We analyzed optical coherence tomographic (OCT) characteristics of different types of coronary thrombi that had been confirmed at postmortem histologic examination. We examined 108 coronary arterial segments of 40 consecutive human cadavers. OCT images of red and white thrombi were obtained and the intensity property of these thrombi was analyzed. Red and white thrombi were found in 16 (17%) and 19 (18%) of the 108 arterial segments, respectively. Red thrombi were identified as high-backscattering protrusions inside the lumen of the artery, with signal-free shadowing in the OCT image. White thrombi were identified as low-backscattering projections in the OCT image. There were no significant differences in peak intensity of OCT signal between red and white thrombi (130+/-18 vs 145+/-34, p=0.12). However, the 1/2 attenuation width of the signal intensity curve, which was defined as the distance from peak intensity to its 1/2 intensity, was significantly different between red and white thrombi (324+/-50 vs 183+/- 42 microm, p<0.0001). A cut-off value of 250 microm in the 1/2 width of signal intensity attenuation can differentiate white from red thrombi with a sensitivity of 90% and specificity of 88%. We present the first detailed description of the characteristics of different types of coronary thrombi in OCT images. Optical coherence tomography may allow us not only to estimate plaque morphology but also to distinguish red from white thrombi.

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Sphingosine 1-phosphate (Sph-1-P), the initial product of Sph degradation by Sph kinase, was shown to be a strong inhibitor of cell motility and phagokinesis of B16 melanoma and other types ofcells at 10-100 nM concentration. It also inhibited "chemoinvasion' of tumor cells through a thick layer of Matrigel on a filter membrane. Such inhibitory effects were produced minimally or not at all by Sph, N-methyl derivatives of Sph, or other related sphingolipids and phospholipids. Sph-1-P did not inhibit cell proliferation or protein kinase C (PKC) activity, in contrast to Sph and N-methyl-Sph, which inhibit PKC activity and cell growth in general. of mouse 3T3 cells through a protein kinase C (PKC)-independent pathway (5); they subsequently attributed this growth-stimulatory effect to formation of Sph-1-P (6). Sph or its catabolites may enhance cytoplasmic Ca2+ release (7); this was also attributed to Sph-1-P (6), in analogy to the effect of inositol 1,4,5-trisphosphate on Ca2+ movement (8). Although Sph-1-P was assumed to induce proliferation in 3T3 cells, particularly in synergy with epidermal growth factor and insulin (6), the physiological role of this endogenous product of Sph metabolism in cells remains unknown. We now report that Sph-1-P, either in chemically synthesized form or rapidly converted from exogenous Sph, strongly and specifically inhibits chemotactic motility and invasiveness of tumor cells at very low (nanomolar) concentration but does not affect cell proliferation or PKC activity even at much higher concentrations. These findings suggest that Sph-1-P controls motility of normal cells and invasiveness of tumor cells, without affecting cell proliferation, via changes in transmembrane signaling independent of the PKC pathway. In contrast, N-methyl derivatives of Sph are known to inhibit cell proliferation, via blocking of the PKC pathway or some unknown mechanism (9, 10).* Preparation of Sph Derivatives and Other Lipids. Sph-1-P was prepared both enzymatically and chemically. Enzymatic preparation was by hydrolysis of sphingosylphosphocholine by Streptomyces chromofuscus phospholipase D as described (11) (both reagents were obtained from Sigma).Sph-1-P was chemically synthesized [Weiss (12) succeeded in synthesizing sphinganine 1-phosphate, but failed to synthesize sphingenin 1-phosphate] from 1-0-and N-protected Sph by acylating 3-OH with pivaloyl chloride. Then 2-N-t-Boc-3-O-pivaloyl-D-erythro-Sph was prepared by selective deprotection of the primary OH group with p-toluenesulfonic acid (tosyl) chloride, followed by phosphorylation of the C1 primary OH group. Deprotection of the C3 allytic OH group and primary NH2 gave the desired Sph-1-P. The enzymatically synthesized product was a mixture of L-threo and D-erythro isomers, whereas the chemically synthesized product consisted only of D-erythro isomer, according to NMR data (27). Enzymatically and chemically synthesized Sph-1-P showed identical mass spectra and indistinguishable biological properties. In this study, therefore, we used enzy...

