Fetal Rat Liver Cells Research Articles

Therapeutic Cell Repopulation of the Liver: From Fetal Rat Cells to Synthetic Human Tissues.

Progenitor cells isolated from the fetal liver can provide a unique cell source to generate new healthy tissue mass. Almost 20 years ago, it was demonstrated that rat fetal liver cells repopulate the normal host liver environment via a mechanism akin to cell competition. Activin A, which is produced by hepatocytes, was identified as an important player during cell competition. Because of reduced activin receptor expression, highly proliferative fetal liver stem/progenitor cells are resistant to activin A and therefore exhibit a growth advantage compared to hepatocytes. As a result, transplanted fetal liver cells are capable of repopulating normal livers. Important for cell-based therapies, hepatic stem/progenitor cells containing repopulation potential can be separated from fetal hematopoietic cells using the cell surface marker δ-like 1 (Dlk-1). In livers with advanced fibrosis, fetal epithelial stem/progenitor cells differentiate into functional hepatic cells and out-compete injured endogenous hepatocytes, which cause anti-fibrotic effects. Although fetal liver cells efficiently repopulate the liver, they will likely not be used for human cell transplantation. Thus, utilizing the underlying mechanism of repopulation and developed methods to produce similar growth-advantaged cells in vitro, e.g., human induced pluripotent stem cells (iPSCs), this approach has great potential for developing novel cell-based therapies in patients with liver disease. The present review gives a brief overview of the classic cell transplantation models and various cell sources studied as donor cell candidates. The advantages of fetal liver-derived stem/progenitor cells are discussed, as well as the mechanism of liver repopulation. Moreover, this article reviews the potential of in vitro developed synthetic human fetal livers from iPSCs and their therapeutic benefits.

Biliary Epithelial Cells Play an Essential Role in the Reconstruction of Hepatic Tissue with a Functional Bile Ductular Network

One of the most challenging issues in liver tissue engineering is the incorporation of a functional bile duct (BD) network in the hepatic tissue. In this study, we investigated the feasibility of a three-dimensional (3D) coculture of adult rat primary biliary epithelial cells (ABECs) and rat fetal liver cells (FLCs) to form functional BD structures. 3D hetero-spheroids containing various ratios of the two cell types were successfully obtained in low-adherence, round-bottom, 96-well plates. Histological analyses and functional analysis using fluorescein diacetate demonstrated that the ABECs critically contributed to the reconstruction of continuous ductular networks. Metabolized fluorescein was transported toward cystic structures in a time- and ABEC ratio-dependent manner. In particular, the duct-like structures containing fluorescein accumulations interlinked the large cystic accumulations. Furthermore, transplantation experiments demonstrated that ABECs, but not FLCs, enabled the establishment of BD networks in vivo. This study provided important insights into the development of transplantable liver tissues with bile excretion functionality.

M6P/IGF2R modulates the invasiveness of liver cells via its capacity to bind mannose 6-phosphate residues

Background & AimsThe mannose 6-phosphate/insulin-like growth factor II receptor (M6P/IGF2R), a multifunctional protein, plays a central role in intracellular targeting of lysosomal enzymes and control of insulin-like growth factor II (IGF-II) bioactivity. Importantly, the gene encoding this receptor is frequently inactivated in a wide range of malignant tumors including hepatocellular carcinomas. Thus, M6P/IGF2R is considered a putative liver tumor suppressor. The aim of this study was to establish the impact of the receptor on the invasive properties of liver cells.MethodsReconstitution experiments were performed by expression of wild type and mutant M6P/IGF2R in receptor-deficient FRL14 fetal rat liver cells. RNA interference was used to induce M6P/IGF2R downregulation in receptor-positive MIM-1–4 mouse hepatocytes.ResultsWe show that the M6P/IGF2R status exerts a strong impact on the invasiveness of tumorigenic rodent liver cells. M6P/IGF2R-deficient fetal rat liver cells hypersecrete lysosomal cathepsins and penetrate extracellular matrix barriers in a cathepsin-dependent manner. Forced expression of M6P/IGF2R restores intracellular transport of cathepsins to lysosomes and concomitantly reduces the tumorigenicity and invasive potential of these cells. Conversely, M6P/IGF2R knock-down in receptor-positive mouse hepatocytes causes increased cathepsin secretion as well as enhanced cell motility and invasiveness. We also demonstrate that functional M6P-binding sites are important for the anti-invasive properties of M6P/IGF2R, whereas the capacity to bind IGF-II is dispensable for the anti-invasive activity of the receptor in liver cells.ConclusionsM6P/IGF2R restricts liver cell invasion by preventing the pericellular action of M6P-modified proteins.

