The human hippocampal formation mediates short-term memory of colour-location associations

The medial temporal lobe (MTL) has long been considered essential for declarative long-term memory, whereas the fronto-parietal cortex is generally seen as the anatomical substrate of short-term memory. This traditional dichotomy is questioned by recent studies suggesting a possible role of the MTL for short-term memory. In addition, there is no consensus on a possible specialization of MTL sub-regions for memory of associative information. Here, we investigated short-term memory for single features and feature associations in three humans with post-surgical lesions affecting the right hippocampal formation and in 10 healthy controls. We used three delayed-match-to-sample tasks with two delays (900/5000 ms) and three set sizes (2/4/6 items). Subjects were instructed to remember either colours, locations or colour-location associations. In colour-only and location-only conditions, performance of patients did not differ from controls. By contrast, a significant group difference was found in the association condition at 5000 ms delay. This difference was largely independent of set size, thus suggesting that it cannot be explained by the increased complexity of the association condition. These findings show that the hippocampal formation plays a significant role for short-term memory of simple visuo-spatial associations, and suggest a specialization of MTL sub-regions for associative memory.     

Extracellular actin impairs glomerular capillary repair in experimental mesangioproliferative glomerulonephritis

Exogenous administration of actin prevents tumour growth in mice by specifically antagonizing angiogenin, a potent inducer of neovascularization. To investigate whether the angiogenin/actin system is also of importance in renal disease, we examined the effect of actin during glomerular capillary repair in anti-Thy-1.1 mesangioproliferative glomerulonephritis. Male Wistar rats were injected intravenously with actin, a control protein, i.e. albumin, or vehicle alone at 8, 16, 24, 32, 40 and 48 h after disease induction. On day 8, actin-treated rats showed significantly more microaneurysms and persistent mesangiolysis as compared to both control groups. This was associated with increased proteinuria in actin-treated rats. Moreover, actin-treated rats showed increased counts of glomerular macrophages (+40%) and polymorphonuclear leukocytes (+100%) on day 3 as well as a decrease in glomerular endothelial area on days 3 and 8. However, no difference in early glomerular endothelial as well as non-endothelial cell proliferation was noted in actin-treated rats as compared to controls. Actin treatment had no apparent influence on mesangial cell activation (i.e. de novo expression of alpha-smooth muscle actin) or glomerular accumulation of fibronectin or type IV collagen. Additional in vitro studies demonstrated that extracellular actin inhibits the angiogenin but not VEGF(165)-induced proliferation of (glomerular) endothelial cells. Moreover, actin inhibited other, yet unidentified, serum-derived angiogenic factors. In conclusion, exogenous actin impairs glomerular capillary repair in experimental mesangioproliferative glomerulonephritis possibly due to interference with angiogenic factors such as angiogenin. Our combined in vivo and in vitro observations suggest that the release of intracellular actin during mesangiolysis is an endogenous pathway by which glomerular capillary damage is augmented.     

The effects of platelet-derived growth factor antagonism in experimental glomerulonephritis are independent of the transforming growth factor-beta system

Platelet-derived growth factor B-chain (PDGF-B)- and transforming growth factor beta (TGF-beta)-mediated accumulation of extracellular matrix proteins contributes to many progressive renal diseases. In vivo, specific antagonism of either PDGF-B or TGF-beta in experimental mesangioproliferative glomerulonephritis resulted in an almost complete inhibition of matrix protein accumulation, which suggests an interaction between signaling pathways of these two growth factors. Because nothing is known on the nature of this possible interaction, PDGF-B was antagonized in the rat anti-Thy 1.1 model of glomerulonephritis by use of specific aptamers and its effects on the TGF-beta system were investigated. Antagonism of PDGF-B led to a significant reduction of glomerular matrix accumulation compared with scrambled aptamer-treated nephritic controls. PDGF-B antagonism had no effect on the overexpression of glomerular TGF-beta mRNA, TGF-beta protein, or the expression of TGF-beta receptor type I and II mRNA. By immunohistology, it was possible to detect overexpression of the cytoplasmic TGF-beta signaling molecules Smad2 (agonistic) and Smad7 (antagonistic) in glomeruli of nephritic control rats which peaked on day 7 after disease induction, i.e., the peak of mesangial cell proliferation in this model. However, immunohistology and Western blot analysis again revealed no difference in the glomerular expression of both Smad proteins between PDGF-B antagonized and nonantagonized nephritic animals. In addition, no difference in the glomerular expression of phosphorylated Smad2 (P-Smad2) was detected between the differently treated nephritic groups. These observations suggest that the effects of PDGF-B antagonism are independent of TGF-beta in mesangioproliferative glomerulonephritides.     

