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Biomedical subjects

B Schmitz

Publications and source records attributed to B Schmitz.

At least 109 records · Page 6Linked to original sources
PubMed1996

Interactions between endogeneous lectins and fucosylated oligosaccharides in megakaryocyte-dependent fibroblast growth of the normal bone marrow.

In order to elucidate the involvement of adhesion mechanisms in the process of megakaryocyte-dependent fibroblast growth, we applied BSA-coupled polymers of glucose, galactose, fucose, mannose, and several lectins (AAA, LCA, LTA, UEA-I) to cocultures of CD61 -positive (CD61+)/MACS-enriched megakaryocytes and human bone marrow fibroblasts. Fibroblast monocultures served as controls. After 6 days, glucose, as well as galactose-treated cultures showed a significant reduction of fibroblast growth in cocultures and fibroblast monocultures. In contrast, application of mannose caused no reducing effect on fibroblast numbers. Administration of fucose, AAA, LTA or UEA-I revealed a strong impairment of fibroblast growth in the megakaryocyte-fibroblast cocultures. Adhesion experiments using MACS-enriched, fluorescein-labelled megakaryocytes cultured in the presence of carbohydrates and lectins on a near-confluent layer of fibroblasts were additionally performed. Following fucose-BSA, alpha Fuc-1,2Gal beta-HSA or UEA-I treatment a significant reduction of megakaryocyte adhesion to the fibroblast layer could be observed. In the case of AAA a weak impairment of megakaryocyte adhesion could be noticed. Selective pretreatment of either fibroblasts or megakaryocytes with fucose-BSA or alpha Fuc-1,2Gal beta-HSA was consistent with the finding of a prominent involvement of fucosylated residues located on megakaryocytes in this interaction. In conclusion, our studies are in keeping with the assumption that fucosylated and fucose-binding structures are playing a key role in adhesion mechanisms between megakaryocytes and fibroblasts and thus influence significantly the megakaryocyte-dependent growth of bone marrow fibroblasts.

Adult↗
PubMed1995

O-linked N-acetylglucosamine is upregulated in Alzheimer brains.

We present evidence that the expression of the novel intracellular carbohydrate modification of proteins--O-glycosidically linked N-acetylglucosamine (O-GlcNAc)--is significantly upregulated in Alzheimer brains over that of age matched control brains. This increase is specific for proteins associated with the detergent insoluble cytoskeleton and not for proteins of the detergent soluble fraction and is not due to an increase in protein expression in this fraction. The possible involvement of abnormal phosphorylation in the disease state and the interplay between phosphorylation and the O-GlcNAc modification suggests that the increased level of intracellular O-GlcNAc expression may be implicated in the pathogenesis of Alzheimer's disease.

Acetylglucosamine↗
PubMed1995

Beta-amyloid precursor protein is modified with O-linked N-acetylglucosamine.

The beta-amyloid precursor protein (APP) has been implicated in the etiology of Alzheimer's disease (Kang et al.: Nature 325:733-736, 1987; Selkoe: Science 248:1058-1060, 1990; Selkoe: In Cowan et al. (eds): "Annual Review of Neuroscience." Palo Alto, CA: Annual Reviews, Inc., pp 489-519, 1994) and numerous studies have shown that beta-amyloid is involved in amyloid plaque formation (Rumble et al.: N Engl J Med 320:1446-1452, 1989; Sisodia et al.: Science 248: 492-495, 1990). Evidence is presented that APP is modified with N-acetylglucosamine linked to cytoplasmic serine or threonine residues (O-GlcNAc). This is the first report of a plasma membrane protein modified with this carbohydrate. It has been postulated that this modification, which is ubiquitous in all organisms studied to date except bacteria (Haltiwanger et al.: Biochem Soc Trans 20:264-269, 1992; Dong et al.: J Biol Chem 268:16679-16687, 1993; Elliot et al.: J Neurosci 13:2424-2429, 1993; Kelly et al.: J Biol Chem 268:10416-10424, 1993), may function as an alternative to phosphorylation (Dong et al., 1993) and is involved in the multimerization of proteins (Haltiwanger et al., 1992; Dong et al., 1993). O-GlcNAc occurs at "PEST" sequences (Rogers et al.: Science 234:364-368, 1986) and it has been suggested that this modification within such a sequence leads to increased proteolytic stability of the molecule (Dong et al., 1993).(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylglucosamine↗
PubMed1995

Up-regulation of a chondroitin sulphate epitope during regeneration of mouse sciatic nerve: evidence that the immunoreactive molecules are related to the chondroitin sulphate proteoglycans decorin and versican.

