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  • 1
    ISSN: 1432-0428
    Keywords: BB rats ; insulitis ; macrophages ; T-lymphocytes silica ; electron microscopy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary We have analysed whether infiltration of macrophages and lymphocyte subtypes into pancreatic islets of diabetes prone BB rats occurs at random or whether insulitis requires a specific sequence of events. Serial sections from more than 700 islets of diabetes prone BB rats (70–150 days of age) were analysed for infiltrating immunocytes and expression of major histocompatibility complex antigens by 4–11 different monoclonal antibodies. In parallel, electron microscopy was performed in a fraction of islets. Part of the animals had been treated with macrophage toxic silica particles. A specific non-random sequence of events was identified and 4 stages of islet inflammation were recognised. Stages 1 a and 1 b are defined by macrophage (ED1+, W3/25+. Ox3/6/17+, ED2−) infiltration and concomitant enhanced major histocompatibility complex class I antigen expression initially at one pole or at the periphery of islets. T-, NK- and B-lymphocytes are absent (〈1 cell per mean islet section). In stage 2, more macrophages are infiltrating and concomitantly Ox19+-T-lymphocytes and Ox8+-granular (NK-) lymphocytes are observed. In stage 3, additional massive infiltration of Ox12+-B-lymphocytes is noted. Silica treatment of BB rats largely prevented macrophage infiltration. Concomitantly islets were free of lymphocytes. Thus, macrophage infiltration clearly precedes T- and NK-lymphocyte and later B-lymphocyte infiltration. Lymphocytes do not infiltrate islets in the absence of prior macrophage invasion.
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  • 2
    ISSN: 1432-0428
    Keywords: Type 1 (insulin-dependent) diabetes ; islet cell antibody ; seasonal efects ; epidemiology ; HLA-dependence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary The prevalence of islet cell antibodies in children with Type 1 (insulin-dependent) diabetes was determined in a cohort of 678 children. The natural course of islet cell antibodies was followed in 375 children at 1 year, 252 and 135 children after 2 and 3 years respectively. Islet cell antibodies were determined by indirect immunofluorescence on cryostat sections of human pancreas. At diagnosis of diabetes 85% of the children had detectable islet cell antibodies (mean titre 10.4). After 3 years 62% of the children were still islet cell antibody positive (mean titre 2.9) indicating a greater persistence of islet cell antibodies than described in earlier studies. In this large cohort a significant correlation between islet cell antibody prevalence or persistence and sex, age or HLA-DR type was not observed except for a faster loss of islet cell antibodies in very young boys and in patients lacking HLA-DR types 3 and 4. Complement fixing islet cell antibodies correlated with high titre islet cell antibodies. Greater persistence of islet cell antibodies was seen for cases with high antibody titre and in children with diagnosis of diabetes during the first half of the year.
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  • 3
    ISSN: 1432-0428
    Keywords: Streptozotocin (low dose) ; prazosin ; vasoactive amine antagonists ; insulitis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary Pancreatic islet inflammation and subsequent diabetes was induced by multiple low doses of streptozotocin in male C57 Bl/6J mice. The development of hyperglycaemia was almost completely prevented by treating the animals with the α1-adrenoceptor antagonist prazosin (20 mg·kg−1. day−1) as well as by the vasoactive amine antagonists methysergide (50 mg·kg−1·day−1), disodium cromoglycate (100mg·kg−1·day−1), pizotifen (5 mg·kg−1·day−1) or cyproheptadine (20 mg·kg−1·day−1). Treatment with vasoactive amine antagonists largely inhibited infiltration of pancreatic islets by L3T4+-lymphocytes and to a lesser extent by Lyt2+-cells. The infiltration of macrophages was not affected except after pizotifen treatment. These results indicate that α1-adrenoceptor activation is required for disease development and that vasoactive amine release is a prerequisite for lymphocytic insulitis but not for macrophage infiltration of islets.
