Biomedical subjects

S A Santoro

Publications and source records attributed to S A Santoro.

6 recordsLinked to original sources

PubMed1992

Differential regulation of the alpha 2 beta 1 and alpha IIb beta 3 integrin genes during megakaryocytic differentiation of pluripotential K562 cells.

Expression of the alpha 2 beta 1 and alpha IIb beta 3 integrin genes is differentially regulated during megakaryocytic differentiation of pluripotent K562 cells induced with phorbol 12,13-dibutyrate. Upon megakaryocytic differentiation, steady-state alpha 2 mRNA increased markedly from the undetectable level present in the uninduced cells. The level of beta 1 mRNA did not change. Expression of alpha IIb beta 3 is regulated differently. beta 3 mRNA was undetectable in uninduced cells but increased significantly following induction. alpha IIb mRNA was detectable at a low level prior to induction, but at an increased level following differentiation. Altered mRNA stability did not contribute to changes in mRNA levels. Nuclear run-off experiments revealed a 20-fold increase in alpha 2 gene transcription upon megakaryocytic differentiation, but no change in transcription of the beta 1 gene. Transcription of both the alpha IIb and beta 3 genes increased 10- and 5-fold, respectively. Thus, the increase in alpha 2 beta 1 protein which accompanies the megakaryocytic differentiation of K562 cells is a consequence of the increased steady-state level of alpha 2 mRNA due to transcriptional activation of the alpha 2 gene. The long-lived beta 1 mRNA is not altered during differentiation. In contrast, increased alpha IIb beta 3 protein appears due to increased steady-state levels of both alpha IIb and beta 3 mRNAs that result from transcriptional activation of both integrin genes.

Cell Differentiation↗

PubMed1992

Induced cell surface expression of functional alpha 2 beta 1 integrin during megakaryocytic differentiation of K562 leukemic cells.

The pluripotential hematopoietic cell line K562 was studied as a model of inducible integrin expression accompanying differentiation. Differentiation along the megakaryocytic pathway was induced with phorbol 12,13-dibutyrate and differentiation along the erythroid pathway with hemin. Induction of megakaryocytic differentiation was associated with changes in cell morphology and with increased cell-cell and cell-substrate adhesion and spreading. Erythroid differentiation was not associated with changes in morphology or adhesion. Cell surface expression of the IIb-IIIa and alpha 2 beta 1 integrins increased markedly with phorbol treatment but decreased with hemin treatment. Phorbol-treated K562 cells, but not control cells or hemin-treated cells, adhered to collagen substrates in a Mg(2+)-dependent manner which was specifically inhibited by a monoclonal antibody directed against the alpha 2 beta 1 integrin. Northern blot analysis revealed that megakaryocytic differentiation of K562 cells was accompanied by de novo expression of the alpha 2 integrin mRNA with no change in the level of mRNA for the beta 1 subunit. K562 cells provide a model of differentiation-dependent, regulated integrin expression in which expression is up- or down-regulated depending upon the differentiation pathway selected.

Antibodies↗

PubMed1992

New RGD analogue inhibits human platelet adhesion and aggregation and eliminates platelet deposition on canine vascular grafts.

