Non-invasive assessment of muscle oxygenation may aid in optimising transfusion threshold decisions in ambulatory paediatric patients.

OBJECTIVE: To assess the potential utility of a novel non-invasive muscle oxygen measurement to determine the presence of muscle hypoxia in patients with anaemia. BACKGROUND: Recent assessment of the risk/benefit ratio of blood transfusion has led to clinical strategies optimising transfusion decisions. These decisions are primarily based on haematocrit (Hct) but not oxygen delivery, the primary function of red blood cells (RBCs). We hypothesised that muscle oxygenation (MOx) would correlate with Hct in patients with anaemia and may be a physiologically relevant determinant of the transfusion threshold. METHODS/MATERIALS: MOx was non-invasively determined in children in the Cancer and Blood Disorders Center ambulatory clinic at Seattle Children's Hospital using a custom-designed optical probe and spectrometer. MOx was compared with contemporaneous Hct. In subjects receiving RBCs, MOx and Hct were also determined following transfusion. RESULTS: MOx ranged from 36·7 to 100%, and Hct ranged from 17·0 to 38·6% in 27 measurements from 16 patients. High MOx values were associated with high Hct. Mean MOx for patients with normal Hct for age (n = 5) was 95·9 ± 2·9%. RBC transfusion increased mean Hct from 19·1 ± 1·5% to 29·3 ± 2·0 and mean MOx from 67·9 ± 21·1% to 89·9 ± 9·8%. Among six transfusion episodes (in five patients) with initial Hct < 22, only three had a pre-transfusion MOx of <70%. Patients with the lowest pre-transfusion MOx had the largest increase in MOx after transfusion. CONCLUSIONS: These preliminary data suggest that MOx may aid in making transfusion decisions when used in combination with Hct.

Okadaic acid induces dephosphorylation of histone H1 in metaphase-arrested HeLa cells.

It is shown here that treatment of metaphase-arrested HeLa cells with okadaic acid (0.15-2.5 microM) leads to dephosphorylation of histone H1. This effect is presumably due to the specific ability of okadaic acid to inhibit protein phosphatases 1 and/or 2A, because okadaic acid tetraacetate, which is not a phosphatase inhibitor, has no effect. Dephosphorylation of H1 does not occur if okadaic acid-treated cells are simultaneously treated with 20 nM calyculin A, or if the okadaic acid concentration is 5.0 microM or greater. The mechanism behind this phenomenon is not known. However, the results suggest that the chain of events leading to histone dephosphorylation may be negatively controlled by a protein phosphatase 2A, while the phosphatase which actually dephosphorylates H1 could be a protein phosphatase 1. It remains to be determined whether the phosphatase involved here is the same enzyme as that which dephosphorylates H1 at the end of normal mitosis.

Rapid analysis of mitotic histone H1 phosphorylation by cationic disc electrophoresis at neutral pH in minigels.

A new method is described for analysis of histone H1 and other basic proteins by cationic disc electrophoresis in polyacrylamide gels at neutral pH. The multiphasic buffer (disc) system uses Na+ as leading ion, L-histidine as trailing ion, and Hepes as buffering counterion. These "Hepes/histidine gels" have three advantages over conventional acid-urea gels for studies of H1 phosphorylation and dephosphorylation: speed, convenience, and the need for only small amounts of cells or chromatin. Core histones and their acetylated forms can also be separated in gels containing 0.4% Triton X-100. The difference in electrophoretic mobility between mitotic (superphosphorylated) and interphase H1 from HeLa cells is approximately twice as great at neutral pH as at pH 4.5, making it possible to separate these two H1 forms rapidly and easily in Hepes/histidine "minigels" only 5-cm long. Total histones can be rapidly prepared by simply neutralizing 0.2 N HCl extracts, and the entire analysis, from harvesting cells to destaining gels, can be carried out in 1 day. The stacking effect of the disc system produces sharp bands and high resolution even with relatively dilute samples.