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C Cillo

Publications and source records attributed to C Cillo.

35 records · Page 2Linked to original sources
PubMed1985

Early hemopoietic progenitor cells: direct measurement of cell cycle status.

We have investigated the cell cycle status of murine hemopoietic progenitors using vital DNA staining and flow sorting. Suspended Balb/c bone marrow cells were stained with Hoechst 33342 dye and separated first on light scattering properties; this procedure allowed a 5-fold enrichment in progenitor cells. A second sorting based on DNA content indicated that 80% of these cells were in G0/G1 and 20% in S-G2 + M. When G0/G1 and S-G2 + M cells were assayed separately in methylcellulose cultures, or with the in vivo colony forming assay, the G0/G1 cells were shown to be markedly enriched in CFU-S, BFU-E, and GM-CFU as compared to S-G2 + M cells with the final recovery increased 20-fold. Comparison of different strains or age groups yielded results identical to those obtained with Balb/c with the exception of C57B1/6. In the latter strain only a 3-fold enrichment could be observed in the G0/G1 fraction. These results demonstrate that the majority of early hemopoietic progenitors are in the G0/G1 phase of the cell cycle.

Animals↗
PubMed1985

Phenotypic and functional heterogeneity of human peripheral blood T lymphocytes producing colony stimulating factor. A clonal and precursor frequency analysis.

In this report we describe the precursor frequency and the subset distribution of peripheral blood human T cells producing lymphokine(s) acting on the proliferation and differentiation of bone marrow precursors of the granulocyte/macrophage series, as assessed in a liquid microculture assay. Because the sensitivity of this system was similar to that of the classic colony formation assay in semi-solid (methylcellulose) medium, it is likely that the lymphokine activity measured in this assay corresponds to the colony-stimulating factor (CSF) activity. Single human T lymphocytes, isolated from peripheral blood by E rosetting and Ficoll-Hypaque gradients, were seeded into microculture wells by limiting dilution or micromanipulation techniques and were incubated under culture conditions that allow clonal expansion of essentially all T cells. After 15 to 20 days, microcultures were stimulated with PHA and CSF activity was assayed in culture supernatants 24 hr thereafter. About 45% (1/2.3) of peripheral blood T cells were found to give rise to CSF-producing progenies. Moreover, when fluorescence-activated cell sorter-purified T4+ and T4- (or T8- and T8+) were analyzed, the frequency of the precursors of CSF-producing cells was 1/1.5 in the T4+ subset, whereas approximately one-third of the T8+ cells had this functional potential. To additionally characterize T cells responsible for CSF production, unfractionated T cells as well as T4+ and T4- cells were cloned by single cell micromanipulation. The resulting clones were analyzed simultaneously for the production of IL 2, production of CSF and for cytolytic activity in a lectin-dependent assay. It was found that 25/48 clones obtained from unfractionated T cells produced CSF, whereas 23/48 and 19/48 produced IL 2 or had cytolytic activity respectively. Six of the 25 CSF-producing clones had only this functional capability, whereas the remaining clones in addition displayed cytolytic activity (4/25), IL 2 production (10/25), or both (5/25). A similar functional heterogeneity was observed among T4+ and T4- clones, thus indicating that T cells producing CSF are functionally heterogeneous within both the T4+ and T8+ subsets.

Cell Differentiation↗
PubMed1984

Antigenic heterogeneity of clones and subclones from human melanoma cell lines demonstrated by a panel of monoclonal antibodies and flow microfluorometry analysis.

Cells from two melanoma cell lines, Me43 and GLL-19, were cloned in methylcellulose cultures and 20 randomly selected colonies from each line were picked up by micromanipulation, expanded in liquid cultures, and considered as clones of the original cell lines. The antigenic cell surface phenotype of these clones defined by panel of 12 monoclonal antibodies (MAb) was analyzed by flow microfluorometry (FMF) using a fluorescence-activated cell sorter (FACS II) and compared with the known stable phenotype of the parent cell line. The antibody panel consisted of eight MAb against melanoma-associated antigens, two MAb against monomorphic determinants of HLA-DR (la) and HLA-ABC, respectively, one MAb against the common acute lymphoblastic leukemia antigen (CALLA) and one MAb against carcinoembryonic antigen used as control. A remarkable heterogeneity in terms of qualitative and quantitative expression of the cell surface antigens studied was observed among and within the different clones. The single-cell origin of the clones was assessed by comparing the clonogenic cell frequency, determined by limiting dilutions in microculture plates, with the cloning efficiency observed in Petri dishes. Both techniques using methylcellulose medium gave the same percentages of growing colonies. Cells from four Me43 clones were recloned in methylcellulose and the phenotype of five randomly selected subclones from each clone was analysed using the same panel of monoclonal antibodies. Each subclone also displayed heterogeneity with individual phenotypes different from that of the original clone and from the parental Me43 cell line. The antigen expression by individual cells in situ within clones was analyzed on frozen sections from colonies using the same panel of MAb and a biotin-avidin immunoperoxidase method. The results confirmed the marked heterogeneity of antigen expression within and among colonies, as indicated by the FMF analysis.

Antibodies, Monoclonal↗
PubMed1980

Fluctuations of BFUe and CFUe cycling after erytrhoid perturbations: correlation with variations of pool size.

