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Biomedical subjects

D Schlöndorff

Publications and source records attributed to D Schlöndorff.

At least 19 recordsLinked to original sources

Multipotent mesenchymal stem cells reduce interstitial fibrosis but do not delay progression of chronic kidney disease in collagen4A3-deficient mice.

Multipotent mesenchymal stem or stromal cells (MSC) have shown to improve outcome of acute renal injury models, but whether MSC can delay renal failure in chronic kidney disease is not known. We injected primary MSC or saline into mice that lack the alpha3-chain of type IV collagen (COL4A3), a model of chronic kidney disease with close similarities to human Alport disease. Weekly injections of MSC from week 6 to 10 of life prevented the loss of peritubular capillaries and reduced markers of renal fibrosis, that is, interstitial volume, numbers of smooth muscle actin-positive interstitial cells, and interstitial collagen deposits as compared to saline-injected COL4A3-deficient mice. However, renal function, that is, blood urea nitrogen, creatinine levels, proteinuria as well as survival of COL4A3-deficient mice were not affected by MSC injections. Although MSC were found to localize to kidneys of COL4A3-deficient mice after injection, differentiation into renal cells was not detected. However, MSC expressed growth factors, that is, vascular endothelial growth factor (VEGF) and bone morphogenetic protein-7 under basal culture conditions. In fact, VEGF mRNA levels were increased in kidneys of MSC-injected COL4A3-deficient mice and MSC supernatants enhance endothelial cell proliferation in vitro. Thus, weekly injections with MSC prevent loss of peritubular capillaries possibly owing to local production of growth factors rather than by differentiation into renal cells. The maintenance of interstitial vasculature is associated with less interstitial fibrosis but, is insufficient to delay renal failure and survival of COL4A3-deficient mice.

Animals↗

Progression of kidney disease: blocking leukocyte recruitment with chemokine receptor CCR1 antagonists.

Chronic kidney disease (CKD) is usually associated with interstitial leukocytic cell infiltrates, which may contribute to disease progression by production of proinflammatory, proapoptotic, and profibrotic mediators. Recruiting leukocytes into the kidney involves local expression of chemotactic cytokines, that is, chemokines, that interact with respective chemokine receptors on the leukocyte's outer surface. Thus, specific chemokine receptor antagonists may represent an attractive therapeutic concept to interfere with renal leukocyte recruitment. Among the proinflammatory chemokine receptors, chemokine receptor (CCR)-1 has nonredundant roles for leukocyte adhesion to activated vascular endothelium and for transendothelial migration. In fact, blocking CCR-1 with specific small-molecule antagonists was shown to retard progression in various types of rodent CKD models. Here we discuss the perspective of CCR-1 as a new potential target for the treatment of CKD.

Animals↗

Expression of the chemokine receptor CXCR1 in human glomerular diseases.

Leukocyte infiltration, a hallmark of renal diseases, is orchestrated in part by the actions of chemokines. The chemokine CXCL8/interleukin (IL)-8 is expressed during renal diseases and allograft rejection, whereas the corresponding receptor CXCR1 has not been described previously. Expression of CXCR1 was characterized in peripheral blood using multicolor fluorescence-activated cell sorter analysis (FACS). CXCR1 was localized in 81 formalin-fixed, paraffin-embedded renal specimens by immunohistochemistry using a monoclonal antibody against human CXCR1. Included were biopsies with crescentic glomerulonephritis (CGN, n = 22), immunoglobulin (Ig) A nephropathy (n = 15), membranoproliferative glomerulonephritis (MPGN, n = 17), lupus nephritis (n = 12), membranous nephropathy (n = 11), and non-involved parts of tumor nephrectomies (n = 4). Consecutive tissue sections of human tonsils, allograft explants, and renal biopsies were stained for CD15- and CD68-positive cells. Expression of CXCR1 and CXCL8/IL-8 mRNA was quantified by real-time reverse transcriptase-polymerse chain reaction of microdissected renal biopsies (n = 35) of the same disease entities. By FACS CXCR1 expression was found on polymorphonuclear CXCR1 expression by polymorphonuclear leukocytes (PMNs), natural killer cells, and a subpopulation of monocytes. By immunohistochemistry, CXCR1 expression was found on infiltrating inflammatory cells (predominantly PMNs), as well as on intrinsic renal cells (arterial smooth muscle cells, endothelial cells of peritubular capillaries). The distribution pattern of CXCR1 differed between disease entities. The highest numbers of glomerular CXCR1-positive cells were present in biopsies with MPGN, followed by lupus nephritis, and CGN. CXCR1 might be involved in the recruitment of PMNs to the glomerular tuft, which could be targeted by CXCR1-blocking agents.