After transection of adult mouse sciatic nerve, the expression of a chondroitin sulphate epitope recognized by the monoclonal antibody 473-HD (mAb 473-HD) was found to be up-regulated. The epitope was localized immunocytochemically mainly in Schwann cell basal laminae and, more weakly, also in the endoneurium. In cultures of mouse dorsal root ganglion cells, Schwann cells expressed high levels but fibroblasts only low levels of the epitope. To identify the molecule(s) carrying this chondroitin sulphate epitope, human sciatic nerves were extracted with phosphate-buffered saline and shown to contain two chondroitin sulphate proteoglycans of apparent molecular weights of 130 and 900 kDa. The 900 kDa and, more weakly, the 130 kDa proteoglycan were reactive with mAb 473-HD, which was found to recognize chondroitin-6-sulphate as epitope. Following chondroitinase ABC treatment of the 130 kDa proteoglycan, a core protein of approximately 45 kDa was seen and shown to react with polyclonal antibodies against the chondroitin-dermatan sulphate proteoglycan decorin from human fibroblasts. Chondroitinase ABC treatment of the 900 kDa proteoglycan yielded a core protein with a molecular weight of approximately 400 kDa that was recognized by polyclonal antibodies against recombinantly expressed fusion proteins from human versican. After transection of adult mouse sciatic nerves, the distal nerve stumps showed up-regulation of the chondroitin-6-sulphate epitope of the 900 kDa proteoglycan, whereas the core protein of this proteoglycan did not show any detectable change in the level of expression. In contrast, the core protein of the 130 kDa proteoglycan was up-regulated in expression. These observations suggest that versican- and decorin-like molecules may contribute to successful regeneration in the peripheral nervous system of mammals.

Animals↗
PubMed1995

Functional involvement of sciatic nerve-derived versican- and decorin-like molecules and other chondroitin sulphate proteoglycans in ECM-mediated cell adhesion and neurite outgrowth.

We have previously described two proteoglycans from human sciatic nerve which are immunochemically related to the chondroitin sulphate proteoglycans versican and decorin. The chondroitin sulphate of the versican-like molecule and the core protein of the decorin-like molecule have been found previously to be up-regulated after lesioning the adult mouse sciatic nerve. To investigate if the versican- and decorin-like molecules are involved in cell-extracellular matrix interactions, we studied the effect of both molecules on cell adhesion. The versican- and decorin-like molecules, substrate-coated in a mixture with fibronectin, but not with laminin or collagen types I or IV, inhibited the adhesion of several cell lines, neonatal dorsal root ganglion neurons and Schwann cells. The inhibitory activity was concentration-dependent and mediated by the chondroitin sulphate. Furthermore, when different proteoglycans were incubated with fibronectin, only the versican- and decorin-like molecules and the chondroitin sulphate proteoglycan aggrecan, but not the heparan sulphate proteoglycan perlecan, were able to inhibit fibronectin-mediated cell adhesion. The versican- and decorin-like molecules, substrate-coated alone or in a mixture with fibronectin or laminin, were at most slightly inhibitory to neurite outgrowth from PC12 phaeochromocytoma cells and neonatal dorsal root ganglion neurons. In a solid-phase ligand-binding assay the versican- and decorin-like molecules interacted with fibronectin, but not with laminin or collagen types I and IV. Binding of the versican-like molecule to fibronectin and inhibition of cell adhesion by this molecule was mediated via the heparin and cell-binding domains of fibronectin. These observations suggest that binding of the two proteoglycans to fibronectin is involved in the modulation of adhesion of cells to fibronectin.

Animals↗
PubMed1995

Comparison of valvular resistance, stroke work loss, and Gorlin valve area for quantification of aortic stenosis. An in vitro study in a pulsatile aortic flow model.

