Surface phenotype analysis of human monocyte to macrophage maturation.

Cells of the mononuclear phagocyte system arise from circulating blood monocytes. Upon emigration from the vasculature, monocytes differentiate into macrophages, a process that monocytes similarly undergo in vitro. We have established primary cultures from elutriated or adherence-purified blood monocytes and analyzed the antigenic modulation during monocyte to macrophage transformation, which could be followed by the expression of specific antigens and which required as yet unknown inducer signals present in the serum. It is shown that in the absence of serum monocytes only survive in vitro when cultured adherent to plastic but rapidly die in suspension culture. Starting at 0.5%, serum induced maturation dose-dependently, with the optimal concentration being 2 to 5%. Of those antigens not present on monocyte, the low-affinity Fc receptor (CD16), the alpha-chain of the vitronectin receptor (CD51), gp65-MAX.1, and gp68-MAX.3 were expressed only upon serum-induced macrophage differentiation, whereas the transferrin receptor (CD71), MAX.26, and to some degree also gp65-MAX.11 appeared to be independent of maturation and were also found on primary cultures of adherent monocytes under serum-free conditions. In addition, the rapid induction of HLA class II antigens (within 24 hr) was similar with and without serum, as was the continued high-density expression in long-term culture. The monocyte-specific CD14 antigen was down-regulated in the absence of serum but kept its level of expression on differentiated macrophages. In comparison, alveolar and peritoneal macrophages, respectively, differed in their antigenic phenotype: Alveolar macrophages expressed high HLA class II antigens but low CD14, whereas for peritoneal macrophages the opposite was found. Both interferon-gamma and -alpha suppressed macrophage maturation in vitro but had contrary effects on HLA class II and CD16 expression: Interferon-gamma up-regulated the two types of antigens, which, in contrast, were down-regulated by interferon-alpha.     

Ecotaxis: the principle and its application to the study of Hodgkin''s disease.

A study of the function, characterization and distribution of T and B lymphocytes in five children with Hodgkin's disease is presented. The results, indicating that lymphocyte depletion in the peripheral blood does not necessarily reflect an overall lack of circulating lymphocytes, are presented to demonstrate that failure of ecotaxis (normal lymphocyte migration and distribution) can occur in man. The underlying reasons for such failure and their relevance to the pathogenesis of Hodgkin's disease are discussed.     

A quantitative study of mast cells in Hodgkin's disease.

Mast cells were counted in 45 specimens from patients with Hodgkin's disease and in five lymph nodes showing follicular hyperplasia. A consistent finding was that of few mast cells in the lymphocyte depleted and lymphocyte predominant Rye subtypes of Hodgkin's disease and in the mixed cellularity variant; however, mast cells were much more prevalent in nodular sclerosing Hodgkin's disease and in the hyperplastic nodes. The mast cells were selectively stained by means of the astra blue technique.     

Monocyte PGE2 secretion in Hodgkin's disease and its relation to decreased cellular immunity.

The secretion of PGE2 from monocytes of newly diagnosed patients with Hodgkin's disease (HD) was compared to that of patients in remission, who were not receiving either chemotherapy or radiotherapy, and normal controls. We found that monocyte monolayers of some patients, both newly diagnosed and those in remission, secreted markedly elevated levels of PGE2. The lymphocyte proliferative response to PHA was increased to a similar extent in both newly diagnosed patients and those in remission when cultured in the presence of indomethacin. PGE2 concentrations in the medium of mononuclear cultures correlated with the lymphocyte proliferative response to PHA (P less than 0.05). However, no correlation of monocyte PGE2 production with decreased E rosette forming lymphocytes, anergy or clinical stage could be demonstrated. We suggest that PGE2 secretion by monocytes is indicative of an 'activated' state of these cells. It is, however, unlikely that PGE2 is the only molecular species responsible for the decreased cellular immune function in HD. 'Activated' monocytes may be part of the immune response in this disease and may be responsible for the decreased cellular immunity.     

Development, differentiation, and maturation of fetal mouse yolk sac macrophages in cultures

The development and differentiation of macrophages in the fetal mouse yolk sac were studied morphologically in four different culture experiments. In the culture of mouse embryos with yolk sac, the development of fetal macrophages was demonstrated to precede that of promonocytes and monocytes in the yolk sac. In vitro differentiation of the fetal macrophages was consistent with the results of our previous in vivo observation indicating that fetal macrophages were differentiated from primitive macrophages, but not from the monocytic cell series. Differentiation of primitive macrophages into fetal macrophages, before the development of promonocytes and monocytes, was reproduced in the culture of cell suspensions from the fetal mouse yolk sacs, with a mouse bone marrow stromal cell clone (ST2) particularly with those at 8 days of gestation. In the soft agar or liquid culture of yolk sac cells with LP3-conditioned medium, monocyte-macrophage colonies were effectively induced, but not fetal macrophage colonies. The results provide evidence for the existence, in yolk sac hematopoiesis, of two distinct macrophage populations: a fetal macrophage population and a monocyte-derived macrophage population. The data indicate an obvious difference in development and differentiation between the two populations and the temporal precedence of fetal macrophages appearing before monocyte-macrophages.     