Flow Cytometry↗

Expression of DARC, CXCR3 and CCR5 in giant cell arteritis.

OBJECTIVES: Leucocyte infiltration is the hallmark of vasculitis, chemokines being mainly responsible for leucocyte migration into inflamed tissues. The objective was to evaluate the local expression of chemokines and chemokine receptors in biopsies of patients with giant cell arteritis (GCA) compared with arteries from patients with polymyalgia rheumatica (PMR). We studied the expression of CCR5, CXCR3 and that of the Duffy antigen/receptor of chemokine (DARC), a chemokine internalizing receptor (interceptor), in parallel to the expression of the CCR5 ligand RANTES/CCL5. METHODS: Paraffin-embedded tissue sections from six patients with GCA and five patients with PMR were available for immunohistological analysis of chemokine receptor expression. RANTES/CCL5 mRNA was detected in tissue sections by in situ hybridization. RESULTS: In patients with biopsy-proven giant cell arteritis, CCR5 and CXCR3 were highly expressed by infiltrating leucocytes in involved tissue sections. Predominant clustering of CCR5+ and CXCR3+ leucocytes was found in the adventitia and was co-localized with the expression of CCL5/RANTES mRNA. Interestingly, we found marked expression of DARC on adventitial high endothelial venules in vasculitis lesions of patients with GCA, while in arteries from patients with PMR DARC was only expressed on a low number of vessels with flat lining endothelium. CONCLUSIONS: The co-localization of infiltrating CCR5+ and CXCR3+ leucocytes together with CCL5/RANTES and DARC in vasculitis lesions suggests a role for these chemokine receptors in leucocyte infiltration, possibly supported by DARC-mediated vascular presentation of chemokines.

Biomarkers↗

Modulation of HIV-1 enhancer activity and virus production by cAMP.

The effect of cAMP on the transcriptional activity of the HIV-1 long terminal repeat/enhancer was investigated and compared to the effect of cAMP on virus replication. In culture cAMP repressed virus replication in vivo using different cell types. Transient transfection studies with HIV-1 enhancer-derived luciferase reporter gene constructs identified the minimal DNA sequence mediating the negative regulatory effect of cAMP on HIV-1 transcription. A single nuclear factor kappaB element from the HIV-1 enhancer mediates the repressive effect on transcription. AP-2 is not involved in cAMP repression. Stable transfection of Jurkat T cells with the co-activators CREB binding protein (CBP) and p300 completely abolished the cAMP repressive effect, supporting the hypothesis that elevation of intracellular cAMP increases phosphorylation of CREB, which then competes with phosphorylated p65 and Ets-1 for limiting amounts of CBP/p300 thereby mediating the observed repressive effect on transcription. These findings suggest an important role of cAMP on HIV-1 transcription.

Binding Sites↗

CC chemokine receptor 5 and renal-transplant survival.

