Biomedical Reviews 1999; 10:55-68.

Dedicated to Rita Levi-Montalcini

©The Bulgarian-American Center, Varna, Bulgaria

ISSN1310-392X

EXPRESSION OFTHETRKNEUROTROPH1N RECEPTORS IN NONNEUROWAL TISSUES

AND CANCER

Hirotaka Koizumi

Department of Pathology, St Marianna University School of Medicine, Kawasaki, Japan

In this article, detailed immunolocalization of the expression of receptor tyrosine kinase (Trk) in a wide range of normal and cancer

nonneuronal tissues is described. Trk immunoreactivity is detected at various levels in all tissues examined, except for the heart and

liver. The gastric parietal cells reveal strong TrkA and TrkC expression and all of the Trk immunoreactivities are expressed in the

putative intestinal neuroendocrine cells. In the pancreas, TrkA and TrkC are detected in the subintralobular ducts, while TrkB is

found specifically in the pancreatic beta cells. The lymph node and spleen exhibit TrkB in monocytes/macrophages. The adrenal

cortex show selective TrkA with TrkC in the medulla. In the reproductive system, TrkA immunoreactivity is detected in the prostatic

epithelial cells, TrkC in the ovarian theca and granulosa cells, TrkA and TrkC in the secretory-phase endometrium, and TrkA in the

mammary ducts. The kidney reveal strong TrkA and TrkC in its tubules, but no Trk receptors are present in the glomeruli. In the

skin, TrkA and TrkB/TrkC are expressed in the basal and granular layers of the epidermis, respectively. The expression patterns

of TrkA in 337 nonneuronal invasive carcinomas of 15 different human tissues are also demonstrated iminunohistochemically.

Overall, 133 (39%), 101(30%) and 103 (31%) tumors exhibited strong, moderate and no TrkA immunoreactivity, respectively.

Esophageal and thyroid carcinomas expressed high levels of TrkA, whereas the levels in gastric and colon cancers are low. TrkA

expression is detected not only in carcinomas originating from TrkA-positive normal counterpart tissues, including the esophagus,

breast, lung and uterus, but also in those from TrkA-negative cells of the thyroid, liver and ovary. Immunostainingforbeta-NGF, the

specific UgandforTrkA, inesophagealandbreast carcinomas demonstrates its immunoreactivity in stromalfibroblasts and some TrkA-

expressing tumor cells. These results suggest that paracrine and autocrine regulation via stromal/tumoral NGF-tumoral TrkA

interaction may be involved in growth of certain nonneuronal carcinomas. Bioined Rev 1999; 10: 55-68.

INTRODUCTION

The Trk family of receptor tyrosine protein kinase (receptozyme),

TrkA, TrkB, and TrkC, play significant roles in the development

and maintenance of neural tissue by mediating the trophic

effects of the nerve growth factor (NGF) family of neurotrophins

(1-4). NGF specifically recognizes TrkA, a receptor identified in

all major NGF targets, including sympathetic, trigeminal, and

dorsal root ganglia as well as in cholinergic neurons of the basal

forebrain and striatum (5-7). Brain-derivedneurotrophic factor

(BDNF) and neurotrophin-4/5 (NT-4/5) activate the TrkB receptor

(8-11), while a higher concentration of NT-4/5 can also activate

the TrkA receptor (12). TrkB transcripts encoding this receptor

are found throughout the central and peripheral nervous system.

Neurotrophin-3 (NT-3) primarily activates the TrkC whose gene is

also expressed widely in the mammalian neural tissues (13).

Unlike the other neurotrophins, NT-3 appears to be somewhat

nonspecific, since it can activate TrkA and TrkB receptors in

Received 17 October 1999 and accepted 30 November 1999.

