Isolation and characterization of a spontaneously transformed malignant mouse mammary epithelial cell line in culture

A method is described that permits the selection of spontaneously transformed mammary epithelial colonies from an untransformed mouse mammary epithelial cell line, NMuMG, and utilizes a long-term anchorage-independent growth of the transformants on soft agarose. These transformed cells (NMuMG-ST) are shown to be distinguishable from the untransformed cells by morphology, growth characteristics, induced carcinomas when transplanted into nude mice and ability to metastasize. This transformed phenotype displayed focal, multilayer growth and higher saturation density in comparison with the untransformed phenotype. Transplanted tumors as well as metastatic lung tumors in nude mice were adenocarcinomas morphologically similar to typical mammary tumors in humans. This selection procedure of mutant mammary cells from an immortalized cell line derived from normal mammary glands could be very useful to identify the genomic biomarkers in the growth regulation and malignant progression of breast cancer.     

Thyrotropic action of human chorionic gonadotropin

Hyperthyroidism or increased thyroid function has been reported in many patients with trophoblastic tumors. In these cases, greatly increased human chorionic gonadotropin (hCG) levels and suppressed TSH levels suggest that hCG has thyrotropic activity. Recent investigations have clarified the structural homology not only in the hCG and TSH molecules but also in their receptors, and this homology suggests the basis for the reactivity of hCG with the TSH receptor. The clinical significance of the thyrotropic action of hCG is now also recognized in normal pregnancy and hyperemesis gravidarum. Highly purified hLH binds to recombinant hTSH receptor and is about 10 times as potent as purified hCG in increasing cAMP. The beta-subunits of hCG and hLH share 85% sequence identity in their first 114 amino acids but differ in the carboxy-terminal peptide because hCG beta contains a 31-amino acid extension (beta-CTP). A recombinant mutant hCG that lacks beta-CTP showed almost identical potency to LH on stimulation of recombinant hTSH receptor. If intact hCG were as potent as hLH in regard to its thyrotropic activity, most pregnant women would become thyrotoxic. One of the roles of the beta-CTP may be to prevent overt hyperthyroidism in the first trimester of pregnancy when a large amount of hCG is produced by the placenta. Nicked hCG preparations, obtained from patients with trophoblastic disease or by enzymatic digestion of intact hCG, showed approximately 1.5- to 2-fold stimulation of recombinant hTSH receptor compared with intact hCG. This suggests that the thyrotropic activity of hCG may be influenced by the metabolism of the hCG molecule itself. Deglycosylation and/or desialylation of hCG enhances its thyrotropic potency. Basic hCG isoforms with lower sialic acid content extracted from hydatidiform moles were more potent in activating adenylate cyclase, and showed high bioactivity/immunoactivity (B/I) ratio in CHO cells expressing human TSH receptors. This is consistent with the finding that the beta-CTP truncated hCG with higher thyrotropic potency is substantially deglycosylated and desialylated in the beta-subunit relative to intact hCG because all four O-linked glycosylation sites occur within the missing C-terminal extension. The desialylated hCG variant also interacts directly with recombinant hTSH receptors transfected into human thyroid cancer cells. There is thyroid-stimulating activity in sera of normal pregnant women, and this correlates with serum hCG levels. The thyroid gland of normal pregnant women may be stimulated by hCG to secrete slightly excessive quantities of T4 and induce a slight suppression of TSH, perhaps being about 1 mU/L less than nongravid levels, but not high enough to induce overt hyperthyroidism. Maternal thyroid glands may secrete more thyroid hormone during early pregnancy in response to the thyrotropic activity of hCG that overrides the normal operation of the hypothalamic-pituitary-thyroid feedback system. Biochemical hyperthyroidism associated with hyperemesis gravidarum has been attributed to hCG. In patients with hyperemesis gravidarum, thyrotropic in serum correlated with hCG immunoreactivity, and the severity of vomiting as indicated by clinical and biochemical parameters correlated with the degree of thyroid stimulation. To understand the thyrotropic action of hCG, it is necessary to know whether hCG activates the same domain of the TSH receptor as does TSH. The identification of the molecular structure of the hCG isoform with the highest thyrotropic potency will resolve the enigma of gestational thyrotoxicosis and the hyperthyroidism associated with trophoblastic disease and hCG-producing tumors.     

