E2F1 has both oncogenic and tumor-suppressive properties in a transgenic model.

Using a transgenic mouse model expressing the E2F1 gene under the control of a keratin 5 (K5) promoter, we previously demonstrated that increased E2F1 activity can promote tumorigenesis by cooperating with either a v-Ha-ras transgene to induce benign skin papillomas or p53 deficiency to induce spontaneous skin carcinomas. We now report that as K5 E2F1 transgenic mice age, they are predisposed to develop spontaneous tumors in a variety of K5-expressing tissues, including the skin, vagina, forestomach, and odontogenic epithelium. On the other hand, K5 E2F1 transgenic mice are found to be resistant to skin tumor development following a two-stage carcinogenesis protocol. Additional experiments suggest that this tumor-suppressive effect of E2F1 occurs at the promotion stage and may involve the induction of apoptosis. These findings demonstrate that increased E2F1 activity can either promote or inhibit tumorigenesis, dependent upon the experimental context.

Deregulated expression of E2F1 induces hyperplasia and cooperates with ras in skin tumor development.

In cell culture studies, overexpression of the E2F1 transcription factor has been shown to stimulate proliferation, induce apoptosis, and cooperate with an activated ras gene to oncogenically transform primary rodent cells. To study the effect of increased E2F1 activity on epithelial growth and tumorigenesis in vivo, transgenic mice expressing E2F1 under the control of a keratin 5 (K5) promoter were generated. Expression of E2F1 in the epidermis results in hyperplasia but does not inhibit terminal differentiation. In a transgenic line expressing high levels of E2F1, mice have decreased hair growth likely as a result of aberrant apoptosis in developing hair follicles. Coexpression of a cyclin D1 transgene with E2F1 augments epidermal hyperplasia and further disrupts hair follicle development suggesting that hypophosphorylated Rb antagonizes the proliferative and apoptotic-promoting activities of E2F1. Finally, the E2F1 transgene is found to cooperate with a v-Ha-ras transgene to induce skin tumors in double transgenic animals. These findings confirm that many of the activities ascribed to E2F1 through in vitro studies can be reproduced in vivo and demonstrate for the first time that deregulated E2F activity can contribute to tumor development.

Azasterol inhibitors in yeast. Inhibition of the 24-methylene sterol delta24(28)-reductase and delta24-sterol methyltransferase of Saccharomyces cerevisiae by 23-azacholesterol.

The effects of 23-azacholesterol on sterol biosynthesis and growth of Saccharomyces cervisiae were examined. In the presence of 0.2, 0.5, and 1 micron 23-azacholesterol, aerobically-growing yeast produced a nearly constant amount of ergosta-5,7,22,24(28)-tetraenol (approx. 36% of total sterol) and slowly accumulated zymosterol with a concommitant decline in ergosterol synthesis. Growth and total sterol content of yeast cultures treated with 0.2-1 micron 23-azacholesterol were similar to that of the control culture. Yeast cultures treated with 5 and 10 micron 23-azacholesterol produced mostly zymosterol (58-61% of total sterol), while ergosta-5,7,22,24(28)-tetraenol production declined to less than 10% of total sterol. The observed changes in the distribution of sterols in treated cultures are consistent with inhibition of 24-methylene sterol 24(28)-sterol reductase (total inhibition at 1 micron 23-azacholesterol) and of 24-sterol methyltransferase (71% inhibition at 10 micron 23-azacholesterol). Yeast cultures treated with 10 micron 23-azacholesterol were found to contain 4,4-dimethylcholesta-8,14,24-trienol and 4alpha-methylcholesta-8,14,24-trienol, which were isolated and characterized for the first time.

Lip cancer in South Australia, 1977-1996.

Lip cancer (140 ICD-9, C00 ICD-10) is a form of oral cancer occurring at the junction between the oral cavity and the skin. Lip cancer has a distinctive global epidemiology that is notably different from cancer occurring at other intraoral sites. This study reviews and analyses the epidemiological data for lip cancer from the South Australian Central Cancer Registry between 1977 and 1996. During this 20-year period, 2716 cases of lip cancer (2095 male, 621 female) and 35 deaths from this disease (23 males, 12 females) were reported. The average age of diagnosis was 58.3 years in males and 66.0 years in females. Very high age-standardised incidence rates (over 15.0 per 100000 per annum in males and 4.0 per 100000 per annum in females) were found, giving the South Australian population amongst the highest incidence of lip cancer in the world. Also of considerable concern was the finding that, contrary to global trends, these rates showed a significant increase over the 20-year period in both sexes. Possible reasons for these findings are discussed.

