In recent years, investigators have unraveled a previously unrecognized role for granulocyte colony-stimulating factor (G-CSF) in the regulation of T-cell and dendritic cell functions. The ...experimental evidence in favor of G-CSF-mediated immune regulation includes the ability to skew T-cell cytokine secretion to T-helper type 2 responses, and to promote regulatory T-cell and tolerogenic dendritic cell differentiation. Accordingly, beneficial effects of G-CSF have been detected in animal models of immune-mediated diseases, including posttransplantation graft-versus-host disease, experimental autoimmune encephalomyelitis, lupus nephritis, inflammatory bowel disease, and diabetes. The growing body of evidence supporting a novel role for G-CSF in the induction of T-cell tolerance is reviewed herein.
The activation of human platelets by α-thrombin is mediated in part by cleavage of the protease-activated receptor (PAR) 1 and 4 and by the glycoprotein Ibα, (GpIbα), which binds with high affinity ...to α-thrombin. Recent studies have shown that the thrombin domain referred to as heparin binding site (HBS) is involved in the interaction with the platelet GpIbα. The HBS is rich in basic amino acids. To identify the key amino acid residues involved in the binding to GpIbα, we have performed alanine scanning mutagenesis of the basic HBS R93, R97, R101, R233, K236, K240, R233/K236/Q239, as well as of the neutral Q239 residues, located in different regions of the domain. For comparison, mutation at R67 within the fibrinogen recognition site (FRS) of thrombin was performed as well. Solid-phase binding experiments showed that the K d of thrombin−GpIb interaction was reduced 22-fold for R93A, 8-fold for R97A, 13-fold for R101A, 29-fold for R233A, 21-fold for K236A, 5-fold for K240A, and 31-fold for the triple mutant R233A/K236A/Q239A, while the Q239A and R67A forms did not show any significant affinity change. The platelet activating capacity of these mutants was evaluated as well. Using gel-filtered platelets, the EC50 value of thrombin-induced aggregation was from 5- to 13-fold higher in the HBS mutants than in the WT form, and was linearly and positively correlated with the corresponding K d values pertaining to thrombin binding to GpIb. Measurements of PAR-1 hydrolysis on the platelet membrane showed that the HBS mutants R233A, R101A, R93A, K236A, and R233/K236/Q239 forms had a reduction of the apparent k cat/K m value. These results are a consequence of a defective binding to GpIb, which is known to optimize the interaction with PAR-1 in situ. A confirm of this hypothesis came from the demonstration that the k cat/K m value pertaining to the hydrolysis by the HBS-mutated thrombins of the synthetic PAR-1 38−60 peptide in solution was similar to that one obtained with the WT form. In conclusion, these experiments provide a structural and functional mapping of the thrombin HBS subregions involved in the binding to the platelet GpIbα and in the cell activation.
Background. Several studies have demonstrated that bone marrow contains a subpopulation of stem cells capable of participating in the hepatic regenerative process, even if some reports indicate quite ...a low level of liver repopulation by human stem cells in the normal and transiently injured liver.
Aims. In order to overcome the low engraftment levels seen in previous models, we tried the direct intraperitoneal administration of human cord blood stem cells, using a model of hepatic damage induced by allyl alcohol in NOD/SCID mice.
Methods. We designed a protocol based on stem cell infusion following liver damage in the absence of irradiation. Flow cytometry, histology, immunohistochemistry and RT-PCR for human hepatic markers were performed to monitor human cell engraftment.
Results. Human stem cells were able to transdifferentiate into hepatocytes, to improve liver regeneration after damage and to reduce the mortality rate both in both protocols, even if with qualitative and quantitative differences in the transdifferentiation process.
Conclusions. We demonstrated for the first time that the intraperitoneal administration of stem cells can guarantee a rapid liver engraftment. Moreover, the new protocol based on stem cell infusion following liver damage in the absence of irradiation may represent a step forward for the clinical application of stem cell transplantation.
The receptor for hepatocyte growth factor (HGF) is a transmembrane tyrosine kinase that is encoded by the proto-oncogene c-met. Recently, c-MET was detected in Reed-Sternberg (RS) cells from ...Epstein-Barr virus-positive (EBV(+)) Hodgkin disease (HD). The c-MET, EBER-1, and LMP-1 expression in 45 lymph node biopsies and 12 bone marrow biopsies obtained from patients with HD was analyzed. In addition, HGF levels in serum samples from 80 healthy individuals and 135 HD patients in different phases of disease. In all 45 lymph node and 12 bone marrow samples examined, RS cells expressed c-MET but not HGF(+). These results were independent of the EBV infection. Interestingly, several HGF(+) dendritic-reticulum cells were found scattered around c-MET(+) RS cells. The mean +/- SEM serum HGF levels in HD patients at diagnosis and at the time of relapse were 1403 +/- 91 (95% confidence interval CI, 1221-1585) and 1497 +/- 242 pg/mL (95% CI, 977-2017), respectively. HGF values were significantly higher than those of healthy individuals (665 +/- 28 pg/mL; 95% CI, 600-721; and P <.001 for both groups of patients) and of HD patients in remission (616 +/- 49 pg/mL; 95% CI, 517-714; and P <.001 for both groups of patients). A significant correlation was found between serum HGF levels and B symptoms at diagnosis (P =.014). In conclusion, this study indicates that HGF and c-MET constitute an additional signaling pathway between RS cells and the reactive cellular background, thereby affecting adhesion, proliferation, and survival of RS cells. Furthermore, the serum concentration of HGF in HD patients may be a useful tool in monitoring the status of disease.
In recent years, several investigators have unraveled a previously unrecognized role for G-CSF in the regulation of T cell and dendritic cell functions. The experimental evidence in favor of ...G-CSF-mediated immune regulation includes the ability to switch T cell cytokine secretion profile to Th2 responses and the promotion of regulatory T cell and tolerogenic dendritic cell differentiation. Interestingly, G-CSF is beneficial in animals for the prevention and/or treatment of immune-mediated diseases, e.g., graft-vs-host disease, multiple sclerosis, systemic lupus erythematosus, inflammatory bowel disease, and diabetes, suggesting a potential role in human autoimmune diseases. This review summarizes the growing body of evidence that supports a critical role for G-CSF as a novel mediator of T cell tolerance.
Intracoronary administration of bone marrow stem cells can improve left ventricular function after acute myocardial infarction (AMI) 1 ; however, it is invasive and therefore not widely applicable. ......we have recently shown that the CD34 cell count is favourably correlated with left ventricular function changes after AMI. 2 Granulocyte-colony stimulating factor (G-CSF), a potent mobiliser of CD34 stem cells, might improve left ventricular function through their increased concentration. ...the restenosis rate in our study was 25%, with a late loss comparable with that found in previous trials on primary PCI (eg, 0.47 mm in this study v 0.82 mm in the controlled abciximab and device investigation to lower late-angioplasty complications (CADILLAC) trial), 5 and in a very recent early trial of G-CSF in AMI. 6 Kang et al, 3 however, performed IRA stenting (and intracoronary injection of peripheral mononuclear cells) after 4 days of G-CSF administration.
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