Dysplasia in Random Biopsies from Barrett's Surveillance Is an Important Marker for More Severe Pathology.

BACKGROUND AND AIM: Although endoscopic recognition of dysplasia in Barrett's esophagus is difficult, experience in recognition of early neoplastic lesions is supposed to increase the detection of early neoplastic lesions. The aim of this study was to assess the significance of dysplasia in random biopsies in Barrett's esophagus, in the absence of reported visible lesions as well as the difference in final outcome of pathology. METHODS: We retrospectively identified all patients with Barrett's esophagus with suspicion of dysplasia or early adenocarcinoma who were referred to our center between February 2008 and April 2016. We analyzed all endoscopy reports, pathology reports, and referral letters from 19 different hospitals. Patients were divided into two groups, based on the presence or absence of visible lesions reported upon referral. RESULTS: In total, 170 patients diagnosed with dysplasia or adenocarcinoma were referred to our tertiary center. Ninety-one of these referred patients were referred with dysplasia or adenocarcinoma in random biopsies, without a reported lesion during endoscopy in the referral center. During endoscopic work-up at our center, a visible lesion was detected in 44 of these 91 patients (48.4%). After endoscopic work-up and treatment, adenocarcinoma was found in an additional 21 patients. Two of these patients were initially referred with low-grade dysplasia, and 19 patients were initially referred with high-grade dysplasia. The final pathology was upstaged in 35.8% of the patients. CONCLUSIONS: The presence of any grade of dysplasia in random biopsies during surveillance in referral centers is a marker for more severe final pathology. Training in recognition of early neoplastic lesions in Barrett's esophagus imaging is recommended for endoscopists performing Barrett's surveillance.

Therapeutic drug monitoring of the HIV protease inhibitor atazanavir in clinical practice.

BACKGROUND: Therapeutic drug monitoring (TDM) is being applied for a number of antiretroviral agents. Little is known about the use of TDM for atazanavir. METHODS: This is a retrospective cohort analysis on the use of TDM of atazanavir at three clinical sites in The Netherlands. Patients were divided into three groups: (i) all patients with evaluable data of plasma atazanavir concentrations and its relationship with hyperbilirubinaemia; (ii) patients who started atazanavir without documented evidence of protease inhibitor (PI) mutations; (iii) patients who started atazanavir with documented evidence of PI mutations. The genotypic inhibitory quotient (GIQ) was calculated by dividing the mean atazanavir plasma trough concentration by the number of PI mutations. RESULTS: A total of 108 patients were included; 70 (65.8%) were using atazanavir/ritonavir (300/100 mg once daily). No significant relationship was observed between atazanavir plasma trough concentration and antiviral response in patients starting atazanavir without PI mutations (group 2; n = 82). In contrast, a significant relationship was observed between atazanavir GIQ and treatment response in patients starting atazanavir while having PI mutations (group 3; n = 26). The cut-off value for GIQ most predictive of virological failure was 0.23 mg/L/mutation: patients (n = 8) with a GIQ equal to or below this value had 50% virological failure whereas patients (n = 18) with a GIQ above 0.23 mg/L/mutation had only 11% virological failure (chi(2): P = 0.030). Atazanavir plasma trough concentrations were significantly related with the occurrence of increased total bilirubin concentrations. CONCLUSIONS: TDM of atazanavir might be beneficial for patients with documented PI resistance or patients with hyperbilirubinaemia.

Viral dynamics during tenofovir therapy in patients infected with lamivudine-resistant hepatitis B virus mutants.