Dietary fat and breast cancer metastasis by human tumor xenografts

Human breast cancer cell lines growing as xenografts in athymic nude mice have been used to examine the effects of dietary fat and fatty acids on tumor progression. The estrogen independent MDA-MB-435 cell line has the advantage that it metastasizes consistently to the lungs and forms quantifiable secondary nodules when injected into the mammary fat pads. With these breast cancer cells, the stimulating effects of polyunsaturated omega-6 fatty acids on both primary tumor growth and metastasis were demonstrated; in contrast, the long-chain omega-3 fatty acids were inhibitory. The model can also be adapted to examine dietary fatty acids, and inhibitors of their metabolism, as experimental adjuvant therapy after surgical excision of the primary tumors. Unfortunately, estrogen dependent human breast cancer cells do not metastasize, or do so rarely, in nude mice; in consequence, it is not possible to use the model to study estrogen-fatty acid interactions on the metastatic process. In addition to metastasis from a primary location, intravenous injection of MDA-MB-435 cells into the nude mouse host, particularly when combined with studies using Matrigel-based in vitro invasion assays, permits further dissection of the steps in the metastatic cascade which are influenced by dietary fatty acids. The results obtained by these several approaches have demonstrated distinct roles for the cyclooxygenase and lipoxygenase-mediated products of omega-6 fatty acid metabolism, and suggest new approaches to experimental breast cancer therapy.     

Networking rural health resources. The woman who cried wolf

OBJECTIVE: To study the genetic stability and to establish a method for detection of micrometastasis using microsatellite DNA in human breast cancer xenograft in nude mice. METHODS: Fresh tissue of human breast cancer was xenotransplanted in nude mice or thotopically. Genomic DNA extracted from tissues of human breast cancer, xenotransplanted tumors and metastatic foci in nude mice were PCR amplified at three microsatellite loci (D14S68, D18S69, D20S199) and were analysed by electrophoresis and silver stain on PAGE. RESULTS Microsatellite DNA in genome of the xenotransplanted tumors and metastatic foci in nude mice were identical with that of the human breast cancer. CONCLUSION: This study has demonstrated in nude mice the xenotransplanted tumors and metastatic foci that originated from human breast cancer. The genetic stability in human breast cancer is evident in the processes of xenotransplantation, serial passages in nude mice, metastasis and in vitro culture. This method is sensitive and specific for the discrimination of metastatic tumor cells.     

Immunological detection of membrane-associated human luteinizing hormone correlates with gene expression in cultured human cancer and fetal cells

We have demonstrated the expression of membrane-associated hCG and its subunits and fragments by cells from 78 human cancer cell lines of different types and origins, indicating that such expression is a common phenotypic characteristic of cultured human malignant cells. Because human (h) LH beta has 80% homology with hCG beta and is coded by one of the seven genes in the gene cluster located in chromosome 19, it was important to determine whether hLH and its beta-subunit are also expressed as membrane-associated proteins by cells from human cancer cell lines. Thus, 11 cancer cell lines of different types and origins were adapted to grow in serumless medium, with Nutridoma-HU or SP as serum substitute, and analyzed by flow cytometry using two monoclonal antibodies directed to different conformational epitopes of intact hLH and a monoclonal antibody reacting with an epitope of hLH beta-free. The cells were also analyzed simultaneously for the expression of hCG and its subunits and fragments. Determination of translatable levels of hLH beta and hCG beta messenger RNAs (mRNAs) was performed in cells from some of the cancer cell lines, including the JEG-3 choriocarcinoma cell line, and in cells from a human fetal lung cell line. The analytical flow cytometry studies showed that in addition to the expression of membrane-associated hCG in all of its forms, expression of membrane-associated intact (holo) hLH and its free beta-subunit occurred in every case. These findings were corroborated by the presence of translatable levels of hLH beta and hCG beta mRNAs in all of the cancer cell lines analyzed, indicating that the expression of these membrane-associated glycoproteins is a phenotypic characteristic of human cancer cells and that the activation of the hCG beta-hLH beta gene cluster is nonselective. The presence of translatable levels of hCG beta-hLH beta mRNAs in the cultured human fetal lung cells punctuates once more the in vivo and in vitro biochemical similarities between fetal and cancer cells.     

