Ultrastructural peroxidase cytochemistry of three established human myelogenous leukemia cell lines, HL-60, KG-1 and ML-2

We studied the ultrastructural peroxidase cytochemistry of three established human acute myelogenous leukemia cell lines (HL-60, KG-1 and ML-2) to clarify the cytochemical differences and the differences in maturation/differentiation stages. HL-60 cells contained azurophil granules which exhibited strong peroxidase reaction. The more central region of individual granules is less reactive than the outer portion of these granules. The secretory apparatus of HL-60 cells, i.e. rough endoplasmic reticulum, perinuclear cisterna and Golgi apparatus, also contained peroxidase-reactive material. ML-2 cells contained azurophil granules with peroxidase reaction; however, nuclear membranes were peroxidase-negative and thin cisternae of rough endoplasmic reticulum exhibited only a faint peroxidase reaction. KG-1 cells were peroxidase-negative. From these findings KG-1 is classified as the least mature of the cell lines studied. ML-2 cells are considered early promyelocytes and HL-60 cells as late promyelocytes.     

Inhibition of the ecto-beta subunit of F1F0-ATPase inhibits proliferation and induces apoptosis in acute myeloid leukemia cell lines

Background

Tissue factor pathway inhibitor-2 (TFPI-2) is an extracellular matrix associated broad-spectrum Kunitz-type serine proteinase inhibitor. Recently, down regulation of TFPI-2 was suggested to be involved in tumor invasion and metastasis in some cancers.

Methods

This study involved 12 normal cervical squamous epithelia, 48 cervical intraepithelial neoplasia (CIN), and 68 cervical cancer. The expression of TFPI-2, Ki-67 and vascular endothelial growth factor (VEGF) were investigated by immunohistochemistry staining. The apoptolic index(AI) was determined with an in situ end-labeling assay(TUNEL). And the marker of CD34 staining was used as an indicator of microvessel density (MVD).

Results

TFPI-2 expression has a decreasing trend with the progression of cervical cancer and was significantly correlated with FIGO stage, lymph node metastasis and HPV infection. In addition, there were significant positive correlations between the grading of TFPI-2 expression and AI(P = 0.004). In contrast, the expression of TFPI-2 and VEGF or MVD was negatively correlated (both p < 0.001). However, we did not establish any significant correlation between Ki-67 and TFPI-2 expression in cervical cancer.

Conclusions

The results suggested that the expression of TFPI-2 had a decreasing trend with tumor progression of cervical cancer. There was a close association between the expression of TFPI-2 and tumor cell apoptosis and angiogenesis in patients with cervical cancer. TFPI-2 may play an inhibitive role during the development of cervical cancer.     

骨髓成纤维样基质细胞系对急性髓细胞白血病细胞系增殖与分化影响的研究

目的：以急性髓细胞白血病（acute myeloid leukaemia,AML）细胞系为研究对象,观察正常人骨髓成纤维样细胞系（human bone marrow fibro—blastoid stromal cell line,HFCL）对急性单核细胞白血病细胞系U937、急性粒细胞白血病敏感细胞HL-60和多药耐药细胞HL-60／VCR增殖和分化的影响。方法：建立U937、HL-60和HL-60／VCR细胞与HFCL细胞的共培养模型,实验分对照组、直接接触组和transwell组。采用台盼蓝拒染法测定生长曲线;硝基四氮唑蓝（NBT）确定细胞分化;流式细胞仪检测细胞周期和CD11b、CD13、CD14和CD33细胞表面抗原进一步鉴定细胞分化;west-ernblot检测增殖细胞核抗原（PCNA）和P-糖蛋白（P-gP）的表达。结果：与HFCL细胞共培养96h后,U937、HL-60和HL-60／VCR细胞的生长受抑,且与HFCL细胞直接接触组的抑制作用大于用tr-answell组,P〈0．01。同时发现AML细胞系与HFCL细胞共培养后,G0／G1期细胞比例均增高,而S期细胞均减少,P〈0．01;尤其是直接接触组CD11b和CD14表达也均增高（P〈0．01）,CD13和CD33变化不大,NBT阳性细胞轻度增高,且差异有统计学意义。Western blot检测结果显示,3种AML细胞系PCNA表达下调;以直接接触组为甚。提示介导着白血病细胞和骨髓基质细胞间的相互作用的整合素p1（VLA-4）和p2（LFA-1）,在HFCL细胞对AML细胞生长作用影响中起着不可忽视的作用。结论：HFCL对3种具有代表性的AML细胞系U937、HL-60和HL-60／VCR有增殖抑制和诱导分化作用,除了能抑制AML细胞的生长,抑制PCNA的表达,出现G0／G1期阻滞外,还能诱导其部分向单核细胞分化。     

Adenosine 3':5' cyclic monophosphate and myeloid leukemic cell proliferation in vitro

In order to elucidate the possible role of adenosine 3′:5′ cyclic monophosphate (cAMP) in the modulation of proliferation of human leukemic myeloblasts, we have determined cAMP levels of samples from patients with leukemia before and after in vitro suspension culture, in the presence and absence of leucocyte conditioned medium (LCM). cAMP levels correlated with the magnitude of increase of cell numbers after culture with LCM (rank-order correlation coefficient (r_s) = 0.57, P < 0.05) and with tritiated thymidine (³H TdR) incorporation during culture (r_s) = 0.68, P < 0.01). Additions of the phosphodiesterase resistant analogue 8-bromo cAMP to cultures caused modest increases of ³H TdR incorporation of 8 of 9 acute myeloid leukemia samples after 72h with LCM. Stimulation was optimal at 10^?7 M but in the absence of LCM occurred less often and required higher concentrations (10^?5 M). Inhibition was noted at concentrations of 10^?4 M and greater. Cells cultured in the presence of LCM had lower levels of cAMP (P < 0.01) particularly at points with maximal stimulation of ³H TdR incorporation (P < 0.01). These differences between paired points (with and without LCM) were small and no immediate effect of LCM upon cAMP levels, after 1 and 2-h incubations, was noted. These data suggest that cAMP promotes initiation of leukemic cell proliferation in vitro.     

BO-0742, a derivative of AHMA and N-mustard,has selective toxicity to drug sensitive and drug resistant leukemia cells and solid tumors

This is a preclinical study of BO-0742, a derivative of 3-(9-acridinylamino)-5-hydroxymethyl-aniline (AHMA) and N-mustard, as an anti-cancer agent. MTS assays revealed a broad spectrum of anti-cancer activities in vitro, with the greatest cytotoxicity against leukemia and neuroblastoma including those with drug resistant characteristics, and a good therapeutic index with leukemia being 10–40 times more sensitive to BO-0742 than hematopoietic progenitors. Administration of BO-0742 at an optimal dose schedule based on its pharmacokinetics significantly suppressed the growth of xenografts of human breast and ovarian cancers in mice. Thus, BO-0742 is a potent anti-cancer agent worthy of further clinical development.