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Biomedical subjects

Christian Breymann

Publications and source records attributed to Christian Breymann.

13 recordsLinked to original sources
PubMed2006

Living autologous heart valves engineered from human prenatally harvested progenitors.

BACKGROUND: Heart valve tissue engineering is a promising strategy to overcome the lack of autologous growing replacements, particularly for the repair of congenital malformations. Here, we present a novel concept using human prenatal progenitor cells as new and exclusive cell source to generate autologous implants ready for use at birth. METHODS AND RESULTS: Human fetal mesenchymal progenitors were isolated from routinely sampled prenatal chorionic villus specimens and expanded in vitro. A portion was cryopreserved. After phenotyping and genotyping, cells were seeded onto synthetic biodegradable leaflet scaffolds (n=12) and conditioned in a bioreactor. After 21 days, leaflets were endothelialized with umbilical cord blood-derived endothelial progenitor cells and conditioned for additional 7 days. Resulting tissues were analyzed by histology, immunohistochemistry, biochemistry (amounts of extracellular matrix, DNA), mechanical testing, and scanning electron microscopy (SEM) and were compared with native neonatal heart valve leaflets. Fresh and cryopreserved cells showed comparable myofibroblast-like phenotypes. Genotyping confirmed their fetal origin. Neo-tissues exhibited organization, cell phenotypes, extracellular matrix production, and DNA content comparable to their native counterparts. Leaflet surfaces were covered with functional endothelia. SEM showed cellular distribution throughout the polymer and smooth surfaces. Mechanical profiles approximated those of native heart valves. CONCLUSIONS: Prenatal fetal progenitors obtained from routine chorionic villus sampling were successfully used as an exclusive, new cell source for the engineering of living heart valve leaflets. This concept may enable autologous replacements with growth potential ready for use at birth. Combined with the use of cell banking technology, this approach may be applied also for postnatal applications.

Absorbable Implants↗
PubMed2006

Engineered living blood vessels: functional endothelia generated from human umbilical cord-derived progenitors.

BACKGROUND: Tissue-engineered living blood vessels (TEBV) with growth capacity represent a promising new option for the repair of congenital malformations. We investigate the functionality of TEBV with endothelia generated from human umbilical cord blood-derived endothelial progenitor cells. METHODS: Tissue-engineered living blood vessels were generated from human umbilical cord-derived myofibroblasts seeded on biodegradable vascular scaffolds, followed by endothelialization with differentiated cord blood-derived endothelial progenitor cells. During in vitro maturation the TEBV were exposed to physiologic conditioning in a flow bioreactor. For functional assessment, a subgroup of TEBV was stimulated with tumor necrosis factor-alpha. Control vessels endothelialized with standard vascular endothelial cells were treated in parallel. Analysis of the TEBV included histology, immunohistochemistry, biochemistry (extracellular matrix analysis, DNA), and biomechanical testing. Endothelia were analyzed by flow cytometry and immunohistochemistry (CD31, von Willebrand factor, thrombomodulin, tissue factor, endothelial nitric oxide synthase). RESULTS: Histologically, a three-layered tissue organization of the TEBV analogous to native vessels was observed, and biochemistry revealed the major matrix constituents (collagen, proteoglycans) of blood vessels. Biomechanical properties (Young's modulus, 2.03 +/- 0.65 MPa) showed profiles resembling those of native tissue. Endothelial progenitor cells expressed typical endothelial cell markers CD31, von Willebrand factor, and endothelial nitric oxide synthase comparable to standard vascular endothelial cells. Stimulation with tumor necrosis factor-alpha resulted in physiologic upregulation of tissue factor and downregulation of thrombomodulin expression. CONCLUSIONS: These results indicate that TEBV with tissue architecture and functional endothelia similar to native blood vessels can be successfully generated from human umbilical cord progenitor cells. Thus, blood-derived progenitor cells obtained before or at birth may enable the clinical realization of tissue engineering constructs for pediatric applications.

Biomechanical Phenomena↗
PubMed2006

Variable efficacy of recombinant human erythropoietin in anemic pregnant women with different forms of heterozygous hemoglobinopathy.

