin expression. Conversely, only microscopic abscess formation was significantly more with colorectal AWMC than MC with no difference in EGFR, MMP-13 and E-cadherin expression between both groups. Colorectal AWMC showed a better survival than MA with no difference with OA. In a univariate analysis, EGFR, MMP-13, and E-cadherin expressions did not show a significant impact on disease-free or overall survival in patients with colorectal AWMC. Colorectal AWMC remains a vague entity that resembles OA in some clinicopathological and molecular respects as well as MA. © 2015 APMIS. Published by John Wiley & Sons Ltd.
 
The aim of the present study was to compare, both radiographically and histologically, malformed vertebral lumbar corpora in trisomies 21, 18 and 13 with earlier reported normal corporal development in the axial lumbar region. Axial skeletons of human fetuses (GA 15-22 wk) derived from therapeutically induced abortion were investigated in connection with requested autopsy. The number of lumbar vertebral corpora examined for each genotype was as follows: 20 from trisomy 21, 10 from trisomy 18, and 10 from trisomy 13. After radiography in frontal, lateral and axial projections, the individual vertebral corpora were decalcified and horizontally embedded in paraffin. The blocks were serially sectioned and stained with toluidine blue and alcian blue/van Gieson. The radiographic characteristics of the vertebral corpora varied from an almost normal appearance of the corporal bone to complete clefting of the bony corpora. Histological examination showed accumulations of cartilage centrally, in some cases associated with amorphous material. Pronounced metachromatic differences were observed in the cartilaginous ground substance. The study showed identical phenotypic characteristics in the corpora from trisomy 21, trisomy 18, and trisomy 13. It is characteristic of all three genotypes that there are central anomalies, corresponding to the location of the notochord in normal corpora, and marked regional differences in metachromasia in the ground substance of the cartilage.
 
Human collagenase-3 (MMP-13) is a matrix metalloproteinase originally identified in breast carcinomas. Recent studies have revealed that this enzyme is also produced by a variety of malignant tumors including head and neck carcinomas, chondrosarcomas and basal cell carcinomas of the skin. In all cases, the expression of collagenase-3 is associated with aggressive tumors. Different cytokines, growth factors and tumor promoters are able to up-regulate collagenase-3 expression in tumor cells or in stromal cells surrounding epithelial tumor cells. Functional analysis of the collagenase-3 gene promoter has allowed the identification of AP-1 and OSE-2 elements mediating, at least in part, its expression in both normal and pathological conditions.
 
During a 7-year period bronchoalveolar lavage (BAL) was performed as a routine diagnostic procedure at fiberoptic bronchoscopy in 172 consecutive patients with diffuse pulmonary lesions. In 42 patients, BAL was technically insufficient or data were incomplete. These patients were excluded. The remaining 130 patients consisted of 78 men and 52 women with a median age of 43 years (range 19-79); 59 were smokers. They were divided into 6 groups: I. sarcoidosis (n = 77); II. cryptogenic fibrosing alveolitis (n = 16); III. secondary fibrosing alveolitis (n = 7); IV. malignancy (n = 7); V. allergic alveolitis (n = 6); VI. miscellaneous (n = 17). Group VI was not included in the statistical evaluation, which involved only the 113 patients in groups I-V. BAL was performed in a segment of the right middle lobe with 150-200 ml isotonic saline. The return fluid (BALF) was filtered through two layers of cotton gauze, and total and differential cell counts were assessed. Median BALF return volume was 67% (range 35-90). Eighty percent of the procedures were performed by the same operator. Total cell count displayed no significant difference amongst the five diagnostic groups (p = 0.06). Differential cell count displayed differences amongst the groups respecting macrophages (p = 0.002), lymphocytes (p = 0.0004), neutrophils (p = 0.0001) and eosinophils (p = 0.04). Patients with sarcoidosis had a higher percentage of lymphocytes, patients with secondary fibrosis a higher percentage of neutrophils, and patients with cryptogenic fibrosis a higher percentage of eosinophils than the other groups. Malignant cells were observed in BALF in 14.3% of patients with malignant lesions. Among the patients with sarcoidosis, 75% had a lymphocyte-dominated BALF (> 10%) compared with 31% of the patients with cryptogenic fibrosis, 14% of the patients with secondary fibrosis, and 43% of the patients with malignancy. Dominance of neutrophils (> 10%) and/or eosinophils (> 5%) in BALF was observed in cryptogenic and secondary fibrosis. In most patients, BAL cannot provide a definite diagnosis, but may support the clinical suspicion of a specific diagnosis. In clinical practice, BAL seems to be of limited value in the diagnostic evaluation of radiologically detected diffuse, non-infectious pulmonary lesions.
 
Top-cited authors
Bente Pakkenberg
  • Region Hovedstaden
Karsten Nielsen
  • Aarhus University Hospital
Lise Korbo
  • Bispebjerg Hospital, Copenhagen University
Jens R Nyengaard
  • Aarhus University
Thomas Bendtsen
  • Aarhus University Hospital