Biomedical subjects

G Schuler

Publications and source records attributed to G Schuler.

At least 145 records · Page 8Linked to original sources

PubMed1999

Cloning, recombinant expression and biochemical characterization of the murine CD83 molecule which is specifically upregulated during dendritic cell maturation.

Human CD83 (hCD83) is a glycoprotein expressed predominantly on the surface of dendritic cells (DC) and represents the best marker for mature DC. Here, we report the cloning of the cDNA encoding mouse CD83 (mCD83) from a murine bone marrow-derived DC (BM-DC) cDNA library. DNA sequence analysis revealed a 196 amino acid protein including a signal peptide of 21 amino acids which shares 63% amino acid identity with hCD83. Using Northern blot analyses, mCD83 mRNA was found to be strongly expressed in mouse BM-DC and its expression was upregulated following stimulation with LPS or TNF-alpha. Transfection experiments using COS-7 cells revealed that mCD83 is glycosylated. Furthermore, the extracellular CD83 domain was recombinantly expressed in Escherichia coli and one-dimensional NMR data strongly support that the protein is structurally folded.

Amino Acid Sequence↗

PubMed1999

Catheter-based percutaneous myocardial laser revascularization in patients with end-stage coronary artery disease.

OBJECTIVES: This study evaluates the feasibility and safety of a catheter-based laser system for percutaneous myocardial revascularization and analyses the first clinical acute and long-term results in patients with end-stage coronary artery disease (CAD) and severe angina pectoris. BACKGROUND: In patients with CAD and intractable angina who are not candidates for either coronary artery bypass grafting (CABG) or percutaneous transluminal coronary angioplasty (PTCA), transmyocardial laser revascularization (TMR) has been developed as a new treatment that results in reduced angina pectoris and increased exercise capacity. However, surgical thoracotomy is required for TMR with considerable morbidity and mortality. METHODS: A catheter-based system has been developed that allows creation of laser channels in the myocardium from within the left ventricular cavity. Laser energy generated by a Holmium: YAG (Cardiogenesis Corporation, Sunnyvale, California) laser was transmitted to the myocardium via a flexible optical fiber capped by an optic lens. The optical fiber was maneuvered to the target area under biplane fluoroscopy through a coaxial catheter system permitting movement in three dimensions. RESULTS: Thirty-four patients with severe CAD not amenable to either CABG or PTCA and refractory angina pectoris (Canadian Cardiologic Society [CCS] Angina Scale Class III-IV) were included in the study. Ischemic regions were identified by coronary angiography and confirmed by thallium scintigraphy. The percutaneous myocardial revascularization (PMR) procedure was successfully completed in all patients. In 29 patients, one vascular territory of the left ventricle and in 5 patients, two vascular territories were treated. Eight to fifteen channels were created in each ischemic region. Major periprocedural complications were limited to an episode of arterial bleeding requiring surgical repair. There was one death early after PMR, due to a myocardial infarction (MI) in a nontreated region. Clinical follow-up at 6 months (17 patients) demonstrated significant improvement of angina pectoris (CCS class before PMR: 3.0+/-0.0, six months after PMR: 1.3+/-0.8, p<0.0001) and increased exercise capacity (exercise time on standard bicycle ergometry before PMR: 384+/-141 s, six months after PMR: 514+/-158 s, p<0.05), but thallium scintigraphy failed to show improved perfusion of the laser treated regions. CONCLUSIONS: Percutaneous myocardial revascularization is a new safe and feasible therapeutic option in patients with CAD and severe angina pectoris not amenable to either CABG or PTCA. Initial results show immediate and significant improvement of symptoms and exercise capacity but evidence of improved myocardial perfusion is still lacking.

Aged↗

PubMed1999

Human monocyte derived dendritic cells express functional P2X and P2Y receptors as well as ecto-nucleotidases.

We investigated the expression and function of P2 receptors and ecto-nucleotidases on human monocyte derived dendritic cells (DC). In addition we analyzed the effect of extracellular ATP on the maturation of DC. By RT-PCR, DC were found to express mRNA for several P2X (P2X1, P2X4, P2X5, P2X7) and P2Y (P2Y1, P2Y2, P2Y4, P2Y5, P2Y6, P2Y10, P2Y11) receptors. As shown by FURA-2 measurement, triggering of P2 receptors resulted in an increase in free intracellular Ca2+. In combination with Tumor necrosis factor-alpha, ATP increased the expression of the DC surface markers CD80, CD83 and CD86 indicating a maturation promoting effect. DC expressed the ecto-apyrase CD39 and the ecto-5'-nucleotidase CD73 as demonstrated by RT-PCR. Extracellular ATP was rapidly hydrolyzed by these ecto-enzymes as shown by separation of 3H-labeled ATP metabolites using a thin layer technique. These data suggest that ATP acts as a costimulatory factor on DC maturation.