OBJECTIVE: The aim of this study was to determine the response to recombinant human erythropoietin (rhEPO) in anemic pregnant women with heterozygous hemoglobinopathies. METHODS: A prospective study including 19 consecutive pregnant women with anemia and heterozygous hemoglobinopathy was performed. Treatment was divided into two phases: the initial low-dose phase and the subsequent high-rhEPO phase. In the initial phase, 3 x 10,000 U of rhEPO was administered with intravenous iron sucrose. In patients showing a poor response (Hb increase <1 g/dl) to low-dose rhEPO, the rhEPO dose was increased to 20,000 U per treatment in the subsequent phase. RESULTS: All patients showed stimulation of erythropoiesis as evidenced by an increase in hemoglobin. In 13 patients, a good response to therapy was observed (mean Hb increase 1.6 +/- 0.5 g/dl). In 6 patients, resistance to rhEPO was noted (mean Hb increase 0.5 +/- 0.5 g/dl). The mean gestational age at the start of therapy was 28 weeks of gestation and at the end 32 weeks. The mean duration of a complete therapy was 3.5 weeks (range 2-4.5 weeks). If calculated for body weight, the initial low- rhEPO dose of 160.4 +/- 30.6 U/kg body weight/treatment was increased to 320.9 +/- 61.2 U/kg body weight/treatment in the subsequent phase. CONCLUSION: Response to rhEPO treatment differs widely in anemic pregnant patients with heterozygous hemoglobinopathy. Resistance was observed in anemic pregnant patients with the beta-thalassemia trait originally from the Mediterranean region.

Adult↗
PubMed2005

Preserved placental oxygenation and development during severe systemic hypoxia.

Local tissue oxygenation profoundly influences placental development. To elucidate the impact of hypoxia on cellular and molecular adaptation in vivo, pregnant mice at embryonic days 7.5-11.5 were exposed to reduced environmental oxygen (6-7% O2) for various periods of time. Hypoxia-inducible factor (HIF)-1alpha mRNA was highly expressed in the placenta, whereas HIF-2alpha was predominantly found in the decidua, indicating that HIF-1 is a relevant oxygen-dependent factor involved in placental development. During severe hypoxia, HIF-1alpha protein was strongly induced in the periphery but, however, not in the labyrinth layer of the placenta. Accordingly, no indication for tissue hypoxia in this central area was detected with 2-(2-nitro-1H-imidazol-1-yl)-N-(2,2,3,3,3-pentafluoropropyl)acetamide staining and VEGF expression as hypoxic markers. The absence of significant tissue hypoxia was reflected by preserved placental architecture and trophoblast differentiation. In the search for mechanisms preventing local hypoxia, we found upregulation of endothelial nitric oxide synthase (NOS) expression in the labyrinth layer. Inhibition of NOS activity by N(omega)-nitro-L-arginine methyl ester application resulted in ubiquitous placental tissue hypoxia. Our results show that placental oxygenation is preserved even during severe systemic hypoxia and imply that NOS-mediated mechanisms are involved to protect the placenta from maternal hypoxia.

Animals↗
PubMed2005

Living patches engineered from human umbilical cord derived fibroblasts and endothelial progenitor cells.

OBJECTIVE: A major shortcoming in contemporary congenital heart surgery is the lack of viable replacement materials with the capacity of growth and regeneration. Here we focused on living autologous patches engineered from human umbilical cord derived fibroblasts and endothelial progenitor cells (EPCs) as a ready-to-use cell source for paediatric cardiovascular tissue engineering. METHODS: EPCs were isolated from 20 ml fresh umbilical cord blood by density gradient centrifugation and myofibroblasts were harvested from umbilical cord tissue. Cells were differentiated and expanded in vitro using nutrient media containing growth factors. Before seeding, cell-phenotypes were assessed by immuno-histochemistry. Biodegradable patches fabricated from synthetic polymers (PGA/P4HB) were seeded with myofibroblasts followed by endothelialization with EPCs. All patches were cultured in a perfusion bioreactor. A subgroup of patches was additionally stimulated by cyclic strain. Analysis of the neo-tissues comprised histology, immuno-histochemistry, extracellular matrix (ECM) analysis and biomechanical testing. RESULTS: Endothelial phenotypes of EPCs before seeding were confirmed by Ac-Dil-LDL, CD 31, von-Willebrand-Factor and eNOS staining. Histology of the seeded patches demonstrated layered viable tissue formation in all samples. The cells in the newly formed tissues expressed myofibroblast markers, such as desmin and alpha-SMA. The EPCs derived neo-endothelia showed constant endothelial phenotypes (CD 31, vWF). major constituents of ECM such as collagen and proteoglycans were biochemically detected. Stress-strain properties of the patches showed features of native-analogous tissues. CONCLUSIONS: Living tissue engineered patches can be successfully generated from human umbilical cord derived myofibroblasts and EPCs. This new cell source may enable the tissue engineering of versatile, living, autologous replacement materials for congenital cardiac interventions.