Identification of ergosta-8,24(28)-dien-3 beta,6 alpha-diol in A delta 8 goes to delta 7 sterol isomerase-blocked yeast mutant.

In addition to the monohydroxysterols found in the delta 8 goes to delta 7 isomerase-blocked Saccharomyces cerevisiae mutant erg 2, a noval dihydroxysterol, ergosta-8,24(38)-dien-3 beta,6 alpha-diol, was isolated. This sterol accumulated to the extent of 2.1% of the total sterol fraction when this mutant was treated with 23-azacholesterol, a known inhibitor of the 24-methylene-sterol-24(28)-reductase.

Sterol composition ofNeurospora crassa.

The sterols extracted from freeze-dried, log-phase cultures ofNeurospora crassa were separated by thin layer and analyzed by capillary gas liquid chromatography and mass spectrometry. Ergosterol, episterol, and ergosta-7,22 dienol were the major sterols of the free sterol fraction. The major esterified sterols, which constituted 4.95% of the total sterol fraction, were ergosterol, episterol, and ergosta- 7,22,24(28)-trienol, with lesser quantities of 4α-methyl-ergosta-8,24(28)-dienol. With the exception of lanosterol, all sterols were alkylated at the C-24 position.

Does fetal gut obstruction cause hydramnios and growth retardation?

In order to study the effects of impaired fetal intestinal absorption of amniotic fluid, two series of neonates (551 from Liverpool and 172 from Rome) with different types of congenital gut obstruction were divided into two groups and compared. Group A consisted of patients with complete obstruction at or above the proximal jejunum (within 15 cm of the ligament of Treitz). Patients of group B had either incomplete obstruction at group A level or either incomplete or complete obstruction at a lower level. Maternal hydramnios and fetal growth retardation rates were found to be significantly higher in group A than in group B. Maternal hydramnios was associated with an increased prematurity rate (P less than .001). Fetal growth retardation was not related to the presence of additional anomalies. In group A growth retardation was more frequent in babies born after 37 weeks of gestation (P less than .05). No differences were found between the Liverpool and Rome series of patients. These findings suggest that fetal gut function not only contributes to the control of amniotic fluid volume but also, in the final stages of pregnancy, to normal fetal growth. Maternal hydramnios may be the cause of premature delivery of fetuses with upper gut obstructions.

Attachment to and phagocytosis of mineral by alveolar bone osteoclasts.

The mechanisms by which the osteoclast attaches to and resorbs bone are not fully understood. Morphologic techniques have primarily been used to examine these mechanisms, but many studies have been based on decalcified material. In this study, the attachment of the osteoclast to alveolar bone and its relationship to the mineral component during the active resorption associated with tooth eruption in the rat was examined using transmission electron microscopy and techniques designed to minimize demineralization effects during processing. Large conglomerates of bone mineral were found within both the ruffled border and the vacuoles adjacent to this area. These deposits were clearly more extensive than the isolated mineral fragments described in other sites by previous investigators. Examination of demineralized sections showed that collagen was largely absent within the ruffled border, and not present within vacuoles. These observations suggest that phagocytosis of bone mineral may supplement its extracellular dissolution in situations associated with rapid bone turnover, such as tooth eruption. Two types of clear zone attachment to mineral were also observed, permitting further speculation on the mechanism by which osteoclasts attach to and move along the bone surface.

Early responses by osteoclasts in vivo and dentinoclasts in vitro to corticosteroids.

