Wednesday, October 31, 2012

Ayman_Hyder - Universitätsklinikum Schleswig - Holstein · Klinik für Allgemeine Chirurgie (Kiel)


Universitätsklinikum Schleswig - Holstein · Klinik für Allgemeine Chirurgie (Kiel)            


  • Journal Article:EGF and HB-EGF enhance the proliferation of programmable cells of monocytic origin (PCMO) through activation of MEK/ERK signaling and improve differentiation of PCMO-derived hepatocyte-like cells.
    ABSTRACT:Hepatocyte-like cells (NeoHepatocytes) generated from a peripheral blood monocyte-derived stem cell-like cell (the PCMO) are a promising alternative for primary hepatocytes in cell transplantation studies to cure liver diseases. However, to be therapeutically effective NeoHepatocytes are needed in l... [more]Hepatocyte-like cells (NeoHepatocytes) generated from a peripheral blood monocyte-derived stem cell-like cell (the PCMO) are a promising alternative for primary hepatocytes in cell transplantation studies to cure liver diseases. However, to be therapeutically effective NeoHepatocytes are needed in large quantities. It was the aim of the present study to investigate i) whether the proportion of actively proliferating NeoHepatocytes can be enhanced by supplementing the PCMO differentiation medium (containing M-CSF, IL-3, and human serum) with either EGF or HB-EGF and ii) which signaling pathway underlies the promitotic effect. EGF and HB-EGF enhanced cell proliferation of PCMOs as demonstrated by increased expression of cycle control genes (ABL, ANAPC2, CDC2, CDK4, CDK6), phosphorylation of the retinoblastoma protein, and increased PCMO cell numbers after stimulation with EGF or HB-EGF. EGF also raised the number of monocytes expressing the proliferation marker Ki67. PCMOs expressed the EGF receptors EGFR (ERBB1) and ERBB3, and expression of both increased during PCMO generation. Phosphoimmunoblotting of PCMOs indicated that both EGF and HB-EGF activated MEK-1/2 and ERK1/2 in a concentration-dependent fashion with the effect of EGF being more prominent. EGF treatment further decreased expression of p47phox and increased that of Nanog indicating enhanced dedifferentiation and pluripotency, respectively. Treatment with both EGF and HB-EGF resulted in NeoHepatocytes with improved functional parameters. The results suggested that the addition of EGF or HB-EGF to PCMO differentiation medium superactivates MEK/ERK signaling which then increases both PCMO proliferation, number, and functional differentiation of PCMO-derived NeoHepatocytes.
    Cell communication and signaling : CCS. 08/2012; 10(1):23. · 5.50 Impact Factor
  • Journal Article:Peripheral Blood Monocytes Can Be Induced to Acquire Stem Cell-Like Properties
    ABSTRACT:Adult stem or programmable cells hold great promise in diseases in which damaged or non-functional cells need to be replaced. We have recently demonstrated that peripheral blood monocytes can be differentiated in vitro into cells whose phenotypes resemble specialized cell types like hepatocytes and ... [more]Adult stem or programmable cells hold great promise in diseases in which damaged or non-functional cells need to be replaced. We have recently demonstrated that peripheral blood monocytes can be differentiated in vitro into cells whose phenotypes resemble specialized cell types like hepatocytes and pancreatic beta cells. During phenotypic conversion the monocytes downregulate monocyte/macrophage differentiation markers being indicative of partial dedifferentiation and are partially reprogrammed to acquire a state of plasticity along with expression of various markers of pluripotency. These cells were termed “programmable cells of monocytic origin” (PCMOs). Current efforts focus on establishing culture conditions that increase both the plasticity and proliferation potential of PCMOs in order to be able to generate large amounts of blood-derived cells suitable for both autologous and allogeneic therapies
    Stem Cells and Cancer Stem Cells. 04/2012; 6(4):367-375.
  • Journal Article:Peptidoglycan recognition protein 3 (PglyRP3) has an anti-inflammatory role in intestinal epithelial cells.