Platelet adhesion and aggregation are mediated by fibrinogen via the receptor glycoprotein IIb/IIIa, which recognizes the arginine-glycine-aspartic (RGD) amino-acid sequence. We investigated the ability of 8-guanidino-octanoyl-Asp-Phe (SC-49992), an intravenously infused, stable RGD analogue, to inhibit human platelet function in vitro and to reduce in vivo canine platelet deposition on prosthetic grafts. Human platelet aggregation induced by 10 mumol/L adenosine diphosphate was inhibited in a concentration dependent manner with an ED50 of 1 microgram/ml of SC-49992. Adenosine diphosphate-induced secretion, which is dependent on fibrinogen occupancy of the glycoprotein IIb/IIIa receptor, was reduced in a concentration dependent manner, also with an ED50 of 1 microgram/ml. Thrombin-induced secretion, which is independent of fibrinogen binding, was unaffected. Activation-dependent platelet adhesion to fibrinogen substrates was reduced in a concentration-dependent manner by SC-49992. Platelet adhesion to fibronectin substrates was also reduced by the analogue, but to a lesser extent. SC-49992 effectively eluted glycoprotein IIb/IIIa bound to RGD derivatized sepharose. Eight thrombosis-prone dogs had polytetrafluoroethylene femoral artery grafts placed. Dogs received the RGD analogue or a normal saline infusion during their first graft procedure. One week later a second contralateral femoral graft with infusion of the other agent was performed. Aggregometry during RGD analogue infusion demonstrated inhibition of induced aggregation, whereas normal saline infusion had no effect. As measured by the adherence of platelets labeled with indium III 8-guanidino-octanoyl-Asp-Phe reduced platelet deposition on vascular grafts by more than 90% (p = 0.0006, log transformed data, paired t test.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1992

Preparation of soluble peptide libraries: application to studies of platelet adhesion sequences.

We describe chemical synthesis of peptide mixtures that equally express many sequence combinations. Using 65 couplings of single amino acids, five mixtures were prepared with the sequences Tyr-Gly-Arg-Gly-Yyy-Xxx-Xxx, where Yyy is Ser, Asp, Arg, Asn, or Glu, and Xxx is any amino acids. Compositional and sequence analyses supported full representation of all amino acids, except isoleucine was deficient in the sixth position. The data suggest formation of a repertoire of 1,900 sequence combinations (5 x 19 x 20). The mixture with Asp as the fifth residue inhibited platelet adhesion to fibronectin more effectively than the other mixtures. Peptide libraries offer a new tool for investigating bioactive peptides.

Amino Acid Sequence↗

PubMed1979

Fibronectin and the multiple interaction model for platelet-collagen adhesion.

A rapid, sensitive, and reproducible assay to determine the adhesion of platelets to collagen has been developed. Collagen fibers and adherent platelets are retained on polycarbonate membrane filters. Chemical modification of collagen by acetylation and of platelets by treatment with chymotrypsin markedly reduces adhesion. The role of fibronectin in the collagen-platelet interaction has been examined. Treatment of platelets with purified antibody or Fab' fragments to fibronectin only slightly reduces adhesion. Preincubation of platelets with high concentrations of gelatin reduces adhesion by only 22% but fails to inhibit aggregation. Thus, fibronectin has only a limited role in the adhesion of platelets to collagen and is either not involved in the adhesion that leads to aggregation or is only one of several adhesion mechanisms, any of which alone can initiate aggregation.

Animals↗

PubMed1977

Collagen-mediated platelet aggregation. Evidence for multivalent interactions of intermediate specificity between collagen and platelets.

We have shown previously that periodate oxidation of collagen carbohydrate does not affect its ability to aggregate platelets. We now describe an additional characterization of periodate-modified collagen which demonstrates that collagen devoid of intact carbohydrate is fully capable of fibril formation, and we confirm its capacity to initiate platelet aggregation. Furthermore, we demonstrate that the platelet aggregating abilities of Types I, II, and III fibrillar collagen are quite similar despite differences in carbohydrate content and amino acid sequence. We also demonstrate that monomeric, pepsin-solubilized Type I human collagen is ineffective inhibiting aggregation by performed fibrils derived from the same molecule, thus establishing that the affinity of platelets for collagen depends upon prior polymerization of collagen. We interpret these and other findings to demonstrate that the hydroxylysyl glycoside regions of collagen are not highly specific sites involved in platelet-collagen interactions leading to "physiological" aggregation, and that the possibility must be considered that multiple interactions involving collagen sites of comparatively low structural specificity may be the initiating events in release of platelet ADP and the ensuing aggregation.

Amino Acids↗