Acquired heterophile antigens on the surface of human cell lines.

Chicken IgM antibodies raised against sheep erythrocytes (SE) reacted with a heterophile carbohydrate antigen designated as "HC". This antigen was visualized by SE rosette formation with antibody-sensitized and formaldehyde-fixed target cells. Cells from eight lymphoblastoid lines and ten lines of diverse histogenetic origin all expressed HC on their surface to a greater or lesser degree when grown in fetal calf serum (FCS). Lymphoblastoid cell lines, HSB2 and Namalva, lost most of their HC antigen after three cell divisions when grown in medium which contained normal human serum (NHS) instead of FCS. The acquired origin of HC was demonstrated by the conversion of HSB2-HC- into HSB2-HC+ cells after incubation in the presence of FCS or fetuin for 18 h at 37 degrees C. The acquisition of HC antigen depended on the concentration of fetuin as well as the time and temperature of incubation. The presence of NHS in the culture medium reduced the degree of HC incorporation. The level of HC expressed on the cell surface was reduced by treating HSB2 cells with sodium periodate but not by proteolytic enzymes. Competitive inhibition with hog blood group substance and sheep IgG suggested similar specificity of HC on target cells which had been sensitized with either fetuin or sheep IgM (Eur. J. Immunol. 1977.7:204). The avidity of chicken antibodies to HC in either of these systems was several orders of magnitude lower than the binding to SE.     

Functional characteristics and differential expression of class II DR,DS, and SB antigens on human melanoma cell lines

Although most cultured melanoma cell lines express DR Class II molecules, many of these do not also express the DS (MB) Class II molecules as detected by a monoclonal antibody specific for DS. Cells lacking either DR or DS molecules or both could only be induced to express DR antigens in rare cases by combined incubations with azacytidine and Interleukin-2 conditioned medium, although the expression of DR molecules on fibroblasts or U937 monocytes could more easily be induced under the same culture conditions. Melanoma cells expressing DR antigens could function in antigen presentation for the histocompatibility antigens themselves and for DR specific presentation of TNP determinants to allogeneic T-cells sensitized to TNP modified lymphocytes and showing restriction in their responses to the specificity of the DR molecules expressed on the original, autologous senzitizing cells. DR positive melanoma cells could not, however, be demonstrated to function in the presentation of any of the soluble antigens tested. All DR positive melanoma cells also expressed SB antigens, but these were not detected on DR negative melanoma cells. These studies collectively indicate that the expression of Class II histocompatibility antigens on diverse cell types is subject to differential regulatory control and is associated with differences in their functional activities.     

Heterophile Hanganutziu-Deicher antigen in ganglioside fractions of human melanoma tissues

Gangliosides were purified from melanoma tissue extracts obtained from 5 patients. GM3 and GD3 were identified as the major components in ganglioside fractions of the melanoma tissues. Following thin-layer chromatography, enzyme immunostaining with Hanganutziu-Deicher (H-D) antigen-specific chicken antisera demonstrated the presence of NeuGc-neolactotetraosylceramide (H-D5) and NeuGc-lacto-N-norhexa-osylceramide (H-D7) in all 5 melanoma extracts.     

Expression and activation of pro-gelatinase A by human melanoma cell lines with different tumorigenic potential

The production of various proteolytic enzymes by tumor cells facilitate the invasion of solid tumors into surrounding tissues. We examined three cell lines (M1Dor, M4Be and M3Da) derived from malignant melanoma which exhibited different abilities to grow in nude mice following subcutaneous grafting. By in vitro invasion assay using Boyden-chambers technique, we found that none of those cell lines were able to invade the Matrigel. Several studies have substantiated the role of matrix metalloproteinases (MMP), mainly gelatinases MMP-9 and MMP-2, in melanoma cell invasion. Each cell line constitutively produced MMP-2 (but not MMP-9) in its latent form only, with stronger production for the most tumorigenic cell line in vivo (M3Da). Integrity of the MMP-2 activation process was studied since MMP-2 was also recovered as zymogen at the cell plasma membrane. All cell lines secreted TIMP-1 and TIMP-2 in a constitutive manner and again, but TIMP-2 production as well as MT1-MMP expression were found inversely related to their tumorigenic potential. Plating cells onto type I or type IV collagen did not trigger pro-MMP-2 activation; on the contrary, conversion of pro-MMP-2 to its active form could be evidenced when melanoma cell lines were seeded in a three dimensional type I collagen lattice. This revised version was published online in July 2006 with corrections to the Cover Date.     

