Distribution of surface charge and concanavalin a-binding sites on normal and malignant transformed cells

Measurement of the rate of agglutination with the positively charged poly- -lysine of normal lymphocytes, Moloney-virus-transformed lymphoma cells, normal fibroblasts and SV40 transformed fibroblasts, has shown that the normal cells were agglutinated at a higher rate than the transformed cells. The labeling density of cationized ferritin in electron micrographs of sectioned cells, also indicated a higher charge density for the normal lymphocytes and fibroblasts. The normal cells showed a more regular clustered distribution of cationized ferritin than the transformed cells, and pre-fixation of cells with glutaraldehyde before labeling with cationized ferritin resulted in a random distribution in both types of cells. The transformed cells had a higher agglutinability than the normal cells by Concanavalin A (ConA) and this difference was also found after treatment of the cells with neuraminidase. Labeling with ConA-ferritin showed the same distribution on the sectioned normal and transformed cells. The results indicate that there was a difference in the redistribution of surface charge by cationized ferritin in normal and transformed cells and that there was no detectable difference in redistribution of ConA-binding sites with ConA-ferritin.     

Ultrastructural identification of round-granule EC cells in rabbit fundic mucosa

In endocrine (EC) cells of rabbit fundic mucosa, it is practically impossible to obtain unequivocal ultrastructural identification of all cells found in order to perform morphometric analysis. In addition to classic EC cells with pleomorphic granules, a cell type with entirely round granules is encountered which can be confused with non-EC cells. To solve this problem, all EC cells in our study were first identified by their 5-HT (immunocytochemistry) and argentaffinity. Examination of the fine structures of reactive cells then revealed that the round granules of EC cells were differentiated from those of non-EC cells by the existence of a dense core surrounded by a less dense halo, a feature providing unequivocal ultrastructural identification. EC cells with round granules showed less argentaffinity and less immunoreactivity to 5-HT as compared with classic EC cells. After labelling with [3H]L-dopa, EC cells with round-granules displayed an overall staining index higher than that of classic EC cells and comparable with that of D cells; however, the nuclear staining index was higher than that of D cells.     

The influence of 2-mercaptoethanol (2-ME) treatment of IgG antibody on its ability to induce cytotoxicity in nonsensitized lymphocytes

Administering dextran 2 hr before giving antigen (sheep red blood cells) to X-irradiated mice repopulated with B and T cells caused alterations in antibody synthesis. Besides a slight increase in the number of cells making IgM antibodies, cells producing IgG antibodies were detected at a time when none are usually present in untreated control animals.Repopulating X-irradiated mice with B cells treated with dextran either in vivo or in vitro and immunizing with sheep red blood cells resulted in twice the background number of sheep red blood cell-specific IgM-plaque-forming cells at 4.5 days of immunization as well as a small number of IgG-plaque-forming cells at 8.5 days. At these times, control animals given only B cells and sheep red blood cells possessed a background number of IgM-plaque forming cells and no IgG plaque-forming cells.Incubating B cells with θ-specific antiserum and complement to remove residual T cells prior to transplantation obliterated dextran's stimulus. Dextran's alterations of immunological responses towards unrelated antigens therefore appears to be manifested through T cells. The latter must be in company with B cells at the time of exposure to dextran. Thus, T cells upon contact with dextran apparently release B cell stimulatory factor (s) responsible for increasing the number of IgM-forming cells and for triggering IgG-forming cells.     

Cellular interaction between fixed and living cells. Transfer of radioactive materials from living cells to fixed cells

Transfer of radioactive materials to fixed cells from an overlying layer of living cells has been examined to determine whether fixed cells can act as acceptors of glycosyltransferases of living cells. After the incubation of living cells were removed by EDTA treatment, and the radioactivity associated with the fixed cells was determined. Lipids, proteins and carbohydrates were found to be transfered from the living cells to the fixed cells. The amount of radioactivity transferred to the fixed cells was dependent on the number of both fixed and living cells and increased with the time of incubation. When fixed cells were treated with chloroform-methanol before the addition of living cells, the transfer of both lipids and proteins to the fixed cells decreased drastically, but only a slight decrease incarbohydrate transfer was observed. Most of the radioactive materials transferred from living cells labeled with glucosamine or fucose to chloroform-methanol-treated fixed cells were solubilized by trypsin but not by the detergents tested. Approximately 55% of the materials transferred from the cells labeled with glucosamine could be solubilized by hyaluronidase and chondroitinase, and the rest was solubilized by neuraminidase and a glycosidase mixture. The treatment of chloroform-methanol-extracted fixed cells with trypsin caused a significant decrease in the transfer from cells labeled with glucosamine. When nucleotide sugars were used as the radioactive precursor, no significant amount of radioactivity was transferred to the fixed cells.     