Direct Oxygen Supply with Polydimethylsiloxane (PDMS) Membranes Induces a Spontaneous Organization of Thick Heterogeneous Liver Tissues from Rat Fetal Liver Cells in Vitro

Oxygen is a vital nutrient for growth and maturation of in vitro cells (e.g., adult hepatocytes). We previously demonstrated that direct oxygenation through a polydimethylsiloxane (PDMS) membrane increases the oxygen supply to cell cultures and improves hepatocyte functions. In this study, we removed limits on oxygen supply to fetal rat liver cells through the use of direct oxygenation through a PDMS membrane to investigate in vitro growth and maturation. We chose fetal liver cells because they are considered a feasible source of liver progenitor cells for regenerative medicine therapy due to their highly efficient maturation and proliferation. Cells from 17-day-old pregnant rats were cultured under 5% and 21% oxygen atmospheres. Some cells were first cultured under 5% oxygen, and then switched to a 21% oxygen atmosphere. When oxygen supply was enhanced by a PDMS membrane, the rat fetal liver cells organized into a complex tissue composed of an epithelium of hepatocytes above a mesenchyme-like tissue. The thickness of this supportive tissue was directly correlated to oxygen concentration and was thicker under 5% oxygen. When cultures were switched from 5% to 21% oxygen, lumen-containing structures were formed in the thick mesenchymal-like tissue and the albumin secretion rate increased. In addition, cells adapted their glycolytic activity to the oxygen concentrations. This system promoted the formation of a functional and organized thick tissue suitable for use in regenerative medicine.

Use of liposome encapsulated hemoglobin as an oxygen carrier for fetal and adult rat liver cell culture

Engineering liver tissue constructs with sufficient cell mass for transplantation implies culturing large numbers of hepatocytes in a reduced volume; however, providing sufficient oxygen to dense cell cultures is still not feasible using only conventional culture medium. Liposome-encapsulated hemoglobin (LEH), an oxygen-carrying blood substitute originally designed for short-term perfusion, may be a good candidate as an oxygen carrier to cultured liver cells. In this study, we investigated the feasibility of maintaining long term hepatocyte cultures using LEH. Primary fetal and adult rat liver cells were directly exposed to LEH for 6 to 14 days in static culture or in a perfused flat plate bioreactor. The functions and viability of adult rat hepatocytes exposed to LEH were not adversely affected in static monolayer culture and were even improved in the bioreactor. However, some cytotoxicity of LEH was observed with fetal rat liver cells after 4 days of culture. LEH, though a suitable oxygen carrier for long-term culture of mature hepatocytes, is not suitable in its present form for perfusing fetal hepatocyte cultures in direct contact with the liposomes; either the LEH will have to be made less toxic or a more sophisticated bioreactor that prevents the direct contact between hepatocytes and perfusates will have to be designed if fetal cells are to be used for liver tissue engineering.