SummaryErythroblastic islands are anatomical units consisting of a central macrophage surrounded by erythroblasts. We studied the adhesion molecules involved in the formation of these structures. Central macrophages of erythroblastic islands isolated from the spleens of phlebotomized mice were dearly stained for vascular cell adhesion molecule 1 (VCAM-1). The surrounding erythroblasts of the erythroblastic islands strongly expressed the a4 integrin of very late activation antigen 4 (VLA-4:a4B1 integrin), the counter receptor of VCAM-1, whereas most reticulocytes and erythrocytes did not. Both monoclonal antibodies (mAbs) against oe4 integrin and VCAM-1 disrupted the erythroblastic islands cultured in the presence of erythropoietin. Moreover, adhesion of splenic erythroblasts to tumor necrosis factor a-stimulated mouse splenic endothelial cells, which showed high expression of VCAM-1 but not intercellular adhesion molecule 1, was inhibited by the anti-VCAM-1 and anti-a4 mAbs. These findings suggest that VLA-4-VCAM-1 interaction plays a crucial role in the formation of erythroblastic islands. Bone marrow resident macrophages (MCFs) establish stroma by extending long cytoplasmic processes and attaching to developing erythroid and myeloid cells (1, 2). These cells are both phenotypica[ly and functionally different from peritoneal M4's and monocytes (3, 4). In long-term bone marrow culture by which hematopoietic stem cells are maintained, immature myelomonocytic cells are attached to and proliferate on the M~s defined by the mAb F4/80 (5). The addition of erythropoietin to these cultures alternatively induces erythropoietic activity on the M~s, which form erythroblastic islands (EI) composed of central M~s and surrounding erythroblasts (Ebs) (6, 7). These lines of evidence suggest that erythropoietin-responsive erythroid precursors adhere to the M~s, where they are induced to proliferate and differentiate into erythrocytes by maintaining contact with the central M~s. To gain insight to the possible function of the M~s in erythropoiesis, we have considered it a priority to identify specific adhesion molecules involved in El formation (8).Recently, Morris et al. (9) characterized the nature of the adhesion using El isolated from fetal liver and found that the adhesion required divalent cations. Soligo et al. (10) demonstrated that a divalent cation-dependent adhesion molecule, very late activation antigen 4 (VLA-4) (a4B1) integrin, was present on Ebs, and that this molecule was localized at sites of intercellular contact between Ebs and M~s in human marrow, suggesting that VLA-4 might be involved in the adhesive interaction. The ligand of VLA-4 for cell-cell interaction is vascular cell adhesion molecule I (VCAM-1), which was first identified on TNF-ce-stimulated human umbilical endothelial cells (11,12). Using the bone marrow stromal cell culture system, VCAM-1 has been shown to be constitutively expressed in stromal cells, and to be involved in B lymphocyte-stromal cell (13) and hematopoietic stem cell-stromal cell i...

ntimal thickening is considered to be an early phase of atherosclerosis; [1][2][3][4][5] in the coronary artery it is related to coronary spasm and is of prognostic importance in patients after cardiac transplantation. 6,7 Evaluation of intimal thickening of the coronary artery is thought to be important clinically and recent advancements in intravascular ultrasound (IVUS) have enabled pathological evaluation of the wall structure of the coronary artery. [8][9][10][11][12][13][14][15] However, intimal thickening is indirectly evaluated as the intima -media thickness (IMT) by IVUS because the boundary of the intima and media cannot be distinguished completely by this method. 16 Optical coherence tomography (OCT) is a new imaging method [17][18][19][20][21][22] with a resolution of approximately 10-20 m, which is approximately 10-fold greater than that of IVUS. 23 OCT can easily identify the internal elastic laminae, which cannot be identified by IVUS. 18,19 We thus hypothesized that OCT could enable more accurate measurement of the IMT and the intimal thickness than IVUS, so the aim of the present study was to test these hypotheses. MethodsWe examined 54 randomly selected segments from 54 coronary arteries from 18 consecutive cadavers (10 males, 8 females; mean age 72±6 years). The cause of death was leukemia (n=3), lung cancer (n=3), acute myocardial infarction (n=2), pulmonary embolism (n=2), intracranial hemorrhage (n=2), liver cirrhosis (n=2), pneumonia (n=2), Circulation Journal Vol.69, August 2005and trauma (n=2). The study protocol was approved by the ethics committee of Kawasaki Medical School, and written informed consent was obtained from each family. An approximate 5-cm length of the proximal site of the 3 major coronary arteries, the left anterior descending coronary artery, left circumflex artery, and right coronary artery, were obtained from cadavers at autopsy within 3 h of death. The surrounding soft tissue was dissected from each specimen. Small arterial perforators and the branches were tied off with sutures, and the distal end of the artery was occluded with a large cork. A 7F sheath was sewn into the proximal end of the artery to complete the closed system. Phosphate-buffered saline (154 mmol/L NaCl, 10 mmol/L NaH2PO4, pH 7.4), which was kept at 37°C, was infused through the side arm of the sheath. The pressure inside the coronary artery was maintained at a physiological level (60-80 mmHg) with a sphygmomanometer connected to the infusion. OCT and IVUS ImagesAn intravascular OCT catheter (ImageWire ® , LightLab Imaging, Westford, MA, USA) and IVUS catheter (Atlantis SR Pro ® 2.5F, 40-MHz, Boston Scientific, Natick, MA, USA) were inserted through the diaphragm of the sheath. Serial OCT and IVUS images were obtained using an automatic pullback device at a rate of 0.5 mm/s. The OCT images were processed and analyzed using proprietary software from LightLab Imaging, Inc and the IVUS images were exported to commercially available image processing software (Netra 3D IVUS system, ScImage) for off-lin...

Both OCT and IVUS underestimated the area of calcification, but OCT estimates of the area of calcification were more accurate than those estimated by IVUS. Thus, OCT may be a more useful clinical tool to quantify calcified plaque.

Macrophages, which are derived from precursor cells in the bone marrow, differentiate specifically under the influence of the local microenvironment. Resident macrophages in hematopoietic tissues can be distinguished from other stromal cells and monocytes by immunostaining with monoclonal antibody F4/80 and anti-Forssman glycosphingolipid antibody, respectively. Erythroid colony-forming units adhere to a resident macrophage and differentiate to erythroblasts in the presence of erythropoietin (EPO), resulting in the formation of an erythroblastic island. Resident macrophages play a supportive role in erythropoiesis, probably by preventing apoptosis of the erythroid precursors via adhesive interaction between very late activation antigen 4 and vascular cell adhesion molecule 1. Herein is proposed a model of erythropoiesis based on cooperative interaction between EPO and resident macrophages.