Histochemical and ultrastructural identification of neurotensin cells in the dog ileum.

In the dog ileum, neurotensin cells stained with immunofluorescence or immunoperoxidase proved distinct from argentaffin (EC) cells, glucagon immunoreactive (GLI) cells and pancreatic peptide immunoreactive (PP) cells. Neurotensin cells showed various degrees of reactivity with Grimelius' silver. With electron microscopy, besides EC cells, large granule cells with a thin peripheral rim of Grimelius-reactivity (L cells) and large granule cells with variable Grimelius-reactivity of the core (N cells) were found. On distributive grounds, L cells were identified with GLI cells and N cells were interpreted as neurotensin cells.     

Induction of proliferation of human follicular (B type) lymphoma cells by cognate interaction with CD4+ T cell clones

We examined stimuli which are required for the induction of in vitro proliferation of follicular lymphoma cells, a low grade non-Hodgkin's B cell lymphoma characterized by a specific chromosomal translocation, t(14;18)(q32;q21), and by in vivo growth of the lymphoma cells in germinal center-like follicles infiltrated with CD4+ T cells. The purified follicular lymphoma cells, which are morphologically uniform, small, and dense, did not respond to stimulation with soluble lymphokines in the absence of T cells. Vigorous in vitro proliferation of follicular lymphoma cells was induced, however, when the follicular lymphoma cells were cultured with a CD4+ T cell clone which recognized alloantigens expressed by the lymphoma cells. This response required B-T cell contact, and was inhibited by anti-class II but not by anti-class I MHC mAb, indicating that these neoplastic B cells behaved as normal B cells and responded to normal activation and differentiation signals from T cells. After the cognate B lymphoma-T cell interaction occurred in culture, addition of IL-2 or IL-4 enhanced the proliferation of the tumor cells. These results, with a monoclonal and homogeneous population of B cells, affirm the idea that cognate interaction between B cells and Th cells is required for the effective activation of resting B cells. Moreover, these results suggest that a critical host-tumor interaction occurs in vivo, and that the polyclonal CD4+ T cells that infiltrate follicular lymphomas play a role in sustaining rather than inhibiting tumor growth in vivo. If so, therapies directed not only against the neoplastic cell but also against specific T cells and their cognate interactions with tumor cells may have a rationale.     

Use of Stable, Sensitized Cells in Indirect Micro Hemagglutination Test for Amebiasis

Human "O" cells were fixed with pyruvic aldehyde, treated with tannic acid, and fixed with glutaraldehyde. The cells were sensitized with amoeba antigen and stored in a refrigerator. The sensitized cells were used periodically for the indirect hemagglutination test with a battery of sera from patients with intestinal amebiasis and confirmed and unconfirmed amebic liver abscess, and also from negative controls. The same battery was tested with cells sensitized with different batches of antigen and also with fresh sheep cells. None of the cells showed any reaction with negative control sera. The fixed cells remained sensitive and stable throughout the study. Reproducibility of the titers with the fixed cells within each day and from day to day was satisfactory. The titers with fixed human "O" cells were slightly lower than were the titers with fresh sheep cells. The advantages of using stable, sensitized cells are pointed out.     

Malignant lymphoma of follicle centre cells with marked nuclear lobation

Four cases of malignant B-cell lymphoma characterized by a conspicuous component of tumour cells with markedly lobatated nuclei are described. Two exhibited a follicular and two a diffuse growth pattern. The tumour cell population formed a continuous spectrum comprising both cells resembling normal follicle centre cells and multilobated lymphoma cells. Cytomorphological analysis of the multilobated cell group indicated a differentiation series from centroblast-like cells with moderately lobated nuclei to large and medium-sized cells with marked nuclear lobation which revealed features of centrocytes. In three cases (1, 3, and 4) the majority of these multilobated cells showed plasmacytoid differentiation in their cytoplasm in conjunction with the synthesis of monotypical cytoplasmic immunoglobulin. No plasmacytoid features were present in a fourth case (2). In only one case (4) monotypical surface immunoglobulin was detectable on the tumour cells. A close relationship between the multilobated tumour cells and follicle centre cells was further substantiated by the finding of a similar cell variant in the follicle centres of a control group of non-neoplastic lymph nodes. It included cells with plasmacytoid differentiation which synthesized polytypical immunoglobulin. We consider this type of B-cell lymphoma with a conspicuous component of cells with lobated nuclei as a variant of malignant lymphoma, centroblastic/centrocytic.     