BACKGROUND: Valvular resistance and stroke work loss have been proposed as alternative measures of stenotic valvular lesions that may be less flow dependent and, thus, superior over valve area calculations for the quantification of aortic stenosis. The present in vitro study was designed to compare the impacts of valvular resistance, stroke work loss, and Gorlin valve area as hemodynamic indexes of aortic stenosis. METHODS AND RESULTS: In a pulsatile aortic flow model, rigid stenotic orifices in varying sizes (0.5, 1.0, 1.5 and 2.0 cm2) and geometry were studied under different hemodynamic conditions. Ventricular and aortic pressures were measured to determine the mean systolic ventricular pressure (LVSPm) and the transstenotic pressure gradient (delta Pm). Transvalvular flow (Fm) was assessed with an electromagnetic flowmeter. Valvular resistance [VR = 1333.(delta Pm/Fm)] and stroke work loss [SWL = 100.(delta Pm/LVSPm)] were calculated and compared with aortic valve area [AVA = Fm/(50 square root of delta Pm)]. The measurements were performed for a large range of transvalvular flows. At low-flow states, flow augmentation (100-->200 mL/s) increased calculated valvular resistance between 21% (2.0 cm2 orifice) and 66% (0.5-cm2 orifice). Stroke work loss demonstrated an increase from 43% (2.0 cm2) to 100% (1.0 cm2). In contrast, Gorlin valve area revealed only a moderate change from 29% (2.0 cm2) to 5% (0.5 cm2). At physiological flow rates, increase in transvalvular flow (200-->300 mL/s) did not alter calculated Gorlin valve area, whereas valvular resistance and stroke work loss demonstrated a continuing increase. Our experimental results were adopted to interpret the results of three clinical studies in aortic stenosis. The flow-dependent increase of Gorlin valve area, which was found in the cited clinical studies, can be elucidated as true further opening of the stenotic valve but not as a calculation error due to the Gorlin formula. CONCLUSIONS: Within the physiological range of flow, calculated aortic valve area was less dependent on hemodynamic conditions than were valvular resistance and stroke work loss, which varied as a function of flow. Thus, for the assessment of the severity of aortic stenosis, the Gorlin valve area is superior over valvular resistance and stroke work loss, which must be indexed for flow to adequately quantify the hemodynamic severity of the obstruction.

Aortic Valve↗
PubMed1995

Secretion of cytokines (interleukins-1 alpha, -3, and -6 and granulocyte-macrophage colony-stimulating factor) by normal human bone marrow megakaryocytes.

The effects of cytokine stimulation [recombinant human interleukin (rhIL)-1 alpha, rhIL-3, rhIL-6, rhIL-11, and rh granulocyte-macrophage colony-stimulating factor (GM-CSF)] on the secretory activity of normal human megakaryocytes were studied by means of the reverse hemolytic plaque assay (RHPA) in enriched cell preparations. This test facilitates an extremely sensitive determination of cytokine secretion at the single-cell level, together with the clear-cut identification of each immunostained (CD61) secretory active megakaryocyte. Moreover, the reverse transcriptase-polymerase chain reaction (RT-PCR) was used to investigate the expression of IL-6, IL-6 receptor (IL-6R), IL-9, IL-10, IL-12, and IL-13 mRNA in highly concentrated megakaryocyte preparations. In comparison with the spontaneous secretion rate, stimulation with rhIL-1 alpha, rhIL-6, and rhGM-CSF failed to induce a significant increase in the release of cytokines by CD61+ cells. On the other hand, both rhIL-3 and, in a less pronounced way, rhIL-11 exerted a marked effect on IL-6 secretion. Additionally, after stimulation with rhIL-3, a significant enhancement of the secretion of IL-3 and GM-CSF, but not of IL-1 alpha, could be observed. Using the RT-PCR, a significant induction of IL-6 expression could be appreciated in the enriched megakaryocyte population (60% to 80%) stimulated with rhIL-3. The results of this study provide persuasive evidence that a number of cytokines are synthesized and secreted by human megakaryocytes and not only by hematopoietic stroma cells. These data suggest the existence of autocrine and paracrine mechanisms that may influence maturation and differentiation of megakaryocytes as well as act on various stroma cells to sustain an appropriate hematopoietic micro-environment.

