transforming-growth-factor-beta and Shock

Ischemia in various organs and tissues takes place during and as a direct result of multiple trauma (MT). Bone marrow-derived endothelial progenitor cells (EPCs) are involved in neovascularization after ischemic incidences. Here, we report that serum derived from patients with MT stimulates differentiation of EPCs in vitro from peripheral blood mononuclear cells (PBMCs). EPCs were identified by DiL-Acetyl-LDL-uptake with concomitant UEA-I-lectin binding. A significant increase in EPC numbers was noted when PBMCs were cultivated for 72 h with the serum of MT patients (n = 25) obtained at 5 days. Furthermore, serum from MT patients enhanced the functional acting of EPCs to form prevascular structures in matrigel. Reverse transcription polymerase chain reaction analysis revealed gene expression of transforming growth factor (TGF)-beta1- and vascular endothelial growth factor (VEGF) receptors 1 and 2. Reverse transcription polymerase chain reaction analysis was based on further cultivated cell preparations, which contained at least 80% EPCs. Moreover, the addition of recombinant VEGF or low concentrations of TGF-beta increased EPC differentiation. In addition, neutralization of TGF-beta1 and of VEGF165 in MT serum using specific antibodies resulted in a significant decrease in EPC differentiation. Our data indicate that TGF-beta1 and VEGF165 play a pivotal role for EPC differentiation induced by serum of polytrauma patients.

Topics: Blotting, Western; Cell Differentiation; Cells, Cultured; Collagen; Culture Media; Dose-Response Relationship, Drug; Drug Combinations; Endothelium; Flow Cytometry; Humans; Ischemia; Laminin; Lectins; Leukocytes, Mononuclear; Neovascularization, Pathologic; Platelet Endothelial Cell Adhesion Molecule-1; Proteoglycans; Recombinant Proteins; Reverse Transcriptase Polymerase Chain Reaction; Serum; Shock; Stem Cells; Time Factors; Transforming Growth Factor beta; Transforming Growth Factor beta1; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2; von Willebrand Factor; Wounds and Injuries

Syndecan-4 is a transmembrane heparan sulfate proteoglycan belonging to the syndecan family. Following intraperitoneal injection of lipopolysaccharide (LPS), syndecan-4-deficient mice exhibited high mortality compared with wild-type controls. Severe endotoxin shock was observed in the deficient mice: systolic blood pressure and left ventricular fractional shortening were lower in the deficient mice than in the wild-type controls 9 h after LPS injection. Although histological examinations revealed no apparent differences between two groups, the plasma level of interleukin (IL)-1beta was higher in the deficient mice than in the wild-type controls 9 h after LPS injection. Consistent with the regulatory roles of syndecan-4, its expression in monocytes and endothelial cells of microvasculature increased in the wild-type mice after LPS administration. Although IL-1beta was produced to the same extent by macrophages from syndecan-4-deficient and wild-type mice after LPS stimulation, inhibition of its production by transforming growth factor-beta1 was impaired in the syndecan-4-deficient macrophages. These results indicate that syndecan-4 could be involved in prevention of endotoxin shock, at least partly through the inhibitory action of transforming growth factor-beta1 on IL-1beta production.

Topics: Animals; Blood Pressure; Cytokines; Endothelium; Flow Cytometry; Glutathione Transferase; Immunohistochemistry; Injections, Intraperitoneal; Interleukin-1; Interleukin-10; Lipopolysaccharides; Liver; Macrophages; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Monocytes; Protein Binding; Proteoglycans; Recombinant Fusion Proteins; Shock; Syndecan-4; Time Factors; Transforming Growth Factor beta; Ventricular Function, Left

Transforming growth factor-beta 1 (TGF beta) is a regulator of protein synthesis in cultured chicken embryo cells (CEC). Preceding a gradual increase in overall protein synthesis, members of the Hsp70 family (Hsp70, Hsc70, and Grp78) and the Hsp90 family (90-2 and 90-3) of molecular chaperones are induced rapidly and represent a new class of TGF beta-inducible proteins (I.M. Takenaka and L.E. Hightower, J. Cell. Physiol., 152:568-577, 1992). Herein, 32P-labeled cDNA probes encoding Hsc70 and Hsp90 were used to show that levels of the corresponding mRNAs increased as a fraction of total RNA and in polysomes within five hours of treatment of CEC with TGF beta. This cytokine did not increase rates of hsc70 and hsp90 gene transcription as measured by run-on transcription assays of isolated nuclei. However, the Hsp RNA inductions were inhibited by dactinomycin, indicating a requirement for newly synthesized RNA. Both Hsc70 and Hsp90 mRNAs had relatively short half-lives, measured by Northern blot analyses of dactinomycin chases, which were not altered substantially in TGF beta-treated cells. In contrast, Hsp mRNA half-lives increased in heat shocked CEC exposed to dactinomycin during recovery, revealing a difference in regulation of these genes in stressed cells compared with TGF beta-treated cells. Our results support the conclusion that hsc70 and hsp90 gene expression is regulated posttranscriptionally in TGF beta-treated CEC, and the mechanism likely involves a nuclear event such as increasing the half-lives of nuclear RNA transcripts, processing, or transport into the cytoplasm.

Topics: Animals; Cells, Cultured; Chick Embryo; Dactinomycin; Drug Stability; Gene Expression Regulation; Heat-Shock Proteins; Hot Temperature; Mitogens; Polyribosomes; RNA, Messenger; Shock; Transcription, Genetic; Transforming Growth Factor beta

Anesthesized male rabbits having a resting mean arterial pressure of 81 +/- 4 mm Hg and superior mesenteric artery blood flow of 91 +/- 7 mL min-1 were subjected to 60 min of splanchnic ischemia followed by 60 min of reperfusion. Upon reperfusion, mean arterial pressure fell. Splanchnic blood flow also decreased but not in parallel with blood pressure; consequently, vascular resistance was increased over the reperfusion period. This increase in splanchnic vascular resistance was not affected by intravenous t-PA (0.5 mg kg-1 + 5 mg kg-1 hr-1) for 30 min prior to and throughout the reperfusion period or by intravenous L-NAME (1 mg kg-1 x 2). However, intravenous infusions of TGF-beta (18 or 54 micrograms kg-1) at the time of reperfusion dose dependently attenuated the increases in vascular resistance (p < 0.05). This effect of TGF-beta was enhanced by coadministration of t-PA and inhibited by the coadministration of L-NAME. We propose that the effects of TGF-beta are ultimately mediated via nitric oxide release, and conclude that this may be useful therapy for the prevention of reperfusion-associated injury following surgery or as an adjunct to thrombolytic therapy.

Topics: Animals; Arginine; Blood Flow Velocity; Blood Pressure; Intestines; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Rabbits; Recombinant Proteins; Reperfusion Injury; Shock; Splanchnic Circulation; Tissue Plasminogen Activator; Transforming Growth Factor beta; Vascular Resistance