The results of previous investigations examining the association between corticosteroids and bone resorption in vitro have been conflicting. Recent studies have shown that application of a triamcinolone-containing medicament to the dental pulp cavity inhibits dentinoclast-mediated tooth resorption in vivo and in vitro. The aims of this study were to quantitate early ultrastructural changes in osteoclasts in vivo in response to a single steroid dose, and to examine the attachment of dentinoclasts grown in steroid-supplemented cultures in vitro. Resorbing bone around erupting rat molars was examined using TEM and morphometrically quantitated for changes in ruffled border and clear zone areas 1 and 4 h after injections of NaCl, calcitonin, hydrocortisone and prednisolone. Dentinoclasts were harvested from resorption areas in mature rat molars, grown in cultures with calcitonin, hydrocortisone or prednisolone, and fixed and viewed with SEM after 30 min and 2 h. Results of TEM studies showed a significant reduction in areas of resorbing structures in osteoclasts treated with calcitonin at 1 and 4 h, and prednisolone at 4 h. At 1 and 4 h, hydrocortisone showed an increase in resorbing structure areas. SEM results indicated all experimental substances reduced dentinoclast spreading and attachment. It was concluded that the direct effect of steroids on clastic cells may be one of inhibition (the degree of which depends on the nature and dose of the steroid) whereas, in vivo, systemic administration may cause more secondary effects (such as PTH stimulation) to compete with this inhibition. The efficacy of steroid-containing medicaments in inhibiting dentinoclastic resorption may be due to their relatively localized area of application.

Oral cancer: role of the basement membrane in invasion.

Invasive growth of cancer cells is a complex process involving specific interactions between tumour cells and the orderly, integrated complexes of the extracellular matrix. Basement membranes have been proposed as one constituent of extracellular matrix which carries responsibility for regulating invasion and metastasis. Using a chemically induced rat tongue carcinoma model, it has been shown that components of the basement membrane and its overall structure are altered during tumour invasion, and methods have been developed to quantitate some of these differences. Since the basement membrane can be specifically characterized by its fibrous protein network of Type IV collagen and laminin, which is embedded in a heparan sulphate-rich proteoglycan matrix, these components have been targeted. In particular, the current paper presents results in the context of current concepts of early changes in neoplastic invasion of underlying connective tissues. In consequence, further elaboration of the underlying mechanisms of epithelial migration in oral cancer may allow an exploration of the use of alterations in expression of basement membrane components as prognostic indicators.

A retrospective analysis of oral hairy leukoplakia in South Australia.

BACKGROUND: The features of oral hairy leukoplakia (OHL) have been widely reported in the literature. However, no studies have described this lesion in the Australian setting. This study retrospectively examines, with respect to specific clinical factors, the prevalence of OHL in a South Australian HIV-infected population. METHODS: Clinical data were collected from the records of 197 HIV-infected patients who had attended the Adelaide Dental Hospital between January 1986 and February 1995. Data were analysed using the chi-square test. RESULTS: The prevalence of OHL in South Australian HIV-infected patients was 45.2 per cent. The study found the presence of OHL was not related to CD4+ T-lymphocyte count or AIDS-defining illness nor did the length of time a patient had been infected with HIV relate to the presence of OHL. An association was observed between a reduced prevalence of OHL in patients who were taking antiviral medication. CONCLUSION: The prevalence of OHL in South Australia is comparable with results of other studies. This study supports the notion that OHL is not an indicator of immunosuppression in South Australian HIV-infected patients. Further longitudinal studies are required to ascertain the relationship of OHL to HIV disease progression.

The localisation of ED1 antibody in the resorbing hard tissues of the periodontium.

A dental hard tissue resorptive model was used to determine the periodontal ligament (PDL) distribution of lysosomal membrane antibody ED1 to cells of the macrophage-phagocyte lineage. Immunolabel was identified in mononuclear cells around inflammatory sites in the PDL, while multinuclear cells were labelled in resorption bays present in both bone and dentine. As repair of the tissues occurred, the label became less obvious. The presence of strong ED1 label in alveolar bone marrow provided evidence supporting the haemopoietic origin of the similarly labelled PDL cells. Also, evidence confirming the current theory that multinucleated resorptive cells differentiate along a monocyte-macrophage pathway was provided. t was concluded that ED1 is a positive PDL marker for mononuclear and multinuclear cells involved in the inflammatory and resorptive processes.

Mer receptor tyrosine kinase inhibition impedes glioblastoma multiforme migration and alters cellular morphology.