Base Sequence↗
PubMed1995

The phenotypic spectrum related to the human epilepsy susceptibility gene "EJM1".

Linkage studies of families ascertained through patients with juvenile myoclonic epilepsy (JME) suggest that an HLA-linked susceptibility gene on chromosome 6, designated "EJM1," predisposes to a group of idiopathic generalized epilepsies (IGEs) comprising JME, juvenile absence epilepsy (JAE), childhood absence epilepsies (CAE), and epilepsies with generalized tonic-clonic seizures (GTCS). To explore the EJM1-related phenotypic spectrum, we conducted linkage studies with HLA-DQ alpha restriction fragment length polymorphisms in 44 families ascertained through patients with CAE or JAE. Our results for the entire group of families provide evidence against a major susceptibility locus for idiopathic absence epilepsies and broader spectra of IGEs in the HLA region. Lod scores less than -2 were obtained for a region from 10 cM up to 23 cM on either side of the HLA-DQ alpha locus, depending on the assumed trait model. Suggestive evidence for linkage was found only for a subgroup of families with JME patients assuming an autosomal dominant mode of inheritance with 70% penetrance. A maximum lod score was obtained when family members with JME, JAE, CAE, and idiopathic GTCS were included into the affection status. Our results demonstrate that (1) the genetic susceptibility to idiopathic absence epilepsies and broader spectra of IGEs is heterogeneous, (2) the gene effect of EJM1 depends on the familial genetic background, and (3) EJM1 confers genetic susceptibility to idiopathic absence epilepsies and broader spectra of IGEs in the presence of family members with JME.

Disease Susceptibility↗
PubMed1995

Evolution of acute focal cerebral ischaemia in rats observed by localized 1H MRS, diffusion-weighted MRI, and electrophysiological monitoring.

Focal cerebral ischaemia was produced in 11 rats by permanent occlusion of the right middle cerebral artery (MCA) using a suture model modified to enable manipulation with the animals in situ in an NMR spectrometer. The development of the ischaemic insults and the resultant infarcts were observed for up to 6 h by localized 1H MRS and diffusion-weighted MRI while performing continuous monitoring of electroencephalogram and extracellular DC potential. The ischaemic areas were depicted as regions of hyperintensity in the diffusion-weighted images. Signals due to lactate became visible in the 1H spectra after MCA occlusion indicating the onset of anaerobic glycolysis. A depletion of N-acetylaspartate was seen in all animals post-occlusion. Transient or stepwise increases of lactate were observed to occur coincidentally with the events of spontaneous transient peri-infarct depolarization detected by the electrophysiological measurements. Expansion of the ischaemic area delineated in the diffusion-weighted images also accompanied peri-infarct depolarizations. These observations are consistent with transient peri-infarct depolarization playing a role in the growth of infarcts.

Acute Disease↗
PubMed1995

Hydrodynamic orientation of crayfish (Procambarus clarkii) to swimming fish prey.

Reversibly blindfolded crayfish (Procambarus clarkii) react to small swimming fish (Astyanax fasciatus mexicanus) approaching or passing nearby with antennal and cheliped movements and body turns (Fig. 3). We studied the accuracy and dynamics of crayfish orientation responses to the previously analyzed hydrodynamic disturbances caused by the fish, mostly produced by tail flicks. Antennal and cheliped movements started slightly before the onset of turning responses (Fig. 4). Antennal sweeps were performed most rapidly. 50% of the appendage sweeps resulted in contacts with the fish (Fig. 5). Most turns were directed toward the stimulus (Fig. 6). Response amplitudes increased with increasing stimulus angle. Turns were accurate for small stimulus angles, but smaller than expected for larger ones. Sweeps of ipsilateral antennae and chelipeds were generally directed backwards, while those of contralateral appendages were smaller and directed forwards. The amplitudes of appendage sweeps first increased with increasing stimulus angle and then decreased again for more caudal stimulus directions. Lateral stimuli (60 degrees-120 degrees) from opposite sides were usually significantly distinguished. The amplitudes of the different elements of orientation behaviour were highly correlated with each other, indicating that they were directed by the same sensory input.