Correspondence and reprint requests to Dr Hirotaka Koizumi. Department of Pathology. StMariannaUniversity School of Medicine,

Miyamae,Kawasaki216-8511, Japan. Tel: 81449778111, Fax: 81449777827,E-mail:hiro@marianna-u.ac.jp

certain cell systems (1). When ectopically expressed in fibro-

blasts, activation of the signal transduction cascade initiated by

each of the Trk receptors results in a similar biological

consequence (10,14). Thus, it would appear likely that the

neuro-trophin requirements of distinct neuronal populations

are dictated by highly specific regulation of Trkreceptor

expression. Recently, we and others have demonstrated

immunocy-tologically or biochemically that various

nonneuronal tissues also express Trks (15-17). We carried out

immunostaining analysis for Trks in 24 types of normal

nonneuronal tissue from adult humans and found that, except for

heart, thyroid and liver, they all expressed these receptors (15),

partly consistent with the results of RNAse protection assay

in rat tissues (17). Moreover, our results demonstrated highly

specific Trk expression by distinct nonneuronal cell populations:

for instance, esophageal and epidermal basal cells selectively

expressed TrkA, gastric parietal cells TrkA and TrkC, pancreatic

beta cell TrkB, adrenal cortical and medullary cell TrkA and

TrkC, respectively, prostatic epithelial cell TrkA and

monocytes/ macrophages of the lymph node and spleen TrkB.

As some of these tissues are known to produce the

corresponding neurotrophins, e.g., the skin (NGF) (18,19),

adrenal glands (NGF andNT-3) (20,21), prostate (NGF) (22,23),

andpancreas, lymph nodes and spleen (BDNF) (16,24), theTrk-

neurotrophin system may be involved in the regulation of specific

functions, such as hormone secretion, of the individual receptor-

expressing cells (15). Furthermore, the fact that Trks are

expressed by a broad spectrum of nonneuronal cells suggests

that they also play roles in phenomena common to such cells,

i.e., cell growth and differentiation, as established for nervous

system (1-4). Indeed, ectopic expression of Trk receptors in

fibroblasts has been shown to result in ligand-mediated cell

growth (10,14), and NGF stimulated nonneuronal cell proliferation

(22,23,25), (26), through TrkA phosphorylation (25). In addition

to the normal tissues, the immunohistochemical expression of TrkA

in over 300 invasive

nonneuronal carcinomas of diverse histologic types is analyzed

(27). TrkA expression is detected not only in carcinomas

originating from TrkA-positi ve normal counterpart tissues, but

also in those from TrkA-negative tissues or cells. These results

suggest that TrkA expression is characteristic of specific subsets of

epithelial malignancies.

In this review, I describe detailed immunolocalization of Trk

receptor family in normal nonneuronal tissues and the expression

profiles of TrkA in nonneuronal carcinomas.

CELLULAR LOCALIZATION OF TRK RECEPTORS IN NORMAL NONNEURONAL TISSUES

Gastrointestinal system

Trk immunostaining is detected in various cell types within the

gastrointestinal system. In the salivary glands, the serous

acinar cells showed TrkB immunoreactivity (IR), whereas the

ductal epithelium stained for both TrkA and TrkC and no Trk IRs

were detected in the mucinous acinar cells (Fig. 1A). In the

esophagus, TrkA immunostaining only was detected in the

basal layers of the stratified squamous epithelium (Fig. 1B,C). In

the stomach, the parietal cells, located in the lower half of the

gastric glands, showed strong IR for TrkA and TrkC, but not for

TrkB (Fig. ID, left). In addition to subepithelial connective

tissue fibroblasts andlymphocytes, no othercellular components of

the gastric mucosa showed Trk immunostaining (Table 1). In the

small intestine and colon, there is little or no staining in the

epithelial cells for Trk proteins, but small cells, located along the

glandular basement membrane show intense immunostaining

for all three Trk receptors (Fig. ID, right). The morphology,

immunoreactivity and distribution of the Trk- positivecells were

consistent with those of the intestinal neuroendocrine cells.