Immunohistological skin alterations in allogeneic bone marrow transplantation

Summary Skin biopsies of 26 patients with leukemia and seven patients with aplastic anemia were investigated before and at different stages after allogeneic bone marrow transplantation (BMT) to establish the immunological criteria which distinguish skin alterations during normal reconstitution from dermal lesions mediated by graft-versushost disease (GvHD). Of the 33 patients studied 27 presented with clinically diagnosed acute and/or chronic GvHD, one patient died of bone marrow rejection. Immunohistological analysis of the respective skin biopsies with selected monoclonal antibodies against human leukocyte antigens (HLA) and differentiation antigens of the lympho-hematopoietic cells revealed low dermal mononuclear cell counts with phenotypically normal constituents in five cases with uncomplicated reconstitution post-grafting. In contrast, increased dermal cellular infiltrates predominantly consisting of Lyt 3⁺, OKT 8⁺ T-lymphocytes, as well as of a large number of Ia-like (immune response associated = HLA-D) determinant⁺ monocytes/macrophages were observed in all patients with active acute/chronic GvH reactivity. As sign of activation simultaneous expression of HLA-D region products was also found on a subset of the invading OKT 8⁺ T-lymphocytes. Progression of GvHD was associated with additional surface staining of keratinocytes for Ia-like determinants. Loss of Ia-like determinant⁺, OKT 6⁺ dentritic epithelial cells in all leukemic patients, as well as in patients with aplastic anemia with or without GvHD suggested damage of Langerhans cells due to the previous radiotherapy and/or specific immunological destruction. In patients with fatal outcome of GvHD prolonged reduction of these dentritic epithelial cells seemed to be indicative of impaired immune reconstitution or bone marrow dysfunction. Thus immunopathological features of skin GvHR may enable early recognition and prognostic evaluation of this disease possibly allowing more effective therapy.Abbreviations GvHD graft-versus-host disease - GvHR graft-versus-host reactivity - SAA severe aplastic anemia - TBI total body irradiation - BMT bone marrow transplantation - HLA human leukocyte antigens - Ia-like antigens Immune-response-associated antigens - PBS phosphate buffered saline - FITC fluorescein isothiocyanate - TRITC tetramethylrhodamine isothiocyanate Supported by DFG Sonderforschungsbereich 120, Projekt A 2 and B 1Dedicated to Prof. Dr. Dr. h.c. H.E. Bock on the occasion of his 80^th birthday     

Relation between regional electrical activation time and subepicardial fiber strain in the canine left ventricle

To determine the relation between regional electrical activation time and fiber strain, epicardial electrical activation and deformation were measured in six open-chest dogs at the left ventricular anterior free wall after 15 min of right atrial, left ventricular free wall, left ventricular apex, or right ventricular outflow tract pacing, when end-diastolic pressure was normal or elevated (volume-loading). Regional electrical activation was measured using a 192-electrode brush. Regional subepicardial fiber strain (e _f) was measured simultaneously in 16 regions, using optical markers which were attached to the epicardial surface and recorded on video. When relating regional e _f during the ejection phase to regional activation time, the best correlation was found when a hemodynamic time reference rather than an electrophysiological one is used. Using the moment of the maximum rate of change of left ventricular pressure as the time reference for electrical activation, regional electrical activation time (t _ea) and the degree of e _f during the ejection phase could be fitted by a linear regression equation e _f=a t _ea+b in which a=–3.46±0.73 s^–1 and b=–0.28±0.05. For electrical activation times ranging from -40 to -80 ms, fiber strain was estimated with an accuracy of ±0.026 (±SE) with this relation. During right atrial pacing, t _ea and e _f were on the average –48 ms and –0.10 respectively. On further investigation, the relation between e _f and t _ea appeared to be influenced by end-diastolic pressure. For normal (1.1 kPa) and elevated end-diastolic pressure (1.8 kPa), the slope of the linear regression line was –3.96 and –2.86 s^–1, respectively. Three conclusions may be drawn. Firstly, the time interval between the moment of regional electrical activation and the moment of the maximum rate of change of left ventricular pressure is an index of regional fiber strain. Secondly, it can be concluded from the above equations that electrical asynchrony of more than 30 ms causes non-uniformities in the degree of e _f of the order of mean e _f during pacing from the right atrium. Finally, differences in fiber strain during asynchronous electrical activation are less pronounced at larger filling pressures.     