Vaccination with vascular progenitor cells derived from induced pluripotent stem cells elicits antitumor immunity targeting vascular and tumor cells

Vaccination of BALB/c mice with dendritic cells (DCs) loaded with the lysate of induced vascular progenitor (iVP) cells derived from murine-induced pluripotent stem (iPS) cells significantly suppressed the tumor of CMS-4 fibrosarcomas and prolonged the survival of CMS-4-inoculated mice. This prophylactic antitumor activity was more potent than that of immunization with DCs loaded with iPS cells or CMS-4 tumor cells. Tumors developed slowly in mice vaccinated with DCs loaded with iVP cells (DC/iVP) and exhibited a limited vascular bed. Immunohistochemistry and a tomato-lectin perfusion study demonstrated that the tumors that developed in the iVP-immunized mice showed a marked decrease in tumor vasculature. Immunization with DC/iVP induced a potent suppressive effect on vascular-rich CMS-4 tumors, a weaker effect on BNL tumors with moderate vasculature, and nearly no effect on C26 tumors with poor vasculature. Treatment of DC/iVP-immunized mice with a monoclonal antibody against CD4 or CD8, but not anti-asialo GM1, inhibited the antitumor activity. CD8⁺ T cells from DC/iVP-vaccinated mice showed significant cytotoxic activity against murine endothelial cells and CMS-4 cells, whereas CD8⁺ T cells from DC/iPS-vaccinated mice did not. DNA microarray analysis showed that the products of 29 vasculature-associated genes shared between genes upregulated by differentiation from iPS cells into iVP cells and genes shared by iVP cells and isolated Flk-1⁺ vascular cells in CMS-4 tumor tissue might be possible targets in the immune response. These results suggest that iVP cells from iPS cells could be used as a cancer vaccine targeting tumor vascular cells and tumor cells.     

Antigenic characterization of a T-CLL with heteroantisera and monoclonal antibodies: evidence for the T cell lineage of an Ia-positive, Fc-IgG--positive, suppressor-cell subpopulation

In a previous report, peripheral blood mononuclear T cells from a patient with T-chronic lymphocytic leukemia (T-CLL) were shown to bear receptors for the Fc portion of IgG (T gamma). Moreover, the ability of these cells to rosette with sheep erythrocytes was strongly inhibited by a preincubation of the cells with theophylline. These data indicated that they represent a highly purified subpopulation of Fc-IgG receptor-positive, low-affinity rosetting cells with in vitro suppressor activity on lectin-induced proliferation of normal lymphocytes. They also were reactive in antibody-dependent cell-mediated cytotoxicity but had no reactivity in natural killer cell assays. These cells were studied in this report with several heteroantisera and monoclonal antibodies. Results indicate that these T-CLL cells express a T cell antigenic pattern (OKT-3+) and the majority are Ia positive. They also react with the OKT-8 reagent (a reagent detecting the subset of T cells that contains the cytotoxic/suppressor cells), whereas they are negative with OKT-4 (which reacts with the subset of T cells that contains helper cells) and OKT-6 (thymocyte) antibodies. Heteroantisera also support the results obtained with monoclonal reagents. Despite some recent evidence showing that a high percentage of T gamma cells may belong to the monocyte-myeloid lineage, these T-CLL cells were negative with OKM-1, a monoclonal antibody reported to detect a monomyeloid antigen. These results suggest that a distinct subpopulation of suppressor T cells can be identified by membrane-marker phenotyping.     