Glioblastoma multiforme (GBM) is an aggressive brain tumor, fatal within 1 year from diagnosis in most patients despite intensive multimodality therapy. The migratory and microscopically invasive nature of GBM as well as its resistance to chemotherapy renders conventional therapies inadequate in its treatment. Although Mer receptor tyrosine kinase (RTK) inhibition has been shown to decrease the long-term survival and improve the chemosensitivity of GBM in vitro, its role in malignant cellular migration has not been previously evaluated. In this study, we report for the first time a role for Mer RTK in brain tumor migration and show that Mer inhibition profoundly impedes GBM migration and alters cellular morphology. Our data demonstrate that Mer RTK inhibition results in altered signaling through focal adhesion kinase (FAK) and RhoA GTPase and a transformation of cytoskeletal organization, suggesting both molecular and structural mechanisms for the abrogation of migration. We also describe a novel and translational method of Mer RTK inhibition using a newly developed monoclonal antibody, providing proof of principle for future evaluation of Mer-targeted translational therapies in the treatment of GBM. Previous findings implicating Mer signaling in glioblastoma survival and chemotherapy resistance coupled with our discovery of the role of Mer RTK in GBM cellular migration support the development of novel Mer-targeted therapies for this devastating disease.

Experimental basis for the management of dental resorption.

Current evidence indicates that the mechanisms by which the mineralized tissues, bone and dentin, are resorbed are similar. Osteoclasts and dentinoclasts are possibly indistinguishable cell types that both resorb mineralized substrata. The principles of treatment of both dental and osseous resorptions should therefore have much in common. This article discusses the basis of hard tissue resorption and explores the rationale for experimental approaches towards the management of dental resorption.

Evidence for capping of Fc gamma receptors on osteoclasts.

Fc receptors, cell surface structures which bind the Fc portion of immunoglobulins, facilitate endocytosis and mediate triggered enzyme release in the phagocytic cells of the mononuclear phagocyte system (MPS). The osteoclast and the macrophage share many similarities, and it has been suggested that they are both derived from a common MPS precursor cell. Nevertheless, Fc gamma receptors, found on both primitive and well-differentiated cells of the mononuclear phagocyte system, have yet to be demonstrated on the osteoclast cell membrane. The aim of the present study was first, to study the distribution of rat endogenous IgG around resorbing bone using immunohistochemistry and the avidin-biotin-peroxidase (ABC) technique; and second, to determine if rat osteoclasts express Fc gamma receptors, using a recently described technique based on the ABC method. Observations that endogenous IgG accumulated at osteoclast/bone interfaces, and that Fc gamma receptors, expressed by osteoclasts, "capped" in the same area, have important implications regarding the origin of this cell and its mechanisms of bone attachment and resorption.

Vulvovaginitis: causes and management.

Over a period of 33 months in a paediatric accident and emergency department, the clinical pattern and possible causes of vulvovaginitis were studied prospectively in 200 girls presenting with genital discharge, irritation, pain, or redness. The major causes were poor hygiene and threadworms. The suspicion of sexual abuse arose in a few girls but no organisms of sexually transmitted disease were found. Urinary symptoms were common but only 20 patients had a significant bacteriuria and 40 had sterile pyuria. Specific skin problems occurred in 28 cases. Simple measures to improve hygiene and treatment of threadworms gave effective relief. Genital irritation caused urinary symptoms with no clinical evidence of infection, and it is advised that antibiotic treatment should await urine culture. Specific skin problems require help from a dermatologist. The possibility of sexual abuse must be considered especially if the vulvovaginitis is persistent or recurrent after adequate treatment.

Hypothermic insult to the periodontium: a model for the study of aseptic tooth resorption.

