The influence of transforming growth factor-beta1 gene polymorphisms on the severity of gingival overgrowth associated with concomitant use of cyclosporin A and a calcium channel blocker.

BACKGROUND: The purpose of this study was to determine whether the prevalence and severity of gingival overgrowth in renal transplant recipients concomitantly treated with cyclosporin and a calcium channel blocker was associated with functional polymorphisms within the signal sequence of the transforming growth factor-(TGF)beta1 gene. METHODS: The extent and severity of gingival overgrowth for 164 renal transplant recipients immunosuppressed with cyclosporin A and concomitantly taking a calcium channel blocker since transplant were entered into the study (86 in Manchester, 78 in Belfast). Two biallelic polymorphisms of the TGF-beta1 gene were studied at position +869, codon 10 (leucine to proline substitution), and position +915, codon 25 (arginine to proline substitution). RESULTS: Subjects who were homozygous for proline at codon 10 had significantly higher overgrowth scores than those who were heterozygous (P= 0.03) or homozygous for leucine (P= 0.01). Subjects who were heterozygous (arginine/proline) at codon 25 had a significantly higher (P= 0.04) gingival overgrowth score than those who were homozygous for arginine. Logistic regression analysis indicated that for codon 25 independent predictors of severe gingival overgrowth were the heterozygous arginine/proline genotype (P= 0.009) and whether the individual was young (P= 0.05). CONCLUSIONS: Polymorphisms in the TGF-beta1 gene influence the expression of gingival overgrowth in renal transplant recipients concomitantly treated with cyclosporin and a calcium channel blocker. The polymorphism in the TGF-beta1 gene at codon 25 represented an independent genetic determinant of severe gingival overgrowth in the susceptible subjects studied.

Bcl-2 expression in sequential biopsies of potentially malignant oral mucosal lesions assessed by immunocytochemistry.

OBJECTIVE: To examine, for the first time Bcl-2 expression in sequential (autogenous) oral mucosal biopsies taken from the same sites in a gender, risk-factor matched, Caucasoid sample, over a 21-year period. DESIGN: Retrospective immunocytochemical longitudinal study of archival serial biopsies. MATERIALS AND METHODS: Computer records were used to identify biopsy specimens derived from 12 patients. These were divided into four groups: (1) Histologically innocuous lesions which remained histologically innocuous. (2) Dysplastic lesions which remained dysplastic. (3) Histologically innocuous lesions which later progressed to squamous cell carcinoma (SCC). (4) Dysplastic lesions which later progressed to SCC. This represented 65 biopsies in total. Bcl-2 expression was studied using mouse antihuman BCL-2 oncoprotein clone 124 (Dako, Denmark). RESULTS: Generally, there was a lack of Bcl-2 immunoreactivity in the epithelium, with one exception in dysplastic epithelium from a group (3) patient. CONCLUSION: These findings suggest that in our series, Bcl-2 is not expressed early in oral premalignant lesions and appears to contradict previous reports. Possible explanations for this disparity are considered.

The calcium channel blocker used with cyclosporin has an effect on gingival overgrowth.

BACKGROUND/AIMS: To investigate whether the choice of calcium channel blocker, used in conjunction with cyclosporin A, affected the prevalence of gingival overgrowth. METHOD: A cohort of 135 renal transplant recipients who had been medicated with cyclosporin A in combination with either nifedipine (89) or amlodipine (46) since transplant, took part in the study. The inclusion criteria were that eligible subjects had been in receipt of a kidney transplant for at least 12 months, had at least 10 teeth and had not received specialist periodontal treatment. The age, gender, current drug regimen and dosage were recorded for each participant and alginate impressions taken of both arches. The presence and severity of gingival overgrowth were scored from plaster models. RESULTS: A higher proportion (72%) of the amlodipine group were categorised as having gingival overgrowth compared with only 53% of the nifedipine group, chi square=4.5, p<0.05. Logistic regression analysis was used to explore the relationship between the presence or absence of gingival overgrowth (dependent variable) and age, gender, time since transplant, dose of cyclosporin A, centre in which the patient was treated, and the calcium channel blocker used (independent variables). Independent predictors of gingival overgrowth in this multivariate analysis were whether the individual was treated with amlodipine or nifedipine (p=0.01) and whether the individual was young or old (p=0.01). Within the multivariate analysis, the odds ratio for amlodipine to be associated with gingival overgrowth compared with nifedipine was 3.0 (confidence interval 1.3-6.9). CONCLUSIONS: The prevalence of gingival overgrowth in renal transplant recipients maintained on cyclosporin A and nifedipine is lower than those treated with cyclosporin A and amlodipine.