In vitro interactions between Sertoli cells and steroidogenic cells

Both the cell and the species specificities of the steroidogenic potentiating activity (SPA) of Sertoli cells on Leydig cells were studied using a coculture system. Coculture of purified pig Leydig cells with rat or pig Sertoli cells in the presence of FSH led in both cases, to a significant increase in hCG receptor number and in hCG-stimulated testosterone production. Similarly, coculture of bovine adrenal cells with rat or pig Sertoli cells enhanced the steroidogenic response of adrenal cells to ACTH and angiotensin II. Such effects were not observed when pig Leydig cells or bovine adrenal cells were cocultured with bovine aortic endothelial cells. Coculture of Sertoli and Leydig cells in the presence of hCG, resulted in a significant increase in FSH receptor number and in FSH-induced plasminogen activator activity. Such effects did not occur when Sertoli cells were cocultured with either adrenal or aortic endothelial cells.     

Selective adhesion of embryonal carcinoma cells and differentiated cells by Ca2+-dependent sites

The specificity of adhesion between embryonal carcinoma cells and fibroblastic cells of various origins was studied. Embryonal carcinoma cells have intercellular adhesion sites requiring Ca²⁺ (CDS). These sites were found to be sensitive to proteases but resistant to them in the presence of Ca²⁺. CDS with a similar protease sensitivity is present in fibroblastic cells. When embryonal carcinoma cells of different lines were mixed, they adhered to each other nonselectively by CDS. Nonselective adhesion by CDS occurred also between fibroblastic cells of various lines. When embryonal carcinoma and fibroblastic cells were mixed, they preferentially adhered to homotypic cells. Fab fragments of antibodies raised against F9 cells (a nullipotent line of embryonal carcinoma) inhibited the adhesion between embryonal carcinoma cells but not between fibroblastic cells. This inhibitory activity of Fab was absorbed with embryonal carcinoma cells with CDS, but not with fibroblastic cells with CDS or embryonal carcinoma cells from which CDS was experimentally removed. SDS-polyacrylamide gel electrophoresis of radioiodinated cell surface proteins showed that the presence of a 140K-dalton component correlated with the presence of CDS in embryonal carcinoma cells, while the presence of a 150K-dalton component correlated with the presence of CDS in fibroblastic cells. These results suggest that CDS in embryonal carcinoma and fibroblastic cells comprise distinct molecules.     

Human cytokine-induced killer cells have enhanced in vitro cytolytic activity via non-viral interleukin-2 gene transfer

Modulation of the immune system by genetically modified immunological effector cells is of potential therapeutic value in the treatment of malignancies. Interleukin-2 (IL-2) is a crucial cytokine which induces potent antitumor response. Cytokine-induced killer cells (CIK) have been described as highly efficient cytotoxic effector cells capable of lysing tumor cell targets and are capable of recognizing these cells in a non-MHC restricted fashion. Dendritic cells (DC) are the major antigen presenting cells. This study evaluated the antitumor effect of CIK cells which were non-virally transfected with IL-2 and co-cultured with pulsed and unpulsed DC. Human CIK cells generated from peripheral blood were transfected in vitro with plasmid encoding for the human IL-2. Transfection involved a combination of electrical parameters and a specific solution to deliver plasmid directly to the cell nucleus by using the Nucleofector(R) electroporation system. Nucleofection resulted in the production of IL-2 with a mean of 478.5 pg/106 cells (range of 107.6-1079.3 pg /106 cells/24 h) compared to mock transfected CIK cells (31 pg/106 cells) (P = 0.05). After co-culturing with DC their functional ability was assessed in vitro by a cytotoxicity assay. On comparison with non-transfected CIK cells co-cultured with DCs (36.5 +/- 5.3 %), transfected CIK cells co-cultured with DC had a significantly higher lytic activity of 58.5 +/- 3.2% (P = 0.03) against Dan G cells, a human pancreatic carcinoma cell line.     

SV40 T antigen and telomerase are required to obtain immortalized human adult bone cells without loss of the differentiated phenotype.

In most human primary bone cells, SV40 T-antigen expression was able to expand life span for a few passages before cells undergo growth arrest, described as crisis. In this study, telomerase activity was reconstituted in human osteoblast precursors (hPOB cells) and marrow stromal cells (Saka cells) transformed with the SV40 T antigen. Bone cells with telomerase activity were able to bypass crisis and show unlimited life span. Despite chromosomal aberrations observed in hPOB-tert cells, these immortalized precursors were able to differentiate into osteoblasts like precrisis hPOB cells. Saka-tert cells enhanced the formation of human osteoclast-like cells in a similar manner as Saka cells. These results demonstrate that reconstitution of telomerase activity in transformed SV40 T-antigen human osteoblast precursors or marrow stromal cells leads to the generation of immortalized cells with a preserved phenotype.     