Convenient and Efficient Enrichment of the CD133+ Liver Cells from Rat Fetal Liver Cells as a Source of Liver Stem/Progenitor Cells

Although the stem cells are commonly isolated by FACS or MACS, they are very expensive and these is no specific marker for liver stem/progentior cells (LSPCs). This paper applied a convenient and efficient method to enrich LSPCs. The fetal liver cells (FLCs) were firstly enriched by Percoll discontinuous gradient centrifugation (PDGC) from the rat fetal liver. Then the FLCs in culture were purified to be homogeneous in size by differential trypsinization and differential adherence (DTDA). Flow cytometric analysis revealed more than half of the purified FLCs expressed alternative markers of LSPCs (CD117, c-Met, Sca-1, CD90, CD49f and CD133). In other words, the purified FLCs were heterogeneous. Therefore, they were sequentially layered into six fractions by Percoll continuous gradient centrifugation (PCGC). Both CD133 and CD49f expressed decreasingly from fraction 1 to 6. In fraction 1 and 2, about 85% FLCs expressed CD133, which were revealed to be LSPCs by high expressions of AFP and CK-19, low expressions of G-6-P and ALB. To conclude, the purity of CD133(+) LSPCs enriched by combination of PDGC, DTDA and PCGC is close to that obtained by MACS. This study will greatly contribute to two important biological aspects: liver stem cells isolation and liver cell therapy.

Hepatoblast and mesenchymal cell-specific gene-expression in fetal rat liver and in cultured fetal rat liver cells

The aim of this study was to determine whether passaged rat fetal liver cells are functional hepatoblasts. Hepatocyte/hepatoblast- and liver myofibroblast-gene-expressions were studied in adult and fetal rat liver tissues as well as in primary and passaged cultures of isolated rat fetal liver cells at both the mRNA and protein level. Desmin- and Alpha-Smooth Muscle Actin (SMA)-positive cells were located in the walls of liver vessels, whereas Desmin-positive/SMA-negative cells were distributed within the liver parenchyma. Primary cultures contained Prox1-positive hepatoblasts, Desmin/SMA-positive myofibroblasts and only a few Desmin-positive/SMA-negative cells. Albumin and alpha-fetoprotein (AFP) could be detected in the primary cultures and to a lesser extent after the first passage. The number of Desmin-positive/SMA-negative cells decreased with successive passage, such that after the second passage, only Desmin/SMA-positive cells could be detected. SMA-gene-expression increased during the passages, suggesting that myofibroblasts become the major cell population of fetal liver cell cultures over time. This observation needs to be taken into account, should passaged fetal liver cells be used for liver cell transplantation. Moreover it contradicts the concept of epithelial-mesenchymal transformation and suggests rather that selective overgrowth of mesenchymal cells occurs in culture.

Soluble Factor–DependentIn VitroGrowth and Maturation of Rat Fetal Liver Cells in a Three-Dimensional Culture System

Fetal liver cell fractions are potent sources of cells for future liver tissue engineering, by virtue of their high proliferation capacity and their potential for hepatic maturation. Recently, some types of hepatic differentiation agents have been identified from findings in stem cell biology. We therefore investigated the in vitro growth and maturation of rat fetal liver cells isolated from 17-day-old pregnant rats in poly-L-lactic acid three-dimensional (3D) macroporous scaffolds in the presence of soluble factors, such as a combination of hepatocyte growth factor, fibroblast growth factor-1, and fibroblast growth factor-4, oncostatin M, and sodium butyrate. Inclusion of all these factors in the 3D culture induced higher levels of hepatic functions and well maintained these enhanced levels during 2 weeks of culture, whereas in the monolayer culture, such functional enhancement was gradually lost after 1 week. The finally achieved functions on a per-cell basis in the 3D culture with all of the soluble factors were comparable to those of adult hepatocytes. We therefore conclude that the 3D culture system shows promise for the in vitro maturation of fetal liver cells as a means of preconditioning of the cells for engineered liver tissue equivalents in future transplantation studies.

