Role of human chromosome 11 in determining surface antigenic phenotype of normal and malignant cells. Somatic cell genetic analysis of eight antigens, including putative human Thy-1.

The expression of eight serologically and biochemically distinct human cell surface antigens defined by monoclonal antibodies was examined on a panel of rodent-human somatic cell hybrids. Cosegregation was observed for human chromosome 11, and surface expression of all eight antigens was studied. Serological analysis of hybrids containing defined segments of human chromosome 11 permitted the regional assignment of genes controlling antigens JF23 (90 kD glycoprotein), G344 (25 kD), T43 (85 kD), A124, and NP13 to chromosome 11pter-q13, and of genes controlling Q14 (130 kD), MC139 (35 kD), and K117 (25 kD) to chromosome 11q13-qter. K117, the putative human Thy-1 antigen, was expressed at high levels in chromosome 11-containing hybrids constructed with mouse neuroblastoma cells, but showed little or no expression in hybrids constructed with mouse L cells. A similar pattern of expression in hybrids was found for MC139, an antigen shared by neuroectoderm-derived cells and normal and malignant T lymphocytes. T43 is a marker of malignant tumors (but not benign tumors) derived from a number of T43- epithelia, and the regional assignment of the T43 locus on chromosome 11 raises questions about its possible involvement in the specific rearrangements of this chromosome seen in human malignancies.

Expression of epidermal growth factor receptor in human cultured cells and tissues: relationship to cell lineage and stage of differentiation.

Mouse monoclonal antibodies have been used to study the distribution of the epidermal growth factor receptor in human cultured cells and tissues. As expected, most epithelial cells expressed epidermal growth factor receptor (EGFr), whereas cells of hematopoietic origin were EGFr-. However, EGFr was found to be differentially expressed on cultured cells of neuroectodermal origin. Normal melanocytes and a proportion of melanomas are EGFr-, whereas a distinct subset of other melanomas, astrocytomas, and neuroblastomas is EGFr+. Expression of the EGFr in melanomas was closely related to the expression of phenotypic traits of differentiation. Less differentiated melanomas have an epithelioid morphology and are nonpigmented, Ia+, and EGFr+; in contrast, more differentiated melanomas have a dendritic morphology and are pigmented, Ia-, and EGFr-. In the melanoma cell panel used, expression of EGFr did not correlate with rate of proliferation. Expression of EGFr in tissues also showed a lack of correlation with the proliferative state of cells. Our findings indicate that expression of EGFr is related to cell lineage and specific stages of cellular differentiation, rather than only to cell proliferation. The data suggest that receptor content may be elevated in a large number of tumor cell lines.

Coordinate changes in neuronal phenotype and surface antigen expression in human neuroblastoma cell variants.

Human neuroblastoma cells growing in culture offer a unique opportunity to study proliferating human cells with a neuronal phenotype. We have previously identified several neuroblastoma cell lines which show spontaneous conversion (N/S interconversion) between two morphologically distinct cell types: neuroblastic (N-type) cells and variant, substrate-adherent (S-type) cells resembling cultured glial or mesenchymal cells. In the present study, we have used molecular markers to confirm the neuronal phenotype of N-type cells and to demonstrate that S-type cells have a nonneuronal phenotype. Furthermore, we have used these markers, including a series of cell surface differentiation antigens, to compare S-type neuroblastoma cells with a wide range of cultured epithelial, mesenchymal, and neuroectodermal cells. The results suggest that N/S interconversion represents an ordered transition between two neuroectodermal differentiation programs rather than random phenotypic instability of cultured cells; S-type variant cells show a molecular phenotype most closely resembling the phenotype of cultured ectomesenchymal cells; and in vitro variant formation of human neuroblastomas may provide an experimental model for the observed in vivo transition of some malignant neuroblastomas into benign ganglioneuromas.

Differential expression of cell surface antigens and glial fibrillary acidic protein in human astrocytoma subsets.

We have characterized five distinct cell surface antigens of human astrocytomas and correlated their expression with the expression of glial fibrillary acidic protein (GFAP) and four previously defined cell surface markers of astrocytomas. One of the newly studied antigens, A4, which was originally detected on rat central nervous system (but not peripheral nervous system) neurons, is expressed on GFAP+ human astrocytoma cells, but not on GFAP- astrocytomas or a wide range of other neuroectodermal, epithelial, and hematopoietic cells. Antigens F19 (Mr 140,000/90,000 glycoprotein) and F24 (Mr 90,000 glycoprotein) also show restricted distribution and are expressed on subsets of neuroectodermal and mesenchymal cells. Antigens G253 (Mr 95,000 glycoprotein) and S5 (Mr 120,000 glycoprotein) are more widely distributed on the cultured cell panel. The distribution of these antigens was determined on a series of 22 astrocytoma cell lines and in normal brain tissue and the results were compared with the distribution of 5 additional glial cell markers: GFAP and cell surface antigens A010 (Mr 110,000 glycoprotein); AJ8 (Mr 100,000 glycoprotein); LK26 (Mr 35,000 glycoprotein); and Thy-1. Distinct patterns of expression on cultured astrocytomas and in neural tissues were identified for all antigenic systems studied, and cell surface expression of antigen A4 was found to correlate closely with GFAP phenotype of cultured astrocytomas. The antigens described in this study provide new markers to study normal glial differentiation and to correlate the phenotypes and biological behavior of distinct subsets of astrocytomas.

