Characteristics of hematopoietic cell line established from human myelomonocytic leukemia

Summary The peripheral blood of an acute myelomonocytic leukemia patient has been cultured for 16 months. The culture is at present at the 140th population doupling level. The cultured cells have the characteristics of so-called lymphoblastoid cells and proliferate actively as individual cells in small clusters, or in large clumps consisting of large mononuclear cells. Some of these cells appeare to be lymphoid, but the majority are immature mononuclear cells with a tendency to lobulate. They gave a weakly positive peroxidase reaction at the beginning of cultivation, and have given a strongly positive esterase reaction persistently. The cytoplasm shows ciliary or tail-like projections as the cell matures. Complement (C₃) receptor and IgG receptors are found on the cell surface, and active phagocytosis is mannifest. Colloidal iron particles or viable red blood cells attached to the cell membrane suggesting possible differentiation to reticulum cells or macrophages. The cultured cells are mostly diploid but some cells show chromosome abnormality.Herpes type virus was found in the nucleus, cytoplasma and on the cell membrane. The transplantation of cultured cells to the cheek pouch of hamsters produced small tumors with histological findings resembling reticulum cell sarcoma.The authors wish to thank Miss Fujiko Yokoyama and Akiko Miyoshi for their technical assistance. This work was supported by a cancer research grant 49101 from Kawasaki Medical School.     

Differentiation in human myeloblastic leukemia studied in cell culture.

Normal adult hemopoiesis orginates in pluripotent stem cells; among the early differentiated descendents of such cells are progenitors committed to the erythropoietic, granulopoietic, or megakaryocytic pathways of myeloid differentiation. These may be detected in cell culture by developmental techniques, in which progenitors form colonies in viscid or semisolid media in response to appropriate stimulation. Certain diseases of hemopoiesis also originate in pluripotent stem cells; these include chronic myeloblastic leukemia, acute myeloblastic leukemia, polycythemia vera, and idiopathic myelofibrosis—the clonal hemopathies. The hypothesis is advanced that the distribution of cell classes among patients with clonal hemopathies is determined both by the differentiation potential of each pluripotent stem cell maintaining an abnormal clone and by random events occurring during clonal expansion. The latter process may account for the large variations observed between patients when committed progenitors are assayed in cultures of marrow from patients with acute myeloblastic leukemia (AML). This variation may also be used to estimate lineage relationships in the clonal hemopathies. When applied to myelopoiesis in AML, obvious differences from the normal are not detected. The analysis is consistent with the view that the blast cell population in AML is distinct from the leukemic myelopoiesis occurring within an abnormal clone. A new assay procedure is described for progenitor cells related to blast cell proliferation. Finally, these concepts are used to develop a model for the pathogenesis and cellular characteristics of AML.     

HL-60细胞中IL6基因的克隆及鉴定

目的：为探索性构建重组人白细胞介素６－绿脓杆菌外毒素融合蛋白（ＩＬ６－ＰＥ４０）以选择性杀伤高表达ＩＬ６受体（ＩＬ６Ｒ）的白血病细胞,本研究试图从人急性早幼粒细胞白血病细胞株（ＨＬ－６０）中克隆Ｎ－末端缺失２４个氨基酸的人ＩＬ６基因（ＩＬ６ｃＤＮＡ）并构建含此基因的重组质粒。方法：根据ＩＬ６基因序列设计合成可扩增ＩＬ６ｃＤＮＡ的特异性引物;利用基因重组技术,构建含ＩＬ６基因的重组质粒ｐＵＣ－ＩＬ６。结果与结论：本文首次从ＨＬ－６０中扩增出预期４８０ｂｐ的ＩＬ６ｃＤＮＡ,将其克隆至ｐＵＣ１８质粒中,命名为ｐＵＣ－ＩＬ６,并经ＥｃｏＲＩ／ＢａｍＨＩ双酶切电泳及序列分析鉴定加以确证,为进一步构建重组人ＩＬ６－ＰＥ４０奠定了坚实基础。     

Induction of granulocyte differentiation in a human leukemia cell line does not require cell division

The human promyelocytic cell line, known as HL-60, provides a model for the study of myeloid differentiation. The HL-60 can differentiate to granulocyte-like cells in the presence of dimethylsulfoxide (DMSO). We investigated the requirement for DNA synthesis in the myeloid differentiation process and found that induction of differentiation of HL-60 cells did not require 3H-thymidine incorporation as judged by autoradiography. The results indicate that the triggering of granulocyte differentiation of HL-60 can occur in the absence of DNA synthesis.     