Soluble Factor–DependentIn VitroGrowth and Maturation of Rat Fetal Liver Cells in a Three-Dimensional Culture System

Soluble Factor–Dependent<i>In Vitro</i>Growth and Maturation of Rat Fetal Liver Cells in a Three-Dimensional Culture System

Involvement of signaling of VEGF and TGF-β in differentiation of sinusoidal endothelial cells during culture of fetal rat liver cells

Embryonic development of the liver is closely associated with vascular organization. However, little is known about the mechanisms of vascular differentiation during liver development. Our previous study showed that the maturation of sinusoidal endothelial cells (SECs) occurred during embryonic day 13.0 (E13.0) to E15.0. To improve our understanding of SEC differentiation, we examined here the expression of maturation markers, SE-1 and stabilin-2, in fetal livers and also attempted to establish an in vitro SEC differentiation system by culturing E13.5 fetal liver cells. Immunohistochemical examination of SE-1 and stabilin-2 expression during fetal rat liver development revealed that these differentiation markers were co-expressed in SECs in the late stage of liver development, although stabilin-2 was expressed in almost all vascular endothelial cells in the early stage. Liver cells from the E13.5 rat fetus were cultured in EBM-2 medium containing vascular endothelial growth factor (VEGF), transforming growth factor beta1 (TGF-beta1) and VEGF plus SB-431542 (an inhibitor of the TGF-beta1 receptor, activin receptor-like kinase 5 [ALK-5]). In vitro SEC differentiation, as indicated by the appearance of cells co-expressing SE-1 and stabilin-2 and of cells with cytoplasmic fenestrae in endothelial sheets, was induced by the addition of both VEGF and SB-431542, an inhibitor of the phosphorylation of Smad2/3 but not that of Smad1/5/8 in the cultured cells. These results indicate for the first time that both VEGF signaling and the blocking of the ALK-5-Smad2/3 signal pathway are important for the fetal differentiation of SECs.

Differentiation potential of the fetal rat liver-derived cells.

Mesenchymal stem cells derived from bone marrow or several fetal tissues can be expanded and differentiated into other cell lines. The fetal liver is the source of early hematopoietic cells and also, as a fetal tissue, may be considered as a source of pluripotent stem cells. The differentiation potential of fetal rat liver cells have been examined. Freshly isolated liver cells from 14-d fetuses were cultured in Dulbecco medium supplemented with 10% FCS. The plastic-adherent cells were then passaged up to 10 times. Freshly isolated cells and cells from every passage were cultured in hematopoiesis-promoting environment that consists of methylcelulose supplemented with FCS, rat IL-3, human IL-6 and Epo. Parallely these cells were incubated in co-culture with rat muscle satellite cells (Dulbecco medium with 10% FCS and 10% HS) to examine their myogenic potential. Culture in methylcelulose resulted in a high number of GM and Mix colonies in case of freshly isolated liver cells and the number of colonies decreased according to the number of passages. In case of cells from 4th passage, there ware no hematopoietic colonies in culture. In contrast--freshly isolated cells were not able to fuse with rat satellite cells and form the myotubes. This ability appeared in plastic-adherent cells just from the second passage and increases to 5th passage. The cells from every next passage up to 10th when co-cultured with satellite cells participated in myotube formation at the same high level. This result may suggest that in the 14-d rat liver there exist at least two subpopulations of cells: the non-adherent hematopoietic cell population, and the population of plastic-adherent cells capable of differentiating into myotubes. Since the attempts to redifferentiate hematopoietic subpopulation into myopoiesis, or myopoietic subpopulation into hematopoiesis failed, it may be concluded that at least under our experimental conditions the fetal liver cells do not reveal the "plasticity" features.

Hepatocyte transplantation

Hepatocyte transplantation

Early T cell development can be traced in rat fetal liver.

The degree of T cell commitment reached by cell precursors present in the fetal liver is a controversial issue. In the present work, the occurrence of fully T cell-committed progenitors among CD45+Thy-1+CD44+ 13-day-old rat fetal liver cells was demonstrated when limiting numbers of these cells in vitro reconstituted SCID mouse fetal thymic lobes providing single lineage-containing lobes for T, natural killer or dendritic cells. In addition, expression of rat pre-TCRalpha chain mRNA was detected in the CD45+ but not in the CD45- fetal liver cells and fully rearranged TCR VBeta8-Cbeta mRNA transcripts were specifically detected in the former population, demonstrating early transcription of some rearranged TCRVBeta genes in the rat fetal liver of 13 days of gestation. Finally, fetal liver organ cultures provided low numbers of TCR gamma delta T cells and CD2+CD8+NKR-P1A- intracytoplasmic CD3+ immature T cells, which intracellularly reacted with a mAb specific to the TCRalpha Beta molecule. These results prove T, NK and DC cell lineage determination at a prethymic stage in the fetal liver.