BACKGROUND: About 1% of white populations are homozygous carriers of an allele of the gene for the CC chemokine receptor 5 (CCR5) with a 32 bp deletion (CCR5Delta32), which leads to an inactive receptor. During acute and chronic transplant rejection, ligands for CCR5 are upregulated, and the graft is infiltrated by CCR5-positive mononuclear cells. We therefore investigated the influence of CCR5Delta32 on renal-transplant survival. METHODS: Genomic DNA from peripheral-blood leucocytes of 1227 renal-transplant recipients was screened by PCR for the presence of CCR5Delta32. Demographic and clinical data were extracted from hospital records. Complete follow-up data were available for 576 recipients of first renal transplants. Graft survival was analysed by Fisher's exact test and Kaplan-Meier plots compared with a log-rank test. FINDINGS: PCR identified 21 patients homozygous for CCR5Delta32 (frequency 1.7%). One patient died with a functioning graft. Only one of the remaining patients lost transplant function during follow-up (median 7.2 years) compared with 78 of the 555 patients with a homozygous wild-type or heterozygous CCR5Delta32 genotype. Graft survival was significantly longer in the homozygous CCR5Delta32 group than in the control group (log-rank p=0.033; hazard ratio 0.367 [95% CI 0.157-0.859]). INTERPRETATION: Patients homozygous for CCR5Delta32 show longer survival of renal transplants than those with other genotypes, suggesting a pathophysiological role for CCR5 in transplant loss. This receptor may be a useful target for the prevention of transplant loss.

Adult↗

Expression and characterization of the chemokine receptors CCR2 and CCR5 in mice.

The chemokine receptors CCR2 and CCR5 play important roles in the recruitment of monocytes/macrophages and T cells. To better understand the role of both receptors in murine models of inflammatory diseases and to recognize potential problems when correlating these data to humans, we have generated mAbs against murine CCR2 and CCR5. In mice CCR2 is homogeneously expressed on monocytes and on 2--15% of T cells, closely resembling the expression pattern in humans. In contrast to humans, murine NK cells are highly CCR5 positive. In addition, CCR5 is expressed on 3--10% of CD4 and 10--40% of CD8-positive T cells and is weakly detectable on monocytes. Using a model of immune complex nephritis, we examined the effects of inflammation on chemokine receptor expression and found a 10-fold enrichment of CCR5(+) and CCR2(+) T cells in the inflamed kidneys. The activity of various chemokines and the antagonistic properties of the mAbs were measured by ligand-induced internalization of CCR2 and CCR5 on primary leukocytes. The Ab MC-21 (anti-CCR2) reduced the activity of murine monocyte chemotactic protein 1 by 95%, whereas the Ab MC-68 (anti-CCR5) blocked over 99% of the macrophage-inflammatory protein 1alpha and RANTES activity. MC-21 and MC-68 efficiently blocked the ligand binding to CCR2 and CCR5 with an IC(50) of 0.09 and 0.6--1.0 microg/ml, respectively. In good correlation to these in vitro data, MC-21 almost completely prevented the influx of monocytes in thioglycollate-induced peritonitis. Therefore, both Abs appear as useful reagents to further study the role of CCR2 and CCR5 in murine disease models.

Animals↗

Depletion of CCR5-expressing cells with bispecific antibodies and chemokine toxins: a new strategy in the treatment of chronic inflammatory diseases and HIV.

The chemokine receptor CCR5 is expressed on the majority of T cells and monocytes in the inflammatory infiltrate of diseases such as rheumatoid arthritis, renal diseases, and multiple sclerosis. In contrast, little expression of CCR5 is found on peripheral blood leukocytes. A specific depletion of CCR5(+) cells could therefore be a useful strategy to reduce the cellular infiltrate in chronic inflammations. Moreover, CCR5 is the major coreceptor for M-tropic HIV-1 strains. Depletion of CCR5(+) leukocytes may help to eliminate cells latently infected with HIV-1. We designed two constructs that specifically destroy chemokine receptor-positive cells. The first construct, a bispecific Ab, binds simultaneously to CCR5 and CD3. Thereby it redirects CD3(+) T cells against CCR5(+) target cells. The Ab specifically depletes CCR5(+) T cells and monocytes, but is inactive against cells that do not express CCR5. Furthermore, ex vivo the bispecific Ab eliminated >95% of CCR5(+) monocytes and T cells from the synovial fluid of patients with arthritis. Also, we designed a fusion protein of the chemokine RANTES and a truncated version of Pseudomonas. exotoxin A. The fusion protein binds to CCR5 and down-modulates the receptor from the cell surface. The chemokine toxin completely destroyed CCR5(+) Chinese hamster ovary cells at a concentration of 10 nM, whereas no cytotoxic effect was detectable against CCR5(-) Chinese hamster ovary cells. Both constructs efficiently deplete CCR5-positive cells, appear as useful agents in the treatment of chronic inflammatory diseases, and may help to eradicate HIV-1 by increasing the turnover of latently infected cells.