人胚胎干细胞H1培养条件的优化

目的:采用不同培养基和饲养层培养人胚胎干细胞H1,建立适合H1细胞增殖的最佳条件并分析其基本生物学特性。方法:鼠源性饲养层采用ICR品系小鼠胚胎成纤维细胞(MEF),人源性饲养层采用人胚胎成纤维细胞系(HFF-1)。H1基本培养基配制分别采用传统DMEM/F12和改良培养基Knock-outTM DMEM。实验共分为MEF DMEM/F12、MEF K-DMEM、HFF DMEM/F12、HFF K-DMEM组。H1基本生物学特性检测采用免疫荧光、RT-PCR、碱性磷酸酶和核型分析。结果:MEF DMEM/F12组中H1克隆形态规则,不发生分化,增殖速度快;而MEF K-DMEM组细胞克隆传代后第4日发生分化;HFF DMEM/F12组和HFF K-DMEM组细胞传代后第3日就显示出分化趋势,克隆变扁。MEF DMEM/F12组中H1细胞保持正常核型和基本生物学特性。结论:不同的人胚胎干细胞系最佳培养条件是不同的,建立的MEF DMEM/F12组培养条件最适合H1细胞增殖。     

Cytogenetic abnormalities in a human null cell leukemia line (REH)

The chromosomes of a null cell line (REH₆) derived from peripheral blood leukocytes of a patient with acute lymphoid leukemia (ALL) were examined with conventional and R-banding techniques on fresh and established cells bearing ALL-associated antigen(s). Five marker chromosomes were found in both fresh and established cells. The constitution of these abnormalities is related to and explained by breaks and translocations involving five chromosomes. Furthermore, one X chromosome is completely missing. The modal chromosome number in vitro was 45 up to 10 months, and 46 from 10 to 43 months after establishment of the cell line.     

Infection of a human leukemia K-562 cell line with Semliki Forest virus

Summary Infection of the human leukemia hematopoietic stem cell line, K-562, with Semliki Forest virus (SFV) can be characterized by three stages: (1) an early virus-proliferating stage lasting 1 to 4 days post-infection (pi) in which infectious virus is produced in high titers (10³ pfu/cell) but in which there is minimal cytopathic effect. All cells appear viable by trypan blue dye exclusion, although they do not proliferate, and DNA and cell protein synthesis decrease to less than 3% of uninfected controls within 24 hours; (2) an intermediate stage extending from day 5 to about day 24–30 pi in which the amount of infectious virus declines to low levels. During this stage, viral protein synthesis decreases to undetectable levels, although viral gylcoproteins are readily demonstrated by immunofluorescence and by immunoblot; however, capsid protein appears to degrade within 21 days pi. Cell numbers remain constant but the viability of the non-proliferating cells determined by trypan blue exclusion could not be determined with confidence; (3) a final long-term stage in which viral glycoproteins, E1 and E2, are detectable by immunoblots and immunofluorescence for many months but the cells are metabolically inactive and do not synthesize viral proteins. These non-viable cells do not lyse for as long as 2 years.     

Transmission of human T-cell leukemia virus type I to an S+L- cat kidney cell line

The S+L- cat kidney cell line CCC was cocultivated with lethally irradiated human lymphoid cell lines that were producing human T-cell leukemia virus type I (HTLV-I). Eight of nine S+L- CCC sublines that had been cocultivated with nine different HTLV-producing T-cell lines gave positive reactions for HTLV antigens by indirect immunofluorescence assay. One subline CCC/2M was cloned. The percentages of fluorescent cells differed markedly in different sublines and clones. Southern blot hybridization with HTLV probes and electrophoresis of immunoprecipitates indicated that defective HTLVs were often transmitted into S+L- cat cells. S+L- CCC cells were permissive for HTLV and the properties of HTLV-infected cat cells were heterogeneous.     

Adhesion of enterotoxigenic Escherichia coli to the human colon carcinoma cell line Caco-2 in culture.