    ABSTRACT:Intestinal epithelial cells produce cytokines in response to bacterial peptidoglycan (PGN), which is detected by several classes of pattern-recognition receptors (PRRs) as peptidoglycan recognition proteins (PGlyRPs), Toll-like receptor 2 (TLR2) and NOD receptors. All types of PGlyRPs recognize bact... [more]Intestinal epithelial cells produce cytokines in response to bacterial peptidoglycan (PGN), which is detected by several classes of pattern-recognition receptors (PRRs) as peptidoglycan recognition proteins (PGlyRPs), Toll-like receptor 2 (TLR2) and NOD receptors. All types of PGlyRPs recognize bacterial peptidoglycan and function in antibacterial innate immunity. In this study, we investigated the role of PGlyRP3 in the response of intestinal epithelial cells (Caco-2) to PGN from pathogenic (Staphylococcus aureus), opportunistic pathogenic (Micrococcus luteus) and non-pathogenic (Bacillus subtilis and Lactobacillus rhamnosus GG) bacteria found in the gut as commensals or in gastroenteritis. All PGNs induced the proinflammatory cytokines IL-12p35, IL-8 and TNF-α and, time-dependently, PGlyRP3, at both the transcription and protein levels. In this context, no differences were observed among the distinct PGN obtained from different bacterial sources. The inflammatory response to PGN is mediated via the TLR2 pathway, since blocking this pathway by inhibiting MyD88 reduced the expression of proinflammatory cytokines. In addition, PGlyRP3 overexpression suppressed, while PGlyRP3 knocking down enhanced the expression of PGN-induced inflammatory cytokines. It is concluded that PGN stimulates inflammatory responses in the intestinal epithelia through activation of the TLR pathway. PGlyRP3 is also stimulated by PGN and has, in contrast to activation of the TLR pathway, an anti-inflammatory effect.
    Immunobiology. 11/2011; 217(4):412-9. · 3.21 Impact Factor
  • Journal Article:Prebiotic oligosaccharides reduce proinflammatory cytokines in intestinal Caco-2 cells via activation of PPARγ and peptidoglycan recognition protein 3.
    ABSTRACT:Prebiotic oligosaccharides modulate the intestinal microbiota and beneficially affect the human body by reducing intestinal inflammation. This immunomodulatory effect was assumed to be bacterial in origin. However, some observations suggest that oligosaccharides may exert an antiinflammatory effect ... [more]Prebiotic oligosaccharides modulate the intestinal microbiota and beneficially affect the human body by reducing intestinal inflammation. This immunomodulatory effect was assumed to be bacterial in origin. However, some observations suggest that oligosaccharides may exert an antiinflammatory effect per se. We hypothesized that oligosaccharides affect the intestinal immunity via activation of peptidoglycan recognition protein 3 (PGlyRP3), which reduces the expression of proinflammatory cytokines. Caco-2 cells were treated with the oligosaccharides, α3-sialyllactose, or fructooligosaccharides (Raftilose p95), and the effects of these treatments on PGlyRP3 and PPARγ expression, the release and expression of some proinflammatory cytokines, and NF-κB translocation were tested. Both oligosaccharides had antiinflammatory activity; they significantly reduced IL-12 secretion in Caco-2 cells and gene expression of IL-12p35, IL-8, and TNFα. They also reduced the gene expression and nuclear translocation of NF-κB. Both oligosaccharides dose and time dependently induced the production of PGlyRP3, the silencing of which by transfection of Caco-2 cells with specific small interfering RNA targeting PGlyRP3 abolished the antiinflammatory role of both oligosaccharides. Incubation of Caco-2 cells with both oligosaccharides induced PPARγ. Antagonizing PPARγ by culturing the cells with GW9662 for 24 h inhibited the oligosaccharide-induced PGlyRP3 production and the antiinflammatory effect of the oligosaccharides. We conclude that oligosaccharides may exert an antiinflammatory effect by inducing the nuclear receptor PPARγ, which regulates the antiinflammatory PGlyRP3.
    The Journal of nutrition. 03/2011; 141(5):971-7. · 3.92 Impact Factor
  • Journal Article:PPARγ-dependent peptidoglycan recognition protein 3 (PGlyRP3) expression regulates proinflammatory cytokines by microbial and dietary fatty acids.
