Faculty Bibliography

A large number of cancer stem cells (CSCs) have been isolated and identified; however, none has been cultured in an unlimited manner in vitro without losing tumorigenicity and multipotency. In this study, we successfully clonogenically cultured a newly identified CD34+ liver CSC (LCSC) on feeder cells up to 22 passages (to date) without losing CSC property. Cloned CD34+ LCSC formed a round packed morphology and it could also be cryopreserved and recultured. Stem cell markers, CD34, CD117, and SOX2; normal liver stem cell markers, alpha fetoprotein, CK19, CK18, and OV6; putative CSC markers, CD44, CD133, EpCAM, and CD90; as well as CD31 were expressed in cloned CD34+ LCSC. SOX2 was the major factor in maintaining this LCSC before colonization, and interestingly, OCT4, SOX2, NAONG, Klf4, c-Myc, and Lin28 were upregulated in association with symmetric self-renewal for colony growth of CD34+ LCSC on feeder cells. Gene expression patterns of in vitro differentiation were consistent with our in vivo finding; furthermore, the tumorigenicity of cloned CD34+ LCSC was not different from uncloned CD34+ LCSC sorted from parental PLC. These results show that our cloned CD34+ LCSC maintained CSC property, including self-renewal, bipotency, and tumorigenicity after long-term culture, demonstrating that this LCSC can be cultured in an unlimited manner in vitro. Thus, establishing pure population of CSCs isolated from the patients will provide an opportunity to explore the mechanisms of tumorigenesis and cancer development, and to identify unique biomarkers presenting potential indicators of drug efficacy against CSCs for establishment of a novel strategy for cancer therapy.

A large number of cancer stem cells (CSCs) were identified and characterized; however, the origins and formation of CSCs remain elusive. In this study, we examined the origination of the newly identified CD34(+) liver CSC (LCSC). We found that CD34(+) LCSC coexpressed liver stem cell and myelomonocytic cell markers, showing a mixed phenotype, a combination of hepatobiliary stem/progenitor cells (HSPCs) and myelomonocytic cells. Moreover, human xenografts produced by CD34(+) LCSCs and the parental cells, which CD34(+) LCSC was isolated from, coexpressed liver cancer and myelomonocytic markers, also demonstrating mixed phenotypes. The xenografts and the parental cells secreted albumin demonstrating their hepatocyte origin and also expressed cytokines [interleukin (IL)-1b, IL-6, IL-12A, IL-18, tumor necrosis factor-alpha (TNF-alpha), and CSF1] and chemokines (IL-8, CCL2, and CCL5). Expression of these cytokines and chemokines responded to the stimuli [interferon-gamma (INF-gamma), IL-4, and lipopolysaccharide (LPS)]. Furthermore, human xenografts and the parental cells phagocytized Escherichia coli. CD34(+) LCSC coexpressed CD45, demonstrating that its origin appears to be from a hematopoietic precursor. The percentage of cells positive for OV6, CD34, and CD31, presenting the markers of HSPC, hematopoietic, and myelomonocytic cells, increased under treatment of CD34(+) LCSC with a drug. Cytogenetic analysis showed that CD34(+) LCSC contained a greater number of chromosomes. HBV DNA integrations and mutations in CD34(+) LCSC and the parental cells were identical to those in the literature or the database. Thus, these results demonstrated that CD34(+) LCSCs were formed by fusion of HSPC with CD34(+) hematopoietic precursor-derived myeloid intermediates; it appears that this is the first report that human CSCs have been formed by the fusion. Therefore, it represents a significant step toward better understanding of the formation of human CSC and the diverse origins of liver cancers.

