Molecular Identity of Hematopoietic Precursor Cells Emerging in the Human Embryo

It is now accepted from studies in animal models that hematopoieticstem cells emerge in the para-aortic mesoderm-derived aorta-gonad-mesonephros region of the vertebrate embryo. We have previously identified the equivalent primitive hematogenous territory in the 4- to 6-week human embryo, under the form ofCD34⁺CD45⁺Lin highproliferative potential hematopoietic cells clustered on the ventralendothelium of the aorta. To characterize molecules involved in initialstem cell emergence, we first investigated the expression in thatterritory of known early hematopoietic regulators. We herein show thataorta-associated CD34⁺ cells coexpress the tal-1/SCL,c-myb, GATA-2, GATA-3, c-kit, and flk-1/KDR genes, as do embryonic andfetal hematopoietic progenitors later present in the liver and bonemarrow. Next, CD34⁺CD45⁺ aorta-associatedcells were sorted by flow cytometry from a 5-week embryo and a cDNAlibrary was constructed therefrom. Differential screening of thatlibrary with total cDNA probes obtained from CD34⁺embryonic liver cells allowed the isolation of a kinase-related sequence previously identified in KG-1 cells. In addition to emerging blood stem cells, KG-1 kinase is also strikingly expressed in alldeveloping endothelial cells in the yolk sac and embryo, which suggestsits involvement in the genesis of both hematopoietic and vascular celllineages in humans.     

Mouse yolk sac and intraembryonic tissues produce factors able to elicit differentiation of erythroid burst-forming units and colony-forming units, respectively.

This work was aimed at elucidating the environmental conditions that account for the production of embryonic erythrocytes in the mouse yolk sac (YS), while the adult-type hemoglobin and erythrocytes are generated in the fetal liver. Differentiation of YS hemopoietic stem cells (YS-HSC) of 9.5-day mouse embryos (prior to the colonization of the liver rudiment by HSC) was investigated in vitro. The influence of well-characterized erythroid growth factors, burst-promoting activity (BPA) and erythropoietin (EPO), which trigger the differentiation of early erythroid burst-forming units (BFU-E) and late erythroid colony-forming units (CFU-E), respectively, was tested on the YS-HSC in two different systems of culture: (i) organ culture and (ii) clonal culture in methylcellulose. When studied in organ culture, where the YS microenvironment was maintained, YS-HSC required only additional EPO to attain complete maturation into adult erythrocytes within 7 days. In contrast, YS hemopoietic single cells grown in methylcellulose, and thus released from the influence of the YS, required the presence of both BPA and EPO to generate BFU-E-derived colonies synthesizing high concentrations of hemoglobin. It is concluded that 9.5-day YS from mouse embryos is by itself able to promote the first differentiation steps of the adult lineage YS-HSC due to an intrinsic production of a BPA-like activity. In contrast, these experiments demonstrate that EPO or an EPO-like activity is not produced by YS tissues. As demonstrated earlier, if embryonic tissue is added to YS organ culture, although separated from it by a filter preventing cell contact, YS-HSC differentiate into adult erythrocytes producing adult-type hemoglobins. This shows that, in contrast to YS tissues, the early embryo produces EPO or a factor that can substitute for EPO.     

Proximal anastomosis in aortobifemoral bypass: end-to-end or end-to-side?

When performing an aortobifemoral bypass, the surgeon may choose between an end-to-end or end-to-side aortic anastomosis. Although several authors have claimed the superiority of the former, controversy still exists. The aim of this study was to compare the early and late results of these two techniques in patients for which both procedures were possible. Of 158 patients, aortic anastomosis was performed end-to-end in 47 (group I) and end-to-side in 111 (group II). Both groups were similar in all other aspects. The type of proximal anastomosis did not affect the rate of perioperative mortality or early thrombosis. There were no secondary aortic aneurysms or aortoenteric fistulae in either group. Actuarial primary (without reoperation) and secondary (after thrombectomy) five year patency rates were 90.2 and 98.9 per cent in group I, and 90.8 and 98.5 per cent in group II, respectively. Five-year limb survival was 95.3 and 95.7 per cent, respectively. As we could not find any difference between the results in the two groups, we suggest choosing the simplest procedure which maintains adequate pelvic and colonic blood supply, according to angiographic findings.     