Animals↗
PubMed1995

[The efficiency of emergency therapy in patients with head-brain, multiple injury. Quality assurance in emergency medicine].

For cardio-pulmonary resuscitation there are standardized treatment concepts, but there have been few prospective investigations examining the efficacy of prehospital advanced trauma life support and its effect on the outcome in patients with severe head injury and multiple trauma treated within the German emergency system. The results of this study underline the importance of intensive prehospital treatment and highlight some problems that should be taken into account in future in the training of emergency physicians. METHODS. A total of 179 patients with cerebral trauma were investigated. Data obtained included demographic and logistic data of the patients and the emergency physicians, diagnoses and treatment at the scene of the accident and state of the patient on admission in each case. Having divided the patients into three groups by severity of the trauma, we distinguished between sufficient and insufficient treatment and assessed infusion therapy, ventilatory support, positioning and immobilization, and analgesic and sedative therapy. For statistical analysis of the data we used chi 2-test and Fisher's exact test. P < 0.05 was considered significant. RESULTS. There were 102 patients who had sustained a cerebral trauma without other life-threatening lesions (score 1), 40 with multiple trauma (score 2) and 37 with multiple trauma (score 3). On average 2.4 IV lines were established and the patients received 1186 +/- 765 cc of crystalloid in addition to 801 +/- 411 cc of colloid fluids. In all groups, patients who received adequate infusion therapy had a better outcome; even in the group with score 1 significantly fewer had a fatal outcome. In all, 167 (93%) patients had endotracheal tubes placed, and in 150 cases (84%) ventilatory therapy was considered sufficient. The proportion of score 1 patients with sufficient ventilatory support who had a fatal outcome was significantly lower than that in the group with insufficient treatment. In patients with multiple trauma we could not separate the benefits of sufficient respiratory therapy and infusion therapy. In only 54% of the cases a vacuum mattress was used and in only 41% the patients were positioned with the upper part of the body elevated by 30 degrees. These were 28 patients (16%) who received neither analgesics nor sedatives. Regardless of the quality of prehospital treatment of isolated head injury, a Glasgow Coma Scale (GCS) score lower than 5 involved a very high mortality and all patients with a GCS score of 9 or more survived. In the group with GCS scores between 5 and 8, however, significantly more of the patients who received adequate treatment survived (82.5% vs 40%). CONCLUSIONS. The present study confirms that sufficient advanced trauma life support can improve the outcome of trauma victims with cerebral trauma. Adequate infusion and respiratory therapy reduce the mortality among such patients significantly. In patients with multiple trauma a clear positive effect of generous infusion therapy also is evident. The clearest effect of sufficient prehospital treatment is seen in patients with isolated cerebral trauma and a GCS score between 5 and 8. These results demonstrate the importance of advanced trauma life support and show emphatically that the so-called scoop-and-run strategy should be abandoned when resources are available for extended preclinical emergency treatment. On the other hand, we detected some problem areas in the prehospital treatment of trauma victims, such as positioning, immobilization and drug therapy with analgesics and sedatives. These findings allow us to pinpoint specific points that should be stressed in the training of emergency physicians and paramedics.

Adult↗
PubMed1995

Megakaryocytes and fibroblasts--interactions as determined in normal human bone marrow specimens.

An in vitro study was performed to investigate possible interactions between megakaryocytes and bone marrow fibroblasts, both obtained from healthy donors. We were able to demonstrate that the proliferation of fibroblasts increased significantly by co-culturing these cells with megakaryocytes for 6 days. Addition of neutralizing antibodies for PDGF and TGF beta 1, caused a significant reduction of fibroblast growth. Inhibition of cell to cell contacts via tissue culture inserts generated a conspicuous impairment of fibroblast proliferation compared with megakaryocyte-fibroblast co-cultures, where contact was allowed. Hence, our findings suggest that a close spatial relationship between megakaryocytes and fibroblasts is needed for the activation of growth in normal human bone marrow. Neighbouring of megakaryocytes and fibroblasts seems to be necessary in order to achieve a certain threshold of local growth factor concentration. Our results are in keeping with the assumption that PDGF and TGF beta 1, are secreted by normal human megakaryocytes in very low concentrations and promote significantly fibroblast proliferation.