    ABSTRACT:PGlyRPs recognize bacterial peptidoglycan and function in antibacterial innate immunity. Focusing on the interference between nutrition and recognition pattern proteins, free fatty acids (FFA) of dietary and bacterial sources may exert their immunological response through modulating the expression l... [more]PGlyRPs recognize bacterial peptidoglycan and function in antibacterial innate immunity. Focusing on the interference between nutrition and recognition pattern proteins, free fatty acids (FFA) of dietary and bacterial sources may exert their immunological response through modulating the expression level of the PGlyRPs in enterocytes. PGlyRP3 was the only PGlyRPs member expressed in Caco2 cells. In silico analysis showed that the promoter of PGlyRP3 has some PPRE regions that, as tested by EMSA, bind physically to the PPARγ-RXRα complex. PGlyRP3 gene expression was induced by PPARγ ligands including GW1929 and some FFA. Overexpression of PGlyRP3 in Caco2 cells down regulated the expression of the inflammatory cytokines IL-8, IL-12 and TNF-α, while its silencing increased the expression of these cytokines. FFA that induced the PGlyRP3 inhibited the tested cytokines. Silencing of PGlyRP3 gene caused the same FFA to increase the cytokine gene expression. A negative regulation of NF-κB pathway, including up-regulation of Iκβ-α and down regulation of NF-κB and COX-2, is involved in the anti-inflammatory effects of PGlyRP3. In conclusion, PPARγ mediates a modulation of PGlyRP3 gene expression, which is involved in inhibiting inflammation through negative regulation of NF-κB pathway.
    Immunobiology. 11/2010; 216(6):715-24. · 3.21 Impact Factor
  • Journal Article:The generation of programmable cells of monocytic origin involves partial repression of monocyte/macrophage markers and reactivation of pluripotency genes.
    ABSTRACT:We have recently demonstrated that peripheral blood monocytes can be differentiated in vitro into hepatocyte-like cells using appropriate differentiation media. Phenotype conversion required prior in vitro culture in the presence of M-CSF, IL-3, and human serum, during which the cells acquired a sta... [more]We have recently demonstrated that peripheral blood monocytes can be differentiated in vitro into hepatocyte-like cells using appropriate differentiation media. Phenotype conversion required prior in vitro culture in the presence of M-CSF, IL-3, and human serum, during which the cells acquired a state of plasticity, so were termed "programmable cells of monocytic origin" (PCMO). Here, we have further characterized the process of PCMO generation with respect to markers of monocyte-to-macrophage transition and pluripotency. During a 6-day culture period, various monocyte/macrophage differentiation markers were down-regulated being indicative of a process of partial dedifferentiation. Dedifferentiation and hepatic redifferentiation also proceeded in highly purified monocyte preparations, albeit with different kinetics, suggesting that the presence of nonmonocytes, or soluble factors derived from them, is not essential in order for monocytes to acquire a multipotent state. PCMOs expressed various markers of human embryonic stem cells with early induction of NANOG and OCT4. Expression of the pluripotency-associated OCT4A isoform was paralleled by a global rise in histone H3 methylation on Lys-4, a marker of active chromatin, and coincided with peak sensitivity to tissue-specific differentiation. These results show that peripheral blood monocytes can be induced in vitro to transiently acquire stem cell-like properties and concomitantly a state of increased differentiation potential toward the hepatocytic phenotype.
    Stem cells and development. 03/2010; 19(11):1769-80. · 4.15 Impact Factor
  • Journal Article:Metabolic aspects of neonatal rat islet hypoxia tolerance.
    ABSTRACT:Sensitivity of pancreatic islets to hypoxia is one of the most important of the obstacles responsible for their failure to survive within the recipients. The aim of this study was to compare the in vitro hypoxia tolerance of neonatal and adult rat islet cells and to study the glucose metabolism in t... [more]Sensitivity of pancreatic islets to hypoxia is one of the most important of the obstacles responsible for their failure to survive within the recipients. The aim of this study was to compare the in vitro hypoxia tolerance of neonatal and adult rat islet cells and to study the glucose metabolism in these cells after exposure to hypoxia. Islet cells from both age categories were cultured in different hypoxic levels for 24 h and insulin secretion and some metabolites of glucose metabolism were analysed. Glucose-stimulated insulin secretion decreased dramatically in both cell preparations in response to the decrease in oxygen level. The reduction of insulin secretion was more detectable in adult cells and started at 5% O(2), while a significant reduction was obtained at 1% O(2) in neonatal cells. Moreover, basal insulin release of neonatal cells showed an adaptation to hypoxia after a 4-day culture in hypoxia. Intracellular pyruvate was higher in neonatal cells than in adult ones, while no difference in lactate level was observed between them. Similar results to that of pyruvate were observed for adenosine triphosphate (ATP) and the second messenger cyclic adenosine monophosphate (cAMP). The study reveals that neonatal rat islet cells are more hypoxia-tolerant than the adult ones. The most obvious metabolic observation was that both pyruvate and lactate were actively produced in neonatal cells, while adult cells depended mainly on lactate production as an end-product of glycolysis, indicating a more enhanced metabolic flexibility of neonatal cells to utilize the available oxygen and, at the same time, maintain metabolism anaerobically.