Follicular Dendritic Cell (FDC) Precursors in Primary Lymphoid Tissues

The origin of follicular dendritic cells (FDC) is unresolved, and as such, remains controversial. Based on the migration of Ag-transporting cells (ATC) into lymphoid follicles and the phenotypic similarity between FDC and ATC, one hypothesis is that ATC may represent emigrating FDC precursors. This contrasts with the view that FDC originate from local stromal cells in the secondary lymphoid tissues. Mice homozygous for the severe combined immunodeficiency (prkdc(scid)) mutation (scid) lack FDC. Thus, they provide a powerful tool for assessing de novo generation of FDC. To test whether FDC precursors could be found in bone marrow or fetal liver, scid/scid mice were reconstituted with either: 1) bone marrow cells from (BALB/c x C57BL/6)F1 donors, 2) bone marrow cells from ROSA BL/6 F1 (lacZ-transfected) mice, 3) rat bone marrow cells, or 4) rat fetal liver cells. Six to eight weeks after reconstitution with F1 bone marrow, cells reactive with the FDC-labeling mAb, FDC-M1, also expressed donor class I molecules on their surfaces. Similarly in mice reconstituted with lacZ-transfected bone marrow cells, these cells were also positive for the lacZ gene product. Furthermore, in spleens of animals reconstituted with either rat bone marrow or rat fetal liver, rat FDC were identified using the specifically labeling mAb, ED5. In all cases, host FDC were also present, indicating that scid/scid mice have FDC precursors that will mature in the presence of allogeneic or xenogeneic lymphoid cells. In summary, FDC can be derived from progenitor cells present in primary lymphoid tissues.

Perinatal induction of immunotolerance to cardiac and pulmonary allografts

Objectives: Tolerance appears to be more easily induced in the fetus before full immunocompetence is established, but elucidation of this process is needed. A model of perinatal tolerance induction to neonatal skin allografts followed by cardiac and pulmonary allografts is described. Methods: Sixty Lewis (RT1 1) rat fetuses were inoculated intraperitoneally at 18 days' gestation with 1 × 10 7 ACI (RT1 a) rat fetal liver cells (group I); 20 Lewis fetuses were inoculated with 2 × 10 7 ACI fetal liver cells (group II). Control groups consisted of Lewis fetuses inoculated with saline solution ( n = 25, group III) and fetuses that were not inoculated ( n = 25, group IV). Twenty-five of the 50 surviving group I rats received ACI skin (<24 hours old) and heart (8 to 10 weeks old) allografts (group IA); the remaining 25 rats received only ACI heart grafts (group IB). Groups II, III, and IV received ACI skin and cardiac allografts. Recipients tolerant to both skin and cardiac grafts received orthotopic ACI lung grafts and third-party skin grafts. Tolerance was indicated by graft survival for more than 100 days. Limiting dilution and flow cytometric analyses were performed. Results: Abortion rates in groups I, II, III, and IV were 17% (10/60), 65% (13/20), 8% (2/25), and 4% (1/25), respectively. Specific tolerance to skin, cardiac, and lung allografts was observed in seven of 25 group IA recipients (28%) and seven of seven group II recipients (100%) compared with no tolerance in any group IB, III, or IV recipients ( p = 0.03, χ 2 test). A 100-fold reduction of precursor cytotoxic T lymphocytes and significant splenocyte and bone marrow chimerism in tolerant versus nontolerant rats were noted ( p = 0.0001, Student's t test). Conclusions: Using donor-strain fetal liver cells and neonatal skin grafts, we achieved higher frequencies of tolerance to solid organ grafts in adulthood with lower cell inocula and abortion rates than previously described. Chimerism and depressed precursor cytotoxic T lymphocyte frequencies in tolerant recipients suggest that hematopoietic stem cell engraftment and clonal deletion/anergy are involved in induction of perinatal tolerance(J Thorac Cardiovasc Surg 1997;114:64-75)