Weak TrkA and TrkC staining was detected in all the muscle

layers and in the ganglion cells present in Meissner's and

Auerbach' s plexuses throughout the gastrointestinal tract (Table

Figure 1. The immunolocalization of Trk receptors in the human nonneuronal tissues. Immunostaining result(s) showing only the

representative type(s) of Trk receptor in each tissue. A. TrkB IR in the serous acinar cells of the salivary glands. B. TrkA

immunoreactivity in the basal layers of the esophageal squamous epithelium. C. No TrkB immunoreactivity in the esophagus. D, left.

Strong TrkA immunoreactivity in the parietal cells of the stomach. D, right. TrkB immuno reactive cells in the small intestine. TrkB

immunoreactivity was detected in small cells present along the glandular basement membrane (arrows). E. Strong TrkB

immunoreactivity in the pancreatic islet. F. Glucagon-immunoreactive beta-islet cells in the serial section. The distribution of

TrkB-positive and glucagon-positive cells was virtually identical. G. TrkB-immunoreactive cells present around the white pulp of

the spleen, which showed a nearly identical distribution pattern to lysozyme-positive monocytes/macrophages in the serial section

(H. left, TrkB; right, lysozyme). I. TrkA immunoreactivity in the adrenal cortex (upper panel) and TrkC in the medulla (lower

panel). J. TrkA immunoreactivity in the prostatic epithelial cells, which was clearly present in luminal secretory cells, but faintly in

the basal cells K. Trk immuno reactivities in the secretory-phase endometrium. K. left. TrkC immunoreactivity at the luminal

surface of the endometrial epithelium. K. right. TrkA immunoreactivity in the glandular epithelium. L. TrkC immunoreactivity

in the mammary duct. Dot-like immunoreactivity was apparent in the luminal epithelium as well as in the intraductal phagocytes.

Very weak immunoreactivity was present in the my o epithelium. M. Strong TrkC immunoreactivity in the collecting tubules of kidney.

The distal tubules showed weaker immunostaining. N. Immunoreactivitiesfor TrkA (upper panel) and TrkB (lower panel) in the skin

epidermis. O. TrkA immunoreactivity in the skeletal muscle. Immunoperoxidase staining.

Biomed Rev 10, 1999

Koizumi 56

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Trk neurotrophin receptors 57

1). No Trk IR is found in the hepatocytes, Kupffer cells, sinusoi-dal cells or bile ductal epithelium of the liver.

In the exocrine pancreas, moderate to strong TrkA and TrkC IRs are present in the intercalated and intralobular ducts, but they were absent from the interlobular and main pancreatic ducts and the acini (15). In the endocrine pancreas, very intense TrkB IR is detected in cells preferentially located around the periphery of the islets (Fig. IE), their localization is consistent with that of glucagon-secreting cells (28). In serial sections, immunolocalization of TrkB is nearly identical with that of glucagon (Fig. IF), but not with those of insulin. No TrkA and TrkC immunostaining is seen in the pancreatic islets.

Cardiopulmonary system

In the lung, low levels of TrkA and TrkC are present in some alveolar cells and in the bronchial gland mucinous cells. No Trk immunoreactivity is present in the cardiac muscles (Table 1).

Hematolymphoid system

Only TrkB is detectable in the lymph node and spleen. The TrkB immunoreactive cells are moderate- to large-sized, cytoplasm rich, and of irregular shape, consistent with the morphological features of monocytes and macrophages. They resided preferentially in the subcapsular and interfollicular sinuses of the node and around the white pulp of the spleen (Fig. 1G). Double immunostaining showed that the nodal TrkB-positive cells are only colabeled with CD 68, the marker for monocytic lineage (29), and not with other markers for hematopoietic cells, such as CDS, CD20, CD45 and immunoglobulin light chains (15). In addition, the splenic TrkB-positive cells show a virtually identical distribution pattern to that of lysozyme-containing monocytes/macrophages on serial sections (Fig. 1H). Like these hematolymphoid tissues, the bone marrow show selective TrkB immunostaining in hematopoietic cells (Table 1). Considering that BDNF and NT-3 are expressed by lymph node and spleen (16), our data indicate that monocytes/macrophages may be the selective targets of BDNF and NT-3 in these tissues. In the thymus, intense TrkA staining was seen in the subcapsular epithelial cells, and the thymocytes themselves stained occasionally for this protein (15). Since the thy mic subcapsular epithelium is a region of known p75 neurotrophin receptor (p75