MB1, a quail leukocyte-endothelium antigen: partial characterization of the cell surface and secreted forms in cultured endothelial cells.

We describe here conditions allowing the selective growth in culture of embryonic capillary endothelial cells from quail yolk sac. Such cultures were set up to characterize an antigen present on the endothelial cell surface and to study whether it was secreted in the culture medium. This antigen, MB1, was previously evidenced by a monoclonal antibody raised to quail IgM heavy chain. It is present at the surface of all endothelial and hemopoietic cells (except mature erythrocytes) starting from the hemangioblast, the early mesodermal precursor of blood and vascular endothelial cells. The MB1 epitope is also found on quail plasma molecules of 80 and 125-200 kDa. By immunoprecipitation of either surface or metabolically labeled endothelial cellular material, we have chemically characterized MB1-bearing components as glycoproteins of apparent molecular mass ranging from 80 to 200 kDa and provided evidence for their release into the culture medium. This is consistent with the hypothesis that, in the quail, vascular endothelium participates in the secretion of the alpha-MB1-positive plasmatic components.     

Femoropopliteal in situ saphenous vein bypass: Technical aspects and factors determining patency

The results and the factors influencing long-term patency rates in two technical variations of femoropopliteal «in situ» saphenous vein bypass procedures were evaluated retrospectively. Technique I (n=33) consisted of a limited approach to the saphenous vein, destruction of the valves with a Cartier stripper and no tributary ligation. Technique II (n=55) included complete exposure of the saphenous vein valvular destruction using Hall’s stripper and ligation of all tributaries. Early thrombosis was observed in 21% and 3.6% of cases in techniques I and II, respectively. The frequency of local complications was identical in both techniques (18%). Overall actuarial primary patency at 4 years was 70%. Secondary patency (including patency following reoperations for graft failure) was 88 % overall, 78.5% in technique I and 95% in technique II (p<0.05). The site of the distal anastomosis significantly influenced the four year cumulative patency rate (upper popliteal ?100%, lower popliteal ?93.6%, tibioperoneal vessels ?70.5%, p<0,05), whereas the number of patent leg vessels, the clinical stage of disease and the site of the proximal anastomosis (common femoral or superficial fernoral artery) did not. Results obtained with the in situ saphenous vein bypass using technique II were better than with technique I. Close follow-up and preventive reoperations clearly enhanced patency rates. The site of distal anastomosis significantly affected long-term results.     

Six-year follow-up of a cohort of 203 patients with diabetes after screening for silent myocardial ischaemia.

AIMS: To determine the prognosis of patients with Type 1 or Type 2 diabetes, 6 years after screening for silent myocardial ischaemia (SMI). METHODS: Two hundred and three asymptomatic patients with diabetes underwent systematic SMI screening. From the results of this screening, they were allocated to one of three groups: patients (n = 171) with negative screening; patients (n = 32) with positive screening; and patients (n = 21) with positive screening and coronary stenosis. Six years after the initial assessment, all patients were re-assessed. All events [death, cardiac death, non-fatal major cardiac events (NFMCEs)--acute myocardial infarction, ventricular rhythm disorders, heart failure, unstable angina] were recorded. RESULTS: Fifteen patients were lost to follow-up. Patients (n = 20) with positive SMI screening and coronary stenosis had a higher risk of NFMCEs (35% vs. 7%, P < 0.001), and a higher mortality rate (35% vs. 15%, P < 0.05) compared with patients (n = 157) with negative screening. SMI-positive patients (n = 31) had a higher NFMCE rate compared with negative SMI screening patients, although overall mortality rate was no different. Cancer was the leading cause of death (36.4%). In multivariate analysis, major cardiac events (cardiac death and NFMCE) were related to baseline age, body mass index and coronary stenosis (P < 0.01). CONCLUSIONS: Patients with diabetes and SMI have a very poor prognosis as assessed by cardiac events or death, especially in the presence of coronary stenosis.