Ruptured popliteal artery aneurysm

BACKGROUND: Previous studies in which monoclonal antibodies (MoAbs) were used against different epitopes of human chorionic gonadotropin (hCG) demonstrated the presence of membrane-associated hCG and its subunits by cancer cells of different types and origins and by human embryonic and fetal cells. To elucidate the mechanism of action of a synthetic vaccine against hCG, experiments were conducted to determine the presence or absence of direct dose dependent cytolytic activity by hCG MoAbs, including those elicited by the vaccine. METHODS: Human adenocarcinoma cells from the uterine cervix (ATCC HeLa CCL 2.0) grown in defined media at 37 degrees C were treated for 2-3 days with different selected doses of each of 12 MoAbs directed against different epitopes of hCG. Three of these MoAbs were against three different epitopes of the synthetic hCGbeta vaccine. RESULTS: There was a direct dose dependent effect by a MoAb directed against the natural hCGbeta carboxy terminal peptide (CTP), by a MoAb directed against hCGalpha, and by one of the three MoAbs produced by the synthetic hCGbeta-CTP, which is the main component of the World Health Organization (WHO) vaccine being tested for fertility control and for cancer treatment or prevention. CONCLUSIONS: For the first time (to the authors' knowledge), these results show the existence of hCG MoAbs that have direct dose related cytotoxicity at 37 degrees C and explain the mechanism of action of the WHO anti-hCG vaccine.     

Development of seven new human prostate tumor xenograft models and their histopathological characterization

Seven human prostate tumor models were established by transplanting tumor fragments in NMRI athymic nude mice. Once established, the tumors were serially transplantable in both NMRI and BALB/c nude mice. The xenografts originated from primary prostatic carcinomas (prostatectomy specimens), transurethral resection material, and metastatic lesions (pelvic lymph nodes and scrotal skin). Histological examination revealed that, in the course of several mouse passages (8 to 23), tumors retained their resemblance to the original patient material. The PC-295, PC-310, PC-329, and PC-346 tumors are dependent on androgens for their growth. The PC-324, PC-339, and PC-374 tumors are androgen independent, although growth of PC-374 tumors still seemed androgen sensitive. All tumors are diploid, except for the PC-374, which is tetraploid. The diploid PC-295 tumor has an additional small population of tetraploid cells. All xenografts displayed a heterogeneous expression pattern of the androgen receptor except for the PC-324 and PC-339 tumors in which the androgen receptor could not be detected. Prostatic acid phosphatase and prostate-specific antigen were retained during serial transplantation in all tumors but the PC-324 and PC-339. This panel of permanent human prostate tumor models comprises tumors representing both the androgen-dependent and -independent stages of human prostate cancer with various degrees of differentiation and, therefore, is of great value for the study of many aspects of growth and progression of human prostate cancer.     

