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Biomedical subjects

D Shouval

Publications and source records attributed to D Shouval.

At least 145 records · Page 8Linked to original sources
PubMed1986

Continuous production of erythropoietin by an established human renal carcinoma cell line: development of the cell line.

Establishment of a stable, transformed human renal carcinoma cell line that produces erythropoietin in vitro and has maintained this function continuously since 1981 and for greater than 150 passages in monolayer culture was accomplished by transplantation of human renal clear cell carcinoma tissue from a patient with erythrocytosis into an immunosuppressed athymic mouse. In addition to its immunocrossreactivity with native human urinary erythropoietin, the tumor erythropoietin demonstrates biological activity in the in vitro mouse erythroid colony-forming unit assay and in tumor-bearing nude mice. The cloned renal carcinoma cell line has an abnormal human karyotype and has ultrastructural features characteristic of human renal clear cell carcinoma. This cell line provides a reproducible model system for the production of an erythropoietin-like material and for the study of its synthesis and secretion.

Animals↗
PubMed1985

Conservation from rat to human of cytosolic phosphoenolpyruvate carboxykinase and the control of its gene expression.

Structural conservation of cytosolic phosphoenolpyruvate carboxykinase protein and mRNA sequence was found in all species examined from rodents to human. The mitochondrial isoenzyme, in all species tested, represents a distinct protein. Moreover, irrespective of the ratio of cytosolic to mitochondrial isoenzyme, cytosolic phosphoenolpyruvate carboxykinase activity in the human as in the rat is controlled at the level of gene expression and through the same multiple hormonal stimulation. This evolutionary conservation of the cytosolic phosphoenolpyruvate carboxykinase structure and mode of regulation supports the enzymes' physiological importance in mammals.

Animals↗
PubMed1985

Development of a novel C1q immunoadsorbent for removal of circulating immunecomplexes: quantitative isolation of hepatitis B virus surface antigen and immunecomplexes.

A technique for large scale production of human C1q from plasma by affinity chromatography on an anti-C1q column is described. Affinity purified C1q was covalently coupled to a newly developed agarose polyacrolein microsphere beads immunoadsorbent. This immunoadsorbent was utilized for quantitative removal of artificially formed bovine serum albumin (BSA)-anti-BSA immune complexes (IC). The C1q affinity column was then used for the isolation of immunecomplexes containing hepatitis B virus (HBV) surface antigen (HBsAg) from serum of an HBsAg carrier. Identical columns may be utilized for quantitative removal of a variety of IC from blood of patients with infectious and autoimmune diseases, as well as neoplastic diseases. Furthermore, dissociated immunecomplexes will provide an additional source for purification of specific antigens.

Animals↗
PubMed1985

Papular acrodermatitis of childhood associated with hepatitis A virus infection.

A 2-year-old girl developed an erythematous papular eruption on her face and extremities a week after an epidemic of hepatitis A had occurred in her school. Clinical and laboratory signs of acute hepatitis, together with serologic verification, confirmed hepatitis A infection. That diagnosis should be considered in the etiology of papular acrodermatitis of childhood.

Acrodermatitis↗
PubMed1985

Antigenic characterization of human hepatocellular carcinoma. Development of in vitro and in vivo immunoassays that use monoclonal antibodies.

Several libraries of monoclonal antibodies have been produced by immunization of Balb/c mice with single cell suspensions of nontrypsin-treated human hepatocellular carcinoma cell (HCC) lines in order to study the antigenic properties of transformed hepatocytes. The antibodies were characterized with regards to specificity for hepatoma-associated antigens and their capability for use as reagents in radioimmunoassays (RIAs) and tumor localization in vivo. Three such antibodies namely, P215457, PM4E9917, P232524 of the IgG2a, IgG2a, and IgG1 isotypes, respectively, not only recognized separate and distinct antigenic determinants on four human hepatoma cell lines but also reacted with epitopes present on chemically induced rat hepatoma cell lines. In contrast, only 1 of 38 other human malignant and transformed cell lines demonstrated reactivity with the three antibodies; normal human tissues were also found to be unreactive. Monoclonal antibody P215457 densely stained the plasma membrane by indirect immunofluorescence, showed rapid binding activity to HCC cells in suspension, and precipitated a 50,000-mol wt cell surface protein; antibody PM4E9917 also stained the plasma membrane and precipitated a 65,000-mol wt protein, whereas P232534 recognized cytoplasmic antigenic determinants. With these antibodies "simultaneous sandwich" RIAs were established that detect soluble hepatoma-associated antigens in culture supernatants. Finally, the Fab fragment of P215457 was found to be useful in tumor localization in vivo. This antibody fragment when labeled with 131I was shown to localize by radionuclide-imaging studies in human hepatoma grown in nude mice. Thus, these investigations demonstrate that monoclonal antibodies may be produced against epitopes that reside almost exclusively on transformed hepatocytes and such antibodies may be successfully employed in the development of in vitro and in vivo immunoassays.

Animals↗
PubMed1984

Detection and characterization of hepatitis B virus DNA in serum of HBe antigen-negative HBsAg carriers.