5'-Nucleotidase↗

PubMed1999

M1204, a novel 2',5' oligoadenylate synthetase with a ubiquitin-like extension, is induced during maturation of murine dendritic cells.

A novel molecule expressed by spleen dendritic cells (DC) was isolated using a subtractive hybridization approach. The full-length M1204 clone has 3063 bp, with 1415 bp spanning a single open reading frame, coding for a protein of a predicted size of about 50 kDa. This sequence has strong homology to 2', 5' oligoadenylate synthetase and contains a ubiquitin-like domain. In Northern blot analyses the mRNA is strongly expressed in spleen DC, whereas, in bone marrow-derived DC, the amount of mRNA increases during the maturation process. None of the other leukocytes nor several hemopoietic cell lines tested express this mRNA, but clear expression occurs in many organs, the highest levels being in thymus, lung, and bone marrow. In situ hybridization, combined with immunocytochemical staining of tissue sections of lung and spleen, shows colocalization of M1204 with the 2A1 and NLDC DC markers. In Western blot experiments, an antiserum raised against the recombinant M1204 recognizes a single band in bone marrow-derived DC and in the lung. The expressed oligoadenylate synthetase domain is active in synthesizing 2',5' diadenylate, which by itself may inhibit viral protein synthesis and may also function as a substrate for 2',5' oligoadenylate synthetase. Since the oligoadenylate/RNase L system provides early protection against virus infection, we hypothesize that M1204 prevents virus-induced cell death in DC.

2',5'-Oligoadenylate Synthetase↗

PubMed1999

Apoptosis in skeletal myocytes of patients with chronic heart failure is associated with exercise intolerance.

OBJECTIVES: The purpose of the study was to investigate if apoptosis occurs in skeletal muscle myocytes and its relation to exercise intolerance in patients with chronic heart failure (CHF). BACKGROUND: Intrinsic abnormalities of skeletal muscle frequently limit exercise tolerance in CHF patients. Recently, apoptosis has been detected in cardiac myocytes of patients with CHF, suggesting that apoptosis may contribute to the reduced contractile force. The presence and regulation of apoptosis in skeletal myocytes of patients with CHF remains to be defined. METHODS: Skeletal muscle biopsies (m. vastus lateralis) of 34 CHF patients (New York Heart Association functional class II-III) and eight age-matched healthy control subjects were analyzed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling for the presence of apoptosis, and by immunohistochemistry and videodensitometrical quantification for inducible nitric oxide synthase (iNOS) and Bcl-2 expression. Maximal oxygen consumption (VO2max) was determined by ergospirometry. RESULTS: Apoptosis was detected in 16/34 (47%) patients with CHF and in none of the healthy subjects. Patients with apoptosis-positive skeletal muscle myocytes exhibited a significantly lower VO2max (12.0 +/- 3.7 vs. 18.2 +/- 4.4 ml/kg/min; p = 0.0005), a higher iNOS expression (6.8 +/- 3.6 vs. 3.7 +/- 2.6% iNOS-positive stained tissue area; p = 0.015) and a lower Bcl-2 expression (1.0 +/- 0.3 vs. 1.4 +/- 0.4% Bcl-2-positive tissue area; p = 0.03) as compared with patients with apoptosis-negative biopsies. CONCLUSIONS: These results indicate that apoptosis is frequently found in skeletal muscle obtained from CHF patients, which is associated with significant impairment of functional work capacity. In skeletal muscle of these patients, iNOS and Bcl-2 are possibly involved in the regulation of apoptosis.

Adult↗

PubMed1999

Interleukin-10-treated human dendritic cells induce a melanoma-antigen-specific anergy in CD8(+) T cells resulting in a failure to lyse tumor cells.