Melanoma angiogenesis and metastasis modulated by ribozyme targeting of the secreted growth factor pleiotrophin

Clinical and experimental evidence suggests that spreading of malignant cells from a localized tumor (metastasis) is directly related to the number of microvessels in the primary tumor. This tumor angiogenesis is thought to be mediated by tumor-cell-derived growth factors. However, most tumor cells express a multitude of candidate angiogenesis factors and it is difficult to decipher which of these are rate-limiting factors in vivo. Herein we use ribozyme targeting of pleiotrophin (PTN) in metastatic human melanoma cells to assess the significance of this secreted growth factor for angiogenesis and metastasis. As a model we used human melanoma cells (1205LU) that express high levels of PTN and metastasize from subcutaneous tumors to the lungs of experimental animals. In these melanoma cells, we reduced PTN mRNA and growth factor activity by transfection with PTN-targeted ribozymes and generated cell lines expressing different levels of PTN. We found that the reduction of PTN does not affect growth of the melanoma cells in vitro. In nude mice, however, tumor growth and angiogenesis were decreased in parallel with the reduced PTN levels and apoptosis in the tumors was increased. Concomitantly, the metastatic spread of the tumors from the subcutaneous site to the lungs was prevented. These studies support a direct link between tumor angiogenesis and metastasis through a secreted growth factor and identify PTN as a candidate factor that may be rate-limiting for human melanoma metastasis.     

Establishment of a highly differentiated thyroid cancer cell line of Hürthle cell origin

Hürthle cell carcinomas (HCC) of the thyroid are a variant of follicular thyroid tumors. In contrast to follicular thyroid carcinoma, HCC rarely take up radioiodine and frequently metastasize to the lymph nodes. Histologically they are indistinguishable from Hürthle cell adenomas except for evidence of invasion and metastasis. How these carcinomas develop and why they behave differently than other follicular tumors is not known. Although some differentiated thyroid cancer cell lines exist, none are from Hürthle cell tumors. We have established a well-differentiated thyroid cancer cell line from a metastasis of a HCC, designated XTC.UC1. In vitro, XTC cells display epitheloid morphology, grow with a population doubling time of 4.3 +/- 0.3 days, migrate, and invade through reconstituted basement membranes. The cells are immunoreactive for and release thyroglobulin, respond to thyrotropin (TSH) with increase of intracellular cyclic adenosine monophosphate (cAMP), proliferation, and invasion of reconstituted basement membrane, thus exhibiting characteristics of well-differentiated thyroid carcinoma. In vivo, xenografted XTC cells grow with a doubling time of 9.8 +/- 0.8 days. Tumors spontaneously metastasize to the lymph nodes and less frequently to the lungs and the liver. The cells retained their differentiated function in vivo as assessed by human thyroglobulin (hTG) secretion and immunohistochemistry. This is a first report of the establishment of a unique, highly differentiated thyroid carcinoma cell line derived from an HCC. Based on the ability to invade through reconstituted basement membrane in vitro and the potential to metastasize in vivo, this cell line may provide a unique model to study invasion and metastazation of well-differentiated thyroid cancer.     

Testicular seminoma with human chorionic gonadotropin production

Testicular seminoma with elevated serum human chorionic gonadotropin level (hCG-positive seminoma) is regarded as more malignant than marker-negative seminoma, although its prognosis is still unclear. To clarify the malignant potential of seminoma with hCG production, the serum levels of the beta subunit of hCG (beta-hCG) and lactic acid dehydrogenase (LDH) were examined in 35 and 40 patients, respectively, and the immunohistochemical expression of beta-hCG examined in 45 tumors. The elevation of the LDH serum level correlated to the invasive status, metastatic status and poor outcome, while that of the serum beta-hCG level correlated only to the metastatic status. Immunohistochemical expression of beta-hCG was observed in syncytiotrophoblastic giant cells in 11 tumors and a few mononuclear seminoma cells in 36 tumors. Expression was not associated with the malignancy potential, except where the expression in mononuclear cells inversely correlated to the invasive status. These results suggest that most seminomas produce a slight amount of hCG; that an elevated hCG serum level indicates the presence of metastatic tumors and mainly reflects an increase in tumor volume but not in cellular malignancy potential; and that the LDH serum level, rather than hCG, is more useful as a prognostic indicator for patients with seminoma.     