Tenofovir, an antihuman immunodeficiency virus (HIV) drug, has activity against lamivudine-resistant hepatitis B virus (HBV) mutants. To describe the efficacy of tenofovir in patients with lamivudine-resistant hepatitis B we applied two investigative approaches based on mathematical models of viral dynamics: the individual nonlinear fitting and the mixed-effect group fitting approaches. Eleven chronic HBV patients on lamivudine for a median of 176 weeks (range: 72-382) with YMDD mutation-related HBV-DNA breakthrough received 'add-on' tenofovir 300 mg once-daily, while maintaining their existing therapy. Sequential sera were taken at day 1 (t = 0 and t = 8 h), days 2, 4, 7, 10, 14, 21, 28 and every 4 weeks thereafter, and HBV-DNA levels were assessed using a validated quantitative polymerase chain reaction (PCR) assay. Median baseline log HBV-DNA was 8.62 (range: 6.48-9.76 log HBV-DNA). Tenofovir treatment resulted in a mean (+/-SD) log HBV-DNA decline of 1.37 +/- 0.51 in the first phase, 2.54 +/- 0.91 after 4 weeks, and 4.95 +/- 0.90 log HBV-DNA after 24 weeks. The median effectiveness of blocking viral replication in the individual fit model was 93% (range: 73-99) for eta = 0 and 93% (range: 59-99) for eta = 1. There was only a small difference between the efficacy parameter 'epsilon' of the individual nonlinear fitting and mixed-effect group fitting on the biphasic exponential model. These data show that tenofovir has good efficacy in blocking viral replication in HBV patients with lamivudine-induced drug-resistant HBV mutants, but effectiveness varies greatly among individuals. Both models can be used to describe viral decay during tenofovir therapy.

[Liver transplantation in a HIV-positive patient receiving potent antiretroviral therapy; the first case in The Netherlands]. / Orthotope levertransplantatie bij HIV-positiviteit en krachtige antiretrovirale behandeling; de eerste patient in Nederland.

In a 49-year-old woman infected with HIV who was receiving highly-active antiretroviral treatment (HAART), terminal liver failure developed. She also had an acute exacerbation of hepatitis B. She was treated by means of liver transplantation and was in good condition two years later. At that time she was treated with tacrolimus, lamivudine, tenofovir, nelfinavir and hepatitis-B immunoglobulin. HIV-RNA and the DNA of hepatitis-B virus could not be detected, her CD4-count was not abnormal and the liver transplant functioned well. No opportunistic infections had developed. HIV infection has long been considered an absolute contraindication to solid organ transplantation, due to the increased risk of infection and rapid progression to AIDS. With HAART, restoration of immune function is possible. Currently, international experience with liver transplantation for HIV-positive patients that are not infected with hepatitis C has shown promising results. Specifically, the risks of transplant rejection, opportunistic infections and progression to AIDS are not increased. Therefore, criteria have been defined for solid organ transplantation in HIV-positive recipients.

Ultrastructural localisation of sialoadhesin (siglec-1) on macrophages in rodent lymphoid tissues.

In previous studies it has been demonstrated that sialoadhesin is a macrophage-restricted adhesion receptor for lymphocytes and myeloid cells. It is under normal circumstances expressed by subpopulations of macrophages in lymphoid and haemopoietic tissues. In this study different immunoelectronmicroscopical techniques are used to investigate the ultrastructural localisation of sialoadhesin within the lymph node and spleen of rodents. The results show that sialoadhesin is selectively expressed by a subset of macrophages in peripheral lymphoid tissues. Sialoadhesin was localised predominantly on the plasma membrane and in particular in areas of intimate contact with lymphocytes, thereby visualizing putative local interaction between these cells. Interestingly, sialoadhesin was also detected in intracellular vesicles that were apparently taken up by macrophages. These findings are consistent with the putative role of sialoadhesin in local cell-cell interactions in lymphoid tissues. Surprisingly, sialoadhesin was also found at contact points of macrophages with other macrophages, sinus-lining cells and reticulum cells, suggesting that sialoadhesin also mediates interactions with these cell types.

Reversible depletion of synovial lining cells after intra-articular treatment with liposome-encapsulated dichloromethylene diphosphonate.

We studied the depletion and repopulation of synovial lining cells in mice. A single intra-articular injection of liposomes encapsulating the drug dichloromethylene diphosphonate (CL2MDP) in the mouse knee joint caused selective elimination of synovial lining cells. Depletion of cells occurred within a few days as evidenced by light microscopic, electronmicroscopic and immunohistochemical studies. Maximal depletion was seen on day 7. Repopulation was observed in the following weeks, starting at the bone side of the joint. Until day 30, full recovery (60% recovery) was not observed in the lining lying adjacent to the dermis. Side effects on cartilage metabolism, such as inhibition of proteoglycan synthesis or degradation of proteoglycans from the matrix was minor but significant, 1 and 2 days after liposome treatment but thereafter full recovery was observed. Selective elimination of lining cells from the joint enabled us to study the in vivo role of these cells in the onset and subsequent pathology of experimental arthritis. An immune-complex-mediated experimental arthritis elicited in lining cell depleted joints that had received CL2MDP-liposomes 7 days earlier prevented inflammation as compared to controls.

