Human eosinophils express transforming growth factor alpha.

Transforming growth factor alpha (TGF-alpha) is a pleuripotential cytokine with diverse biological effects, including the ability to influence the proliferation of normal cells or neoplastic epithelial cells. Eosinophils are a subset of granulocytes that normally enter the peripheral tissues, particularly those beneath gastrointestinal, respiratory, and urogenital epithelium, where they reside in close proximity to the epithelial elements. In this study, we demonstrate that the great majority of eosinophils infiltrating the interstitial tissues adjacent to two colonic adenocarcinomas and two oral squamous cell carcinomas labeled specifically by in situ hybridization with a 35S-riboprobe for human TGF-alpha (hTGF-alpha). No other identifiable leukocytes in these lesions contained detectable hTGF-alpha mRNA. We also examined leukocytes purified from a patient with the idiopathic hypereosinophilic syndrome. 80% of these eosinophils, but none of the patient's neutrophils or mononuclear cells, were positive for hTGF-alpha mRNA by in situ hybridization, and 55% of these eosinophils were positive by immunohistochemistry with a monoclonal antibody directed against the COOH terminus of the mature hTGF-alpha peptide. Finally, the identification of the purified eosinophil-associated transcript as hTGF-alpha was confirmed by polymerase chain reaction product restriction enzyme analysis followed by Southern blot hybridization. In contrast to eosinophils from the patient with hypereosinophilic syndrome, the peripheral blood eosinophils from only two of seven normal donors had detectable TGF-alpha mRNA and none of these eosinophils contained immunohistochemically detectable TGF-alpha product. Taken together, these findings establish that human eosinophils can express TGF-alpha, but suggest that the expression of TGF-alpha by eosinophils may be under microenvironmental regulation. Demonstration of TGF-alpha production by tissue-infiltrating eosinophils and the eosinophils in the hypereosinophilic syndrome identifies a novel mechanism by which eosinophils might contribute to physiological, immunological, and pathological responses.

Eosinophils, tissue eosinophilia, and eosinophil-derived transforming growth factor alpha in hamster oral carcinogenesis.

Eosinophilia in tissues and/or circulating blood is known to be associated with a wide variety of malignancies but the role of the eosinophil in neoplastic conditions is not known. Using the cheek pouch of the Syrian hamster as an experimental model for oral carcinogenesis, it has recently been shown that eosinophils at sites of developing oral cancer express the multifunctional cytokine, transforming growth factor alpha (TGF-alpha). This study investigated the time course of eosinophil infiltration, tissue eosinophilia associated with malignant epithelium, and eosinophil-derived TGF-alpha mRNA during the 16-week 7,12-dimethylbenz[a]anthracene (DMBA)-induced oral cancer development process. The results reveal that the occasional eosinophil is normally present in the lamina propria of hamster oral mucosa. With progressive DMBA treatments, there is an increase of eosinophils infiltrating into the lamina propria. By weeks 12-16, the number of eosinophils is significantly higher in DMBA-treated pouches than in control pouches treated with the vehicle mineral oil alone. Analysis of the infiltrating eosinophils into fully developed hamster oral carcinomas reveals that tissue eosinophilia is associated with 78% of the stromal areas associated with malignant epithelium, while only 7% of sites associated with non-tumor oral epithelium (normal, hyperplastic-dysplastic) exhibited eosinophilia. Furthermore, the majority of the eosinophils associated with malignant epithelium were found to contain TGF-alpha mRNA. The number of TGF-alpha mRNAs containing eosinophils associated with malignant oral epithelium is significantly higher than that associated with nonmalignant oral epithelium. Together, these results suggest that eosinophils are recruited to tumor-developing sites, that they predominantly associate with malignant epithelium, and that most tumor-associated eosinophils express the cytokine TGF-alpha.

Use of intracellular H3 messenger RNA as a marker to determine the proliferation pattern of normal and 7,12-dimethylbenz[a]anthracene-transformed hamster oral epithelium.

