Evidence suggests that haematopoietic stem cells might have unexpected developmental plasticity, highlighting therapeutic potential. For example, bone-marrow-derived hepatocytes can repopulate the liver of mice with fumarylacetoacetate hydrolase deficiency and correct their liver disease. To determine the underlying mechanism in this murine model, we performed serial transplantation of bone-marrow-derived hepatocytes. Here we show by Southern blot analysis that the repopulating hepatocytes in the liver were heterozygous for alleles unique to the donor marrow, in contrast to the original homozygous donor cells. Furthermore, cytogenetic analysis of hepatocytes transplanted from female donor mice into male recipients demonstrated 80,XXXY (diploid to diploid fusion) and 120,XXXXYY (diploid to tetraploid fusion) karyotypes, indicative of fusion between donor and host cells. We conclude that hepatocytes derived form bone marrow arise from cell fusion and not by differentiation of haematopoietic stem cells.

Proteins are inherently flexible at ambient temperature. At equilibrium, they are characterized by a set of conformations that undergo continuous exchange within a hierarchy of spatial and temporal scales ranging from nanometers to micrometers and femtoseconds to hours. Dynamic properties of proteins are essential for describing the structural bases of their biological functions including catalysis, binding, regulation and cellular structure. Nuclear magnetic resonance (NMR) spectroscopy represents a powerful technique for measuring these essential features of proteins. Here we provide an introduction to NMR-based approaches for studying protein dynamics, highlighting eight distinct methods with recent examples, contextualized within a common experimental and analytical framework. The selected methods are (1) Real-time NMR, (2) Exchange spectroscopy, (3) Lineshape analysis, (4) CPMG relaxation dispersion, (5) Rotating frame relaxation dispersion, (6) Nuclear spin relaxation, (7) Residual dipolar coupling, (8) Paramagnetic relaxation enhancement.

The appearance of bipotential oval cells in chronic liver injury suggests the existence of hepatocyte progenitor͞stem cells. To study the origin and properties of this cell population, oval cell proliferation was induced in adult mouse liver by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) and a method for their isolation was developed. Transplantation into fumarylacetoacetate hydrolase (Fah) deficient mice was used to determine their capacity for liver repopulation. In competitive repopulation experiments, hepatic oval cells were at least as efficient as mature hepatocytes in repopulating the liver. In mice with chimeric livers, the oval cells were not derived from hepatocytes but from liver nonparenchymal cells. This finding supports a model in which intrahepatic progenitors differentiate into hepatocytes irreversibly. To determine whether oval cells originated from stem cells residing in the bone marrow, bone marrow transplanted wild-type mice were treated with DDC for 8 months and oval cells were then serially transferred into Fah mutants. The liver repopulating cells in these secondary transplant recipients lacked the genetic markers of the original bone marrow donor. We conclude that hepatic oval cells do not originate in bone marrow but in the liver itself, and that they have valuable properties for therapeutic liver repopulation. In mammals, hepatic cells, especially hepatocytes can rapidly proliferate after acute liver injury to repair liver damage (1). Usually, undifferentiated liver progenitors do not take part in such acute liver regeneration. However, hepatocyte progenitors are still required in some chronic injury responses, especially when the ability of differentiated hepatocytes to divide is impaired (reviewed in refs. 2 and 3). The existence of endogenous stem cells inside mammalian liver was first suggested by the observation that treatment with carcinogens results in the emergence of oval cells in the portal region of the hepatic lobule (4-9). Oval cells are Ϸ10 m in size, have a high nuclear͞ cytoplasmic ratio, and express markers of immature liver cells such as ␣-fetoprotein (AFP) (10, 11). In addition, oval cells express markers of both the biliary epithelium (CK19) and hepatocyte lineages (albumin). In vitro, these cells can be differentiated into both of these hepatic epithelial lineages under appropriate conditions.Although the anatomical location of the oval cell proliferation suggests the existence of a hepatocyte progenitor in the Canal of Hering (12), the precise origin of oval cells remains uncertain. Most studies imply the existence of an undifferentiated oval cell precursor in this location that proliferates and gives rise to oval cells (13)(14)(15)(16). In analogy to the hematopoietic system, oval cells would then represent a committed bipotential progenitor. Recent studies in both rat and human have suggested that oval cells could be derived from precursors in the bone marrow (17). Others, however, have postulated that oval cells may arise by dedifferentiation of hepatocytes,...

Liver repopulation with bone marrow-derived hepatocytes (BMHs) can cure the genetic liver disease fumarylacetoacetate hydrolase (Fah) deficiency. BMHs emerge from fusion between donor bone marrow-derived cells and host hepatocytes. To use such in vivo cell fusion efficiently for therapy requires knowing the nature of the hematopoietic cells that fuse with hepatocytes. Here we show that the transplantation into Fah(-/-) mice of hematopoietic stem cells (HSCs) from lymphocyte-deficient Rag1(-/-) mice, lineage-committed granulocyte-macrophage progenitors (GMPs) or bone marrow-derived macrophages (BMMs) results in the robust production of BMHs. These results provide direct evidence that committed myelomonocytic cells such as macrophages can produce functional epithelial cells by in vivo fusion. Because stable bone marrow engraftment or HSCs are not required for this process, macrophages or their highly proliferative progenitors provide potential for targeted and well-tolerated cell therapy aimed at organ regeneration.