Decrease of pancreatic somatostatin in neonatal nesidioblastosis.

The inappropriate insulin release that is characteristic of severe neonatal hypoglycemia and nesidioblastosis was thought to be principally due to a marked proliferation of B-cells. A possible deficiency of somatostatin, one of the factors controlling insulin release, has only recently been considered. We report have a significant decrease of pancreatic somatostatin cells and somatostatin content in nesidioblastosis, as compared with appropriate controls. Pancreatic tissue from five babies with severe hypoglycemia, hyperinsulinemia, and nesidioblastosis was examined for insulin, somatostatin, and glucagon by immunocytochemistry and radioimmunoassay. There was a variable insulin cell hyperplasia among the nesidioblastotic specimens but no significant difference from controls was detected. In contrast, there was a consistent and highly significant (P less than 0.02) decrease (of more than 50%) of somatostatin cells (controls, 1.29 +/- 0.22%; nesidioblastosis, 0.53 +/- 0.14%, mean +/- SEM). Similarly, there was no significant alteration in insulin content, although a slight increase was found in non-microadenomatous areas of the diseased pancreata (controls, 42.3 +/- 4.1; nesidioblastosis, 58.6 +/- 9.4 nmol/g wet weight of tissue, mean +/- SEM). However, somatostatin content was almost 60% below control values (controls, 0.365 +/- 0.038; nesidioblastosis, 0.16 +/- 0.039 nmol/g), a statistically significant difference (P less than 0.01). Thus, a marked reduction in the content of somatostatin was present in the pancreata of these infants with nesidioblastosis, resulting in a distinct alteration of the normal pancreatic hormone balance.

Expression of cytokeratins in normal and neoplastic colonic epithelial cells. Implications for cellular differentiation and carcinogenesis.

Cells of the normal colonic mucosa express several types of cytokeratins, the characteristic intermediate filament proteins of epithelial cells. An immunohistochemical study was designed to examine the expression of two distinct groups of cytokeratins, recognized by monoclonal antibodies AE1 and AE3, in the colonic mucosa and to compare the findings with those obtained with a large number of polypoid lesions (adenomatous and hyperplastic) and carcinomas of the colon. AE1 and AE3 immunostaining was found in the surface epithelium and upper portions of the crypts of Lieberkühn (functional zone) of normal colonic mucosa, whereas the lower portions of the crypts (proliferative compartment) were unreactive with both AE1 and AE3. Polypoid lesions of the colonic mucosa can be placed into two categories based on their patterns of cytokeratin expression. Solitary tubular adenomas and hyperplastic polyps are composed of AE1 and AE3 nonexpressing cells with only few, patchy areas of AE1 and AE3 expressing cells present within glands and in the surface epithelium. In contrast, villous adenomas show strong AE1 and AE3 reactivity throughout the glands. Furthermore, tubular and villous adenomas, and even histologically normal mucosa in patients with familial polyposis, show AE1/AE3 expression throughout the glands and surface epithelium. Colonic carcinomas show a predominance of AE1/AE3 expressing cells. Thus, cytokeratins recognized by monoclonal antibodies AE1 and AE3 represent molecular markers of cellular maturation in the normal colonic mucosa, that are expressed in colonic carcinomas and, in addition, serve as markers that distinguish colonic mucosa and adenomas with a high risk for development of cancers from those with a lower risk.

Immunohistochemical analysis of nerve growth factor receptor expression in normal and malignant human tissues.

Nerve growth factor (NGF) is a polypeptide important for normal development of the nervous system and promotion of survival and differentiation of sensory and sympathetic neurons in culture. The cellular effects of NGF are mediated by a specific cell surface molecule, nerve growth factor receptor (NGF-R). In the present study we have used a monoclonal antibody against human NGF-R to examine, by the avidin-biotin-immunoperoxidase method, the receptor distribution in a wide range of normal tissues and in more than 200 malignant tumors. Our results show that (a) human NGF-R is expressed in the peripheral nervous system but not in any of the central nervous system areas tested; (b) NGF-R expression is not restricted to neural tissues but is also found in a number of normal epithelial, mesenchymal, and lymphoid tissues; (c) NGF-R expression changes during normal development; and (d) NGF-R expression in malignant tumors generally parallels its normal tissue distribution. Thus, NGF-R is detected in a proportion of neuroectoderm-derived tumors, carcinomas, and lymphomas, and also in a characteristic group of small round-cell tumors (Ewing's sarcomas and embryonal rhabdomyosarcomas). These findings suggest a normal regulatory role for NGF in both neuronal and non-neuronal cells and identify a range of human tumors in which the NGF/NGF-R system may contribute to the malignant phenotype.