Selection of asymptomatic HIV carriers for antiviral therapy.

In order to find suitable markers for selection and monitoring of antiviral therapy in asymptomatic HIV-infected patients, we evaluated 18 anti-HIV positive individuals at three monthly intervals by HIV culture, HIV antigen, and core (p24) antibody testing as well as by measurement of lymphocyte subsets. Consistent results were obtained with HIV antigen, p24 antibody testing and T4 cell enumeration, whereas results of virus detection were variable. Therefore cumbersome and expensive virus culture is not of use in selecting patients for antiviral therapy. On the basis of our results and recent literature we currently propose using absence of p24 antibodies, presence of HIV antigen and low or falling T4 cells as eligibility criteria for antiviral therapy in asymptomatic infected individuals.

Rat thymus macrophages: an immunohistochemical study on fetal, neonatal and adult thymus.

The pre- and postnatal development of the macrophage population of rat thymus is investigated applying enzyme-histochemical and immunohistochemical techniques, both on tissue sections and cell suspensions. A set of three monoclonal antibodies (ED1, ED2 and ED3), each of which recognizes cells of the monocyte-macrophage lineage in the rat, enabled us to distinguish between macrophages in the various compartments of the thymus. The medulla is characterized by ED1-positive dendritic cells, the corticomedullary region comprises numerous monocyte-like ED1-positive macrophages and the cortex contains a particular subpopulation of branched ED2-positive macrophages. Both the medullary dendritic cells and the cortical branched cells show Ia-membrane staining. ED3-positive cells are only occasionally present. During fetal life ED1-positive monocyte-like macrophages and dendritic cells are present. Just after birth ED2-positive cortical macrophages start to develop. Their number increases strongly during the first week after birth. The role of the various subpopulations of thymic macrophages is discussed.

Changes occurring in the epithelium covering the bronchus-associated lymphoid tissue of rats after intratracheal challenge with horseradish peroxidase.

Changes occurring in the epithelium covering bronchus-associated lymphoid tissue (BALT) in the rat after several intratracheal administrations of horseradish peroxidase (HRP) were studied using morphological and ultrastructural methods. The epithelium is invaded by W3/25-positive (T-helper) lymphocytes, the BALT epithelial cells become Ia-positive and develop microvilli; there is an apparent loss of cilia. The number of non-ciliated cells in stimulated BALT increases. The non-ciliated cells can be subdivided into two cell types, one with electron-dense cytoplasm and cytoplasmic granules and the other without granules. The electron-density of the latter cell type is intermediate between that of the ciliated cells and that of the granule-containing non-ciliated cells. The granule-containing cell types may be responsible for the uptake of antigens, while the other non-ciliated cell may be involved in the production of the secretory component and the passage of secretory IgA.

Non-lymphoid cells of bronchus-associated lymphoid tissue of the rat in situ and in suspension. With special reference to interdigitating and follicular dendritic cells.

Immunohistochemical, enzyme-histochemical and electron-microscopical methods were used to study non-lymphoid cells of control and stimulated rat bronchus associated lymphoid tissue (BALT) in situ and in suspensions. Particular attention was paid to the so-called antigen-handling cells, i.e., the interdigitating cells (IDC), which are situated in the T-cell areas, the follicular dendritic cells (FDC), which appear to be restricted to germinal centers, and macrophages, present both in T-cell and B-cell areas. The interdigitating cells were distinguished by being Ia-positive and by the presence of acid phosphatase and non-specific esterase activity in an area near the nucleus. Follicular dendritic cells could be observed in situ by using a monoclonal antibody and by the in vitro trapping of HRP-anti-HRP complexes. Several types of macrophages were found. At the electron-microscopical level no well-developed IDC and FDC could be detected in control BALT. However, in BALT of lipopolysaccharide-stimulated and mycoplasma-infected rats, well-developed IDC and FDC were found. It can be concluded that IDC's and FDC's can be found in BALT.

Morphology, kinetics and secretory activity of antibody-forming cells.