Knochenmarktransplantation bei akuter Leukämie,chronisch myeloischer Leukämie,schwerer aplastischer Anämie und Neuroblastom Stadium IV. Einfluß antiviraler Prophylaxe mit Anti-CMV-Hyperimmunglobulin und Azyklovir

Summary Bone marrow transplantation was performed between IV/82 and X/85 in 64 patients with acute leukemia (n=36), chronic myelogenous leukemia (CML;n=13), severe aplastic anemia (n=12), and neuroblastoma stage IV (n=3). Of these patients 57 received allogeneic marrow from HLA-ABCDR identical, MLC-negative sibling donors. Six transplants were performed with syngenic marrow and one with autologous marrow. Of the 64 patients 48 survived 40-1,250 days after transplantation, resulting in a survival rate (SR) of 75% and a survival probability (SP) of 71%. Of the 36 patients suffering from acute leukemia (SR=64%, SP=51%), patients with acute myelogenous leukemia (AML) in first complete remission (n=11; SR=81%, SP=76%), as well as patients with acute lymphatic leukemia (ALL) in 1st to 4th complete remission at the time of transplantation (n=14; SR=81%, SP=76%) show a favorable prognosis. A poor survival rate was seen for patients with AML when transplanted in second or partial remission (1/5; SR=20%), as well as for patients suffering from ALL and transplanted during relapse or partial remission (1/6; SR=16%). Of 13 patients suffering from CML 12 survived the transplantation free of relapse (SR=93%, SP=92%), and one patient died from varicella zoster pneumonia. Of the transplanted patients with severe aplastic anemia, 12 of 13 are surviving with complete hematologic reconstitution; one patient, however, died on day 10 from a sepsis. In our patient group, the SR as well as the SP has been improved through changes in the irradiation protocol concomitant with prophylactic application of anti-CMV hypergammaglobulin, as well as through additional oral medication of Azyklovir. The 41 patients (BMT No. 7–47) with total body irradiation at one time-show an SR of 44% and an SP of 41%. The following 46 patients (BMT No. 48–93) have reached an SR of 83% and an SP of 74% under the regimen of fractionated total body irradiation, plus prophylaxis with anti-CMV hypergammaglobulin and Azyklovir. Within this group, no fatal CMV pneumonia was encountered as opposed to six patients lost from CMV pneumonia in the first group.

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Abkürzungsverzeichnis AHTCG Anti-human-T-cell-globulin - ALL Akute lymphatische Leukämie (acute lymphatic leukemia) - AML Akute myeloische Leukämie (acute myelogenous leukemia) - ANV Akutes Nierenversagen - ARA-C Cytosin-Arabinosid - ARDS Akutes respiratorisches Distress-Syndrom - ATG Antithymozytenglobulin - CML Chronisch myeloische Leukämie (chronic myelogenous leukemia) - CMV Cytomegalievirus (cytomegalic virus) - CsA Cyclosporin A - Ext. Extensive Form - GKB Ganzkörperbestrahlung - GvHD Graft-versus-Host-Erkrankung - HLA Human leukocyte antigen - HUS Hämolytisch-urämisches Syndrom - IP Idiopathische interstitielle Pneumonie - KMT Knochenmarktransplantation - Lim. Limitierte Form - MLC Mixed lymphocyte culture - Mtx Methotrexat - OPSI Overwhelming-postsplenectomy-syndrome - SAA Schwere aplastische Anämie - SP Survival probability - SR Survival rate - TR Teilremission - ÜR Überlebensrate - ÜW Überlebenswahrscheinlichkeit - VR Vollremission - VZV Varizella-Zoster-Virus Herrn Prof. Dr. H.D. Waller zum 60. Geburtstag gewidmet     