Tumor suppressor activity of the TGF-beta pathway in human cancers

Immunodeficient animals, principally nude mice, when used in appropriately designed studies have been shown to be useful for the experimental analysis of human breast cancer metastasis. As with many other human tumors, the implantation of breast cancer cells into an anatomically appropriate tissue (the mammary fatpad) results in increased tumor take and incidence of metastasis for certain cell lines compared with subcutaneous injection. Testing a number of widely available human breast cancer cell lines identified the MDA-MB-435 cell line as the most metastatic, producing lung and lymph node metastases in a high proportion of nude and severe combined immunodeficient (SCID) mice after injection in the mammary fatpad. Mixing human breast cancer cells with normal fibroblasts or with Matrigel also increases the tumor incidence and growth rates in nude mice. Different routes of injection can be used to assess the ability of human breast cancer cells to form metastatic lesions in the lungs (i.v. injection), the liver (injection in the spleen), the brain (direct or intracarotid artery injection) and the bone marrow and bone (injection into the left ventricle of the heart). These different approaches demonstrate the potential of experimental studies of human breast cancer growth and metastasis using immunodeficient mice; this model is valuable for experiments that test the role of metastasis-associated genes and the efficacy of novel forms of therapy.     

Ectopic expression of carbohydrate antigens that determine the metastatic potential of human colorectal carcinoma

Highly malignant and metastatic tumor cells are thought to arise within primary tumors and become predominant during cancer progression. We demonstrated, by the analysis of > 500 surgical specimens, that colorectal carcinomas with increased metastatic potential were characterized by an increased expression of sialyl-Le(x) antigens expressed on mucins. The biological role of sialyl-Le(x) antigens expressed on mucins produced by colon carcinoma cells has been investigated using variant cell lines selected for their expression of this antigen. KM12-HX and KM12-LX, high and low expresser variant cells, differed in their metastatic potential in nude mice after intrasplenic injection. KM12-HX cells contain higher levels of polyA+mRNA for alpha(1-3/4) fucosyltransferase than KM12-LX cells. Sialyl-Le(x) antigenic carbohydrate chains were attached to mucins as well as glycoproteins with various M(r). KM12-HX cells adhered more strongly than KM12-LX cells to human umbilical vein endothelial cells treated with tumor necrosis factor-alpha and to mouse hepatic sinusoidal endothelial cells. We have retrospectively evaluated post-operative survival of colon carcinoma patients for their sialyl-Le(x) antigen levels in the primary tumors according to the percentage of stained cells by specific antibodies. The adjusted survival rate of the patients with high levels of sialyl-Le(x) antigen in their primary tumors was much lower due to recurrence and metastasis than that of the patients with tumors containing low levels of sialyl-Le(x) antigen. The results suggested that sialyl-Le(x) antigen has a potential to be used as a predictive marker for colorectal cancer metastasis.     

Overexpression of bcl-2 protects prostate cancer cells from apoptosis in vitro and confers resistance to androgen depletion in vivo