Biocompatible Materials↗
PubMed2005

Effect of altitude on thrombopoietin and the platelet count in healthy volunteers.

Although there is evidence that altitude increases the platelet count, its effect on the platelet precursor stimulating factor, thrombopoietin (TPO), is unclear. Unlike erythropoietin, TPO appears largely unresponsive to exogenous signals. In a study in 16 healthy volunteers, we report the effects of altitude exposure at between 1000 and 1822 m for 1 or 2 weeks on TPO, the platelet count (+ indices), erythropoietin, hemoglobin, hematocrit and erythrocytes (+ indices). There were significant post-exposure increases in TPO (57.9 vs 37.1 U/l; P=0.0006), platelet count (219.1 vs 208.0 x 10(3)/ml; P=0.031) and erythropoietin (16.1 vs 9.9 U/l; P=0.0032). There was a positive correlation between the increases in TPO and platelet count (r=0.52, P=0.043). Hemoglobin and hematocrit remained unchanged. Our results provide clear evidence for a relationship, presumably driven by hypoxia, between altitude exposure, TPO production and the platelet count.

Adult↗
PubMed2004

Umbilical cord blood derived endothelial progenitor cells for tissue engineering of vascular grafts.

BACKGROUND: A substantial limitation regarding present pediatric cardiac surgery is the lack of appropriate materials for the repair of congenital defects. To address this shortcoming, tissue engineering is a scientific field that aims at in vitro fabrication of living autologous grafts with the capacity of growth, repair, and regeneration. Here we focused on tissue engineered vascular grafts using human umbilical cord blood derived endothelial progenitor cells (EPCs), as a noninvasive cell source for pediatric applications. METHODS: EPCs were isolated from 20 ml fresh human umbilical cord blood by Ficoll gradient centrifugation and cultured in endothelial basal medium containing growth factors. After proliferation and differentiation cells were analyzed by immunohistochemistry and seeded onto three-dimensional (3D) biodegradable vascular scaffolds (porosity > 95%, n = 22). Twenty-four hours after seeding the vascular grafts were positioned into a pulse-duplicator-in vitro system and grown for 48 hours under biomimetic conditions. A second group was grown 6 days statically and an additional 6 days biomimetically. Controls were cultured statically. Analysis of the grafts included immunohistochemistry, histology, and scanning electron microscopy. RESULTS: Preseeding differentiated EPCs indicated constant endothelial phenotypes including acetylated low-density lipoprotein, cluster of differentiation 31, von Willebrand factor, and endothelial nitric oxide synthetase. Seeded EPCs established favorable cell-to-polymer attachment and proliferation into the 3D tubular scaffolds. Both conditioned and static cellular constructs demonstrated positive staining for cluster of differentiation 31, von Willebrand factor, and expression of endothelial nitric oxide synthase. CONCLUSIONS: Human umbilical cord derived EPCs indicated exceptional growth characteristics used for tissue engineering of vascular grafts. These cells demonstrated a constant endothelial phenotype and related functional features. Based on these results EPCs seem to be a promising autologous cell source with regard to cardiovascular tissue engineering, particularly for the repair of congenital defects.

Absorbable Implants↗
PubMed2004

Human umbilical cord cells for cardiovascular tissue engineering: a comparative study.

OBJECTIVE: Tissue engineering of viable, autologous cardiovascular replacements with the potential to grow, repair and remodel represents an attractive approach to overcome the shortcomings of available replacements for the repair of congenital cardiac defects. Currently, vascular myofibroblast cells represent an established cell source for cardiovascular tissue engineering. Cell isolation requires the invasive harvesting of venous or arterial vessel segments prior to scaffold seeding, a technique which may not be preferable, especially in pediatric patients. This study evaluates cells isolated from human umbilical cord artery, umbilical cord vein and whole cord as alternative autologous cell sources for cardiovascular tissue engineering. METHODS: Cells were isolated from human umbilical cord artery (UCA), umbilical cord vein (UCV), whole umbilical cord (UCC) and saphenous vein segments (VC), and were expanded in culture. All three expanded cell groups were seeded on bioabsorbable copolymer strips and grown in vitro for 28 days. Isolated cells were characterized by flow cytometry, histology, immunohistochemistry, proliferation assays and compared to VC. Morphological analysis of the seeded polymer strips included histology, immunohistochemistry, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transmission electron microscopy (TEM), scanning electron microscopy (SEM) and uniaxial stress testing. RESULTS: UCA, UCV and UCC demonstrated excellent cell growth properties comparable to VC. Following isolation, all three cell groups showed myofibroblast-like morphology and characteristics by staining positive for alpha-smooth muscle actin (ASMA) and vimentin. Histology and immunohistochemistry of seeded polymers showed good tissue and extracellular matrix formation containing collagen I, III and elastin. TEM showed viable myofibroblasts and the deposition of collagen fibrils and progressive growing tissue formation, with a confluent surface, was observed in SEM. No difference was found among the mechanical properties of UCA, UCV, UCC and VC tissue engineered constructs. CONCLUSIONS: Tissue engineering of cardiovascular constructs by using UCA, UCV and UCC is feasible in an in vitro environment. Cell growth, morphology, characteristics and tissue formation were comparable between UCA, UCV, UCC and VC. UCC represent an attractive, readily available autologous cell source for cardiovascular tissue engineering offering the additional benefits of utilizing juvenile cells and avoiding the invasive harvesting of intact vascular structures.