NTR) localization (30), the coexpression of low- and high-

affinity NGF receptors by these epithelial cells might be related to their well-characterized neuroendocrine phenotype (31) and to their ability to synthesize different thymic hormones (30).

Endocrine system

In endocrine tissues, the thyroid follicular cells reveal no Trk immunostaining, exceptforsomecellsinphysiologicallyatrophic follicles that showed TrkA and TrkC immunostaining (Table 1). TrkA IR was detected throughout the adrenal cortex, although several cells stained quite intensely. Virtually no TrkB or TrkC immunostaining is seen in the cortex. The medulla expresses

only for TrkC IR (Fig. II). The distinct distribution of TrkA and TrkC in the adrenal glands indicates that endogenous NGF and NT-3 (20,21) might be capable of exerting selective regulatory effects on the secretion of cortical and medullary adrenal hormones, respectively. As mentioned above, the pancreatic alpha cells are TrkB-positive.

Reproductive system

In the male reproductive system, selective immunostaining of TrkA is found in the epithelial cells of the prostate; it was clearly present in the luminal secretory cells, but not or only faintly in the basal cells (Fig. 1J). Pflugetal also reported TrkA expression in the human prostate using a pan-Trk antibody in conjunction with Scatchard plot and receptor phosphorylation analyses (25). Since prostatic stromal cells produce NGF that has been shown to bind to prostatic epithelial cells (25), this strongly suggests that the growth of the latter is regulated by a paracrine mechanism. In the testis, the fibrous septa surrounding the seminiferous epithelium also showed TrkA IR (also see this volume of Biomedical Reviews). In the female reproductive system, the ovarian theca and granulosa cells stain for TrkC (15). This implies that ovary-secreted neurotrophins, such as NT-3 (33,34), may be involved in the direct regulation of gonadal endocrine function. The proliferative-phase endometrium completely lack Trk immunostaining, whereas the secretory-phase epithelium expresses strong TrkC IR on its luminal surface. TrkA is also detected during this phase in parts of the glandular epithelium and in the stromal cells (Fig. IK). These results suggest that estrogen may upregulate expression of TrkA and TrkC in the endometrium in the same way as it does for TrkA in PC12 cells in vitro and in rat dorsal root ganglion neurons in vivo (35-37). Since NGF significantly enhances estrogen binding in PC 12 cells (35,37), it is feasible that TrkA, expressed in the secretory-phase endometrium, may function to increase endometrial sensitivity to estrogen. A similar interaction among estrogen, TrkC, and NT-3 might augment the secretory activity of the endometrial epithelium. In the breast, luminal epithelium of the mammary ducts show cytoplasmic, dot-like reactivity for TrkA and TrkC. These immunoreactive dots are also found in the cytoplasm of the intraductal phagocytes. Weak TrkA and TrkC immunostaining is also seen in the ductal myoepithelium (Fig. 1L).

Urinary system

In the kidney, intense TrkA and rather weaker TrkC

immunostaining are found in the epithelium of the collecting

tubules (Fig. 1M). Similar but less intense immunostaining is

also detected in the proximal and distal tubules. In contrast,

none of the Trk IR were present in the cells of the glomeruli (15).