Dendritic cells (DC) are critically involved in the initiation of primary immune processes, including tumor rejection. In our study, we investigated the effect of interleukin-10 (IL-10)-treated human DC on the properties of CD8(+) T cells that are known to be essential for the destruction of tumor cells. We show that IL-10-pretreatment of DC not only reduces their allostimulatory capacity, but also induces a state of alloantigen-specific anergy in both primed and naive (CD45RA+) CD8(+) T cells. To investigate the influence of IL-10-treated DC on melanoma-associated antigen-specific T cells, we generated a tyrosinase-specific CD8(+) T-cell line by several rounds of stimulation with the specific antigen. After coculture with IL-10-treated DC, restimulation of the T-cell line with untreated, antigen-pulsed DC demonstrated peptide-specific anergy in the tyrosinase-specific T cells. Addition of IL-2 to the anergic T cells reversed the state of both alloantigen- or peptide-specific anergy. In contrast to optimally stimulated CD8(+) T cells, anergic tyrosinase-specific CD8(+) T cells, after coculture with peptide-pulsed IL-10-treated DC, failed to lyse an HLA-A2-positive and tyrosinase-expressing melanoma cell line. Thus, our data demonstrate that IL-10-treated DC induce an antigen-specific anergy in cytotoxic CD8(+) T cells, a process that might be a mechanism of tumors to inhibit immune surveillance by converting DC into tolerogenic antigen-presenting cells.

Antigen Presentation↗

PubMed1999

An advanced culture method for generating large quantities of highly pure dendritic cells from mouse bone marrow.

As dendritic cells (DC) are rare populations in all organs, their generation from hematopoietic precursors in large quantities has proven critical to study their biology. From murine bone marrow about 5 x 10(6) cells at 70% purity are obtained per mouse after 8 days of culture with GM-CSF. We have improved this standard method and routinely achieve a 50-fold higher yield, i.e., 1-3 x 10(8) immature and mature DC per mouse at 90-95% purity. The major modifications were: (i) the avoidance of any active depletion of bone marrow cell subpopulations to circumvent loss of precursors, (ii) a lower plating density of bone marrow cells, (iii) a prolonged culture period of 10-12 days, (iv) the reduction of the GM-CSF dose from day 8 or 10 onwards to reduce granulocyte contaminations. The final non-adherent population at day 10-12 constitutes a mixture of immature and mature DC. Further maturation of DC could be induced by high doses of LPS or TNF-alpha for the last 24 h, where 50-70% of the non-adherent fraction represented mature DC with high levels of NLDC-145, CD86 and CD40. This method allows by simple means the generation of high numbers of murine DC with very low B cell or granulocyte contaminations. It will be valuable to study DC biology notably at the molecular level.

Animals↗

PubMed1999

Generation of large numbers of fully mature and stable dendritic cells from leukapheresis products for clinical application.

Dendritic Cell (DC)-based vaccination approaches in man require a reproducible DC generation method that can be performed in conformity with GMP (Good Manufacturing Practice) guidelines and that circumvents the need for multiple blood drawings to generate DC. To this end we modified our previously described method to generate mature DC from CD14 + monocytes by a two step method (priming in GM-SF + IL-4 followed by maturation in monocyte conditioned medium) for use with leukapheresis products as a starting population. Several adaptations were necessary. We established, for example, a modified adherence step to reliably enrich CD14 + DC precursors from apheresis mononuclear cells. The addition of GM-CSF + IL-4 at the onset of culture proved disadvantageous and was, therefore, delayed for 24 h. DC development from apheresis cells occurred faster than from fresh blood or buffy coat, and was complete after 7 days. Monocyte conditioned medium when added on day 6 resulted in fully mature and stable DC (veiled, highly migratory and T cell sensitizing cells with a characteristic phenotype such as 85% CD83 + , p55/fascin + , CD115/M-CSF-R - , CD86 + ) already after 24 h. The mature DC progeny were shown to remain stable and viable if cultured for another 1-2 days in the absence of cytokines, and to be resistant to inhibitory effects of IL-10. Freezing conditions were established to generate DC from frozen aliquots of PBMC or to freeze mature DC themselves for later use. The approach yields large numbers of standardized DC (5-10 x 10(8) mature CD83 + DC/leukapheresis) that are suitable for performing sound DC-based vaccination trials that can be compared with each other.

Cell Adhesion↗

PubMed1999

The transcriptional program in the response of human fibroblasts to serum.

The temporal program of gene expression during a model physiological response of human cells, the response of fibroblasts to serum, was explored with a complementary DNA microarray representing about 8600 different human genes. Genes could be clustered into groups on the basis of their temporal patterns of expression in this program. Many features of the transcriptional program appeared to be related to the physiology of wound repair, suggesting that fibroblasts play a larger and richer role in this complex multicellular response than had previously been appreciated.