Pinocchio cells: morphologically atypical immunologically heterogeneous lymphocytes induced by treatment with interleukin 2.

We describe a novel cell type, the Pinocchio cell, that appears in the peripheral blood of all patients receiving treatment with interleukin 2, up to 20,000 cells/microliter. This cell is characterized by a prominent and granular proboscis with which it attaches to tumor cells and mediates tumor cell lysis. Pinocchio cells are immunologically heterogeneous and express antigens of both T and NK cells; Pinocchio cells are adherent in culture and are more cytolytic than non-adherent cells against NK-sensitive and resistant tumor cells. Incubation of normal whole human blood for 1 h induces Pinocchio morphology of mononuclear white blood cells.

Definition of an extracellular matrix protein in rostral portions of the human central nervous system.

We have identified and characterized an extracellular matrix (ECM) glycoprotein of cultured astrocytomas, NEC1, that is expressed in normal human brain parenchyma. Detailed immunohistochemical analysis reveals a region-specific NEC1 pattern along the rostrocaudal axis of the central nervous system (CNS), with strong expression throughout the white matter of telencephalon and diencephalon, scant expression in some areas of mesencephalon, and no expression in pons, cerebellum, medulla, spinal cord, and peripheral nervous system. NEC1 is not co-distributed with any known neural cell type, suggesting that expression of specific ECM proteins in the CNS is segmentally controlled.

Expression of p21ras in normal and malignant human tissues: lack of association with proliferation and malignancy.

Proteins encoded by cellular ras oncogenes (p21ras) are expressed in a wide variety of malignant tumors, including carcinomas, lymphomas, and neuroectodermal tumors. The function of p21ras in these tumors and the distribution and role of p21ras in corresponding normal tissues are unclear. This immunohistochemical study examined the relationship between p21ras expression and malignant transformation, cellular differentiation, and proliferative activity in vivo. p21ras was found to be widely expressed in normal tissues, but within those tissues expression was often sharply restricted to cells at specific stages of differentiation; terminally differentiated cells generally showed stronger reactivity with antibodies to p21ras than did rapidly proliferating cells. Fetal and adult tissues had corresponding patterns of p21ras expression, and the distribution of p21ras in neoplasms paralleled the pattern in normal tissue from which they were derived. Thus, p21ras seems to play a role in many fully differentiated cell types, and levels of p21ras expression do not correlate with proliferative activity in normal cell or, in contrast to past reports, with the transformed phenotype.

Expression of human nerve growth factor receptor on cells derived from all three germ layers.

Nerve growth factor (NGF) is a protein which promotes the survival and differentiation of neuronal cells in vitro and plays an important role in neuronal development. In this study, we have examined the expression of the receptor for NGF (NGFR) in human neuronal and nonneuronal cells, both in tissue culture and in vivo. In addition to cell lines derived from neuroblastoma, astrocytoma, and melanoma, all of which share a common neuroectodermal origin, NGFR was detected in a number of cultured cells of mesenchymal, epithelial, and hematopoietic derivation. Immunohistochemical analysis showed that NGFR is expressed in several nonneural human tissues, and the cell types in which NGFR was found include derivatives from all three germ layers. Thus, our findings demonstrate that NGFR is much more widely expressed in human cells and tissues than was previously thought.

Cell surface antigen of human neuroblastomas is related to nuclear antigen of normal cells.

The localization of MC25, an antigen first detected on the surface of human neuroblastoma cells, was determined in cultured cells and tissues. Neuroblastoma cell lines (15/17) express the antigen on the surface and in the cytoplasm (scMC25+), whereas 156/160 cell lines derived from other normal and malignant human cell types are scMC25-. However, MC25 is found in the nucleus of scMC25- cells (nMC25+), presenting a discrete granular pattern. In scMC25+/nMC25- neuroblastoma lines, apparent antigen shifting from the cell surface/cytoplasm to the nucleus accompanies variant formation, which represents a transition in the neuronal differentiation program of these cells. Results of immunohistochemical studies with human tissues parallel the findings with cultured cells. Almost all cell types are scMC25-/nMC25+; basal cells of the epidermis are the only cells constitutively expressing cMC25; and a population of neurons are the only scMC25-/nMC25- cells. Alternative localization of MC25 to different cellular compartments and antigen shifting are reminiscent of the behavior of certain developmentally regulated antigens in Drosophila and Xenopus.

Regulation of Thy-1 gene expression in transgenic mice.

Genomic DNA fragments encompassing the human Thy-1 or mouse Thy-1.1 gene have been microinjected into pronuclei of mouse embryos homozygous for the Thy-1.2 allele. In the resulting transgenic mice, the human gene is expressed in a pattern characteristic of normal human tissues, and is not influenced by the pattern of endogenous mouse Thy-1 expression. The mouse Thy-1.1 gene fragment is expressed in a pattern typical of mouse Thy-1, although it is more limited in its distribution. The results indicate the presence of multiple cis-acting regulators of Thy-1 gene expression that have changed in both their character and arrangement over the course of Thy-1 gene evolution.