Although there are no published reports describing Trk

expression in the human kidney, a study in fetal mice showed a

similar renal localization of TrkC transcripts using in situ

hybridization (38). As no NGF expression or activity are

58 Koizumi

Biomed Rev 10, 1999

Trk neurotrophin receptors 59

Biomed Rev 10, 1999

- , absent; +, weakly positive; ++, moderately positive; + + +, strongly positive. *TrkA was present in the basal layer of the stratified

squamous epithelium. 'Cells with neuroendocrine cell-like morphology. **TrkC immunostaining was detected at the luminal surfaces

of most of the glandular epithelium and TrkA was found in the cytoplasm of some of them. "TrkA and TrkB/TrkC were present in the

basal and granular layers, respectively, of the epidermis.

Biomed Rev 10, 1999

Koizumi

detected in the kidney (39,40) and NT-3 IR is found in the

distal tubular cells of rat (20), it is possible that NT-3, which

can bind to both TrkA and TrkC (41), may play a role either in

survival of tubular epithelial cells or in the reabsorption

function of the renal tubules.

Musculocutaneous system

The skeletal muscle is stained for TrkA and TrkC, but not for

TrkB (Fig. 1O). In the skin, TrkA and TrkB/TrkC are present in

the basal and upper, or granular, layers of the epidermis,

respectively (Fig. IN). TrkA was also detectable in the hair

follicles and sweat glands. Since the basal layer of the epidermis

also expresses p75NTR (42) and keratinocytes and dermal

fibroblasts are both capable of producing NGF (43,44), it seems

likely that the behavior of human skin epidermis might be

influenced by NGF-mediated autocrine/paracrine regulation via its

low- and high-affinity receptors. Moreover, the observation that

transgenic mice, deficient for TrkA, have numerous scabs over

their entire body and ulcerated paws (45), suggests that the NGF-

TrkA interaction may play crucial roles in regulating both cell

growth and the regeneration of normal epidermis (also see this

volume of Biomedical Reviews).

I have shown highly specific Trk receptor expression by

distinct nonneuronal cell populations. However, I do not yet

know whether the expressed receptors actually function as

signal transducing molecules in these cells. RNase protection

analyses have shown that rat nonneuronal tissues express both

the full-length and truncated TrkA, whereas they only express

the truncated TrkB and TrkC transcripts that appear to encode

noncatalyticreceptorproteins (17,46-48). Although the number of

tissues examined was small, these results imply that only the full-

length TrkA is capable of transmitting the ligand's signal into

nonneuronal cells. This is consistent in part with a study

showing Trk phosphorylation by NGF in prostatic epithelial

cells, indicating that TrkA is a signal transducing molecule in

this nonneuronal tissue (25). However, it is still possible that

truncated receptors may present neurotrophins to neurons, or

may interact with another receptor that is itself the signal-

transducing molecule in these cells. Moreover, expression of

the full-length TrkB transcripts in rat hematolymphoid tissues

has recently been reported (14). A precise understanding of the

functional roles of Trk receptors in nonneuronal tissues awaits

further analyses, including RNase protection in a wide range of

organs/tissues, immunohistochemical studies using antibodies

that recognize the truncated peptides, and in vitro expression

studies of the truncated trk genes in various nonneuronal cells.

EXPRESSION OF TRKA, NGF, AND P75NTR IN HUMAN NONNEURONAL CANCERS

TrkA expression in nonneuronal carcinomas

The TrkA expression patterns of nonneuronal invasive

carcinomas of the esophagus, stomach, breast and uterine

cervix are summarized in the upper panel of the Table 2. Of the

100 esophageal squamous cell carcinomas, 86(86%), 11 (11%)

and 3 (3%) tumors are evaluated as TrkA++, TrkA+ and TrkA-,

respectively (Fig. 2A,B). Virtually all the well (17/18, 94%)