Absorbable Implants↗
PubMed2004

Sequence analysis of bile salt export pump (ABCB11) and multidrug resistance p-glycoprotein 3 (ABCB4, MDR3) in patients with intrahepatic cholestasis of pregnancy.

Intrahepatic cholestasis of pregnancy (ICP) is a liver disorder associated with increased risk of intrauterine fetal death and prematurity. There is increasing evidence that genetically determined dysfunction in the canalicular ABC transporters bile salt export pump (BSEP, ABCB11) and multidrug resistance protein 3 (MDR3, ABCB4) might be risk factors for ICP development. This study aimed to (i). describe the extent of genetic variability in BSEP and MDR3 in ICP and (ii). identify new disease-causing mutations. Twenty-one women with ICP and 40 women with uneventful pregnancies were recruited between April 2001 and April 2003. Sequencing of BSEP and MDR3 spanned 8-10 kb per gene and comprised the promoter region and 100-350 bp of the flanking intronic region around each exon. DNA sequencing of polymerase chain reaction fragments was performed on an ABI3700 capillary sequencer. MDR3 promoter activity of promoter constructs carrying different ICP-specific mutations was studied using reporter assays. A total of 37 and 51 variant sites were detected in BSEP and MDR3, respectively. Three non-synonymous sites in codons for evolutionarily conserved amino acids were specific for the ICP collective (BSEP, N591S; MDR3, S320F and G762E). Furthermore, four ICP-specific splicing mutations were detected in MDR3 [intron 21, G(+1)A; intron 25, G(+5)C and C(-3)G; and intron 26, T(+2)A]. Activity of the mutated MDR3 promoter was similar to that observed for the wild-type promoter. Our data further support an involvement of MDR3 genetic variation in the pathogenesis of ICP, whereas analysis of BSEP sequence variation indicates that this gene is probably less important for the development of pregnancy-associated cholestasis.

ATP Binding Cassette Transporter, Subfamily B↗
PubMed2002

Living, autologous pulmonary artery conduits tissue engineered from human umbilical cord cells.

BACKGROUND: Tissue engineering represents a promising approach to in vitro creation of living, autologous replacements with the potential to grow, repair, and remodel. Particularly in a congenital operation, there is a substantial need for such implantation materials. We previously demonstrated fabrication of completely autologous, functional heart valves on the basis of peripheral vascular cells. Presently the feasibility of creating pulmonary artery conduits from human umbilical cord cells was investigated. METHODS: Human umbilical cord cells were harvested and expanded in culture. Pulmonary conduits fabricated from rapidly bioabsorbable polymers were seeded with human umbilical cord cells and grown in vitro in a pulse duplicator bioreactor. Morphologic characterization of the generated neo-tissues included histology, transmission, and scanning electron microscopy. Characterization of extracellular matrix was comprised of immunohistochemistry. Extracellular matrix protein content and cell proliferation were quantified by biochemical assays. Biomechanical testing was performed using stress-strain and burst-stress tests. RESULTS: Histology of the conduits revealed viable, layered tissue and extracellular matrix formation with glycosaminoglycans and collagens I and III. Cells stained positive for vimentin and alpha-smooth muscle actin. Scanning electron microscopy showed confluent, homogenous tissue surfaces. Transmission electron microscopy demonstrated elements typical of viable myofibroblasts, such as collagen, fibrils, and elastin. Extracellular matrix proteins were significantly lower compared with native tissue; the cell content was increased. The mechanical strength of the pulsed constructs was comparable with native tissue; the static controls were significantly weaker. CONCLUSIONS: In vitro fabrication of tissue-engineered human pulmonary conduits was feasible utilizing human umbilical cord cells and a biomimetic culture environment. Morphologic and mechanical features approximated human pulmonary artery. Human umbilical cord cells demonstrated excellent growth properties representing a new, readily available cell source for tissue engineering without necessitating the sacrifice of intact vascular donor structures.