Reduction in gingival overgrowth associated with conversion from cyclosporin A to tacrolimus.

BACKGROUND: Unsightly gingival overgrowth affects many individuals immunosuppressed with cyclosporin A (CsA). Current management involves repeated periodontal surgery and intensive hygienist support. Tacrolimus is an effective alternative immunosuppressive agent for renal transplantation which does not appear to produce gingival enlargement. AIMS: The purpose of the present study was to monitor the gingival response of 4 renal transplant patients (RTPs), with clinically significant CsA-induced gingival overgrowth, after their immunosuppressive therapy was switched to tacrolimus. METHODS: Intra-oral photographs and alginate impressions were taken both prior to the drug conversion and again, 6 to 9 months later. Gingival overgrowth scores were determined, from plaster models on both these occasions. RESULTS: All of the RTPs experienced significant resolution of their gingival enlargement within the time period studied; however, only one had complete regression. CONCLUSION: It is concluded that conversion of RTPs with gingival overgrowth from CsA to tacrolimus may provide an effective management strategy for this clinical problem.

Signal recognition particle receptor (SRPR) is downregulated in a rat model of cyclosporin A-induced gingival overgrowth.

Differential display is a powerful technique which can be used to identify those genes whose expression is altered between two or more tissues under investigation. We have applied differential display to a rat model of cyclosporin A-induced gingival overgrowth (CIGO) to identify genes which are differentially expressed as a result of drug treatment. Ten weanling Wistar rats were fed with a pelleted diet containing cyclosporin A (CsA) at 120 mg/kg for 10 d and then 200 mg/kg for a further 30 d prior to culling. Experimental rats were compared with 10 age/sex-matched rats on a control diet. Significant evidence of overgrowth was observed in the interdental papilla between the mandibular first and second molar teeth in the CsA group. Differential display was performed on total cellular RNA extracted from the mandibular buccal gingiva. A cDNA product was isolated which was underexpressed in the overgrowth tissue and demonstrated a 95% sequence homology to the human signal recognition particle receptor (Human Docking Protein). Preliminary studies indicate that this gene is also underexpressed in human CIGO tissue. The method of approach and the potential implications of our findings are discussed.

Cyclosporin A regulates interleukin-1beta and interleukin-6 expression in gingiva: implications for gingival overgrowth.

BACKGROUND: Gingival overgrowth is a common side effect following the administration of cyclosporin A (CsA); however, the cellular mechanisms remain poorly understood. CsA's immunosuppressant properties involve the regulation of synthesis and cellular response to cytokines. A CsA-induced alteration in the cytokine profile within gingival tissue could provide a mechanism for gingival hyperplasia. The aim of this study was to investigate the effects of CsA on the production of 2 cytokines - interleukin-1beta (IL-1beta) and interleukin-6 (IL-6) - by both gingival fibroblasts and peripheral blood mononuclear cells (PBMC). METHODS: Cells were stimulated for 24 hours in the presence of CsA over a concentration range of 100 to 2,000 ng/ml and the resultant cytokine production determined by ELISA. In addition, levels of both cytokines within normal, inflamed, and overgrown gingival tissue were determined. RESULTS: CsA inhibited IL-6 production by gingival fibroblasts in a dose-dependent manner. In contrast, at a concentration of 2,000 ng/ml, CsA stimulated IL-6 production by PBMC (P <0.05). Fibroblasts derived from overgrown gingiva produced significantly higher levels of IL-6 than their normal counterparts (P <0.05). CsA inhibited IL-1beta production by PBMC over the whole concentration range (P <0.05). IL-1beta was not found in measurable quantities in any of the fibroblast cultures. Levels of IL-6 extracted from overgrown gingival tissue were significantly higher than in inflamed or normal tissue. In contrast IL-1beta levels in overgrown tissue were not statistically significantly greater than those in inflamed tissue. CONCLUSIONS: These results show that CsA does regulate cytokine expression in gingival tissue. This effect may play an important role in the pathogenesis of CsA-induced gingival overgrowth.

Cyclosporin associated gingival overgrowth in renal transplant recipients.