    Transplant international : official journal of the European Society for Organ Transplantation. 09/2009; 23(1):80-9. · 2.92 Impact Factor
  • Journal Article:Gamma-aminobutyric acid up- and downregulates insulin secretion from beta cells in concert with changes in glucose concentration.
    ABSTRACT:The role of gamma-aminobutyric acid (GABA) and A-type GABA receptors (GABA(A)Rs) in modulating islet endocrine function has been actively investigated since the identification of GABA and GABA(A)Rs in the pancreatic islets. However, the reported effects of GABA(A)R activation on insulin secretion fr... [more]The role of gamma-aminobutyric acid (GABA) and A-type GABA receptors (GABA(A)Rs) in modulating islet endocrine function has been actively investigated since the identification of GABA and GABA(A)Rs in the pancreatic islets. However, the reported effects of GABA(A)R activation on insulin secretion from islet beta cells have been controversial. This study examined the hypothesis that the effect of GABA on beta cell insulin secretion is dependent on glucose concentration. Perforated patch-clamp recordings in INS-1 cells demonstrated that GABA, at concentrations ranging from 1 to 1,000 micromol/l, induced a transmembrane current (I(GABA)) which was sensitive to the GABA(A)R antagonist bicuculline. The current-voltage relationship revealed that I(GABA) reversed at -42+/-2.2 mV, independently of glucose concentration. Nevertheless, the glucose concentration critically controlled the membrane potential (V (M)), i.e., at low glucose (0 or 2.8 mmol/l) the endogenous V (M) of INS-1 cells was below the I(GABA) reversal potential and at high glucose (16.7 or 28 mmol/l), the endogenous V (M) of INS-1 cells was above the I(GABA) reversal potential. Therefore, GABA dose-dependently induced membrane depolarisation at a low glucose concentration, but hyperpolarisation at a high glucose concentration. Consistent with electrophysiological findings, insulin secretion assays demonstrated that at 2.8 mmol/l glucose, GABA increased insulin secretion in a dose-dependent fashion (p<0.05, n=7). This enhancement was blocked by bicuculline (p<0.05, n=4). In contrast, in the presence of 28 mmol/l glucose, GABA suppressed the secretion of insulin (p<0.05, n=5). These findings indicate that activation of GABA(A)Rs in beta cells regulates insulin secretion in concert with changes in glucose levels.
    Diabetologia. 04/2006; 49(4):697-705. · 6.81 Impact Factor
  • Journal Article:Effect of the pancreatic digestion with liberase versus collagenase on the yield, function and viability of neonatal rat pancreatic islets.
    Ayman Hyder
    ABSTRACT:Many obstacles hinder the clinical application of pancreatic islet transplantation as a cure for diabetes mellitus. One of them is the suitable isolation method of sufficient number of healthy islets for transplantation. In this context, liberase enzyme was developed as a purified form of the tradit... [more]Many obstacles hinder the clinical application of pancreatic islet transplantation as a cure for diabetes mellitus. One of them is the suitable isolation method of sufficient number of healthy islets for transplantation. In this context, liberase enzyme was developed as a purified form of the traditional collagenase. It was the aim of this study to investigate the effect of liberase-digestion on the yield, function and viability of neonatal rat islets, and to compare the new enzyme with the collagenase. Glucose-stimulated insulin secretion was measured as indication of the function, insulin content as indication for the synthetic activity of islet cells and DNA as an indication of cell viability. The results showed no difference between islets isolated either with collagenase or liberase. Glucose stimulated similarly the insulin secretion in both. Stimulation index tended, without significance, to be higher (55%) in liberase-isolated islets compared with the collagenase islets (49%). The viability of both was similar. The insulin synthesis (content) tended also to be better in liberase-isolated islets. It could be concluded that liberase could be non-significantly preferred in the isolation of neonatal rat islets in comparison with collagenase.