Blood↗

PubMed1999

A novel stent delivery catheter: the Intella Trinity balloon.

The Trinity catheter is a novel multiballoon device designed to facilitate stent delivery by providing for predilatation, stent deployment, and further expansion on a single shaft. The device was tested extensively in a canine model with a variety of stents before being used in a limited human feasibility trial in Europe. A description of the device and its initial human application is presented.

Animals↗

PubMed1999

[Ulcerative stomatitis as clinical clue to inadvertent methotrexate overdose].

Gastrointestinal side effects and the development of toxic liver fibrosis are well-known side effects of low dose methotrexate therapy. A female patient receiving a long term low dose methotrexate therapy for a seronegative chronic polyarthritis developed an ulcerative stomatitis, as clinical clue to an inadvertently given methotrexate overdose. A summary of other side effects of methotrexate and of oral side effects of other cytostatic agents and immunosuppressive drugs is given.

Aged↗

PubMed1999

[Digital ischemia as paraneoplastic marker of metastatic endometrial carcinoma].

Digital ischemia with gangrene of one or several fingertips has been described as a paraneoplastic syndrome associated with various malignant tumors, especially adenocarcinomas. Most often this paraneoplastic syndrome represents the first symptom of an occult neoplasia in an advanced stage. We present the case of an 83 year-old patient with digital ischemia and gangrenous fingertips in association with a latent adenocarcinoma of the uterus and metastatic involvement of the paraaortal lymph nodes. Acute occurrence of digital ischemia and gangrene without pathological laboratory findings and negative past medical history concerning cardiovascular-induced emboli, arteriosclerotic occlusion, or rheumatologic and autoimmune diseases should suggest this paraneoplastic syndrome and lead to thorough search for an underlying tumor.

Aged↗

PubMed1999

Tripolar acytokinetic mitosis and formation of feto-maternal syncytia in the bovine placentome: different modes of the generation of multinuclear cells.

The vast majority of trophoblast giant cells in the ruminant placenta are binuclear and are believed to derive from mononuclear trophoblastic cells by a single acytokinetic mitosis. There is no satisfactory explanation for the generation of the small proportion of trophoblast giant cells with one, three, or more nuclei. In this light-and electronmicroscopic study of bovine placentomal tissue from the second half of gestation, developmental stages of the trophoblast giant cells are investigated. Large mitotic figures indicate mitotic polyploidization, which is proposed to be due to two subsequent acytokinetic mitoses. Tripolar mitoses offer an explanation for the development of trinucleate trophoblast giant cells. Measurements of nuclear volumes in a series of semithin sections revealed that three size classes of trophoblast giant cells occur. The approximately doubling of nuclear volume between each class is thought to reflect different levels of DNA content that result from polyploidization in this cell type. Although trinuclear feto-maternal hybrid cells are the standard outcome of the fusion of binuclear trophoblast giant cells with uterine epithelial cells, some syncytia with at least five nuclei were observed in the uterine epithelium.

Animals↗

PubMed1999

Physical training as an adjunct therapy in patients with congestive heart failure: patient selection, training protocols, results, and future directions.

Exercise intolerance in patients with chronic heart failure (CHF) shows no correlation to the degree of left ventricular dysfunction. This surprising finding has directed attention to peripheral changes in CHF: reduced endothelium-dependent vasodilation and altered skeletal muscle metabolism. Physical exercise training has evolved as an important therapeutic approach to influence these noncardiac causes of exercise intolerance. It has been shown to enhance the oxidative capacity of the working skeletal muscle, to attenuate ergoreflex activity, to correct endothelial dysfunction, and to improve ventilation. All exercise-induced adaptations converge to increase peak oxygen uptake by up to 2 mL/kg.min(-1). Uncertainty remains concerning optimal patient selection, training protocol, and long-term effects on cardiac function. For patients experiencing stable CHF while on optimal cardiac medication, a combination of in-hospital and home-based aerobic endurance training in combination with local muscle strength training seems most promising. Although exercise training offers no causal treatment of CHF, it has great potential as an adjunct therapy directed at improving exercise tolerance and expanding the physical limits of CHF patients.

Arrhythmias, Cardiac↗

PubMed1999

Exercise intolerance in patients with chronic heart failure and increased expression of inducible nitric oxide synthase in the skeletal muscle.