OBJECTIVES: To investigate the prevalence and severity of gingival overgrowth in a group of renal transplant recipients treated in one centre in Northern Ireland. STUDY DESIGN: A consecutive group of patients who had had a renal transplant for at least 6 months and were attending the Renal Unit in Belfast City Hospital took part in the study. These were divided into a group of 84 subjects treated with cyclosporin since their transplant who were compared with a control group of 36 transplant recipients who had never received cyclosporin. Each subject had a periodontal examination and completed a questionnaire. The severity of gingival overgrowth was scored from plaster models. OUTCOME MEASURES: Clinically significant gingival overgrowth was equated with a score of > or = 30 using the index developed by Seymour et al (1985). RESULTS: 41 (49%) of the cyclosporin group had clinically significant gingival overgrowth compared with none of the controls. A significantly higher proportion of males had overgrowth than females. There were significant correlations between age at transplant, plaque, bleeding, pocketing and the severity of gingival overgrowth. Many patients with clinically significant gingival overgrowth were apparently unconcerned about this condition. CONCLUSIONS: It is concluded that gingival overgrowth is a significant problem for renal transplant patients treated with cyclosporin, particularly if they are also treated with a calcium channel blocker. None of the factors measured, in isolation, explained the variable expression of gingival overgrowth in those at risk.

A comparison of the management of potentially malignant oral mucosal lesions by oral medicine practitioners and oral & maxillofacial surgeons in the UK.

This study describes the results of a survey undertaken to assess the management of potentially malignant oral mucosal lesions by oral medicine practitioners and compares their approach with that of oral & maxillofacial surgeons that we have previously described. Significant differences were noted between the two groups in the use of photography to document the lesions and in the use of certain special investigations, which included measurement of serum iron, serum ferritin, serum Vit B12, red cell folate and candidal isolation. The groups also varied in the perceived importance of the age of the patient and anatomical site of the lesion when deciding on the need for further biopsy. There was also significant variation in the use of certain treatment modalities, including excising non-dysplastic and severely dysplastic/carcinoma in-situ lesions and eliminating trauma when treating mild/moderately dysplastic and severely dysplastic/carcinoma in-situ lesions. Significant differences in the frequency and duration of follow-up were noted for non-dysplastic lesions. Finally, the two groups differed significantly when asked to rank the perceived importance of certain factors (the histopathology of the most recent biopsy and the anatomical site of the lesion) when deciding the need to follow-up. Possible reasons for the variation are discussed.

The teaching of molecular biology to undergraduate dental students within the UK.

This work was undertaken to assess the extent and nature of molecular biology teaching to undergraduate dental students in the UK. We surveyed the lecturers of those schools involved in teaching molecular biology and a questionnaire and covering letter were circulated to all 14 UK Dental Schools.

Recrudescent herpes simplex infection mimicking primary herpetic gingivostomatitis.

We report six cases of recrudescent intraoral herpes simplex infection clinically indistinguishable from primary herpetic gingivostomatitis. All infections occurred in healthy children or young adults. Serological analysis demonstrated herpes simplex virus (HSV)-specific IgG at initial presentation, indicating that the infection was not a primary infection. Convalescent sera exhibited HSV-specific IgM and a rising HSV-specific IgG titre. These findings demonstrate that the initial clinical diagnosis of primary herpetic gingivostomatitis was erroneous and that what was actually being observed was widespread recrudescent intraoral herpes simplex infection.

Expression of the apoptosis-suppressing gene BCL-2 in pheochromocytoma is associated with the expression of C-MYC.

It has become increasingly clear that deregulation of programmed cell death is a critical component in multistep tumorigenesis. Previous studies have demonstrated a high frequency of Bcl-2 expression in tumors arising from cells derived from the neural crest and in tumor cell lines of neural origin. The present investigation was undertaken to determine whether similar molecular events occur in human pheochromocytoma. With the aim of determining the potential role of apoptosis in the pathogenesis of this tumor, we assessed proto-oncogene Bcl-2 and c-myc protein products as well as Bcl-2 messenger RNA levels in a collection of such tumors. Western blot analysis revealed that such tumors expressed the 26 kDa Bcl-2 (5 of 8 cases) and the 64 kDa c-Myc (7 of 8 cases) proteins. Northern blot analysis detected the Bcl-2 transcripts in 6 of 8 tumors. Immunoperoxidase staining, using a monoclonal anti-Bcl-2 antibody, was positive in 18 (82%), including 5 malignant tumors, of the 22 specimens examined. This Bcl-2 immunoreactivity was seen in 14 of 18 (78%) sporadic tumors, including 2 that were extra-adrenal, and all familial tumors. Of the 22 tumor samples examined for c-Myc protein, 20 (91%) tumors were positive. Our results suggest that deregulation of programmed cell death may be a critical component in the multistep tumorigenesis of human pheochromocytoma. The genetic complementation of simultaneously deregulated Bcl-2 and c-myc may be implicated in this process.