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  • 4
    ISSN: 1432-0428
    Keywords: Experimental diabetes ; mice ; streptozotocin ; anti-Ia-antibodies ; major histocompatibility complex ; immunomodulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary In male mice of strains C3H and C57BL/6 an experimental immune-mediated diabetes can be induced by multiple low doses of streptozotocin. The delay and partial suppression of hyperglycaemia after anti-I-A monoclonal antibody administration was dose dependent. Even saturation levels of anti-I-A did not cause complete protection from diabetes development. Administration of anti-I-E monoclonal antibody also significantly delayed the onset of hyperglycaemia. Surprisingly, the combined treatment with anti-I-A and anti-I-E did not result in better protection from diabetes. Thus, there is an I-A and I-E independent component of the disease. Furthermore, there is no restriction to either I-A or I-E. Anti-I-A was only effective when given at the beginning of the experiment, which implies that I-A molecules have a primary function during the induction of diabetes. The contribution of I-J to the disease process is different. Administration of a polyspecific alloantiserum to I-J almost completely prevented hyperglycaemia. Injections of monospecific antibodies to I-J determinants enhanced hyperglycaemia, especially when given after the induction of diabetes. This indicates that I-J is involved in initial as well as in later stages of the disease process.
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  • 5
    ISSN: 1432-0584
    Keywords: CML ; Busulfan ; Hydroxyurea ; Interferon-alpha ; Duration of chronic phase ; Prospective study
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary For palliative therapy during the chronic phase of CML busulfan has proved to be the drug of choice. During the past years hydroxyurea and also interferon-alpha have gained increasing significance since they might prolong the duration of the chronic phase. In a multicenter study it is being determined, whether the use of hydroxyurea or of interferon-alpha instead of busulfan prolongs the duration of the chronic phase of Philadelphia positive CML. Additional goals are the examination of whether the types of disease evolution and the terminal phases differ between the treatment groups, and the prospective recognition of prognostic criteria for the duration of the chronic phase of CML. By December 31, 1987, 326 CML-patients had been randomized, 150 for busulfan, 150 for hydroxyurea and 26 for interferon-alpha. The average age is 50 years. 59 patients reached the end of the chronic phase, 55 died. The mean observation time of all patients is 1.34 years. At present no significant difference in survival is recognizable between the busulfan and hydroxyurea groups. Fewer adverse effects have been observed in the hydroxyurea group. Philadelphia chromosome negative patients show a higher average age and tend to have lower white blood cell and platelet counts. The number of patients having received interferon-alpha is still too small to allow evaluation. This report intends to document organization and progress of this study which to our knowledge is, at present, the largest ongoing prospective multicenter study on the therapy of CML.
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  • 6
    ISSN: 1432-1424
    Keywords: electrical coupling ; gap junction ; double whole cell patch clamp ; cAMP ; protein kinase C ; OAG
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary Gap junctional coupling was studied in pairs of murine pancreatic acinar cells using the double whole-cell patch-clamp technique. During stable electrical coupling, addition of OAG (1-oleoyl-2-acetyl-sn-glycerol) induced a progressive reduction of the junctional conductance to the detectable limit (∼3 pS). Prior to complete electrical uncoupling, varius discrete single channel conductances between 20 and 100 pS could be observed. Polymyxin B, a potent inhibitor of the protein kinase C (PKC) system, completely suppressed OAG-stimulated electrical uncoupling. Dialysis of cell pairs with solutions containing PKC. isolated from rat brain, also caused electrical uncoupling. The presence of 0.1mm dibutyryl cyclic AMP and 5mm ATP in the pipette solution, which serves to stabilize the junctional conductance, did not suppress the effects of OAG or isolated PKC. We conclude that an increase of protein kinase C activity leads to the closure of gap junction channels, presumably via a PKC-dependent phosphorylation of the junctional peptide, and that this mechanism is dominant over cAMP-dependent upregulatory effects in the experimental time range (≤1 hr). A correlation of the observed single channel conductances with the appearance of channel subconductance states or various channel populations is discussed.
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  • 7
    ISSN: 1432-1424
    Keywords: MDCK cells ; serotonin ; patchclamp ; K+ channel ; intracellular calcium
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary The present study has been performed to test for the effect of intracellular calcium and of serotonin on the channel activity in patches from subconfluent MDCK-cells. In inside-out patches, inwardly rectifying potassium-selective channels are observed with open probabilities of 0.01±0.01, 0.24±0.03 and 0.39±0.07, at 100 nmol/liter, 1 μmol/liter or 10 μmol/liter calcium activity, respectively. The single-channel slope conductance is 34±2 pS, if the potential difference across the patch (V ϱ) is zero, and approaches 59±1 pS, ifV ϱ is −50 mV, cell negative. In the cell-attached mode, little channel activity is observed prior to application of serotonin (open probability=0.03±0.03). If 1 μmol/liter serotonin is added to the bath perfusate, the open probability increases rapidly to a peak value of 0.34±0.04 within 8 sec. In continued presence of the hormone, the open probability declines to approach 0.06±0.02 within 30 sec. At zero potential difference between pipette and reference in the bath (i.e., the potential difference across the patch is equal to the potential difference across the cell membrane), the single-channel conductance is 59±4 pS. In conclusion, inwardly rectifying potassium channels have been identified in the cell membrane of subconfluent MDCK-cells, which are activated to a similar extent by increase of intracellular calcium activity to 1 μmol/liter and by extracellular application of 1 μmol/liter serotonin.