The aim of the current investigation was to define an animal model for the study of hard tissue resorption by examining the responses of the periodontal ligament (PDL) to both single and multiple episodes of hypothermic injury to the crowns of rat teeth. A group of 12 male rats weighing 200-250 g were anesthetized, and pellets of dry ice (CO2) were applied once to the crowns of the right first maxillary molars for continuous periods of 10 or 20 min. Animals were sacrificed at 2, 7, 14 and 28 days and tissues were processed for routine histological examination. A second group of eight animals and a third group of 12 animals were subjected to three applications of dry ice over a period of 1 week and sacrificed at 2 and 14 days respectively after the final application. In addition to thermal insult, the periodontium of teeth from a fourth group of six rats was subjected to mechanical trauma. Examination of the sections from the group undergoing a single freezing episode revealed that, by 1 week, shallow resorption lacunae had appeared on the root surface. These became more extensive after 14 days. At the same time hyaline degeneration was evident in the PDL. Within this group, teeth subjected to the longer 20-min application times generally showed more extensive injuries. By 28 days, evidence of repair was observed with reparative cementum beginning to line the resorption lacunae in the root dentin. Sections from animals subjected to multiple episodes of thermal trauma and those subjected to additional mechanical insult showed more extensive external root resorption than those from single-injury animals. It was concluded that low temperature stimuli applied to the crowns of rat molars were capable of eliciting a sterile degenerative response in the PDL which, in turn, resulted in external root resorption. Furthermore, the degree of this tissue injury was commensurate with the duration and number of exposures to the trauma. The results also indicated that progression of the resorptive process required periodic exposure to the injury, in the absence of which repair to the damaged root occurred.

Osteoclasts: structure and function.

Osteoclasts are multinucleated giant cells showing specialized membrane structures, clear zones and ruffled borders, which are responsible for the process of bone resorption. These cells arrive at the resorption site via the bloodstream as mononuclear cells, derived from haemopoietic precursors in the spleen or bone marrow, which fuse prior to resorption. The osteoclast may share an early progenitor cell, the granulocyte macrophage colony-forming unit (GM-CFU) with monocytes, macrophages and granulocytes, implying that osteoclasts share the pluripotent haemopoietic stem cell with all other haemopoietic cells. In the past, elucidation of the structure of these cells relied upon traditional ultrastructural techniques. Transmission electron microscopic studies revealed details of the unique ultrastructure of these cells and, in combination with stereological techniques, showed the response of cells to various hormonal stimuli. Scanning electron microscopy not only demonstrated the surface appearance of osteoclasts, and their predilection for spreading on various substratum components, but has also been used as an adjunct in resorption assays in which areas of resorption lacunae are measured as indicators of cell activity. Recent advances in fields such as immunocytochemistry and freeze fracture techniques have contributed towards a more detailed delineation of antigenic profile, cytoskeletal structure and localization of enzymatic pathways. The osteoclast is subject to extensive regulatory mechanisms and it has been established that the osteoblast plays a major rôle in mediating the effects of osteotropic hormones and local mediators on these cells. Hence, research aimed at elucidating the coupling mechanisms between these two cells may result in new therapies for bone disease.

IgE in postsecretory ameloblasts suggesting a hypersensitivity reaction at tooth eruption.

The clinical symptoms associated with the eruption of primary teeth resemble a mild hypersensitivity reaction. Light microscopic examination showed that IgE accumulated in postsecretory ameloblasts. The formation of IgE was elicited by enamel matrix proteins, which are chemotactic for mast cells.

Palatal changes associated with reverse smoking in Filipino women.

OBJECTIVE: To determine baseline data for the presence or absence of reverse-smoking and conventional smoking associated oral palatal mucosal changes in women. DESIGN: A cross-sectional evaluation of the clinical and cytological changes associated with the condition. SETTING: Nine rural barangays in Cabanatuan City, Philippines. SUBJECTS AND METHODS: Nine-one volunteer women smokers (61 reverse and 30 conventional) were examined clinically and photographically. Smears were also taken from three areas and the palate to investigate the cytology of palatal mucosal epithelium. MAIN OUTCOME MEASURES: Variations in colour, texture and topography of the palatal mucosa; determination of epithelial cell characteristics and inflammatory cell populations present in the lesions. RESULTS: Clinical findings showed that subjects could be grouped into three categories: Group A subjects showed pigmentation and some erythema only; Group B subjects included those with ulceration, marked erythema and non-descript mucosal roughening; Group C subjects (comprising the majority of reverse smokers) exhibited various combination of leukoplakia, fissuring, thickening and pigmentation of the palatal mucosa. Additional features, including nodularity, erythema, prominence and reddening of minor salivary gland duct openings were also occasionally observed in this group. Cytologic analysis revealed that, within each smoking group, there was a significant site-dependent difference in the predominant epithelial cell type present. CONCLUSIONS: This study reports the first systematic description of reverse smoking associated palatal mucosal changes in Filipino women. It also provides a basis for classification of the palatal mucosal changes among reverse smokers.