Enterotoxigenic Escherichia coli (ETEC) strains possessing colonization factor antigen I (CFA/I), CFA/II, CFA/III, and antigen 2230 were tested for their ability to adhere to the following cell lines: HeLa, HEp-2, HRT 18, Hutu 80, MDBK, MDCK, Vero, and Caco-2. ETEC strains adhered only to the Caco-2 cell line. Irrespective of the known adhesive factors, the ETEC strains that adhered to the brush border of human enterocytes also adhered to the Caco-2 cell line. The negative variants, which were cured of the plasmid encoding the adhesive factor, did not adhere. Adhesion of ETEC strains no longer occurred when the Caco-2 cells were pretreated with the homologous colonization factor antigen or when the bacterial cells were pretreated with homologous antibodies raised against the adhesive factors. This indicates that this adhesion is specific and that a different receptor exists for each type of adhesion factor. Electron micrographs of cross sections of the monolayer showed that the adhesion of ETEC strains to the brush border microvilli does not induce any lesion. Therefore, the Caco-2 cell line behaves in the same way as human enterocytes do.     

Induction of HLA-DQ antigen expression on a human promyelocytic leukemia cell line HL-60

A variant strain of HL-60, which is positive for HLA-DR antigen, was induced to express HLA-DQ antigen following treatment with phorbol esther. It was preceded by cell cycle arrest in the G1 phase and was accompanied by augmented phagocytosis. This differential expression of HLA class II antigens on this subline may contribute to understanding the functional role of HLA class II antigens in the hematopoietic differentiation of macrophage cells.     

Entry of human immunodeficiency virus (HIV) into MT-2, human T cell leukemia virus carrier cell line

Summary The ultrastructural features of early events in human immunodeficiency virus (HIV) infection of HTLV-I-carrying MT-2 lymphocytes were investigated by electron microscopy. Within 10 min after virus inoculation at 37°C, the virus entered the cell in two ways; (1) the virus attached to the lymphocyte membrane and the viral core entered the cell after fusion of the viral envelope with the cell membrane, and (2) part of the cell membrane to which the virus was attached became invaginated, the virus became trapped in a phagosome and the viral core entered after the fusion of viral membrane with the vacuolar membrane. Thereafter, some cells were observed to form syncytia with multiple nuclei. When the proportion of anti-HIV antibody-reactive cells present exceeded 90%, virus production was strongly activated, and budding on the cell membrane was frequently observed.     

Enrichment of a human leukemia cell line (K562) with a plant histaminase

Inflammation Research -     

Modal karyotype of human leukemia cell line, K562 (ATCC CCL 243)

The chromosome composition of the human leukemic cell line, K562, deposited at the American Type Culture Collection (ATCC) (CCL 243), was studied and further compared with those reported by other laboratories. In CCL 243, the modal karyotype had 55 normal and 14 constitutive marker chromosomes and occurred in nearly 50% of the cultured cell population. Other variant karyotypes were observed either at a low frequency in coexisting minor cell types or only once in all other analyzed metaphases. Normal chromosome #9 and the chronic myeloid leukemia (CML) specific Ph/t(9;22) were absent. It appears that karyotypes of most K562 cultures distributed world-wide are similar to each other, with the stable modal chromosome number, and therefore, the modal karyotype presented here should be useful in karyotypic identification of a cell line.     

Amino acids exhibit anti-inflammatory effects in human monocytic leukemia cell line, THP-1 cells

Objective  

The elemental diet is one of the effective therapies for inflammatory bowel disease. However, the mechanism remains unclear, and there have never been reports about the inhibitory effects of amino acids in human monocytes/macrophages. We investigated the inhibitory effects of amino acids on cytokine production or expression of adhesion molecules that are involved in inflammatory diseases, in human monocytes/macrophages.     

Susceptibility of a line of dolphin kidney cell culture to several herpesviruses.

A cell line was established from cell cultures of kidney cortex of a pantropical spotted dolphin, Stenella attenuate. The replication of 6 strains of herpesviruses was studied in the cells. Five strains of them, herpes simplex virus type I and type II, equine rhinopneumonitis virus, infectious bovine rhinotracheitis virus and Aujeszky's disease virus, were grown fairly well in showing clear cytopathic effects and plaques under agar overlay medium.