One of the major goals in cancer research and diagnosis is to identify in a tissue the population of actively dividing cells and their pattern of growth and to differentiate the proliferation patterns of normal and transformed tissues. We now describe a method for determining the proliferation pattern of any tissue (normal, diseased, or transformed), applicable in any mammalian species. This method is based on the fact that the transcription of histone H3 gene in mammalian cells is tightly coupled to DNA synthesis during cellular division. Resting cells or cells that just exited the cell cycle will have no detectable H3 mRNA. The presence of H3 mRNA in a cell is thus a good indicator of its proliferation status. We carried out in situ hybridization of H3 mRNA in hamster oral epithelia exhibiting a variety of altered growth patterns as a consequence of exposure to the chemical carcinogen, 7,12-dimethylbenz[a]anthracene to demonstrate the usefulness of this technique. This application does not require in vitro manipulation of tissues nor does it require the prior administration of a tracer. The proliferation pattern at a single moment in time instead of an accumulated pattern over a period of time is produced. Finally, since the technique of in situ hybridization can be applied to archival tissues, retrospective studies can be done. This application should find usefulness in a wide variety of experimental research settings, particularly cancer research.

Transforming growth factor alpha in chemically transformed hamster oral keratinocytes.

The cheek pouch of the Syrian hamster is an excellent tissue for the experimental induction of oral cancer by carcinogenic chemicals. Lysate prepared from a cell line (HCPC-1) derived from one of these hamster oral tumors greatly increased the growth of these oral tumor cells in vitro. We now show that the mitogenic substance, transforming growth factor alpha (TGF-alpha), is present in all of the chemically transformed hamster oral tumors examined (in vitro and in vivo). In no adult normal tissue of the Syrian hamster can we detect expression of TGF-alpha. TGF-alpha could be partly or wholly responsible for the mitogenic activity detected in the lysate of the chemically transformed hamster oral keratinocytes. Both normal and chemically transformed hamster oral keratinocytes express the receptor to epidermal growth factor. The consistent detection of TGF-alpha and epidermal growth factor receptor mRNAs in these hamster oral tumor cells suggests that an autocrine growth mechanism might be operative. This hamster cheek pouch oral cancer model can be used for the molecular analysis of how TGF-alpha and epidermal growth factor receptor might be involved in the malignant transformation of epithelial tissues.

Detection of transforming growth factor-alpha messenger RNA in normal and chemically transformed hamster oral epithelium by in situ hybridization.

We have recently demonstrated the consistent detection of transforming growth factor alpha (TGF-alpha) in chemically transformed hamster oral tumors. By Northern blot analysis, no TGF-alpha mRNA can be detected in normal cheek pouch mucosa. The consistent expression of TGF-alpha associated with the malignant transformation in the well-defined hamster oral cancer model prompted us to hypothesize that the aberrant expression of this important cellular gene could be related to a specific stage of epithelial alteration. In situ hybridization was used to test this hypothesis. We now report that by in situ hybridization we can detect TGF-alpha mRNA in normal hamster oral epithelium and also at all stages of transformation. In all epithelium, labeling of TGF-alpha mRNA in the basal layer is more pronounced than that observed in the spinous and squamous layers. There is a significant increase of TGF-alpha mRNA labeling early in 7,12-dimethylbenz(a)anathracene-induced oral carcinogenesis. This increase is associated with morphological changes of epithelial hyperplasia or dysplasia. Although lesions exhibiting full-thickness epithelial dysplasia (carcinoma in situ) showed more labeling of TGF-alpha mRNA than do areas of lesser dysplasia, the transition to full-fledged papillary or invasive squamous cell carcinoma is not associated with further elevations of TGF-alpha expression.

Oral cancer in vivo gene expression profiling assisted by laser capture microdissection and microarray analysis.

Large scale gene expression profiling was carried out on laser capture microdissected (LCM) tumor and normal oral epithelial cells and analysed on high-density oligonucleotide microarrays. About 600 genes were found to be oral cancer associated. These oral cancer associated genes include oncogenes, tumor suppressors, transcription factors, xenobiotic enzymes, metastatic proteins, differentiation markers, and genes that have not been implicated in oral cancer. The database created provides a verifiable global profile of gene expression during oral carcinogenesis, revealing the potential role of known genes as well as genes that have not been previously implicated in oral cancer.

Cellular sources of transforming growth factor-alpha in human oral cancer.