    Cell biology international. 10/2005; 29(9):831-4. · 1.48 Impact Factor
  • Journal Article:Effect of the immunosuppressive regime of Edmonton protocol on the long-term in vitro insulin secretion from islets of two different species and age categories.
    ABSTRACT:The success of the new immunosuppressive regime known as the Edmonton protocol in islet allotransplantation may suggest that it is also possible that this regime may prevent the rejection of xenografts. This protocol applies a combination of Tacrolimus, Sirolimus and Daclizumab at low doses. This co... [more]The success of the new immunosuppressive regime known as the Edmonton protocol in islet allotransplantation may suggest that it is also possible that this regime may prevent the rejection of xenografts. This protocol applies a combination of Tacrolimus, Sirolimus and Daclizumab at low doses. This combination may have some toxicity that affects the function and viability of the pancreatic islets. The choice of species or age category, whose islets can tolerate the toxicity of this immunosuppressive combination, may become important for the graft survival. It was the aim of this study to investigate the long-term effect of this regime on insulin secretion from pancreatic islets isolated from two species (rats and pigs) and from two age categories (day 7 postnatal [P7] and adult rat). Islets were cultured for three weeks in medium containing Tacrolimus in a concentration of 5 ng/ml, while the concentration of Sirolimus was 15 ng/ml. Daclizumab was added at the beginning of culture and once weekly in a final concentration of 10 ng/ml. In immunosuppressive-treated groups, Glucose was able to stimulate increases of insulin secretion over the basal value after 1 and 3 weeks in adult rat islets, and could not stimulate this secretion in P7 islets, while it stimulated the secretion only after 1 week, but not 3 weeks, in porcine islets. The immunosuppressive regimen caused significant reductions of glucose-stimulated insulin secretion magnitude in adult rat and porcine islets after 3 weeks, while it reduced both basal and stimulated secretions after 1 and 3 weeks in P7 rat islets. There was no difference in DNA contents between control and immunosuppressive-treated groups after 1 or 3 weeks in any of the islet preparations. DNA decreased considerably with the time in culture. The change in DNA content over 3 weeks was higher in the Edmonton group of adult porcine and P7 rat than in adult rat islets. Comparison of the responses of islets from different age categories and species leads to conclude that in vitro cultures of adult rat islets are more tolerant to this immunosuppressive combination toxicity than P7 islets, and there is variable responses of islets from different adult species to this toxicity.
    Toxicology in vitro : an international journal published in association with BIBRA. 07/2005; 19(4):541-6. · 2.78 Impact Factor
  • Journal Article:Effect of extracellular pH on insulin secretion and glucose metabolism in neonatal and adult rat pancreatic islets.
    ABSTRACT:Changes in extracellular pH are known to affect glucose-stimulated insulin secretion. In the present study, glucose metabolism in pancreatic islets cultured at different pHs was investigated. Also, for islet transplantation purposes, insulin secretion and glucose metabolism were compared in neonatal... [more]Changes in extracellular pH are known to affect glucose-stimulated insulin secretion. In the present study, glucose metabolism in pancreatic islets cultured at different pHs was investigated. Also, for islet transplantation purposes, insulin secretion and glucose metabolism were compared in neonatal and adult islets at different pHs to determine which islet preparation is more tolerant to acidity and alkalinity. The results revealed a dependency of insulin secretion on the external pH in both neonatal and adult islets. Reduction of insulin secretion was observed at both the acidic and alkaline sides of pH 7.3. Glucose stimulated increases of insulin secretion in all cases. Similar results were obtained for ATP and pyruvate contents. Intracellular insulin increased with the increase of pH value. In contrast, calcium content decreased with the increase of pH. The results demonstrate that neonatal islets are more acid tolerant than adult islets. Both basal and glucose-stimulated insulin secretions, as well as other parameters of neonatal islets were significantly higher than those of adult islets in response to low pH. The differences under alkaline conditions were not significant but give an indication that neonatal islets are more tolerant to alkalinity than are adult islets.
    Acta diabetologica. 01/2002; 38(4):171-8. · 2.78 Impact Factor
  • Journal Article:HOE 077 reduces fibrotic overgrowth around the barium alginate microcapsules.
    Transplantation proceedings. 03/2000; 32(1):206-9. · 1.01 Impact Factor



------------------------------------------ Best Wishes: Dr.Ehab Aboueladab, Tel:01007834123 Email:ehab10f@gmail.com,ehababoueladab@yahoo.com ------------------------------------------
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