Ewing's sarcoma. Characterization in established cultures and evidence of its histogenesis

We have studied the ultrastructure and collagen biosynthetic profiles of three cultured lines of Ewing's sarcoma (ES) to better understand the histogenesis of this tumor. The histology and ultrastructure of the original tumors were characteristic of ES. Light and electron microscopic appearance of the cell lines and of tumors formed in nude mice from injection of these lines were similar to the native tumors. Isozyme studies of the ES cell lines demonstrated that they were not HeLa and were different from other known cell lines. Collagen profiles performed by immunofluorescence, polyacrylamide gel electrophoresis, and immunoprecipitation showed conclusively that ES in tissue culture produces collagen types I, III, and IV. This collagen profile coupled with the morphologic and ultrastructure features of the ES cell lines leads us to conclude that Ewing's sarcoma is derived from a multipotential primitive mesenchymal cell, uncommitted to osteoblastic, fibroblastic, or endothelial origin.     

Type IV collagen in Hodgkin's disease. An immunohistochemical study

A series of 50 lymph nodes affected by Hodgkin's disease (HD) and 10 further nodes exhibiting the features of reactive follicular hyperplasia (RFH) have been studied. A peroxidase-antiperoxidase (PAP) sequence for the demonstration of type IV collagen has been applied to routinely processed paraffin-embedded sections of these specimens. Blood vascular structures of all calibers were clearly demonstrated, as were lymphatic sinuses. Structures recognizable as the latter were generally lacking in HD, but the sinus structures in the cases of RFH were well defined. Blood vessels were highly active on type IV collagen staining in the interfollicular areas of reactive nodes but were relatively scanty in lymphocyte-predominant, mixed-cellularity, and lymphocyte-depleted HD. However, in the nodular sclerosing Rye subtype, blood vessels containing type IV collagen were as numerous as in RFH and could be seen within and around the cellular nodules. A similar number of type IV collagen-containing blood vessels were also apparent in the cellular variant of nodular sclerosing HD. The results are compared with those obtained on immunostaining for Factor VIII-related antigen and on binding of Ulex europaeus lectin I, and type IV collagen is considered to be the most sensitive vascular marker compared with these.     

Characterization of the adherent cells developed in Dexter-type long-term cultures from human umbilical cord blood

We have previously shown that when human umbilical cord blood (UCB) cells are cultured in standard Dexter-type long-term cultures (D-LTC), adherent cells develop forming a discrete net on the bottom of the culture flask. The identity of such cells, however, has not been defined. Accordingly, the major goal of the present study was to characterize the adherent cells developed in standard UCB D-LTC. Cultures were established from 14 UCB samples and from nine bone marrow (BM) samples, as controls. Both UCB and BM cultures were initiated with the same number of mononuclear cells (MNC) (2.5 x 10(6) MNC/ml). After three weeks in culture, adherent cell numbers in UCB D-LTC were 24%-30% of the numbers found in BM cultures. More than 90% of the adherent cells in UCB D-LTC expressed the acid phosphatase enzyme, whereas no alkaline phosphatase-positive cells were observed. This was in contrast to BM D-LTC, in which alkaline and acid phosphatase were expressed by 60%-75% and 20%-45% of the adherent cells, respectively. Immunochemical analysis showed that CD61 (osteoclast marker) and Factor VIII (endothelial cell marker) were not expressed by the adherent cells developed in UCB cultures. Interestingly, the majority of such cells expressed CD1a (dendritic cell marker), CD14, CD68 and CD115 (antigens mainly expressed by macrophagic cells). When the cultures were supplemented with the recombinant cytokines epidermal growth factor, basic fibroblast growth factor, platelet-derived growth factor or granulocyte-macrophage colony-stimulating factor (GM-CSF), only GM-CSF had a significant positive effect on adherent cell number. In order to test for some functional properties of the adherent cells developed in culture, production of stem cell factor (SCF), interleukin 6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) was assessed. IL-6 and TNF-alpha showed elevated levels in UCB D-LTC, whereas SCF levels were always below detection. Finally, analysis of fibroblast progenitors (fibroblast colony-forming units [CFU-F]) showed that these cells were present in BM samples (6 CFU-F/10(5) MNC) and were totally absent in UCB samples. Taken together, the results of the present study indicate that the vast majority of the adherent cells developed in standard UCB D-LTC belong to the macrophage lineage and that fibroblasts seem to be absent. Interestingly, the high proportion of CD1a+ cells suggests that dendritic cells are also present in these cultures.     