Absorbable Implants↗
PubMed2002

Human umbilical cord cells: a new cell source for cardiovascular tissue engineering.

BACKGROUND: Tissue engineering of viable, autologous cardiovascular constructs with the potential to grow, repair, and remodel represents a promising new concept for cardiac surgery, especially for pediatric patients. Currently, vascular myofibroblast cells (VC) represent an established cell source for cardiovascular tissue engineering. Cell isolation requires the invasive harvesting of venous or arterial vessel segments before scaffold seeding, a technique that may not be preferable, particularly in pediatric patients. In this study, we investigated the feasibility of using umbilical cord cells (UCC) as an alternative autologous cell source for cardiovascular tissue engineering. METHODS: Human UCC were isolated from umbilical cord segments and expanded in culture. The cells were sequentially seeded on bioabsorbable copolymer patches (n = 5) and grown in vitro in laminar flow for 14 days. The UCC were characterized by flow cytometry (FACS), histology, immunohistochemistry, and proliferation assays and were compared to saphenous vein-derived VC. Morphologic analysis of the UCC-seeded copolymer patches included histology and both transmission and scanning electron microscopy. Characterization of the extracellular matrix was performed by immunohistochemistry and quantitative extracellular matrix protein assays. The tissue-engineered UCC patches were biomechanically evaluated using uniaxial stress testing and were compared to native tissue. RESULTS: We found that isolated UCC show a fibroblast-like morphology and superior cell growth compared to VC. Phenotype analysis revealed positive signals for alpha-smooth muscle actin (ASMA), desmin, and vimentin. Histology and immunohistochemistry of seeded polymers showed layered tissue formation containing collagen I, III, and glycoaminoglycans. Transmission electron microscopy showed viable myofibroblasts and the deposition of collagen fibrils. A confluent tissue surface was observed during scanning electron microscopy. Glycoaminoglycan content did not reach values of native tissue, whereas cell content was increased. The biomechanical properties of the tissue-engineered constructs approached native tissue values. CONCLUSIONS: Tissue engineering of cardiovascular constructs using UCC is feasible in an in vitro environment. The UCC demonstrated excellent growth properties and tissue formation with mechanical properties approaching native tissue. It appears that UCC represent a promising alternative autologous cell source for cardiovascular tissue engineering, offering the additional benefits of using juvenile cells and avoiding the invasive harvesting of intact vascular structures.

Actins↗
PubMedDate not supplied

Iron deficiency and anaemia in pregnancy: modern aspects of diagnosis and therapy.

The prevalence of iron-deficiency anemia in different regions of the world ranges from 12 to 43%. The increased iron requirement in pregnancy and the puerperium carry with it an increased susceptibility to iron deficiency and iron-deficiency anemia and perioperative or peripartal blood transfusion. Prevention and correction presuppose reliable laboratory parameters and a thorough understanding of the mechanisms of iron therapy. The Hb level alone is insufficient to guide management. A complete work-up (ferritin, transferrin saturation) is essential, preferably with hematological indices such as hypochromic and microcytic red cells and reticulocytes, classified by degree of maturity, in particular, before parenteral therapy is given. Since ferritin acts as both an iron-storage and acute-phase protein, it cannot be used to evaluate iron status in the presence of inflammation. A high ferritin level thus requires the presence of an inflammatory process to be eliminated before it can be taken at face value. If the C-reactive protein level is also raised, the soluble TfR concentration can be used, since it is unaffected by inflammation. Inadequate understanding of the complex chemistry of parenteral iron administration was previously responsible for serious side effects, such as toxic and allergic reactions, and even anaphylactic shock, in particular with dextran preparations. However, the current type II iron complexes that release iron to the endogenous iron-binding proteins with a half-life of about 6 hours are not only effective but carry a minimal risk of allergic accident and overload, especially after a comprehensive pretreatment work-up. Our departmental data collected over 8 years and backed by postmarketing experience in 25 countries indicate that iron sucrose complex therapy is a valid first-line option for the safe and rapid reversal of iron-deficiency anemia.

Anemia↗