Sera from 153 Israeli patients in various stages of hepatitis B virus (HBV) infection with undetectable hepatitis Be antigen (HBeAg) were studied for the presence of HBV DNA in the serum by molecular hybridization. HBV DNA was detected in 10 patients: 3 with acute hepatitis, 4 asymptomatic hepatitis B surface antigen (HBsAg) carriers, 1 with chronic active hepatitis, 1 with cirrhosis, and 1 with mixed cryoglobulinemia. HBV DNA was detected in 7 of 10 HBeAg-positive control samples tested. Hybridization analysis was used for quantitative comparison of HBV DNA levels in serum. HBV DNA levels, found in HBeAg-negative patients sometimes exceeded the levels found in HBeAg-positive patients. Restriction enzyme analysis of serum HBV DNA from four HBeAg-negative samples gave undistinguishable digestion patterns as compared to 3 HBeAg-positive samples. However, heterogeneity in HBV DNA restriction fragments was detected among HBV genomes in sera of HBeAg-positive samples. These data demonstrate that HBV DNA may be present in the serum at various stages of HBV infection, regardless of HBeAg detection. Failure to detect HBeAg in these patients does not necessarily reflect low serum levels of viral particles, or the occurrence of HBV genome variants.

Carrier State↗
PubMed1983

Role in nude mice of interferon and natural killer cells in inhibiting the tumorigenicity of human hepatocellular carcinoma cells infected with hepatitis B virus.

The human hepatoma cell line, PLC/PRF/5, which is persistently infected with hepatitis B virus (HBV), has integrated HBV-DNA, secretes HBV surface antigen (HBsAg), and does not grow readily in congenitally athymic (nu/nu) mice. The present investigation was undertaken to ascertain whether the low tumorigenicity of this cell line was governed by a host immune response and/or was related to expression of HBsAg. Subcutaneous injection of 4-5 X 10(6) cells into BALB/c nude mice produced localized encapsulated tumors with morphologic features of primary hepatocellular carcinoma in 25% of the animals within 29-40 d. No tumor growth was observed at lower cell inocula. In contrast, SK-HEP-1, an HBV-negative human hepatoma cell line, produced tumors at 1-5 X 10(6) cells inocula in 66% of the animals. Immunosuppression of mice with antilymphocyte serum (ALS) or irradiation increased tumor incidence in mice inoculated with 1 X 10(6) PLC/PRF/5 cells to almost 100% and produced local invasiveness. Immunosuppression also reduced the latency, i.e., time to tumor appearance, and increased mean tumor weight. These results suggest that tumorigenicity was limited by the host immune response. The nature of the response was delineated by treating nude mice challenged with tumor cells with sheep anti-mouse interferon globulin (anti-IFN). When 2 X 10(6) cells were injected, tumor growth occurred in 75% of anti-IFN-treated mice, whereas controls injected with the same number of cells, but not receiving anti-IFN, failed to develop tumors. The tumors in the anti-IFN-treated mice were highly invasive and the latency period until tumor appearance was reduced to 3-5 d. An inverse correlation was found between susceptibility of the hepatoma cells to natural killer (NK) activity in vitro and resistance to tumor growth in vivo. In vitro cytotoxicity for PLC/PRF/5 cells was eliminated by anti-NK 1.1 and complement, establishing the effector cell as an NK cell. NK cell activity 14 d after inoculation of mice with PLC/PRF/5 cells was augmented against PLC/PRF/5 target cells but not against SK-HEP-1 cells. Treatment of mice with ALS, irradiation, or anti-IFN abolished NK activity against PLC/PRF/5 cells. Co-cultivation of nude mouse spleen cells with PLC/PRF/5 but not with HBsAg or SK-HEP-1 cells induced secretion of murine IFNalpha. These results suggest that the IFN/NK cell system may play a role in limiting tumorigenicity and invasiveness of HBV-infected human hepatocellular carcinoma cells by a mechanism similar to that found for other cells persistently infected with viruses.

Animals↗
PubMed1982

Immunotherapy in nude mice of human hepatoma using monoclonal antibodies against hepatitis B virus.

The human hepatoma cell line PLC/PRF/5, which contains integrated hepatitis B virus DNA, synthesises and secretes hepatitis B virus surface antigen (HBsAg). When injected into BALB/c nude mice, these cells produce well-vascularized, encapsulated tumours in almost 100% of animals. HBsAg has been demonstrated on the surface and in the cytoplasm of PLC/PRF/5 cells using immunofluorescence techniques. Recently, we reported that monoclonal antibodies to HBsAg (IgG1, IgG2a and IgM isotypes) bind to HBsAg-associated viral determinants of PLC/PRF/5 cells in culture and immunoprecipitate HBsAg secreted into the growth medium. Also IgG2a and IgM antibodies to HBsAg (anti-HBs) produce specific lysis of PLC/PRF/5 cells in culture in the presence of complement, but have no lytic effect on human hepatoma cells which do not express HBsAg. In the present study, we demonstrate that IgM and IgG2a, but not IgG1, monoclonal anti-HBs specifically prevent or suppress tumour formation in a substantial number of athymic nude mice injected with PLC/PRF/5 cells.