Aberrant expression of TGF-alpha is associated with human malignant oral epithelium. Experiments were initiated to determine the cellular sources of transforming growth factor-alpha (TGF-alpha) in human oral cancer. Ten freshly resected human oral cancers and four specimens of normal human oral epithelium were studied by in situ hybridization and immunohistochemistry. Tissues were probed with 35S-labeled sense and antisense riboprobes to (i) human TGF-alpha (hTGF-alpha), (ii) human epidermal growth factor receptor (EGFR) to determine the distribution of TGF-alpha responsive cells, and (iii) histone H3 to examine TGF-alpha and/or EGFR's possible contribution to altered proliferation in transformed epithelium. Results of our experiments showed that TGF-alpha mRNA could be detected in normal and transformed human oral epithelium. More surprising, we have identified the major source of TGF-alpha mRNA to be the infiltrating eosinophils. A monoclonal antibody to the mature human TGF-alpha peptide stained similar areas in normal and malignant specimens. Eosinophils associated with tumors exhibited positive cytoplasmic immunostaining for TGF-alpha protein. Labeling of EGFR mRNA in human oral epithelium demonstrated uniform labeling of basal layers in normal, hyperplastic, and mildly dysplastic epithelium. In severely dysplastic epithelium and carcinomas (particularly moderate to poorly differentiated types), cellular levels of EGFR mRNA were significantly higher. The profile of altered cellular levels of EGFR mRNA correlated well with the profile of altered proliferation as indicated by H3 mRNA labeling. We hypothesize that the overproduction of EGFR mRNA in tumor epithelium--together with the localized delivery of high amounts of TGF-alpha by eosinophils at tumor-developing sites--is responsible for the increased proliferation of the tumor epithelium.

Eosinophil ablation and tumor development.

Tissue eosinophilia in squamous cell carcinoma has long been recognized; however, the role of eosinophils in tumor development remains unclear. Studies have reported both favorable and unfavorable prognoses for patients with tumors exhibiting tumor-associated tissue eosinophilia (TATE). This study seeks to elucidate the potential role of the eosinophil in squamous cell carcinoma development and provide an experimental model for future studies. The carcinogen-induced hamster oral cancer model was found to fulfill these objectives. Eosinophils progressively infiltrate into this carcinogen-induced oral cancer model. We now demonstrate that TATE is completely abolished by the use of an anti-interleukin-5 monoclonal antibody (mAb) preparation, TRFK-5. Clinical observations revealed that TRFK-5-treated hamsters exhibited smaller tumor burden and delayed onset of tumor development. The results suggest that anti-interleukin-5 antibody treatment may delay and/or inhibit tumor development, and that eosinophils may have a tumor-promoting role.

Eosinophil-derived transforming growth factors (TGF-alpha and TGF-beta 1) in human periradicular lesions.

Inflammatory mediators of periradicular lesions are poorly understood. Transforming growth factors-alpha and -beta 1 (TGF-alpha and TGF-beta 1) have been linked with the cellular processes for both soft and hard tissue wound healing. The purpose of this study is to demonstrate the cellular sources of TGF-alpha and TGF-beta 1 mRNA and protein in periapical lesions by in situ hybridization and immunohistochemistry. Nine periapical granulomas and nine periapical cysts were examined. TGF-alpha mRNA and protein were not detectable in the granulomas examined. However, eosinophils surrounding the periapical cysts demonstrated both TGF-alpha mRNA and protein. The vast majority of eosinophils present in the periapical granulomas and cysts also demonstrated TGF-beta 1 mRNA and protein. Other cells producing TGF-beta 1 were lymphocytes, fibroblasts, and monocytes. The presence of wound repair cytokines, such as TGF-alpha and TGF-beta 1, suggests a mechanism by which the host inflammatory response may participate in the repair and remodeling of periapical tissues.

Maxillary odontogenic carcinoma with distant metastasis to axillary skin, brain, and lung: case report.