ADP Ribose Transferases↗

Identification of new regulatory sequences far upstream of the mouse monocyte chemoattractant protein-1 gene.

We systematically searched for sequences influencing the expression of the mouse monocyte chemoattractant protein-1 (MCP-1) gene (Scya2) by mapping DNase I hypersensitive sites (HS) in the chromatin of mesangial cells in a 40-kb interval around the gene. We found nine HS located between -24 kb and +12.7 kb. Three HS coincided with previously known regulatory sequences (HS-2.4, HS-1.0, and HS-0.2). We tested two of the previously unknown HS located far upstream of Scya2 (HS-19.4 and HS-16.3) in transfection experiments using luciferase reporter constructs and mouse mesangial cells as recipients. In transient transfections, both HS had a moderate effect on basal promoter activity as well as promoter activity stimulated by tumor necrosis factor-alpha. In stable transfection experiments, we found much higher activity. A DNA fragment containing HS-19.4 and HS-16.3 caused a considerable increase in the number of stably integrated luciferase copies. We determined the nucleotide sequence of the 5' flanking region to -28.6 kb. Computer-assisted sequence analysis did not yield evidence of an additional gene. These HS are located within the 5' flanking region of a gene cluster consisting of Scya2 (MCP-1), Scya7 (MCP-3), Scya11 (eotaxin), Scya12 (MCP-5), and Scya8 (MCP-2). This report represents the first comprehensive chromatin analysis of the mouse MCP-1 locus leading to the identification of a complex regulatory region located far upstream of Scya2.

Animals↗

Surface expression of CC- and CXC-chemokine receptors on leucocyte subsets in inflammatory joint diseases.

Chemokine receptors play a crucial role in the recruitment of leucocyte subsets into inflamed tissue. Using FACS analysis we have studied the surface expression of different CC- and CXC-chemokine receptors on synovial fluid (SF) and peripheral blood leucocytes from 20 patients with various forms of arthritis. In the SF the majority T cells stained positive for CCR5 (93%) and CCR2 (57%), compared to the peripheral blood (36% and 25%). In addition, most of the T cells expressed CXCR4 in both compartments, with a somewhat higher percentage in the SF (90%) versus peripheral blood (83%). To date little information is available on chemokine receptor expression on monocytes in arthritis. We report a marked increase of CCR5(+) monocytes in the SF (87%) compared to the peripheral blood (22%). In contrast, the frequency of CXCR1(+), CXCR2(+), CXCR4(+) and CCR1(+) monocytes was considerably lower in the SF than in the peripheral blood. Moreover, we report the expression CXCR4 on neutrophils in the SF. Approximately 60% of neutrophils stained positive for CXCR4 in the SF, while in the peripheral blood the number of CXCR4(+) neutrophils was low (24%). Surface expression of CXCR1 and CXCR2 was significantly reduced on SF neutrophils (53% and 68%) compared to the peripheral blood. Chemokine receptors are differentially expressed on leucocyte subsets in arthritis. The identification of their pattern of expression might help to identify suitable targets for therapeutic intervention.

Alleles↗

Increased rate of renal transplant failure in patients with the G20210A mutation of the prothrombin gene.