Specific antibody-forming cells from spleen, bone marrow and popliteal lymph nodes were studied in mice after subcutaneous priming and intravenous boosting with horseradish peroxidase (HRP). Functional antibody-secreting capacity of these cells was correlated with their morphology at the cell population level. For this purpose, cells synthesizing anti-HRP antibody from the same cell suspensions were studied simultaneously by light and electron microscopy and by a plaque assay. It appeared that the population of cells responsible for antibody synthesis as well as antibody secretion was morphologically heterogeneous: besides plasma cells, considerable numbers of antibody-forming lymphocytes, antibody-forming plasmablasts and antibody-forming immature plasma cells were observed. Immature plasma cells constituted the majority of antibody-forming as well as antibody-secreting cells. Among the immature plasma cells in the popliteal lymph nodes proliferation occurred. Evidence is presented that the light-microscopically identified mature plasma cell is not the main antibody-forming cell. It does not show 3H-Thymidine incorporation and should be considered as a non-dividing end-cell.

Mesenteric lymph nodes: cells with surface and sytoplasmic immunoglobulins.

The distribution and morphology of cells with surface and cytoplasmic immunoglobulins were investigated in mesenteric lymph nodes (MLN) from rats, using both frozen and (fixed) paraffin sections, with a two-step immunoperoxidase technique. Anti-IgA, -IgE, -IgG and -IgM sera were used. Surface Ig-cells (sIg) of all four isotypes studied were found in MLN, mainly localized in the interfollicular area and within the follicle corona. The percentages of sIgA, IgE, IgG and IgM were about 15, 5, 45 and 35%, respectively. In addition, sIgM- and sIgG-cells were found around high endothelial venules. The percentages of cells containing IgA, IgG or IgM (cIg-cells) were about 60, 25 and 15%, respectively; only a few cIgE-cells were found. cIg-cells were not only present in the interfollicular areas and the medulla but also within the germinal centers of the follicles. These results are discussed with regard to the interaction between Peyer's patches (PP) and MLN.

Development of bronchus associated lymphoid tissue (BALT) in the rat, with special reference to T- and B-cells.

The development of bronchus associated lymphoid tissue was studied in normal rats, in serial sections, using both routine histological techniques and the two-step immuno-peroxidase method on cryostat sections for demonstration of T lymphocytes, IgM-, IgG- and IgA-bearing B lymphocytes and plasma cells, respectively. BALT first appears 4 days postpartum (p.p.) as a condensation of reticulum cells near a lymph vessel, between the main bronchus and the accompanying artery. Only a few lymphocytes are present at first. Vascularisation is considerable 8 days p.p. and includes high endothelial venules. Leucocytes are seen in transit in both blood- and lymph vessel walls. Lymphocytes populate the area under the epithelium from about 2 weeks p.p. A few IgM, IgG- and IgA-bearing B cells are already present at 4 days, and rapidly increase in numbers; T cells usually appear at day 8. Discrete T- and B-cell areas do not appear until 4 weeks and are only observed regularly after 12 weeks, when secondary follicles appear. BALT starts development at a similar time to other peripheral lymphoid organs but apparently achieves immunological activity later. It is concluded that antigens probably play an important part in organising BALT.

Specific antibody formation in mouse spleen. Histology and kinetics of the secondary immune response against HRP.

Specific antibody formation in the spleen was described in mice primed subcutaneously with horseradish peroxidase (HRP) and boosted intravenously with the same antigen. The first specific antibody-forming cells responding to the booster injection were observed after one day in the periarteriolar lymphocyte sheath (PALS). The number and staining intensity of these cells greatly increased subsequently. Between day 3 and 6 after the booster injection a shift was observed in the distribution of antibody-forming cells from the PALS to both the site of entry of the central artery in the PALS and the site where the blood passes via terminal arterioles to the red pulp. Specific antibody-forming cells became highly concentrated in these areas, which constitute the so-called marginal zone bridging channels. After day 6 the number of antibody-forming cells decreased sharply. On the basis of this distribution pattern it was suggested that after subcutaneous priming and intravenous boosting specific antibody-forming cells migrate from the popliteal lymph nodes to the spleen and gradually leave the spleen at later stages of the response. Specific antibody-forming cells did not occur in the germinal centers during any stage of the response.