Increased histidine decarboxylase expression during in vitro monocyte maturation; a possible role of endogenously synthesised histamine in monocyte/macrophage differentiation

OBJECTIVE: In this study the expression of histidine decarboxylase (HDC), the pivotal enzyme in histamine formation and the effect of endogenously produced histamine on differentiation antigens was examined during in vitro differentiation of human monocytes. MATERIAL AND TREATMENT: Human elutriated monocytes from healthy volunteers were incubated with macrophage colony stimulating factor (M-CSF) and the expression of HDC was followed at both mRNA and protein levels. To study the possible function of histamine we followed the expression of some cell surface markers (CD14, CD16, CD91, CD49d and CD11c) relevant for phagocytic differentiation upon incubation in the presence of different histamine inhibitors, an HDC inhibitor: S(+)-alpha-fluoromethyl-histidine HCl, (alphaFMH), a compound that disturbs the interaction of histamine with intracellular cyp450 moieties: N,N-diethyl-2-[4-(phenylmethyl) phenoxy]-ethanamine HCI, (DPPE); and H1 and H2 receptor antagonists, Triprolidine and Cimetidine. RESULTS: During in vitro culture of elutriated human monocytes, in the presence of M-CSF, the gene expression and biosynthesis of HDC was considerably increased. The various antihistamine agents decreased the expression of the cell surface markers examined in this study. CONCLUSIONS: These data support the elevation of HDC expression during human monocytic differentiation and the possibility that monocyte-derived histamine is partially involved in regulation of M-CSF induced in vitro human monocyte/macrophage phagocytic differentiation.     

A critique and case study of nodular sclerosing Hodgkin's disease.

This paper presents a critique of the present concepts of the pathology of nodular sclerosing Hodgkin's disease and particularly of the criteria required for diagnosis for the condition and its separation from other types of Hodgkin's disease. In addition, the natural history and histopathology of the condition have been studied among 104 cases of Hodgkin's disease presenting at The London Hospital, and the significance of the appearance is discussed.     

Hypercalcaemia in Hodgkin's disease related to prostaglandin synthesis.

A case of paraneoplastic hypercalcaemic syndrome is reported in a patient with Hodgkin's disease. This was detected eight months before widespread lymphadenopathy became apparent. Lymphocyte depleted Hodgkin's disease was diagnosed. PTH (parathyroid hormone) activity was suppressed and PTHRP (PTH related protein) was less than 5 pmol/l. 1,25(OH)2D3 was in the normal range. Plasma calcium values returned to normal after the administration of indomethacin. Thus the pathogenesis of the hypercalcaemia in this patient could be associated with the synthesis of prostaglandins.     

Coexisting Hodgkin's disease and mycosis fungoides. Immunohistochemical proof of its existence

Hodgkin's disease and mycosis fungoides have been rarely reported in the same patient. This coexistence has been debated in the medical literature. We studied such a patient and report, to our knowledge, the first immunophenotypic evidence for such a coexistence. Reed-Sternberg cells and their variants stained with anti-Leu-M1, Hefi-1, anti-Tac, anti-HLA-DR, and OKT9, but were negative for T cell markers 3A1, Leu-1, Leu-2a, and Leu-3a, a phenotype typical of Hodgkin's disease; infiltrating small lymphocytes were predominantly T cells and were phenotypically normal. In the skin lesions, cells with the phenotype of Hodgkin's disease were not present; the infiltrate was composed of helper T lymphocytes that were 3A1-negative, a phenotype characteristic of the malignant cells of mycosis fungoides. Unexpectedly, a dermatopathic lymph node from the same patient showed the presence of the Leu-M1 antigen on the majority of normal-appearing interdigitating reticulum cells; this was not the case with control dermatopathic lymph nodes from patients without a malignancy. The significance, implications, and possible interrelationships of the findings are discussed.     

Coexistence of Hodgkin's disease and Gaucher's disease.

A case of Hodgkin's disease associated with long-standing Gaucher's disease is presented and compared with the only previously reported case. The two diseases coexisted in lymph nodes both above and below the diaphragm, liver, bone marrow, and pancreas. The question of a possible relationship of the two diseases is discussed.