Animals↗
PubMed1982

Selecting binding and complement-mediated lysis of human hepatoma cells (PLC/PRF/5) in culture by monoclonal antibodies to hepatitis B surface antigen.

High-affinity 125I-labelled monoclonal antibodies to hepatitis B virus surface antigen (HBsAg) bind to human hepatocellular carcinoma cell line, PLC/PRF/5, which synthesizes and secretes HBsAg. These monoclonal antibodies of the IgG1, IgG2a, and IgM isotypes are directed against different antigenic determinants on HBsAg and, in the presence of complement, both anti-HBs IgG2a and IgM, but not anti-HBs IgG1, lyse PLC/PRF/5 cells in culture. Although there is a low level of anti-HBs binding (especially with anti-HBs IgM) to human hepatoma cell lines which do not synthesize HBsAg (SK-Hep 1 and Mahlavu cells), this interaction does not lead to complement-mediated cell lysis and is thought to be nonspecific. Minimal binding of 125I-labeled anti-influenza HA antigen IgM binding to PLC/PRF/5 cells was also detected, but this likewise did not lead to complement-mediated cell lysis. These results indicate that a human hepatocellular carcinoma cell line, persistently infected with hepatitis B virus, can be recognized and lysed by monoclonal antibodies directed against specific determinants of HBsAg. Monoclonal antibodies to this viral envelope protein may prove to be useful immunodiagnostic and immunotherapeutic agents when such viral epitopes are expressed on the surface of infected cells.

Animals↗
PubMed1982

Histocompatibility antigens and cell-mediated immunity in carriers of hepatitis B virus; a study of a family.

The cause of the HBsAg carrier state is unknown, and environmental genetic and immunologic factors have been implicated. Herein, a family is described in which the carrier state was associated with the HLA-A28BW15 haplotype and with impaired cell mediated immunity. The relevance of the genetic marker in this family is uncertain. It could be associated with a genetic susceptibility to the virus or alternatively with inherited immunodeficiency. According to the first possibility the impaired immunity could be secondary to the persistent viral infection. According to the second one the acquisition of the carrier state could result from the inherited immune defect.

Carrier State↗
PubMed1981

Integration of hepatitis B virus DNA into the genome of liver cells in chronic liver disease and hepatocellular carcinoma. Studies in percutaneous liver biopsies and post-mortem tissue specimens.

We used recombinant-DNA technology and gel electrophoresis to find hepatitis B virus DNA (HBV-DNA) in liver and tumor tissue from patients with hepatocellular carcinoma and chronic liver disease, and to study the integration of HBV-DNA into the genome of these tissues' cells. In 12 patients with hepatocellular carcinoma who had hepatitis B surface antigen (HBsAg) in their serum, integrated HBV-DNA was identified in the tumors; it was also found in tumors from three of eight patients who were seronegative for HBsAg but positive for antibody to HBsAg. In some cases, integrated HBV-DNA was also present in nontumorous liver tissue that had the same hybridization pattern or one different from that of the tumor. In five carriers of HBsAg who had evidence of the carrier state and chronic liver disease for less than two years, HBV-DNA was present but not integrated in liver tissue. In the two patients who had carried HBsAg for more than eight years, HBV-DNA was integrated into the host genome. These data suggest that integration of HBV-DNA into hepatocytes occurs during the course of persistent HBV infection and precedes development of gross neoplasm.

Biopsy↗
PubMed1981

Tumorigenicity in nude mice of a human hepatoma cell line containing hepatitis B virus DNA.

The human hepatocellular carcinoma cell line PLC/PRF/5, which synthesizes and secretes hepatitis B surface antigen, was grown under optimal conditions in tissue culture, using Eagle's minimal essential medium supplemented with 10% fetal bovine serum and 10(-11) M triiodothyronine on collagen rafts. Injection s.c. of the PLC/PRF/5 cell line into athymic BALB/c nude mice resulted in the growth of a well-circumscribed, moderately differentiated hepatocellular carcinoma. The intervals until tumor appearance and tumor "take" rates were dependent on inoculum dose. Four to 5 x 10(6) cells induced tumor growth in 29% of 14 injected mice within 29 to 40 days, while 7 to 13 X 10(6) cells induced tumors in all 15 mice within 10 to 12 days after inoculation. Hepatitis B surface antigen was detected in the nude mouse serum and tumor tissue, and its concentration roughly correlated with tumor weight. A low level of antibody against hepatitis B surface antigen was detected in five tumor-bearing animals, as well as in one mouse which did not produce a tumor. Hepatitis B core antigen and its antibody and hepatitis B e antigen and its antibody were not detected in 26 mice, using immunohistochemical and radioimmunoassay methods. alpha-Fetoprotein, carcinoembryonic antigen, and alpha-antitrypsin were detected in nude mice tumors, using the peroxidase-antiperoxidase technique. Finally, hepatitis B virus DNA, identified in the nude mouse tumor by molecular hybridization techniques, was compared to PLC/PRF/5 cell line hepatitis B virus DNA.

Animals↗