and moderately (27/28, 96%) differentiated esophageal

cancers, but a smaller proportion of the poorly differentiated

tumors (42/54, 77%) are TrkA++. In contrast to this high level

TrkA expression in the esophageal cancers, most of the

gastric adenocarcinomas (38/49, 78%) are TrkA- and this is

largely attributable to no protein expression in all 19 poorly

differentiated tumors. Twenty three (42%) of the 55 breast

ductal carcinomas are classified as TrkA++, 15 (27%) as

TrkA+ and 17 (31%) as TrkA- tumors (Fig. 2C,D). Analysis of

the tumor differentiation grades reveal 59% (16/27) of the

well-differentiated breast cancers are TrkA++, compared

with only 9% (2/22) and none of the moderately and poorly

differentiated tumors, respectively. Of the 37 uterocervical

squamous cell carcinomas, 7 (19%), 18 (49%) and 12 (32%)

are TrkA++, TrkA-t- and TrkA-, respectively. The TrkA

expression levels in these tumors also paralleled their

differentiation grades: approximately 60% each of the well

and poorly differentiated carcinomas are TrkA++ and TrkA-,

respectively. In addition to these tumors, a small series of

invasive carcinomas of the thyroid, lung, liver, pancreas,

colon, ovary and skin, is surveyed for TrkA expression without

considering the tumor differentiation grades (Table 2, lower

panel; Fig. 2E-G). Varying degrees of TrkA IR are detected in all

these cancers, among which the highest protein expression level

is found in the thyroid carcinomas (TrkA++ in 8/10) and the

lowest in the pancreatic and colon cancers (TrkA- in 8/11 and

10/16, respectively). TrkA+ level expression seemed to be

typical in the other organ cancers (lung: 11/22, liver: 8/11,

ovary: 11/16, skin: 4/10). Overall, 133 (39%), 101 (30%) and

103 (31%) of the 337 nonneuronal carcinomas studied are

TrkA++, TrkA+ and TrkA-, respectively.

The TrkA expression patterns could be divided into three

categories. First, TrkA was detected in a proportion of tumors

derived from TrkA-positive normal tissues and, in these cases,

receptor expression may simply reflect the molecular phenotype of

the target cells for transformation. Subsets of the breast

carcinoma and squamous cell carcinomas of the esophagus,

uterine cervix and skin fell into this category (15). It is therefore

rational that the TrkA expression levels in these cancers paralleled

their tumor differentiation grades (Table 2). Second, lOofthe

16 colon carcinomas are TrkA-, despite the fact that their normal

counterpart glandular cells are TrkA-positive (15). It is

conceivable that these tumor cells lost the ability to express

TrkA following malignant transformation. Although the trkA

gene was first isolated from colon carcinoma cells (49), the above

result suggests TrkA is unlikely to be involved in the pathology of

this tumor in vivo. Some, if not all, pancreatic ductal

carcinomas (TrkA- in 8/11) may also belong to this category, as

normal pancreatic subintralobular ducts expressed TrkA (15).

61 Trk neurotrophin receptors

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Koizumi 62

Third, TrkA++ or TrkA+ expression was detected in all the

thyroid carcinomas (10/10: 100%), which may result from the

reported trkA rearrangement in this tumor (50,51), and the

majority of the liver (8/11: 73%) and ovarian (14/16: 88%)

carcinomas. It is significant that we observed TrkA expression in

these tumors, but it was not detected in their counterpart

normal cells, i.e., thyroid follicular cells, hepatocytes, and ovarian

mesothelial cells (15). In such cases, receptor expression may

be related to the transformed phenotype or to increased

proliferative activity, not to the tissue derivation of the tumor

cells.

Expression of NGF and p75NTR in esophageal and breast

carcinomas

Immunostaining for NGF and p75NGFR was carried out in frozen

sections of 7 esophageal and 10 breast invasive carcinomas.

Normal esophageal tissues adjacent to the tumors showed NGF

IR in the epithelial spinocellular cells and submucosal

fibroblasts, whereas they exhibit p75NTR immunostaining

exclusively at the epithelial basal and parabasal layers. No NGF

immunostaining is seen in the luminal epithelium and

myoepithelium of normal mammary glands, although they stain

positively for p75NTR, and the stromal fibroblasts surrounding the

mammary glands show NGF immunoreactivity. These results

are consistent partly with previous reports (19, 52, 53) (Fig.