Identification of a novel rat B cell subset in the peritoneal cavity of xenogeneic rat to mouse scid chimeras

In the mouse B-1 cells represent a small subset of B cells with a unique phenotype, anatomical distribution pattern, function and developmental origin. 1 In essence, these B cells are self-replenishing cells that are strongly enriched in the peritoneal cavity (PerC) as IgM highIgD lowCD11b lowB220 low cells. Most of the peritoneal B-1 cells also express low levels of CD5 (B-la cells) whereas a minority lack CD5 completely (B-1b cells). So far, we have not been able to unequivocally detect the presence of B-1 lineage cells (or their homologues) in rats. 2–4 In adult animals, CD5-expressing B cells are absent, whereas in neonates all (splenic) B cells seem to express low amounts of CD5, which become gradually lower with age. Possible detection of B-1 cell (homologues) in rats might be hampered by the notion that in this species there are virtually no B cells in the PerC, in marked contrast to mice. 2,3 Here we studied whether the relative absence of B cells from the rat PerC is an inherent property of the leukocytes themselves or due to non-leukocyte factors, such as a difference in microscopical anatomic organization or properties of mesothelial cells. To this end, we constructed xenogeneic rat-to-mouse SCID chimeras by injecting rat fetal liver (FL) cells into lightly irradiated C.B17 SCID mice. As shown by Surh and Sprent rat B cells develop readily in the mouse SCID environment. 5 Here we demonstrate that the PerC of these xenogenic rat-to-mouse SCID chimeras harbor high numbers of B cells. In addition, we show that a relative high proportion of the PerC B cells have a unique, previously undetected phenotype (IgM highIgD lowHIS24/B220 lowThy-1 high), to some extent resembling splenic marginal zone B cells. We speculate that this novel B cell subset might represent the rat homologue of the B-1 cells in the mouse.

Analysis of fetal rat liver using monoclonal antibodies.

In previous studies, we developed several monoclonal antibodies (mAbs) against fetal and adult rat liver to analyze the hemopoietic microenvironment of the fetal liver during the gestational period. In this study, we have developed two new mAbs against fetal rat liver cells and have examined the characteristics at various gestational ages of fetal liver and of adult liver. The characteristics of these monoclonal antibodies were demonstrated by examining several tissues using immunohistochemical staining and flow cytofluorometry. Monoclonal antibodies HAM10 and HAM11 were developed against fetal rat liver cells. These reacted with the cytoplasm of fetal and adult hepatocytes. HAM10 antigen expression was strong at approximately day 18 of gestation in the active period of hemopoiesis in fetal rat liver but was much lower in adult liver. HAM10 antigen expression also increased in liver after partial hepatectomy and was reduced abruptly to a normal level thereafter. HAM11 antigen expression in fetal liver was weaker than that of HAM10 antigen expression. The degree of HAM11 antigen expression increased as gestation proceeded, reaching a maximum in adult liver. Both HAM10 and HAM11 antigens may play a role in the morphogenesis of hepatocytes and in the hemopoietic microenvironment for hemopoietic cells. Moreover, HAM10 antigen is may play a role in hepatocyte proliferation in the fetal liver, whereas HAM11 antigen may contribute to the maturation of fetal- to the adult-type hepatocytes.