This article briefly reviews the biofield hypothesis and its scientific literature. Evidence for the existence of the biofield now exists, and current theoretical foundations are now being developed. A review of the biofield and related topics from the perspective of physical science is needed to identify a common body of knowledge and evaluate possible underlying principles of origin of the biofield. The properties of such a field could be based on electromagnetic fields, coherent states, biophotons, quantum and quantum-like processes, and ultimately the quantum vacuum. Given this evidence, we intend to inquire and discuss how the existence of the biofield challenges reductionist approaches and presents its own challenges regarding the origin and source of the biofield, the specific evidence for its existence, its relation to biology, and last but not least, how it may inform an integrated understanding of consciousness and the living universe.

CD34+ stem cells play an important role during liver development and regeneration. Thus, we hypothesized that some human liver carcinomas (HLCs) might be derived from transformed CD34+ stem cells. Here we determined that a population of CD34+ cells isolated from PLC/PRF/5 hepatoma cells (PLC) appears to function as liver cancer stem cells (LCSC) by forming human liver carcinomas (HLC) in immunodeficient mice with as few as 100 cells. Moreover, the CD34+ PLC subpopulation cells had an advantage over CD34- PLCs at initiating tumors. Three types of HLCs were generated from CD34+ PLC: hepatocellular carcinomas, HCC; cholangiocarcinomas, CC; and combined hepatocellular cholangiocarcinomas, CHC. Tumors formed in mice transplanted with 12 subpopulations and 6 progeny subpopulations of CD34+ PLC cells. Interestingly, progenies with certain surface antigens (CD133, or CD44, or CD90, or EPCAM) predominantly yielded HCCs. CD34+PLCs that also expressed OV6 and their progeny OV6+ cells produced primarily CHC and CC.This represents the first experiment to demonstrate that the OV6+ antigen is associated with human CHC and CC. CD34+PLCs that also expressed CD31 and their progeny CD31+ cells formed CHCs. Gene expression patterns and tumor cell populations from all xenografts exhibited diverse patterns, indicating that tumor-initiating cells (TIC) with distinct antigenic profiles contribute to cancer cell heterogeneity. Therefore, we identified CD34+ PLC cells functioning as LCSCs generating three types of HLCs. Eighteen subpopulations from one origin had the capacity independently to initiate tumors, thus functioning as TICs. This finding has broad implications for better understanding of the multistep model of tumor initiation and progression. Our finding also indicates that CD34+ PLCs that also express OV6 or CD31 result in types of HLCs. This is the first report that PLC/PRF/5 subpopulations expressing CD34 in combination with particular antigens defines categories of HLCs, implicating a diversity of origins for HLC.

Epithelial cell adhesion molecule (EpCAM) is a transmembrane glycoprotein, which is frequently and highly expressed on carcinomas, tumor-initiating cells, selected tissue progenitors, embryonic and adult stem cells. During liver development, EpCAM demonstrates a dynamic expression since it can be detected in fetal liver, including cells of the parenchyma, whereas mature hepatocytes are devoid of EpCAM. Liver regeneration is associated with a population of EpCAM-positive cells within ductular reactions, which gradually lose the expression of EpCAM along with maturation into hepatocytes. EpCAM can be switched on and off through a wide panel of strategies to fine-tune EpCAM-dependent functional and differentiative traits. EpCAM-associated functions relate to cell-cell adhesion, proliferation, maintenance of a pluripotent state, regulation of differentiation, migration and invasion. These functions can be conferred either by the full-length protein and/or EpCAM-derived fragments, which are generated upon regulated intramembrane proteolysis. Control by EpCAM, therefore, not only occurs in the presence or lack of full-length EpCAM at cellular membranes, but also in varying rates of the formation of EpCAM-derived fragments that have their own regulatory properties, and in changes in the association of EpCAM with interaction partners. Thus, spatiotemporal localization of EpCAM in immature liver progenitors, transit-amplifying cells and mature liver cells will decisively impact on the regulation of EpCAM functions and might be one of the triggers that contribute to the adaptive processes in stem/progenitor cell lineages. This review will summarize EpCAM-related molecular events and how they relate to hepatobiliary differentiation and regeneration.