We present a case of odontogenic carcinoma with ghost-cell keratinization of the right maxilla, with a history of 23 years after initial treatment. Within this period, multiple local recurrence, as well as metastasis to axilla, brain, and lung, was noted. Several attempts at resection of the primary lesion were unsuccessful at achieving local control, even when supplemented with chemotherapy and radiotherapy. Metastatic tumors were studied histologically, and their morphology coincided with that of the primary tumor. The medical history of the patient and pathologic findings of the tumor are reviewed.

Intraoral liposarcoma: case report and review of the literature.

The purposes of this article are to present a case report of liposarcoma of the tongue and to review the existing literature regarding liposarcomas with intraoral locations.

Salivary gland changes in chronic fatigue syndrome: a case-controlled preliminary histologic study.

OBJECTIVE: The purpose of this preliminary study is to compare labial salivary gland changes of 11 patients with chronic fatigue syndrome with control subjects. STUDY DESIGN: Changes in labial salivary glands were graded from 0 to 3+ for acinar dilatation, ductal dilatation, periductal fibrosis, plasmacytic infiltrate, lymphocytic infiltrate, mast cell infiltrate, and lymphocytic aggregates or foci. RESULTS: Four of the 11 subjects had 2+ to 3+ changes in at least 4 of the 7 parameters examined. Only the presence of mast cells was statistically significant between the 2 groups. Two of these 4 patients had 1 lymphocytic focus per 4 mm(2) of tissue. CONCLUSIONS: The salivary gland changes in patients with chronic fatigue syndrome show varying degrees of ductal and acinar dilatation, periductal fibrosis, lymphoplasmacytic infiltrates, and occasional lymphocytic foci, all suggestive of primary gland damage. The one parameter that showed statistical significance was the presence of mast cells (Fisher exact test, 0.0125).

Lack of TGF-alpha and TGF-beta 1 synthesis by human eosinophils in chronic oral ulcers.

We recently demonstrated that eosinophils infiltrate prominently into cutaneous wounds in the Syrian hamster and represent a source of transforming growth factor-alpha and transforming growth factor-beta. In this study, we assessed the role of the eosinophil and eosinophil-derived transforming growth factors in human oral ulcers that exhibit delayed healing, descriptively termed traumatic ulcerative granuloma with stromal eosinophilia. Our aim was to determine whether eosinophils, which characteristically infiltrate traumatic ulcerative granuloma with stromal eosinophilia lesions, produced transforming growth factor-alpha or transforming growth factor-beta. Twelve cases of traumatic ulcerative granuloma with stromal eosinophilia were examined for transforming growth factor-alpha and transforming growth factor-beta mRNA and cellular protein by in situ hybridization and immunohistochemistry. Eosinophils in 92% of the cases did not express detectable cellular levels of mRNA for either of the transforming growth factors. In addition, only a small percentage of the many eosinophils infiltrating these lesions produced transforming growth factor-alpha or transforming growth factor-beta. The lack of significant synthesis of transforming growth factors by eosinophils in most of the cases of traumatic ulcerative granuloma with stromal eosinophilia is in striking contrast to the expression of transforming growth factors by the eosinophils that infiltrate the animal wound-healing model. Our findings may help to explain the delayed healing that is typical of TUGSE lesions.

Role of local secretory and motility changes in the pathogenesis of experimental duodenal ulcer.

Changes in gastric acid and pepsin secretions do not fully account for the duodenal ulcerogenic effect of cysteamine or propionitrile in the rat. We investigated the role of pancreatic and biliary secretions as well as motility changes in the duodenum. Bypass of bile to the jejunum and/or ablation of pancreatic secretion or drainage of these secretions through chronic duodenal fistula aggravated the cysteamine-induced duodenal ulcers. Biliary bypass to the proximal duodenum, on the other hand, decreased the incidence and intensity of experimental duodenal ulcers. An increased and dopamine-sensitive myoelectric activity caused by cysteamine or propionitrile was recorded in the proximal duodenum, indicating a state of hypermotility. Indeed, a decreased quantity of bilirubin was recovered through the chronic fistula in the proximal duodenum, suggesting an impaired delivery of bile to the ulcer area after cysteamine administration. Thus, duodenal hypermotility probably prevents the proper mix and neutralization of gastric acid and duodenal (mucosal, biliary and pancreatic) secretions, predisposing to ulceration in the proximal duodenum.

Oral mucous membrane reactions to drugs and chemicals.