Paraffin-embedded tissue as a source of RNA for gene expression analysis in oral malignancy.

OBJECTIVE: To assess the feasibility of using archival oral mucosal tissue to examine gene expression at the ribonucleic acid (RNA) level. MATERIALS AND METHODS: We describe the isolation of RNA from 8 microns sections of formalin-fixed paraffin-embedded oral mucosal tissue. RNA was reverse transcribed and three candidate genes amplified by polymerase chain reaction (PCR). The ribosomal protein S14 gene is a housekeeping gene which has been used as an internal standard in several quantitative PCR protocols. The thymidine kinase (TK) gene is expressed at low levels in most tissues and, with a well-documented genomic organisation, is a useful tool for discrimination between genomic DNA and cDNA. The RI alpha gene is reported to be overexpressed in many cancer cell lines, in malignant tissue and in vitro transformed cells. RESULTS: The S14 gene, the TK gene and the RI alpha gene of the cAMP-dependent protein kinase (PKA) were amplified successfully from formalin-fixed paraffin-embedded tissue sections. The TK primer pair is a useful additional tool in the unambiguous identification of RNA-derived species. CONCLUSION: RNA suitable for reverse transcribed (RT)-PCR was extracted from archival oral mucosal tissue. This should permit rapid sequence analysis of transcribed tumor suppressor genes and oncogenes in this material. Furthermore, the RT-PCR approach described may allow quantification of gene expression in oral mucosal archival material processed in a standard fashion.

Management of potentially malignant oral mucosal lesions by consultant UK oral and maxillofacial surgeons.

This paper describes the results of a recent survey carried out under the auspices of the Professional Education and Evaluation Subgroup of the UK Working Group on Screening for Oral Cancer and Precancer. The aim of this survey was to assimilate information regarding currently used management options of potentially malignant oral lesions as a basis from which to rationalise our future approach to their management. The survey has confirmed that variation exists among oral and maxillofacial consultants in their approaches and a more formal approach to management may therefore be indicated.

Expression of human cytokeratin 14 in normal, premalignant and malignant oral tissue following isolation by plaque differential hybridisation.

Differences in gene transcription between RNA samples extracted from oral normal and squamous cell carcinoma (SCC) tissue were examined using the technique of cDNA library differential plaque screening. A differentially expressed transcript was selected on the basis of it being under-expressed in the cancer tissue and was identified, using DNA sequencing, as cytokeratin 14. The level of cytokeratin 14 transcription in RNA samples extracted from a range of oral SCC and normal tissue, as well as "white patch" lesions, was then investigated. Cytokeratin 14 appeared to be significantly under-expressed in oral cancer specimens studied compared to normal and white-patch tissue (P < 0.01). The trend for higher levels of cytokeratin 14 transcription in the dysplastic "white patch" samples compared to that observed for the malignant tissue (P < 0.05) suggests that the decrease in cytokeratin 14 transcription is a late event in the carcinogenic pathway.

Differential expression of protease inhibitor and small proline-rich protein genes between normal human oral tissue and odontogenic keratocysts.

The technique of differential hybridization was used to compare gene transcription between normal oral mucosa and odontogenic keratocyst lining. Protease inhibitors, elafin and stefin-B as well as beta-actin and two epithelial-specific small proline-rich (spr) proteins, which we have named SPRC and SPRK and which are distinct from salivary proline-rich proteins, were differentially expressed. Increased abundance of alpha I(I) collagen and elafin transcripts was demonstrated in the keratocyst, with decreased abundance of stefin B, SPRC and cytokeratins 4 and 13 transcripts compared to normal palatal mucosa. The deduced protein sequences of SPRC and SPRK were described and compared, and the relative abundance of their respective cDNAs in palatal and keratocyst libraries determined. Identification of factors controlling transcription of these genes could advance our understanding of the development of odontogenic keratocysts.