The myelin basic protein‐specific T cell repertoire in (transgenic) Lewis rat/SCID mouse chimeras: preferential Vβ8.2 T cell receptor usage depends on an intact Lewis thymic microenvironment

In the Lewis rat, myelin basic protein (MBP)-specific, encephalitogenic T cells preferentially recognize sequence 68-88, and use the V beta 8.2 gene to encode their T cell receptors. To analyze the structural prerequisites for the development of the MBP-specific T cell repertoire, we reconstituted severe-combined immunodeficient (SCID) mice with fetal (embryonic day 15-16) Lewis rat lymphoid tissue, and then isolated MBP-specific T cell lines from the adult chimeras after immunization. Two types of chimera were constructed: SCID mice reconstituted with rat fetal liver cells only, allowing T cell maturation within a chimeric SCID thymus consisting of mouse thymic epithelium and rat interdigitating dendritic cells, and SCID mice reconstituted with rat fetal liver cells and rat fetal thymus grafts, allowing T cell maturation within the chimeric SCID and the intact Lewis rat thymic microenvironment. Without exception, the T cell lines isolated from MBP-immunized SCID chimeras were restricted by MHC class II of the Lewis rat (RT1.B1), and none by I-Ad of the SCID mouse. Most of the T cell lines recognized the immunodominant MBP epitope 68-88. In striking contrast to intact Lewis rats, in SCID mice reconstituted by rat fetal liver only, MBP-specific T cell clones used a seemingly random repertoire of V beta genes without a bias for V beta 8.2. In chimeras containing fetal Lewis liver plus fetal thymus grafted under the kidney capsule, however, dominant utilization of V beta 8.2 was restored. The migration of liver-derived stem cells through rat thymus grafts was documented by combining fetal tissues from wild-type and transgenic Lewis rats. The results confirm that the recognition of the immunodominant epitope 68-88 by MBP-specific encephalitogenic T cells is a genetically determined feature of the Lewis rat T cell repertoire. They further suggest that the formation of the repertoire requires T cell differentiation in a syngeneic thymic microenvironment.

Evaluation of oncofetal protein-related mRNA transport activity as a potential early cancer marker in dogs with malignant neoplasms

SUMMARY A 55-kd protein with mRNA transport activity found in fetal rat liver cells and plasma from mice, rats, and human beings with malignant neoplasms has been designated oncofetal protein 55 (ofp55). Monoclonal antibody produced to rat ofp55 cross-reacts with human ofp55. Using this monoclonal antibody in a bioassay measuring mRNA transport stimulated by ofp55, we tested the plasma from 19 dogs with a variety of malignant neoplasms, including carcinomas, sarcomas, lymphomas, and melanomas, and compared the results with plasma from 20 clinically normal dogs without evidence of neoplasia. The mean mRNA transport activity from the group of dogs with malignant neoplasms was 0.43 ± 0.28%/mg of protein. Mean transport activity from the group of control dogs was 0.04 ± 0.02%/mg of protein. These means were significantly different (P &lt; 0.0001). The degree of overlap between these 2 groups in their ofp55-related mRNA transport activity was minimal, and measurement of this protein appears to have potential for the early detection of malignant neoplasms in dogs.

Establishment of a Xenogeneic Chimera without GVHD in NK Cell-Depleted SCID Mice by Grafting Rat Fetal Liver Cells

Rat lymphopoiesis did not develop when naive SCID mice were transplanted with rat fetal liver cells. However, when SCID mice were depleted of NK cells by administration of anti-murine IL-2Rβ mAb before transplantation, remarkable reconstitution of both rat T and B cells was observed in these mice without any evidence of graft-versus-host disease macroscopically or histologically. T cells in these reconstituted mice proliferated well in response to Con A and third-party rat and mouse antigens, whereas no response was seen to the stimulation with either donor rat- or host mouse-type cells. When these xenogeneic chimera mice had been immunized with SRBC, these mice exhibited DTH reaction and antibody production against the homologous antigen. These results indicate that rat fetal liver cells can differentiate to functional T and B cells in the xenogeneic microenvironment of SCID mice, if host NK cells are depleted beforehand. These rat-type T cells develop within SCID thymuses and acquire tolerance to either donor F344 rat or host SCID mouse antigens.