Adult↗
PubMed1995

Resuscitation from cardiac arrest in cats: influence of epinephrine dosage on brain recovery.

The quality of brain recovery after cardiac arrest depends crucially on the speed of cardiac resuscitation because the low cerebral perfusion pressure during the resuscitation procedure facilitates the development of no-reflow. To accelerate return of spontaneous circulation, high dose epinephrine has been recommended but the effect on the dynamics of early brain recovery is still unknown. We, therefore, studied the dynamics of brain resuscitation after cardiopulmonary resuscitation (CPR) with standard and high dose epinephrine using non-invasive NMR techniques. Fifteen min cardiac arrest was induced in normothermic cats by ventricular fibrillation. CPR was performed using an inflatable pneumatic vest for cyclic chest compression. With the beginning of CPR the standard dose group received 0.02 mg/kg epinephrine (n = 6) and the high dose group received 0.2 mg/kg (n = 8). Brain recovery was monitored by magnetic resonance imaging of the apparent diffusion coefficient (ADC) of water for 3 h. Although high dose epinephrine treatment led to a significantly higher blood pressure during early reperfusion, rapidly changing heterogeneities of early brain recovery were observed in both groups. High dose epinephrine thus does not improve the quality of post-cardiac arrest brain recovery during the first 3 h of reperfusion.

Animals↗
PubMed1995

Imaging of the apparent diffusion coefficient for the evaluation of cerebral metabolic recovery after cardiac arrest.

The apparent diffusion coefficient (ADC) of water is a sensitive indicator of water and ion homeostasis of brain. Resuscitation of the brain after cardiac arrest, the most frequent reason for global cerebral ischemia under clinical conditions, depends critically on the reversal of disturbances of water and ion homeostasis. We, therefore, investigated whether ADC imaging can be used to monitor the development and reversal of ischemic brain injury during and after cardiac arrest. Ten adult mongrel normothermic cats were anesthetized with alfentanil and midazolam, immobilized with pancuronium, and mechanically ventilated with O2/N2O. Arterial, left ventricular, central venous, and intracranial pressures were monitored throughout the experiment. Magnetic resonance imaging was performed in a 4.7T MR scanner with a shielded gradient system. Diffusion-weighted images (DWI) were obtained by pulsed gradient spin echoes (diffusion weighting factor b: 0, 500, 1000, 1500 s/mm2). Quantitative ADC images were calculated from DWIs by fitting signal intensities against b-factors. Fifteen minute cardiac arrest was induced in the magnet by electrical fibrillation. Resuscitation was also carried out in the magnet, using a pneumatic vest for remotely controlled closed chest cardiac massage. Seven of 10 animals were resuscitated successfully and subsequently monitored for 3 h. During cardiac arrest, ADC declined from 678 +/- 79 x 10(-6) to 430 +/- 128 x 10(-6) mm2/s (63% of baseline). In the successfully resuscitated animals ADC returned to 648 +/- 108 x 10(-6) mm2/s within 30 min and remained at this level throughout the 3 h of recirculation. Regional evaluations of ADC revealed a transient overshoot in brainstem and basal ganglia to 114% of control at 15 min before returning to baseline values after 40 min. Failure of cardiac resuscitation prevented ADC normalization and led to its further decline to below 50% of control. Postcardiac arrest normalization of ADC maps correlated with homogeneous return of ATP, glucose, and lactate to near normal, whereas failure of ADC normalization was associated with depletion of ATP and glucose and severe lactate accumulation. In conclusion, our data indicate, that normalization of ADC is a reliable indicator of cerebral recovery after resuscitation from cardiac arrest.

Animals↗
PubMed1995

Carbamazepine and PIP-syndrome in temporal lobe epilepsy.

We describe four patients with left temporal lobe epilepsy who developed psychosis, intermittent polydipsia and hyponatremia (PIP-syndrome) while being treated with carbamazepine. These cases illustrate the close links between dysfunction of the limbic system causing epileptic seizures and psychosis on the one hand and dysregulation in hypothalamic centers manifestating in polydipsia and osmodysregulation on the other.