OBJECTIVES: This study was designed to analyze the effect of iNOS on mitochondrial creatine kinase (mi-CK) expression and exercise capacity in chronic heart failure (CHF). BACKGROUND: The molecular mechanisms underlying exercise intolerance in CHF are still unclear. Expression of inducible nitric oxide synthase (iNOS) and reduced phosphocreatine resynthesis have been described in skeletal muscle of patients with CHF. However, it is unknown whether these phenomena are causally related to each other and to exercise tolerance. METHODS: Thirty-eight patients with CHF and 8 healthy controls (C) underwent bicycle ergospirometry and biopsy of the vastus lateralis muscle. Expression of iNOS was quantified by immunohistochemistry and reverse-transcriptase polymerase chain reaction, mi-CK by Western-blot. Intracellular presence of NO was confirmed by immunohistochemical quantification of nitrotyrosine (NT). To corroborate clinical findings, L6 rat skeletal myoblasts were incubated with sodium nitroprusside (SNP). RESULTS: Expression of iNOS was significantly increased in CHF (4.0+/-2.8 vs. 0.8+/-0.7% iNOS positive tissue area, p < 0.001 vs. C) and inversely correlated to maximal oxygen uptake (r=-0.65, p < 0.001). Intracellular NO-accumulation was confirmed by increased NT levels (13.5+/-8.5 vs. 2.0+/-1.7% NT-positive tissue area, p < 0.001 vs. C). Mi-CK was decreased in CHF (0.84+/-0.36 vs. 1.57+/-0.60, p < 0.001 vs. C). The inverse correlation seen between iNOS and mi-CK expression in patients (r=-0.68, p < 0.001) was reproduced in incubation experiments with SNP. CONCLUSIONS: Increased expression of iNOS in skeletal muscle of patients with CHF was inversely correlated with mi-CK expression and exercise capacity. Cell experiments confirmed a causal relationship via NO. These findings extend our knowledge of the pathophysiology of exercise intolerance in CHF.

Aged↗

PubMed1999

The human dendritic cell marker CD83 maps to chromosome 6p23.

The chromosomal localization of the human CD83 gene was determined using somatic cell hybrids, a radiation hybrid mapping panel and FISH analysis on human metaphase chromosomes. PCR-based analysis of a single chromosome hybrid panel identified the presence of the CD83 gene on human chromosome 6 and subsequent analysis of the Genebridge4 radiation panel located the gene between AFMa192wg9 and AFMb322wd1 with a lod score of 9.2. Finally, using FISH analysis the CD83 gene was localized to chromosome 6 band p23.

Antigens, CD↗

PubMed1999

Immunolocalization of progesterone receptors in bovine placentomes throughout mid and late gestation and at parturition.

The corpus luteum is the main source of progesterone (P(4)) responsible for maintenance of gestation in cattle. So far it has not been possible to assign any biological role to placental P(4), which contributes only marginally and temporarily to peripheral maternal blood levels. In order to identify possible P(4) target cells within the placenta, placentomes from 150-, 220-, 240-, and 270-day-pregnant cows and from parturient cows (3 animals per group) were screened immunohistochemically for expression of the progesterone receptor (PR). During gestation, PR-positive staining was found exclusively in the nuclei of caruncular stromal cells (CSC; maternal part of the placentome) and of caruncular vascular pericytes. In placentomes from parturient cows, occasional positive nuclear staining was also observed in the walls of small caruncular arteries. The percentage of PR-positive CSC increased slightly from 51.8 +/- 2.6% on Day 150 to 56.2 +/- 5.6% at Day 270 (p < 0.05) and was 58.9 +/- 1.8% at parturition. These results suggest that in pregnant cattle, CSC are under the control of P(4) of placental rather than luteal origin. Thus, whereas luteal P(4) may regulate "coarse" systemic progestational functions in the maternal compartment in the classical hormonal manner, placental P(4) may act as a paracrine factor involved in the local regulation of caruncular growth, differentiation, and functions.

Animals↗

PubMed1999

A resource of mapped human bacterial artificial chromosome clones.

To date, despite the increasing number of genomic tools, there is no repository of ordered human BAC clones that covers entire chromosomes. This project presents a resource of mapped large DNA fragments that span eight human chromosomes at approximately 1-Mb resolution. These DNA fragments are bacterial artificial chromosome (BAC) clones anchored to sequence tagged site (STS) markers. This clone collection, which currently contains 759 mapped clones, is useful in a wide range of applications from microarray-based gene mapping to identification of chromosomal mutations. In addition to the clones themselves, we describe a database, GenMapDB (http://genomics.med.upenn.edu/genmapdb), that contains information about each clone in our collection.

Chromosome Mapping↗