8株人黑色素瘤细胞系的建立及其生物学特性研究

目的：从人黑色素瘤组织建立一系列黑色素瘤细胞系，命名为黑色素瘤细胞系1-8号（HME1-8），并研究鉴定其体外细胞生物学与免疫学特性。方法：取黑色素瘤组织进行组织块培养法，待长满90%汇合面后消化传代培养，并进行形态学、生长动力学、免疫学及致瘤性等生物学特性研究。结果：该系列黑色素瘤细胞系已在体外培养生存1-2年，传60-100余代，其生物学特性显示：①这些细胞系体外生长的倍增时间为34.2-59.5h；②软琼脂集落形成率平均值在8.6%-26.2%之间；③接种至裸鼠后具有较高的致瘤性；④染色体核型分析为非整倍体，众数为64-117条；⑤ 透射电镜下可观察到细胞含丰富的核糖体及黑色素颗粒；⑥免疫组化分析显示细胞浆内含大量阳性棕色黑色素颗粒；⑦肿瘤抗原检测显示8株黑色素瘤细胞系的Melan-A抗原阳性率为75%, MAGE-1阳性率为50%, MAGE-2阳性率为37.5%，MAGE-3阳性率为62.5%。结论：8株黑色素瘤细胞系经体外长期培养后已永生化，形成了连续传代细胞系，具有明显的黑色素瘤细胞恶性表型特征。     

Turnover of histamine in human melanoma cell lines

Inflammation Research -     

A cytogenetic study of eight human melanoma cell lines.

Eight cell lines established originally at the Queensland Institute of Medical Research from human malignant melanoma explants have been studied by means of cytogenetic techniques. All showed abnormalities characteristic for each individual line and consisting of marker chromosomes and of changes in ploidy due to the addition of extra copies of normal chromosomes. Repeat cultures of some lines after one or two years contained most of the markers which had characterized the original samples; additional chromosome abnormalities were also found. An anlysis of the break points concerned in the production of markers showed preferential involvement of chromosomes 1 and 5, with a prevalence of centromeric breaks on No. 1. These findings add further weight to the evidence suggesting that changes in chromosome 1 may be of special significance in the pathogenesis of some solid tumours.     

Mapping cell surface antigens on mouse pre-B cell lines

Characterization of cell surface molecules expressed by B cell precursors has been hampered by the inability to routinely assay such cells from freshly explanted normal lymphoid tissues. However, a large number of transformed pre-B cell clones have been isolated which presumably include those precursors recently entering the B cell lineage (obtained from fetal liver and more rarely from adult bone marrow) and extending to those clones which are poised to become surface Ig-positive B cells. In this report, we have quantitated expression of 10 different cell surface molecules, 5 of which bear special interest as B cell differentiation antigens, among a large panel of pre-B cell clones.     

Contamination of human melanoma cell lines by mouse L cells.

Five of 8 cell lines from human melanomas originally established elsewhere were found to consist exclusively of mouse cells when examined in our laboratory some months after their receipt. Cytogenetic studies, including G- and C-banding, showed that the mouse cells in all cultures had originated from a single cell line, identified as the L-line. Contamination probably occurred, one year before its discovery, in a laboratory where L cells and human melanoma cells were briefly kept in the same incubator.     

The expression of MHC antigens by human teratocarcinoma derived cell lines

We have used flow microflourimetry to investigate quantitatively the expression of HLA-A, B and C antigens, and β₂ microglobulin, by cell lines derived from human teratocarcinomas. Although low levels of these cell surface molecules were expressed by all the lines examined, there was no evidence of discrete HLA-A, B, C/β₂-microglobulin positive and negative subpopulations in any cultures. In contrast, using similar techniques, no murine embryonal carcinoma cell line was found to express the homologous H-2 antigens. It is suggested that human embryonal carcinoma cells may differ from their mouse counterparts by expressing low levels of major histocompatibility antigens.     

Surface antigens of human melanoma cells defined by monoclonal antibodies. I. Biochemical characterization of two antigens found on cell lines and fresh tumors of diverse tissue origin

Monoclonal antibodies were produced against melanoma cell lines derived from patients presently disease free. Based on tissue distribution of binding, the antibodies obtained could be used to classify melanoma surface antigens into groupings similar to those obtained from studies of autologous and xenogeneic antibodies. Three antibodies (21.43, 15.75 and 15.95) reacted with the immunizing tumor but not with peripheral blood lymphocytes or a lymphoblastoid cell line derived from the tumor donor. When tested on a broader cell panel, antibody 21.43 reacted only with melanoma lines, while antibodies 15.75 and 15.95 reacted with carcinoma cell lines as well as with the majority of melanoma cell lines. Antibody 15.95 precipitated a 49000 dalton glycoprotein from both melanoma and carcinoma cells. Antibody 15.75 precipitated a 74 000 dalton glycoprotein from the surface of melanoma and carcinoma cell lines which is also found on freshly isolated tumor cells from melanoma and carcinoma metastases in vivo.     