Phosphatidylserine-dependent adhesion of T cells to endothelial cells

Phosphatidylserine (PS) was exposed at the surface of human umbilical vein endothelial cells (HUVECs) and cultured cell lines by agonists that increase cytosolic Ca(2+), and factors governing the adhesion of T cells to the treated cells were investigated. Thrombin, ionophore A23187 and the Ca(2+)-ATPase inhibitor 2, 5-di-tert-butyl-1,4-benzohydroquinone each induced a PS-dependent adhesion of Jurkat T cells. A23187, which was the most effective agonist in releasing PS-bearing microvesicles, was the least effective in inducing the PS-dependent adhesion of Jurkat cells. Treatment of ECV304 and EA.hy926 cells with EGTA, followed by a return to normal medium, resulted in an influx of Ca(2+) and an increase in adhering Jurkat cells. Oxidised low-density lipoprotein induced a procoagulant response in cultured ECV304 cells and increased the number of adhering Jurkat cells, but adhesion was not inhibited by pretreating ECV304 cells with annexin V. PS was not significantly exposed on untreated Jurkat cells, as determined by flow cytometry with annexin V-FITC. However, after adhesion to thrombin-treated ECV304 cells for 10 min followed by detachment in 1 mM EDTA, there was a marked exposure of PS on the Jurkat cells. Binding of annexin V-FITC to the detached cells was inhibited by pretreating them with unlabelled annexin V. Contact with thrombin-treated ECV304 cells thus induced the exposure of PS on Jurkat cells and, as Jurkat cells were unable to adhere to thrombin-treated ECV304 cells in the presence of EGTA, the adhesion of the two cell types may involve a Ca(2+) bridge between PS on both cell surfaces. The number of T cells from normal, human peripheral blood that adhered to ECV304 cells was not increased by treating the latter with thrombin. However, findings made with several T cell lines were generally, but not completely, consistent with the possibility that adhesion to surface PS on endothelial cells may be a feature of T cells that express both CD4(+) and CD8(+) antigens. Possible implications for PS-dependent adhesion of T cells to endothelial cells in metastasis, and early in atherogenesis, are discussed.     

The ontogeny of the vascular cambium inGinkgo biloba roots

Observation was made on early ontogeny of vascular cambium in the developing root ofGinkgo biloba L. After completion of root elongation, the vascular meristem gradually acquires cambial characteristics. Strips of the periclinal division of cells in transverse section are observed on the inner side of phloem when the primary xylem and phloem in the stele have been established. The strips are united into a continuous layer between phloem and xylem. In tangenital section, the procambium shows a homogeneous structure, which is initially composed of short cells with transverse end walls and subsequently, of long cells with tapering ends. Then, the procambium is organized into two systems of cells; axial strands of short cells with transverse end walls resulting from the sporadic transverse divisions of long cells, and long cells with tapering ends. Still later, the short cells are divided frequently in a trasverse plane exhibiting one or a few cells in width and several decades of cells in height, while the long cells are elongated. The frequency of transverse divisions of the short cells decreases in subsequent stages. Eventually, the short cells in axial strands are vertically separated from one another by the elongation of neighboring long cells and by the decrease in the frequency of transverse divisions of short cells themselves. Cambial initials occur in two forms; ray initials a few cells in height and one cell in width derived from the short cells, and fusiform initials with tapering ends derived from the long cells.     

Prolactin and growth hormone cells in the human hypophysis: A study with immunoenzyme histochemistry and differential staining