The pool size of erythroid burst-(BFUe) and colony-forming units (CFUe) has been evaluated in normal or hypoxia-induced polycythemic mice at sequential times after either transfusion or administration of purified erythropoietin (Ep). The present investigations were focused on the in vitro tritiated thymidine (3H-TdR) suicide index of the erythroid precursors in these two experimental models. After transfusion an early but transient decline of the DNA sythesis index was observed in the BFUe pool, whereas this parameter showed a later, more prolonged decrease within the CFUe population. Symmetric patterns were documented after Ep injection: an early, transient elevation of the 3H-TdR sensitivity at the BFUe level and a late, more persistent rise within the CFUe compartment. In all experiments no modification of this indes was observed within the pool of the CFUe. These fluctuations of BFUe and CFUe cycling were temporally consistent with modifications of their respective pool size, as previously reported. Thus, variations of the pool size may be at least partially mediated by the cycling activity. Furthermore, early fluctuations of BFUe proliferative rate indicate that, after erythroid perturbations, this pool may initially be sensitive to and regulated by modifications of Ep activity. Later, however, compensatory mechanisms allow the BFUe to escape from this early Ep influence, thus leading its cycling activity and pool size to return to normality and stabilize.

Animals↗
PubMed1979

Early fluctuations of BFU-E pool size after transfusion or erythropoietin treatment.

The kinetics of erythroid burst-(BFU-E) and colony-forming units (CFU-E) have been evaluated in marrow and spleen of normal or polycythemic mice, respectively, after transfusion or administration or purified erythropoietin (Ep). Ep injection induces an early but temporary rise of BFU-E number, versus a later but more prolonged expansion of the CFU-E pool. Symmetric patterns are observed after transfusion, i.e., an early but transient depletion of the BFU-E population, versus a later but more persistent decrease of the CFU-E number. It is suggested that the size of the BFU-E compartment is Ep-dependent in the "early" phase after erythroid perturbation. Later on, however, compensatory mechanisms allow the BFU-E pool to "escape" from the early Ep influence, thus allowing its size to return to and stabilize in the near normal range. It is further suggested that the BFU-E may represent an early target cell of Ep stimulus.

Animals↗
PubMed1978

Role of the hypophysis in erythropoietin production during hypoxia.

Hypophysectomized or sham-operated male rats were exposed to hypoxia (0.42--0.40 or 0.37--0.35 atm for 6, 12, or 24 hr) applied 2 wk to 7 mo after surgery. Erythropoietin (Ep) levels in rat serum were evaluated on the basis of the exhypoxic polycythemic mouse assay. Ep activity evoked by hypoxia was significantly lower in hypophysectomized rats than in sham-operated controls. Progressive increase of the EP response to hypoxia correlated with extension of the time interval between hypophysectomy and hypoxia from 2 wk to 2--4 mo apparently mediated by the simultaneous inverse decline of red cell mass (RCM) values, i.e., of the "relative plethora" induced by a low O2 demand associated with relatively high RCM values. However, after 3--7 mo hypoxic Ep activity was still lower than in sham-operated controls. In these ablated animals the relative plethora became negligible or absent; accordingly, the Ep response apparently had reached plateau levels. These studies indicate that hypophysis (hypophyseal and target hormones, with the exception of estrogens) modulates Ep production under hypoxic conditions, possibly via a permissive enhancement of renal Ep activity.

Animals↗
PubMed1977

Kinetics of erythroid and myeloid stem cells in post-hypoxia polycythaemia.

The number of erythroid burst-(BFU-E) and colony-forming units (CFU-E), as well as of myeloid-macrophage colony-forming units (CFU-C), has been evaluated in tibial marrow and spleen of ex-hypoxic polycythaemic mice, at sequential time intervals after the end of hypoxia. In both marrow and spleen, the kinetics of the CFU-E pool is characterized by a sharp fall from above normal to lower than normal values. BFU-E and CFU-C however rise from below normal to higher than normal levels. These results have been correlated with both the erythropoietin (Ep) and the erythropoietic activity curves. It is apparent that Ep levels largely control both the differentiation and the amplification of the CFU-E pool and it is suggested that Ep may act as a 'survival factor' at the CFU-E level and/or increase the flow of cells from BFU-E to CFU-E. The difference in response between CFU-E and BFU-E favours a clearcut distinction between these populations, whereas the similarity between the BFU-E and CFU-C response suggest a close relationship between these two cell populations. It is also of interest that the murine spleen functions as a large reservoir of erythroid microenvironment for hypoxia-induced stress erythropoiesis.

Animals↗
PubMedDate not supplied

HOX genes in human cancers.

Knowledge about transcription factors is crucial for understanding the molecular basis of neoplasia. Homeobox-containing genes are a family of transcription factors mostly involved in normal development. Class I human homeobox-containing genes (HOX genes) are organized in four clusters on different chromosomes. The order of the genes within each cluster is highly conserved throughout evolution suggesting that the physical organization of HOX genes may be (1) essential for their expression and (2) responsible for major biological functions. We have studied HOX gene expression in several human tissues and organs as well as in their neoplastic counterparts. We have observed (a) characteristic patterns of HOX gene expression for each normal solid organ analyzed, (b) altered HOX gene expression in kidney and colon cancer, (c) a correlation between HOX gene expression and different histological types of primary small cell lung cancer (SCLC) and (d) marked alterations of HOX gene expression among primary and metastatic SCLC variant types. Furthermore, we have shown that differential patterns of HOX gene expression correlate with the adhesion profile (VLA-2, VLA-5, VLA-6 and ICAM-1) and N-RAS mutation in clonal melanoma populations isolated from a single human melanoma metastasis. This suggests that HOX genes act as a network of transcriptional regulators involved in the process of cell to cell communication during normal morphogenesis, the alteration of which may contribute to the evolution of cancer.

Adult↗