Normal (nonneoplastic) human prostatic secretory epithelial cells do not express the bcl-2 protein. However, a recent immunohistochemical survey of neoplastic human prostate tissues showed that a fraction of primary untreated prostate adenocarcinoma cells expressed this apoptosis-suppressing oncoprotein at significant levels (Colombel et al., Am. J. Pathol., 143: 390-400, 1993). Additionally, a number of hormone-refractory prostatic adenocarcinomas obtained from hormonally-treated patients (subsequent to surgical or drug castration therapy) were examined and were found to be uniform in their elevated expression of bcl-2 oncoprotein. The results of this preliminary survey imply that bcl-2 expression distinguishes a subgroup of primary human prostate cancers and that the expression of this protein might be a factor enabling prostate cancer cells to survive in an androgen-deprived environment. The current study was undertaken to determine the degree to which overexpression of bcl-2 can protect human prostate cancer cells from apoptotic stimuli in vitro and in vivo. Human prostate cancer cells (LNCaP) were transfected with a neomycin-selectable eucaryotic expression vector containing cDNA encoding human bcl-2. Transfected clonal variants that express bcl-2 protein (LNCaP/bcl-2) were unaltered with regard to their basal growth rate in 10% serum-containing medium, or with regard to their expression of the differentiated human prostate cell gene products prostate-specific antigen or androgen receptor protein. The bcl-2-transfected clones were altered, however, with regard to their growth rate in charcoal-stripped serum lacking dihydrotestosterone. Additionally, in contrast to the parental or control-transfected cell lines, LNCaP/bcl-2 cells were highly resistant to a variety of apoptotic stimuli in vitro including serum starvation and 10 nM phorbol ester (phorbol 12-myristate 13-acetate) supplementation of the medium. Lastly, the overexpression of bcl-2 by these prostate cancer cells altered their tumorigenic potential in a nude mouse assay. s.c. injections of 10(6) LNCaP/bcl-2 cells into male nude mice resulted in earlier and larger tumor formation compared to an equivalent injection of parental or control-transfected LNCaP cells. When these variant cell lines were injected into castrated male nude mice, only the LNCaP/bcl-2-transformed cells gave rise to tumors. Moreover, LNCaP/bcl-2 tumors grown in intact male nude mice were refractory to the growth-inhibiting effects of castration demonstrated by parental LNCaP cells. Data obtained in this study demonstrate that the bcl-2 oncoprotein can protect prostate cancer cells from apoptotic stimuli in vitro and suggest that such protection correlates with the ability to form hormone-refractory prostate tumors in vivo.     

Rho proteins induce metastatic properties in vivo

Rho proteins have been implicated in the regulation of multiple signal transduction processes. Some of the members of this family, including the rho gene from Aplysia californica and the human genes (rhoA, rhoB and rac-1), are proto-oncogenes since when properly mutated they can induce cell transformation, and the generated rho-transformed cells are tumorigenic when inoculated into mice. In addition to their tumorigenic activity, there is evidence suggesting that Rho proteins may contribute to the metastatic phenotype. However, all the experiments implicating Rho proteins or Rho-regulating proteins in the induction of metastatic potential are either indirect or have been performed in vitro. In this study we investigated whether cells transformed by rho oncogenes do have metastatic potential in vivo. We present evidence that cells transformed by the Aplysia californica rho gene, when injected directly into the blood stream are able to efficiently colonize lungs and secondary organs, consistent with the acquisition of the metastatic potential. Moreover, tumors derived from subcutaneous injections of these rho-transformed cells are also able to metastasize in distant organs, a strong support to the hypothesis that Rho proteins play a role in the metastatic phenotype. Finally, cells transformed by the human oncogenes dbl, vav and ost, three well-known guanine exchange factors for members of the Rho family, or cells transformed by the activated human rac-1 or rhoA genes do also have metastatic potential when injected into the blood stream. These results demonstrate that signaling pathways regulated by Rho proteins play an important role in the acquisition of the metastatic phenotype in vivo.     

Modulation of tumor cell response to chemotherapy by the organ environment

The outcome of cancer metastasis depends on the interaction of metastatic cells with various host factors. The implantation of human cancer cells into anatomically correct (orthotopic) sites in nude mice can be used to ascertain their metastatic potential. While it is clear that vascularity and local immunity can retard or facilitate tumor growth, we have found that the organ environment also influences tumor cell functions such as production of degradative enzymes. The organ microenvironment can also influence the response of metastases to chemotherapy. It is not uncommon to observe the regression of cancer metastases in one organ and their continued growth in other sites after systemic chemotherapy. We demonstrated this effect in a series of experiments using a murine fibrosarcoma, a murine colon carcinoma, and a human colon carcinoma. The tumor cells were implanted subcutaneously or into different visceral organs. Subcutaneous tumors were sensitive to doxorubicin (DXR), whereas lung or liver metastases were not. In contrast, sensitivity to 5-FU did not differ between these sites of growth. The differences in response to DXR between s.c. tumors (sensitive) and lung or liver tumors (resistant) were not due to variations in DXR potency or DXR distribution. The expression of the multidrug resistance-associated P-glycoprotein as determined by flow cytometric analysis of tumor cells harvested from lesions in different organs correlated inversely with their sensitivity to DXR: increased P-glycoprotein was associated with overexpression of mdr1 mRNA. However, the organ-specific mechanism for upregulating mdr1 and P-glycoprotein has yet to be elucidated.     