Growth hormone and prolactin cells were immunostained in human hypophyses with antibody against rat growth hormone or prolactin and the peroxidase-antiperoxidase complex. Growth hormone cells were round and, in normal pituitaries, arranged in sizable groups. Prolactin cells occurred singly and were less numerous; they were often extensively branched. Only a few prolactin cells stained with carmoisine. Incubation of the antibody with an excess of the appropriate antigen greatly diminished or abolished immunostaining; absorption of anti-prolactin with growth hormone often enhanced it. Prolactin cells were somewhat hypertrophied and hyperplastic in a neonate. Many of them stained with carmoisine. An even greater hypertrophy and hyperplasia of these cells (which pushed apart the growth hormone cells) was found in a lactating woman. Immunostained giant prolactin cells were also observed. Staining of the prolactin cells with carmoisine was extensive. Upon prolonged exposure to anti-growth hormone antibody, ACTH/MSH cells also showed immunostaining which was abolished by absorption of the antiserum with growth hormone but not with synthetic 1-24ACTH. Growth hormone cells evidently correspond to the alpha acidophils of Romeis, prolactin cells in lactation to his eta cells; the relation of his epsilon cells to the pleomorphic "resting" prolactin cells is not clear.     

Paracrine role of Sertoli cells

Data from several experimental approaches strongly suggest that Sertoli cells exert a paracrine control of the two main testicular functions, androgen secretion and spermatogenesis. Further evidence supporting this role of Sertoli cells was obtained by coculture of Sertoli cells with other testicular cells. Coculture of pig or rat Sertoli cells with pig Leydig cells produces an increase in the hCG receptor number and an increase in the steroidogenic activity of Leydig cells. Pretreatment with FSH further increases the values of these two parameters. These biochemical changes were associated with ultrastructural changes in Leydig cells. The effects of Sertoli cells on Leydig cells depend upon the ratio of the two cells and on the substrate in which the cells are cultured. Moreover, Leydig cells produce an increase in the FSH receptor number and in the FSH stimulation of plasminogen activator production by Sertoli cells. Coculture of rat or pig Sertoli cells with rat germ cells, induces an increase in the RNA and DNA biosynthetic activities of germ cells. Most of the stimulatory effects seemed to be mediated by diffusible factors, secreted by Sertoli cells, but full expression of the stimulatory action was observed when germ cells were in contact with other cells. In this coculture system, a fraction of rat germ cells containing mainly mature forms of spermatocytes inhibited rat Sertoli cell RNA and DNA synthesis, but had no effect on pig Sertoli cells. On the contrary, a fraction of rat germ cells richer in spermatogonias and preleptotene spermatocytes, stimulated rat Sertoli cell DNA synthesis but was without effect on pig Sertoli cells. These results clearly show that the stimulatory effects of Sertoli cells on Leydig and on germ cells which are not species specific are mediated mainly by diffusible factors, the secretion of which is regulates by FSH.     

Role of the host cell in persistent viral infection: Coevolution of L cells and reovirus during persistent infection

Mutant L cells, designated LR cells, were isolated after “curing” a persistently infected cell line (L/C) with antireovirus serum. The LR cells were shown to be virus-free; no reovirus was detectable by infectious center assays, plaque assays, presence of viral proteins, presence of viral dsRNA and immunofluorescence studies. Persistent infections were readily established in LR cells following infection with either cloned, low passage wild-type reovirus or cloned, low passage reovirus isolated from carrier cultures. Reovirus isolated from carrier cultures, however, grew much better than wild-type reovirus in LR cells and showed complete dominance over wild-type reovirus in coinfection experiments. Infection of LR cells with wild-type reovirus resulted in a low-level persistent infection with inefficient viral replication; these mutant L cells were partially resistant to infection with wild-type reovirus. In contrast, infection of the mutant L cells with virus isolated from the persistently infected cells resulted in a persistent infection accompanied with efficient viral replication. Infection of the original L cells with either wild-type reovirus or reovirus isolated from the persistently infected cells resulted in a lytic infection with no surviving cells. Thus the host cell plays a crucial role in the maintenance of persistent reovirus infection. Our results show that there is a coevolution of both mutant L cells and mutant reovirus during persistent infection.     

Histochemical and ultrastructural identification of neurotensin cells in the dog ileum

Summary In the dog ileum, neurotensin cells stained with immunofluorescence or immunoperoxidase proved distinct from argentaffin (EC) cells, glucagon immunoreactive (GLI) cells and pancreatic peptide immunoreactive (PP) cells. Neurotensin cells showed various degrees of reactivity with Grimelius'silver. With electron microscopy, besides EC cells, large granule cells with a thin peripheral rim of Grimelius-reactivity (L cells) and large granule cells with variable Grimelius-reactivity of the core (N cells) were found. On distributive grounds, L cells were identified with GLI cells and N cells were interpreted as neurotensin cells.