3C,D for p75NTR). NGF IR is present in the stromal fibroblasts, as

well as in various numbers of the cancer cells in tumor

portions of the esophageal and breast tissues, although none of

these tissues exhibit p75NTR immunostaining (Fig. 3A-D). These

findings are common to all the specimens tested. Altogether, I

suggest that paracrine regulation via the stromal NGF-tumoral

TrkA interaction might mediate certain effect, e.g., cell growth, in

these nonneuronal tumors. This concept has already been

suggested for prostate cancer. Djakiew et al (22,23,25,54)

localized Trk- and NGF-immunoreactive proteins in the epithelial

and stromal cells, respectively, in human prostate tissue, and

showed that NGF or its related stromal protein specifically

bound to the epithelial TrkA, thereby inducing tyrosine

phosphorylation of the receptor and tumor proliferation in a

prostate cancer cell line. Similar findings of NGF-

dependent clonal growth of lung cancer cells (26) and NGF

production by tissue fibroblasts (55,56) are reported recently.

Therefore, our findings extend the types of tumor that express

TrkA and stromal NGF and indicate that the paracrine regulation

model of tumor growth might be applicable to a variety of

nonneuronal human carcinomas. I suggest that the low-affinity

p75NTR is either not involved, or plays a minor role in this

paracrine process, as p75NTR immunoreactivity appears to have

been lost by most of the nonneuronal cancers that expressed

TrkA, such as those of the esophagus, prostate, thyroid, breast

and ovary (25,52,57) (Fig. 3C,D). Moreover, immunostaining of

NGF and TrkA in serial sections reveal that NGF is colocalized in

some, although not all, TrkA-expressing tumor cells (Fig.

3E,F). This result raises a possibility that autocrine regulation

via the tumoral NGF-tumoral TrkA interaction mechanism might

also involve in proliferation of some, if not all, nonneuronal

carcinomas.

The mechanism(s) underlying NGF-mediated nonneuronal

tumor growth hypothesized is(are) not well understood, although

the product of the NGF inducible-A (NGFI-A) gene in rat

prostate cells is found to prompt transcription of the Rb gene

(58), the protein product of which is crucial for Gl-S phase

transition of the cell cycle that eventually results in cell division

and proliferation (59). Finally, it is noteworthy that NGF-

stimulated expression of proteolytic enzymes, such as endo-f5-

glucuronidase heparanase, which are important for basement

membrane penetration, in TrkA-negative/p75NTR-positive

neuroectodermal melanoma cells (60,61), suggesting a further

role for NGF in tumor invasion and metastasis.

ACKNOWLEDGMENTS

I wish to thank Drs Eiichi Shibay ama, Mikita Morita, and Shiny a

Mikami for their contributions to the present studies. I am also

grateful to Professors Toshiyuki Uchikoshi and Mamoru -

Tadokoro for constant support and encouragement. The studies

carried out in our laboratory were supported in part by Grant-in-

Aid for Scientific Research from the Ministry of Education,

Science and Culture, Japan.

Table 2. (legend)

hnmunohistochemical detection ot TrkA in paraffin sections of human tumors. Counterpart normal cells of each tumor and their

status of TrkA expression are shown in parentheses following the organ names (Ref 15) (Y, yes; N, no), ep, epithelium. Well, well

differentiated type; moderate, moderately differentiated type; poor, poorly differentiated type; n, number of specimens. Staining

results were scored according to the following scale: -, no staining; +, low percentage (<50%) of positively stained tumor cells;

++, at least 50% of the tumor cells stained positive. *TrkA was present in the parietal cells but absent from the chief cells,

foveolar/neck cells and pyloric glands. 'TrkA was present in the intercalated and intralobular ducts but absent from the

interiobular and main pancreatic ducts.

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