This article reviews recent reports of interest to practitioners concerned with the damaging effects of drugs and exogenous substances on the oral mucosa. Major topics include the incidence of the use of dentally important drugs in selected populations, lichenoid reactions in oral mucosa, diagnosis and management of mucositis related to cancer therapy, and issues relating to the diagnosis of oral foreign body reactions.

Secretory changes associated with chemically-induced duodenal ulceration: simultaneous measurements of acid, pepsin, base and pancreatic enzymes in rats with chronic gastric fistula.

Rats with chronic gastric fistula were used to study gastric and duodenal/pancreatic secretory changes evoked by chemical duodenal ulcerogens. Although the data demonstrate stimulation of gastric output of acid and pepsin by certain doses of cysteamine or propionitrile, cysteamine produced dose-response increases in acid secretion only during the 1st hour following administration. The base output was also elevated at most of the time intervals after cysteamine. Decreased alkaline secretion was only seen following propionitrile injection. Although hyperacidity at the ulcer site may occur during duodenal ulcerogenesis, these results suggest that direct stimulation of acid-pepsin secretion or depression of pancreatic alkaline output are not the primary mechanisms explaining the ulcerogenic action of cysteamine and propionitrile.

Direct measurement of duodenal acid-pepsin exposure at site of ulceration in rats.

A new technique for the preparation of a chronic duodenal fistula in the rat is described. The effect of cysteamine and propionitrile on the acidity of duodenal contents was studied, and the degree of acidity correlated with the respective duodenal ulcerogenicity of these two compounds. After subcutaneous administration of cysteamine, reduced duodenal acid was seen for 2-4 h, followed by large increases continuing up to 12 h and declining thereafter. Oral cysteamine administration produced acid increases of rapid onset and shorter duration, accompanied by increased pepsin activity. The weak duodenal ulcerogen propionitrile did not affect duodenal acidity. Cimetidine markedly reduced the duodenal acidification induced by cysteamine, whereas the dopamine agonist bromocriptine did not affect acid delivery to the duodenum but reduced the pepsin activity in the duodenal contents by 50%. The duodenal fistula rat provides a new model system for studies on the pathogenesis of experimental duodenal ulcer.

Cysteamine induces duodenal ulcer in the mouse.

A new model of duodenal ulcer disease has been developed in the mouse. The ulcers were produced after either oral or subcutaneous administration of cysteamine which has been shown to cause duodenal ulcer in the rat. Cysteamine induced duodenal ulcers in a time- and dose-dependent manner after oral administration. The new mouse model shares many similarities with both the rat model and human ulcer disease. Cysteamine caused a significant increase in gastric acidity and pepsin activity. The mouse can be protected against the cysteamine-induced duodenal ulcer by either the dopamine agonist lergotrile or histamine H2 receptor antagonist cimetidine. This new model of duodenal ulcer disease in the mouse may represent a simple and inexpensive way to screen for new antiulcerogenic drugs.

Biliary and pancreatic secretions influence experimental duodenal ulcer without affecting gastric secretion in the rat.

The effect of the absence of biliary and/or pancreatic secretions in the duodenum or the enhanced presence of bile at the proximal duodenum on the incidence, severity, number, and location of cysteamine-induced duodenal ulcers was investigated in the rat. Cysteamine produced ulcers on the anterior wall of the duodenum in 75% and on the posterior wall ("kissing ulcers") in 50% of the animals. Diversion of biliary and/or pancreatic secretions from the duodenum increased both the severity and the incidence of the posterior duodenal ulcers. Diversion of bile to the proximal duodenum, on the other hand, decreased the severity as well as the incidence of the anterior duodenal ulcers. Mortality in rats receiving cysteamine correlated with the severity of ulcers. Taurocholic acid at nontoxic doses given subcutaneously or orally to nonoperated rats and rats which had bile diverted to the proximal duodenum aggravated the cysteamine-caused duodenal ulcers. Neither proximal nor distal diversion of bile had a major effect on gastric secretion of acid and pepsin in normal or cysteamine-treated rats. We conclude that both bile and pancreatic secretions may directly influence the development of cysteamine-induced duodenal ulcers in the rat.