Anticonvulsants↗
PubMed1995

Influence of cytokines (IL-1 alpha, IL-3, IL-11, GM-CSF) on megakaryocyte-fibroblast interactions in normal human bone marrow.

The evolution of myelofibrosis accompanying chronic myeloproliferative disorders (CMPDs) is often linked with megakaryopoiesis. However, it is not known whether or to what extent megakaryocytes of normal human bone marrow are capable of stimulating fibroblast growth. For this reason, an in vitro study was performed to elucidate possible cytokine-dependent interactions between megakaryocytes and fibroblasts derived from healthy volunteers. Fibroblast growth was significantly promoted by the presence of megakaryocytes and modulated by additional application of various cytokines. While recombinant human (rh) interleukin (IL)-1 alpha had no obvious effect on fibroblast proliferation, a slight increase was detected on adding granulocyte-macrophage colony stimulating factor (rhGM-CSF). Application of rhIL-3 caused a significant increase in the number of fibroblasts. In contrast, administration of rhIL-11 suppressed the megakaryocyte-dependent growth-promoting effect and co-stimulation with rhIL-3 led to a significant decrease of fibroblast number in comparison to rhIL-3-stimulated co-cultures. Inhibition of cell-cell contact in unstimulated, as well as in rhIL-3-stimulated co-cultured led to a conspicuous impairment of fibroblast growth. A similar effect was observed when neutralizing antibodies directed against platelet-derived growth factor (PDGF) and transforming growth factor (TGF)beta 1 were added to rhIL-3-stimulated cultures. Our findings are in keeping with the assumption that interactions between megakaryocytes and fibroblasts involve in cytokine-mediated functional network regulated by factors such as spatial relationship, cytokine stimulation, and low concentrations of mediators, particularly PDGF and TGF beta. In this complex system rhIL-3 seems to play a crucial role in the promotion of these various interrelationships.

Bone Marrow↗
PubMed1995

The initiation of de novo methylation of foreign DNA integrated into a mammalian genome is not exclusively targeted by nucleotide sequence.

The de novo methylation of foreign DNA integrated into the mammalian genome is a fundamental process whose mechanism has not yet been elucidated. We have studied de novo methylation in adenovirus type 12 (Ad12) genomes inserted into the genomes of Ad12-induced hamster tumor cells. De novo methylation of Ad12 DNA, which is not methylated in the virion, is initiated in two paracentrally located regions and spreads from there across the integrated Ad12 genomes. (i) After extensive cultivation of cloned Ad12-induced hamster tumor cell lines, the same segments in integrated Ad12 DNA in different cell lines become methylated or remain unmethylated, depending on their positions in the viral genome. (ii) When Ad12 DNA or Ad12 DNA fragments are transfected into hamster cells and permanent cell lines are established by selection for the cotransfected neomycin phosphotransferase gene, patterns of de novo methylation in terminally or internally located segments of Ad12 DNA are different from those in Ad12-induced tumor cell lines. (iii) A detailed study on the topology of the integrated viral genomes in the Ad12-transformed hamster cell lines T637 and A2497-3 and in the Ad12-induced hamster tumors T191, T1111(1), and T181 has been performed. Some of the integrated viral genomes are inserted into the cellular genome in an orientation colinear with the virion genome; others have been rearranged. An originally internally located Ad12 DNA segment has become transposed to the left-terminal sequences of the viral genome in several cell lines and tumors. In the complete Ad12 genomes, the internally located PstI-D fragment becomes extensively methylated at the 5'-CCGG-3' and 5'-GCGC-3' sequences. When this DNA segment has been juxtaposed to the left-terminal, hypomethylated fragment of Ad12 DNA in rearranged genomes, the PstI-D fragment remains unmethylated. We therefore reason that the initiation of de novo methylation in integrated Ad12 DNA cannot be directed exclusively by the nucleotide sequence. Other parameters, such as site of integration, conformation of integrates, mode of cell selection, or chromatin structure related to transcriptional activity, may play decisive roles.

Adenoviridae↗