Assessment of self-injurious behavior maintained by access to self-restraint materials

Metastatic tumor is one of several etiologies of space-occupying masses in the orbit that accounts for 1%-13% of all orbital masses (1). In the adult patient population, breast cancer is the most common tumor to metastasize to the orbit followed by metastases from the lung, prostate and gastrointestinal tract (2). It is rare for carcinoid tumors to metastasize to the eye or to the orbit. Carcinoid tumors arise from Kulchitsky cells that originate in the neural crest. Histologically, these tumors resemble, but are not as aggressive as, adenocarcinomas. Most carcinoids arise in the gastrointestinal tract or the lung. The most common site for carcinoid metastases is the liver. On anatomical imaging studies, such as CT and magnetic resonance imaging, metastatic orbital carcinoid tumors appear as nonspecific tumor masses. Carcinoid tumors have an affinity for uptake of the radiopharmaceutical 131I-metaiodobenzylguanidine (MIBG) (3). We report a case of a patient with a known carcinoid tumor who developed a left orbital mass that demonstrated abnormal uptake of 131I-MIBG indicative of metastatic carcinoid tumor to the orbit.     

Frequent development of murine T-cell lymphomas with TcR alpha/beta+, CD4-/8- phenotype after implantation of human inflammatory breast cancer cells in BALB/c nude mice

Tumors developed quite frequently in some of the visceral organs, including spleen and liver, in BALB/c nude mice upon subcutaneously xenografting surgical specimens from five different inflammatory breast cancer patients. All of these tumors developed within two and a half months to one year after the subcutaneous inoculation of surgical specimens. From these tumors, five independent transplantable tumors, including tMK-2, tHK-1, tYK-1, tYK-2 and tTY-1 have been established. Chromosome analysis, morphologic studies by light and electron microscopy and phenotype analysis indicated that these tumors are of mouse origin. The tMK-2 tumor was highly metastatic to the spleen and liver when it was subcutaneously transplanted into the right scapular region. In addition, the region where the tMK-2 tumor cells were subcutaneously inoculated showed an apparently inflammatory process represented by erythema. After subcutaneous inoculation into the right scapular region, tHK-1, tYK-1, 2, and tTY-1 tumors also metastasized to some of the visceral organs, including spleen and liver. From these tumors, in vitro cell lines were established. The cells grew in a stromal-cell dependent manner under in vitro culture conditions. The cells were again tumorigenic at the inoculated region and metastasized to various organs, including liver and spleen, of BABL/c nude mice. Histological examination revealed that the tumors showed features of malignant lymphoma. Phenotypically, these five tumors expressed early T lymphocyte markers as revealed by anti-mouse anti-TcR alpha/beta, anti-CD3, CD4 and CD8 monoclonal antibodies. To our knowledge, these cell lines are the first T-cell lines showing the phenotype of extrathymically differentiated T-cells in the liver.     

Reference ranges for haematology parameters in pregnancy derived from patient populations

Using immunodeficient mice human tumor cell lines, which metastasize to specific organs from orthotopic implantation sites but not from ectopic sites, have been established. We have been studying mechanisms of the regulation of tumor metastasis by organ specific microenvironments, and have recently found significant differences in interactions between metastatic tumor cells and fibroblasts from different tissues. Fibroblasts isolated from orthotopic organ tissues of nude mice enhanced invasion of metastatic human colon carcinoma cells. In contrast, fibroblasts derived from a subcutaneous tissue suppressed carcinoma cell invasion. These divergent effects were found in part due to differential expression of cytokines and growth factors by fibroblasts from different tissues.