Pure interferon gamma enhances class II HLA antigens on human monocyte cell lines

Human recombinant interferon gamma (IFN-gamma) with a chemical purity of over 90% was shown to enhance membrane expression of HLA-DR antigens on cells from 3 human myelomonocytic lines (HL-60, U-937 and THP-1). Immunofluorescence techniques using a series of anti-HLA-DR monoclonal antibodies showed that the low, although variable, levels of DR-positive cells were clearly enhanced as soon as 24 h after incubation with IFN-gamma. Only IFN-gamma was able to exert this effect, since incubation with high concentrations of IFN-alpha or -beta did not induce any significant modification of the percentage of HLA-DR-positive cells. In contrast, doses of IFN-gamma as low as 2 units were effective, indicating a highly preferential, apparently selective effect of IFN-gamma for enhancement of HLA-DR expression. Private class II antigen expression was also enhanced by IFN-gamma on the U-937 cell line. Through its enhancing effect on class II public and private HLA antigens on the membrane of human monocytes, the IFN-gamma lymphokine may have a critical role in the modulation of antigen presentation by monocytes and on the regulation of HLA-DR-restricted cell cooperation.     

Classification of human heterophile antibodies

A classification of heterophile antibodies is proposed, which is based on interactions with guinea pig kidney homogenate. The major groups of antibodies combining with guinea pig kidney encompass Hanganutziu-Deicher antibodies, Forssman antibodies, and antibodies to Newcastle disease virus. Antibodies which fail to combine with guinea pig kidney are primarily those of Paul-Bunnell variety.     

Human malignant melanoma cell lines.

A series of 10 cell lines established from human malignant melanomas is described. The morphology varied but, in general, was epithelioid. Five produced visible melanin in culture but others produced melanin precursors. All lines were aneuploid and each had a distinctive human karyotype. One line appeared not to have the same sex as the patient but its karyotype was distinct from that of other lines.     

Expression of Epstein-Barr virus nuclear antigens 3, 4, and 6 are altered in cell lines containing B-type virus

A high proportion of HIV-positive sera were found to react with 130- and 180-kDa antigens which were present in the Jijoye cell line. The majority of the HIV-positive sera which detected these antigens also contained antibodies to Epstein-Barr virus (EBV) nuclear antigen 2B (EBNA2B) suggesting a relationship between B-type EBV strains and the expression of the 130K/180K antigens. Cell lines were established by infection of B lymphocytes with different A- and B-type strains of EBV. Incubation of these lines with sera from individuals infected with either A-type or B-type EBV strains demonstrated that the 130K and 180K antigens were only expressed by cell lines containing B-type virus. Sera from individuals infected with A-type EBV did not react with the 180K antigen in any cell lines but could detect EBNAs 3, 4, and 6 antigens in the A-type cell lines. Restriction enzyme analysis of the BamHI E region of the EBV genome revealed marked differences between the A and B types of the virus. These results demonstrate that expression of antigens encoded from the BamHI E region of EBV (EBNAs 3, 4, and 6) are altered in cell lines transformed by B-type strains of EBV.     

Expression of human GIPC1 in normal tissues, cancer cell lines, and primary tumors

GIPC1/RGS19IP1/GIPC, GIPC2, and GIPC3 are a family of central PDZ-domain proteins with GH1 and GH2 domains. GIPC1 interacts with GTPase-activating protein RGS19/RGS-GAIP, TGFbeta type III receptor, receptor tyrosine kinase TrkA, and integrin alpha6A subunit. Xenopus homologue of human GIPCs interacts with Frizzled-3 class of WNT receptor. We investigated expression of human GIPC1 mRNA in normal tissues, cancer cell lines, and primary tumors. GIP1A probe (nucleotide position 1075-1483 of GIPC1 cDNA) hybridized to GIPC1 mRNA of 1.8 kb in size. GIPC1 mRNA was almost ubiquitously expressed in various normal tissues. Expression level of GIPC1 mRNA was relatively lower in bone marrow and peripheral blood leukocytes. GIPC1 mRNA was relatively highly expressed in gastric cancer cell lines OKAJIMA, TMK1, MKN28, MKN45, MKN74, KATO-III, pancreatic cancer cell line AsPC-1, colorectal cancer cell line SW480, and lung cancer cell line A549. On the other hand, GIPC1 mRNA was almost undetectable in leukemia/lymphoma cell lines HL-60, Raji, and Daudi. Expression of GIPC1 mRNA was down-regulated in 12 out of 14 cases of primary kidney tumors, 10 out of 18 cases of primary colorectal tumors, 3 out of 8 cases of primary gastric cancer, 3 out of 3 cases of primary prostate cancer. Because GIPC1 induces increased expression of TGFbeta type III receptor at the cell surface and enhanced responsiveness to TGFbeta, down-regulation of GIPC1 mRNA in tumors might promote cellular proliferation through interference of TGFbeta signaling.