CD133 is a positive marker for a distinct class of primitive human cord blood-derived CD34-negative hematopoietic stem cells.

The identification of human CD34-negative (CD34(-)) hematopoietic stem cells (HSCs) provides a new concept for the hierarchy in the human HSC compartment. Previous studies demonstrated that CD34(-) severe combined immunodeficiency (SCID)-repopulating cells (SRCs) are a distinct class of primitive HSCs in comparison to the well-characterized CD34(+)CD38(-) SRCs. However, the purification level of rare CD34(-) SRCs in 18 lineage marker-negative (Lin(-)) CD34(-) cells (1/1000) is still very low compared with that of CD34(+)CD38(-) SRCs (1/40). As in the mouse, it will be necessary to identify useful positive markers for a high degree of purification of rare human CD34(-) SRCs. Using 18Lin(-)CD34(-) cells, we analyzed the expression of candidate positive markers by flow cytometric analysis. We finally identified CD133 as a reliable positive marker of human CB-derived CD34(-) SRCs and succeeded in highly purifying primitive human CD34(-) HSCs. The limiting dilution analysis demonstrated that the incidence of CD34(-) SRCs in 18Lin(-)CD34(-)CD133(+) cells was 1/142, which is the highest level of purification of these unique CD34(-) HSCs to date. Furthermore, CD133 expression clearly segregated the SRC activities of 18Lin(-)CD34(-) cells, as well as 18Lin(-)CD34(+) cells, in their positive fractions, indicating its functional significance as a common cell surface maker to isolate effectively both CD34(+) and CD34(-) SRCs.

In vivo dynamics of human cord blood-derived CD34(-) SCID-repopulating cells using intra-bone marrow injection.

The identification of human CD34-negative (CD34(-)) hematopoietic stem cells (HSCs) provides a new concept for the hierarchy in the human HSC compartment. This study investigated the long-term repopulating capacity and redistribution kinetics of human cord blood-derived CD34(-) severe combined immunodeficiency (SCID)-repopulating cells (SRCs) and compared them with those of CD34(+)CD38(+) and CD34(+)CD38(-) SRCs using the intra-bone marrow injection (IBMI) to clarify the characteristics of CD34(-) SRCs. On the basis of the limiting dilution analyses data, estimated numbers of CD34(+)CD38(+), CD34(+)CD38(-), and CD34(-) SRCs were transplanted to NOD/SCID mice by IBMI. The human cell repopulation at the site of injection and the other bones were serially investigated. Interestingly, CD34(+)CD38(+), CD34(+)CD38(-), and CD34(-) SRCs began to migrate to other bones 2 and 5 weeks after the transplantation, respectively. Accordingly, the initiation of migration seemed to differ between the CD34(+) and CD34(-) SRCs. In addition, CD34(+)CD38(+) SRCs only sustained a short-term repopulation. However, both CD34(+)CD38(-) and CD34(-) SRCs had longer-term repopulation capacity. Taken together, these findings showed that CD34(-) SRCs show different in vivo kinetics, thus suggesting that the identified CD34(-) SRCs are a distinct class of primitive HSCs in comparison to the CD34(+)CD38(+) and CD34(+)CD38(-) SRCs.

Parity-violating energy shifts of Murchison L-amino acids are consistent with an electroweak origin of meteorite L-enantiomeric excesses.

In 1996, four alpha-methyl amino acids in the Murchison meteorite--L-isovaline, L-alpha-methylnorvaline, L-alpha-methyl-allo-isoleucine and L-alpha-methyl-isoleucine--were found to show significant enantiomeric excesses of the L form, ranging from 2% to 9%. Their deuterium to hydrogen isotope ratios suggest they formed in the pre-solar interstellar gas cloud rather than during a later aqueous processing phase on the asteroid parent body. In this paper we apply the techniques of the preceding two papers to compute the parity-violating energy shifts of these amino acids. We find that, in the gas phase, the PVESs of the neutral L forms of all four Murchison alpha-methyl amino acids are decisively negative, and there is even some correlation between the magnitudes of the L-excesses and the magnitudes of the PVESs--all of which is at least consistent with an electroweak origin of the Murchison enantiomeric excesses.

Electroweak parity-violating energy shifts of amino acids: the "conformation problem".

The preceding paper described our coupled-perturbed Hartree-Fock (CPHF) and density functional theory (DFT) methods of computing the parity-violating energy shift (PVES). This paper addresses the "conformation problem"-the difficulty determining which hand of amino acids in solution is favoured by the weak force due to the difficulty determining the solution conformation. We attempt to resolve this by using the methods of the preceding paper to compute the PVES of solution and gas-phase amino acid structures determined by other groups from high level optimizations that include solvation. We conclude that the conformational hypersensitivity of the PVES still precludes a definite conclusion as to the sign of the PVES of L-alanine in solution, but that there is no problem in the gas phase: the PVES of gas-phase L-alanine is decisively negative. We show that the PVES is very sensitive to certain torsion angles, but is not hypersensitive to bondlengths or bond angles. In determining structures for PVES computations, there is therefore no need for expensive full optimizations: one can just optimize the crucial torsion angles. We present new computations of gas-phase amino acids PVESs, using partial optimizations with small basis sets, and the results agree well with those from higher level techniques. In the following paper we apply these less costly techniques to larger amino acids. The "conformation problem" has led some to dismiss the PVES as the source of life's handedness, but we believe this is premature: we show here that amino acids are a special case because their favoured conformations are almost achiral.