In patients with thrombophilia caused by reduced physiological anticoagulation, renal transplant failure occurs more frequently. Previous studies showed the importance of the protein C system, a physiological anticoagulatory pathway that inhibits thrombus formation. However, excess activation of the hemostatic system also may result in thrombosis. The G20210A mutation in the prothrombin gene is such a prothrombotic risk factor that results in increased thrombus formation because of elevated factor II levels in plasma. We analyzed graft function in 270 consecutive patients who received 311 renal transplants. The presence of a normal or mutated prothrombin allele was determined by polymerase chain reaction amplification and restriction fragment length polymorphism analysis of genomic DNA. Demographic data were extracted from hospital records. Graft survival was calculated for patients with and without the G20210A mutation. We identified 9 patients heterozygous for the G20210A mutation in the prothrombin gene who had received a total of 12 renal transplants. Of these 12 transplants, 2 grafts were lost within the first year. Median graft survival for patients heterozygous for the 20210A allele was 65.9 months (range, 0 to 101 months) compared with 149 months (range, 0 to 237 months) for patients homozygous for the normal 20210 G allele (P = 0.02). The G20210A mutation represented a 2.95-fold (95% confidence interval, 1.03 to 8.46) increase in risk for graft loss. Only 1 patient with this mutation achieved graft function exceeding 101 months. The G20210A mutation of the prothrombin gene is an independent risk factor for graft failure.

Adult↗

A severe case of pneumatosis cystoides intestinalis with massive accumulation of gas outside the gastrointestinum.

We report an unusually severe case of pneumatosis cystoides intestinalis in an adult female patient undergoing chemotherapy for lymphoblastic crisis in chronic myelogenous leukemia. An impressive accumulation of gas was not only detected within the intestinal wall and mesentery but also in the peritoneum, retroperitoneum, mediastinum and cervical subcutaneous tissue. The patient was almost asymptomatic and fully recovered within a few days of treatment. In addition to chemotherapy, the patient was receiving trimethoprim/sulfamethoxazole for pneumocystis carinii pneumonia and lactulose for constipation. Mucosal damage due to chemotherapy and depletion of tetrahydrofolic acid, changes in bowel flora induced by antibiotic treatment, and the intake of unabsorbable carbohydrates may be responsible for the severity of the disease.

Aged↗

Chemokine-Induced secretion of gelatinase B in primary human monocytes.

Chemokines help control normal leukocyte trafficking as well as their infiltration into tissues during acute and chronic inflammation. Matrix metalloproteinases (MMPs) help support the extravasation and infiltration of leukocytes through limited proteolysis of basement membranes and matrix material. The effect of the chemokines RANTES/CCL5, MCP-1/CCL and SDF-1/CXCL12 on secretion of the matrix metalloproteinase B and its endogenous inhibitor TIMP-1 was studied. RANTES/CCL5 and SDF-1/CXCL12 were found to induce MMP-9 secretion in primary human monocytes while TIMP-1 secretion was not affected. RANTES/CCL5 effects were mediated through CCR1 because the CCR1 antagonist BX471 was found to effectively block RANTES/CCL5-induced MMP-9 secretion.

Chemokines↗

[Patient medication counseling--patient counseling about discharge medication].

BACKGROUND: With discharge from hospital, the correct use of his drugs is the responsibility of the patient. The patient's compliance mainly affects the success of therapy. An essential requirement for good compliance is informing the patient properly about the purpose and correct use of his drugs. In the first pilot study in Germany on this subject, we investigated how the additional patient medication counseling by a clinical pharmacist prior to discharge improves the patient's knowledge of his drugs. STUDY DESIGN: During a 10-week period, 37 patients were randomly assigned to either an intervention group or a control group. Patients of the intervention group were verbally and in writing informed about the purpose and correct use of their drugs. The effect of the additional counseling was determined in both groups by a questionnaire at the time of discharge and a telephone interview 2 weeks later. CONCLUSION: Patient medication counseling by a clinical pharmacist prior to discharge improves the patient's knowledge of his drugs as an essential requirement of good compliance. Taking into consideration the economical consequences of noncompliance and the financial problems of the public health system it is advisable to establish this service in German hospitals.

Adult↗