Lung diseases after bone marrow transplantation

Summary The case histories of 72 subsequently treated patients — 44 with acute leukemia, 10 with chronic myeloid leukemia, 16 with severe aplastic anemia and 2 with neuroblastoma — were analyzed after bone marrow transplantation (BMT) with respect to pulmonary diseases. Thirty-eight patients suffered from a total of 51 pulmonary complications, which led to death in 20. Of 13 patients, 3 died of bacterial pneumonia, all of them during granulocytopenia; 2 of 6 patients died of fungal pneumonia and 2 out of 3 of a mixed bacterialmycotic infection. Adult respiratory distress syndrome (ARDS) led to death in 2 patients. A granulocyte count under 500/µl correlated significantly (P<0.002) with the fatal outcome of bacterial, fungal and ARDS pneumonia as well as with bronchitis. Viral pneumonia led to death in 8 of 9 patients; in each there was a significant correlation (P<0.05) with graft-versus-host disease (GvHD). Patients with repeated episodes of pulmonary illness had significantly more chronic GvHD (P<0.05); several of these patients displayed a reduction in helper T cells and an increase in suppressor T cells in the peripheral blood. The natural killer (NK) cells were reduced and the percentage of activated NK cell level lay between 6% and 69%. B-cells were absent or deficient. These findings explain in part the absence of specific antibody reactivity. Five of these patients also contracted GvHD-associated obstructive bronchiolitis, which did not respond to therapy. Pulmonary infiltrates of unknown origin (including idiopathic interstitial pneumonia) occurred in 8 of the patients (11.1%), with a fatal outcome in 3 patients. Significant changes (P<0.05) in lung function after BMT appeared in the form of reduced vital capacity (VC) increased residual volume (RV) and an increase in RV expressed as the percentage of total lung capacity. Pulmonary diseases were the most common complication and cause of death in our patients after BMT.Abbreviations AEL acute erythroid leukemia - AHTCG anti-human T-cell globulin - ALL acute lymphoblastic leukemia - AML acute myeloid leukemia - ARDS adult respiratory distress syndrome - AUL acute undifferentiated leukemia - BMT bone marrow transplantation - CR complete remission - CML chronic myeloid leukemia - CMV cytomegalovirus - Ext. extensive - GvHD graft versus host disease - Lim. limited - PR partial remission - Rel. relapse - SAA severe aplastic anemia - VZV varicella zoster virus.Lung function tests - DLCO single-breath CO-diffusion capacity - FEV₁ forced expiratory volume in 1 s - KCO Krogh's constant - MBC maximal breathing capacity - RAW airway resistance - RV residual volume - SRAW specific airway resistance - TGV thoracic gas volume - TLC total lung capacity - VC vital capacity Supported in part by SFB 120, A2, B1, C1, F2     

The procoagulant activity of human granulocytes,lymphocytes and monocytes stimulated by endotoxin

Summary Leukocytes from donor blood were separated by Ficoll/Urovison density centrifugation into granulocytes, lymphocytes and monocytes. The cell fractions were suspended in a culture medium to which endotoxin of Salmonella enteritidis was added at a final concentration of 10 µg/ml. Endotoxin-stimulated monocytes developed a very high tissue factor (thromboplastin) activity while in granulocytes an only negligible amount of tissue factor activity was detectable. The tissue factor activity measured in the preparation of the lymphocytes can be explained by contamination with monocytes. Electron microscopic studies showed the lysosomes of all monocytes to be enlarged and activated. Only a fraction of the granulocytes appeared degranulated with prominent vacuoles containing inclusion bodies. Possibly the high tissue factor activity of the monocytes triggers the development of the disseminated intravascular coagulation in the Shwartzman phenomenon.These investigations were supported in part by DFG Hi 224/1     

Genome-wide methylation profiling of glioblastoma cell-derived extracellular vesicle DNA allows tumor classification

BackgroundGenome-wide DNA methylation profiling has recently been developed into a tool that allows tumor classification in central nervous system tumors. Extracellular vesicles (EVs) are released by tumor cells and contain high molecular weight DNA, rendering EVs a potential biomarker source to identify tumor subgroups, stratify patients and monitor therapy by liquid biopsy. We investigated whether the DNA in glioblastoma cell-derived EVs reflects genome-wide tumor methylation and mutational profiles and allows noninvasive tumor subtype classification.MethodsDNA was isolated from EVs secreted by glioblastoma cells as well as from matching cultured cells and tumors. EV-DNA was localized and quantified by direct stochastic optical reconstruction microscopy. Methylation and copy number profiling was performed using 850k arrays. Mutations were identified by targeted gene panel sequencing. Proteins were differentially quantified by mass spectrometric proteomics.ResultsGenome-wide methylation profiling of glioblastoma-derived EVs correctly identified the methylation class of the parental cells and original tumors, including the MGMT promoter methylation status. Tumor-specific mutations and copy number variations (CNV) were detected in EV-DNA with high accuracy. Different EV isolation techniques did not affect the methylation profiling and CNV results. DNA was present inside EVs and on the EV surface. Proteome analysis did not allow specific tumor identification or classification but identified tumor-associated proteins that could potentially be useful for enriching tumor-derived circulating EVs from biofluids.ConclusionsThis study provides proof of principle that EV-DNA reflects the genome-wide methylation, CNV, and mutational status of glioblastoma cells and enables their molecular classification.