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  • 8
    ISSN: 1432-1424
    Keywords: maxi-K(Ca) channel ; open probability ; burst kinetics ; apical membrane ; on-cell patch clamp
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary A Ca and potential-dependent K channel of large unit conductance was detected in the apical membrane of JTC-12.P3 cells, a continuous epithelial cell line of renal origin. The open probability of the channel is dependent on membrane potential and cytoplasmic-free Ca concentration. At cell-free configuration of the membrane patch, the open probability shows a bell-shaped behavior as function of membrane potential, which decreases at larger depolarization. With increasing Ca concentration, the width of the bell-shaped curve increases and the maximum shifts into the hyperpolarizing direction. For the first time the kinetics of this channel was analyzed under cell-attached conditions. In this case the kinetics could sufficiently be described by a simple open-closed behavior. The channel has an extremely small open probability at resting potential, which increases exponentially with depolarization. The low probability induces an uncertainty about the actual number of channels in the membrane patch. The number of channels is estimated by kinetic analysis. It is discussed that this K channel is essential for the repolarization of the membrane potential during electrogenic sodium-solute cotransport across the apical membrane.
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  • 9
    ISSN: 1432-1424
    Keywords: K channels ; rectification ; open probability ; plant vacuoles ; patch clamp
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary Voltage-dependent K channels could be identified in on-cell and excised patch-clamp records on membranes of isolated plant cell vacuoles. The current through a membrane patch is dominated by a channel population with a conductance of about 121 pS in symmetrical 250mm KCl solution. The single channel adopts at least two conducting levels the 121-pS state being most frequently observed. The channel shows outward rectification, representing a cation flux into the vacuoles. The rectification appears to be caused by a vanishing open probability and a short channel lifetime at hyperpolarizing voltages. A selectivity ratio of potassium over sodium of about 6 was derived as an estimate. Occasionally, an additional population of K channels with a single-channel conductance of approximately 18 pS is observed. This channel type exhibits outward rectification as well.
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  • 10
    ISSN: 1432-1440
    Keywords: Immunology ; Diabetes mellitus ; Prediabetes ; Islet cell antibodies ; Insulin autoantibodies
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary The presence of cytoplasmatic islet cell antibodies (ICA) and IgG insulin autoantibodies (IgG-IAA) has been observed in the prediabetic state of type 1 (insulin-dependent) diabetes (IDDM). We therefore analyzed the prevalence of these markers in sera from 1117 healthy HLA-typed first-degree relatives (1° Rel) of IDDM patients. ICA was determined by indirect immunofluorescence on cryostat sections of human pancreas. For IgG-IAA measurement a competitive solid-phase ELISA was used. ICA were present in 3.5% of 1° Rel vs 0.4% of controls (P〈0.025). The highest frequencies of ICA were found in individuals of IDDM multiplex families (7.7%) and HLA-DR1,3 (5.4%), -DR1,4 (5.8%), and -DR3,4 (6.7%) positive subjects. We therefore conclude that the prevalence of ICA is increased in 1° Rel with high genetic risk for diabetes. IgG-IAA occurred in 9.9% of 1° Rel vs 1.4% of controls (P〈0.01). Like ICA, IgG-IAA were significantly increased in a group of subjects being positive for either HLA-DR1,3-DR1,4, or -DR3,4 (16.5%,P〈0.01). In multiplex families, however, prevalence of IgG-IAA was not increased. In contrast to ICA there was an additional influence of age and sex: IgG-IAA were found more often in siblings (mean age, 16.6 years; prevalence, 15.0%) than in parents (mean age, 44.1 years; prevalence, 8.3%) of IDDM patients (P〈0.01). In brothers the prevalence of IgG-IAA is higher than in other 1° Rel. Only a weak association between ICA and IgG-IAA was observed in subjects (n=810) tested for both antibodies. IgG-IAA occurred in 6/35 (17%) ICA positive 1°Rel, while ICA were found in 6/79 (8%) IgG-IAA positive relatives.
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